UNIVERSITY OF CALIFORNIA
SAN FRANCISCO LIBRARY

A I LABORATORY MANUAL

U-

OF

EXPERIMENTAL PHYSIOLOGY

(Including General Physiology)

BY

LOIS McPHEDRAN ERASER, M.A.

Lecturer in Physiology, University of Toronto

F. A. HARTMAN, PH.D.

Professor of Physiology, University of Buffalo

J. J. R. MACLEOD, M.B.

Professor of Physiology, University of Toronto

J. M. D/OLMSTED, PH.D.

Assistant Professor of Physiology, University of Toronto

PM-M

UNIVERSITY OF TORONTO PRESS

1922

COPYRIGHT, CANADA, 1919

BY
UNIVERSITY OF TORONTO PRESS

PREFACE.

Experimental physiology is now recognized as a fundamental
subject in the curriculum of the medical student and as one having
a most important place in the training of the student of biology.
In the medical course, the physiological laboratory serves as the
portal to the clinic; as the testing ground where the student may
try for himself in how far the known laws of physics and chemistry
can be successfully employed to explain the normal working of the
human machine. No more can theoretical study, or demonstra-
tion, by itself supply a correct understanding of the functions spf
the living body than could similar methods in training an engineer
to understand an engine. Attempts to rectify, by operations or
by drugs, functional derangement in the diseased animal without
a practical knowledge of the normal working of the various organs,
both isolated and as a whole must be as unjustifiable as attempting
to repair a complicated piece of machinery would be by any other
than a practical engineer.

In the training of the biologist, experimental physiology finds
its value because it teaches how to interpret the relationship be-
tween structure and function. For the advancement of physio-
logical knowledge it is essential that the functions of the lower
animals should be more intensively investigated by those who
have been trained in the methods of the experimental physiologist.
In the crowded curriculum, either of medicine or of biology, it
is impossible to find an amount of time that is sufficient for the
performance of more than a few of the fundamental experiments
in each of the main subdivisions of the subject of physiology.
It is the duty of the teacher, therefore, to select these experiments
with the greatest care so that the experience which the student
gains in their performance may guide him aright in building up
his knowledge of this subject. The experience gained in the labora-
tory is to serve as the framework upon which the detailed con-
struction is to be completed by fitting on to it in their proper re-
lationship to one another the other facts of physiological know-
ledge, acquired by theoretical study and demonstration.

3

4 EXPERIMENTAL PHYSIOLOGY.

While admitting these principles some have averred that the
technical difficulties of experimental physiology are such as to
make it impossible for the average medical student to secure a
sufficient number of results to justify the expenditure of time and
energy required in the laboratory. This is a fallacious criticism
for it assumes that every experiment to be of value should be
crowned by results that are technically flawless, and it fails to
recognize that the performance of the work, if carefully and in-
telligently done, affords that personal experience without which,
in a highly practical science like medicine, theoretical knowledge
by itself is valueless and without meaning.

There is, however, no subject of the medical curriculum in
which the organization and arrangement of laboratory courses is
more difficult for classes of average size than in physiology. In
physics and chemistry, as well as the various morphological sub-
jects, the material for the laboratory course is constantly available,
and can be stored away for future use, whereas in physiology living
material must be provided afresh for each experiment and there
must be constant supervision of the practical work to see that the
material is properly used. This requires that the student be
adequately directed as to how he should proceed with the experiment
without at -the same time stifling originality on his part by re-
quiring him explicitly to follow detailed directions in every step.
The experiment is of no value unless it is performed with an inquir-
ing attitude of mind, and laboratory directions and instruction
should be no fuller than is necessary to guide the student in its
general performance.

It is with these requirements in view that the present volume
has been compiled, and while the authors realize full well its many
shortcomings they hope that the practical instruction, not only in
their own laboratories, but also in those of other institutions may
be assisted by its publication.

The work is arranged in sections, each of which deals with
some special part of the subject, this plan being adopted because
it has been found the most practicable for courses designed for
large elementary classes, as well as for smaller groups of more
advanced students. The first section deals with the fundamental
experiments in the physiology of isolated muscle and nerve, placed

PREFACE. 5

in this position not because this is the simplest part of the subject
of physiology to understand, but because it is the easiest to provide
material for and therefore the best in which to permit of sufficient
practice, so that the student may become familiar with the methods
of physiological technique. The two following sections deal with
essential experiments illustrating the principles of the heart beat
and the circulation of the blood. In selecting experiments in the
last mentioned group the endeavour has been made to apply them,
as far as possible, to man. A certain number of experiments in
which other mammalian material is used is, however, essential in
order that the student may be in a position to appreciate the
significance of results which will later be demonstrated to him.
These demonstrations are described in the last sections of the
manual in sufficient detail, so that the various steps may be followed
by every student of the general course and the experiments may
be performed by small groups of more advanced students.

In the experiments on the central nervous system the decere-
brate or spinal animal is extensively made use of, as recommended
by C. S. Sherrington whose work in this direction must be con-
sidered as the most important contribution that has recently been
made to the advancement of the teaching of practical physiology.
In the section on the special senses a relatively larger proportion of
theoretical matter is given, along with the directions for the experi-
ments, partly because the latter, being practically all subjective
in nature, cannot be performed unless the student understands fully
what he is looking for, and partly because most of them can, and
should be performed in the study rather than the laboratory.

A great difficulty has always been felt by instructors in physi-
ology in attempting to supply simple experiments bearing on
the chemistry of respiration and yet there is no part of physiology
in which practical experience is more important, if the student is
to understand the principles of this difficult subject. A section of
the manual is devoted to experiments in which simplified and inex-
pensive apparatus for gas analysis is employed.

Throughout the book, besides the directions for the experi-
ments, a brief statement is given of the theoretical matter which
bears on them. This is done to enable the student to appreciate
the object of the experiment and to guide him in the interpretation

6 EXPERIMENTAL PHYSIOLOGY.

of the result; but the latter is not described in detail, this being left
for the student to determine for himself.

While the general plan and arrangement of the book is the work
of the authors jointly, each author is responsible for certain chapters
as indicated by the initials given in the table of contents.

The authors wish to express their thanks to Miss Marion E.
Armour for the care and patience which she bestowed in the pre-
paration of the drawings for the illustrations, many of which are
original while others are copied from other texts, acknowledgment
for which is given in the legends.

They wish also to thank Miss Jean Halliday for her assistance
in arranging the index and correcting the proof. They are indebted
to Dr. J. P. Eisenberger for a final reading of the proof.

In the present edition considerable additions have been made
in order that the manual may cover a wider field of experiments
and, so it is hoped, be of greater assistance in other institutions.
A section on General Physiology has been added as well as chapters
on Electrocardiography and Respiratory experiments. The
methods for air and blood gas analysis have been largely rewritten
since it is now possible to obtain at reasonable cost apparatus
with stopcocks, thus making it unnecessary to use the simpler
apparatus described in the first edition.

We wish to thank Dr. N. B. Taylor for valuable assistance in
the preparation of this edition.

LOIS McPHEDRAN ERASER.

F. A. HARTMAN.
J. J. R. MACLEOD.
J. M. D. OLMSTED.
The University of Toronto.

CONTENTS

Chap.

I.

II.
III.
IV.

Section I.

MUSCLE AND NERVE.
F. A. H.

PREFACE

PRELIMINARY TECHNIQUE

VOLUNTARY MUSCLE

SMOOTH MUSCLE AND CILIATED CELLS

PHYSIOLOGY OF NERVE..

Page.

3

9

14

42

46

V.
VI.

Section IIA.

CIRCULATION.

F. A. H.

CARDIAC MUSCLE 61

BLOOD PRESSURE. EFFECT OF CHEMICAL SUBSTANCES ON THE

BLOOD VESSELS AND HEART 71

Section IIB.

CIRCULATION.

J. J. R. M.

VII. HAEMODYNAMICS . 78

VIII. VASO MOTOR NERVES. DEPRESSOR NERVE. CIRCULATION TIME. 87

IX. POLYSPHYGMOGRAPH TRACINGS 93

Section HI.

REFLEX ACTION.

J. J. R. M.

X. REFLEX ACTION IN THE FROG 98

XI. REFLEX ACTION IN MAN. REACTION TIME IN MAN 104

Section IV.
RESPIRATION.

J. J. R. M.
XII. ANALYSIS OF AIR AND COLLECTION OF ALVEOLAR AIR 107

XIII. RESPIRATORY EXCHANGE IN MAN 118

XIV. DETERMINATION OF THE GASES OF BLOOD BY THE PUMP METHOD. 122
XV. DETERMINATION OF BLOOD GASES BY THE CHEMICAL METHOD. 128

XVI. THE DISSOCIATION CURVE FOR OXYGEN AND THE CO2 COMBINING

POWER OF BLOOD 132

Section V.

SPECIAL SENSES.

L. McP. F.

XVII. VISION 139

XVIII. ERRORS IN REFRACTION 149

XIX. EXAMINATION OF THE REFRACTION OF THE EYE AND OF THE

INTERIOR OF THE EYEBALL 158

XX. RETINA 163

XXI. COLOUR VISION 170

XXII. SIMULTANEOUS CONTRAST. VISUAL JUDGMENTS. . 178

XXIII. HEARING 182

XXIV. SKIN SENSATIONS 192

7

CONTENTS.

Section VI.
DEMONSTRATIONS.
CIRCULATION AND RESPIRATION.

J. J. R. M.

Chap. Page.
XXV. THE CIRCULATION OF THE BLOOD AND LUMPH AND THE RE-
SPIRATION 199

XXVI. THE DIRECT DEMONSTRATION OF VASOCONSTRICTOR BY THE

PLETHYSMOGRAPHIC METHOD 202

XXVII. PERFUSION OF THE EXCISED MAMMALIAN HEART. . 206

XXVIII. MEASUREMENT OF THE BLOOD FLOW THROUGH THE HANDS
AND FEET BY THE CALORIMETRIC METHOD. ELECTROCAR-
DIOGRAMS 210

XXIX. LYMPH FORMATION 218

XXX. EXPERIMENTS TO DEMONSTRATE THE PUMPING ACTION OF THE

HEART AND THE ACTION OF THE VALVES 223

Section VII.

DEMONSTRATIONS.

DIGESTIVE SYSTEM.

J.J. R. M.

XXXI. THE INNERVATION OF THE SALIVARY GLANDS: 226

XXXII. THE CONTROL OF THE PANCREATIC SECRETION. THE SECRE-
TION OF BILE 228

XXXIII. EXPERIMENT ON THE NORMAL SECRETION OF SALIVA AND

GASTRIC JUICE 231

XXXIV. THE MOVEMENTS OF THE OESOPHAGUS AND INTESTINE 236

Section VIII.

DEMONSTRATIONS.

THE CENTRAL NERVOUS SYSTEM.

J.J. R. M.

XXXV. REFLEX ACTION IN THE MAMMALIA 239

XXXVI. CEREBRAL LOCALIZATION. DECEREBRATE RIGIDITY. RECI-
PROCAL INNERVATION. FUNCTIONS OF SPINAL ROOTS IN

THE MAMMAL (Doc) 243

XXXVII. THE DECEREBRATE CAT 1 247

XXXVIII. THE DECEREBRATE CAT II. THE DECEREBRATE PIGEON 251

XXXIX. THE SPINAL CAT 255

Section IX.

GENERAL PHYSIOLOGY.

J. M. D. O.

XL. PROPERTIES OF LIVING PROTOPLASM., CHEMICAL CONSTI-
TUENTS .' 258

XLI. PHYSICAL CHEMISTRY OF CELLS 263

X — II. ENZYMES, DIGESTION, BUFFERS 267

XLIII. MOVEMENT. ENVIRONMENT 269

XLIV. CIRCULATION (CAPILLARY) 273

APPENDIX 277

INDEX.. 281

SECTION I.

CHAPTER I.

PRELIMINARY TECHNIQUE.

Electrical Stimulation. — In experimental physiology it is
necessary in many instances to employ an artificial stimulus in
order to cause activity in the tissue to be studied. Many kinds of
stimuli are capable of doing this, but to most of them there are
objections when used on nerve or muscle. Mechanical, chemical
and thermal stimuli are all more or less injurious. Moderate
electrical stimulation on the other hand brings these tissues into
action without appreciable injury. As this is the means by which
the tissues are usually stimulated, we will discuss briefly the prin-
ciples involved in the more important pieces of electrical apparatus
employed.

For a source of current either a dry cell or a dynamo may
be used. A dry cell is essentially a large zinc cup serving as
container in which is suspended a carbon cylinder. A small amount
of water containing ammonium chloride held by gelatin or some
inert substance fills the rest of the container. A dry cell is
essentially a wet cell, but with the water protected from spilling
by the gelatin "sponge'4'. Whenever the carbon and zinc are con-
nected by a conductor, electrical energy is set free by the solution
and is transmitted along the conductor. It has been found that
if a live muscle is placed in the electrical circuit that the tissue will
be stimulated at the time the circuit is made and again when it is
broken provided the current is great enough.

Two important factors must be kept in mind in the use of an
electric current, the resistance of the tissue and the "pressure"
of the current. The unit of "pressure" is the volt. Therefore the
amount of pressure is called the voltage. By increasing the voltage
the penetrating power is increased and thus the physiological effect
augmented. If the resistance of the tissue is high as in dry skin, a

9

10 EXPERIMENTAL PHYSIOLOGY.

greater voltage is necessary than if the skin has been moistened,
because moist skin is a better conductor. Dry cells are made to
give in the neighbourhood of one or two volts. Greater voltage
in a circuit can be obtained by connecting two or more cells in
series, i.e., zinc to carbon.

In most of your work the current generated by dynamos in
the power plant will be used. At each working place two wires
from the main line are connected with a metal ribbon of high
resistance. Terminals are arranged so that different proportions
of this ribbon can be used in the circuit, thus varying the amount
of voltage. The amount of voltage obtained can be estimated by
adding the total voltages between the two terminals employed.
The following diagram gives the voltages between terminals.
T represents the terminals as arranged on the table.
T8T8T4T4T4T2T2T
abcdef gh

To illustrate the method of finding voltage, connecting a to f
gives 28 volts; f to h, 4 volts; d to e, 4 volts.

Besides the direct current one can use the induced current for
stimulation. It is well known that if one wire is placed near
another wire which carries a current, a brief current is induced in
the first wire at the time of breaking the circuit in the second,
likewise a current is induced at the closing or making of the circuit.
The wire carrying the direct current is called the primary circuit,
while that in which the current is induced is called the secondary
circuit. Greater effects can be produced by having a coil of well
insulated wire surrounding a soft iron core as the primary circuit
element and another coil of insulated wire as the secondary element.
These two coils are arranged on a stand for convenience, the whole
apparatus being termed an inductorium (Fig. 1). The chief
advantages of the induced over the direct current for purposes of
stimulation are; first, the voltage is very much greater so that
high resistances can be overcome and, secondly, it can readily be
altered in strength.

Note that the inductorium is so arranged that by connecting
binding post 1 and 2 a single momentary current is induced in the
secondary coil when the circuit is either made or broken in the
primary coil; and that by connecting binding posts 1 and 3 brief

PRELIMINARY TECHNIQUE. 11

currents are continuously induced in the secondary coil due to the
constant interruption of the current in the primary by the vibrating
spring. The latter effect is called a tetanizing current.

It has been found that the least induced current is produced
when the secondary is at right angles to the primary; from this
position the current is increased more and more as the secondary
becomes more nearly parallel. In addition to the angular relation-
ship, the distance between the two coils determines the strength
of current induced, the closer they are the greater the effect.

FIG. 1. Inductorium: (a) Primary Coil; (b) Secondary Coil; (c) Bar
for short-circuiting the secondary terminals. Connect the source of
supply to 1 and 2 for single shocks and to 1 and 3 for a succession of rapid
shocks.

Experiment 1. — Try the following. — -1. Connect the electrodes to
two terminals on the table so as to obtain 2 volts. Touch the
tongue with the electrodes and then make and break the circuit
by means of the switch. Repeat with increasing voltage.
Describe the effects at make, at break and during the passage
of the current.

2. Connect the binding posts 1 and 2 (Fig. 1) of the primary
coil to 4 volts on the table, with a simple key in the circuit.
Connect the electrodes to the end of the rods upon which the
secondary slides. With the secondary coil far out and parallel

12 EXPERIMENTAL PHYSIOLOGY.

to the primary coil stimulate the tongue by making and break-
ing the primary circuit. Move the secondary closer and closer
to the primary coil and note the effect produced on the tongue.
Again move the secondary out into such a position that it just
clears the primary coil. Now study the effect of tilting the
secondary coil.

The Graphic Record. — By the use of a lever it is possible to
magnify muscle contraction. The movement of this lever is re-
corded on smoked paper. The kymograph is devised for carrying
the smoked paper.

Glazed paper is wrapped tightly around the drum of the kymo-
graph so that the glaze is outward. The gummed end is fastened
to the opposite end so that the paper will not slip and the writing
point of the lever will not catch on the overlap. The drum is
held in the smokiest part of a gas flame while being rotated rapidly
on the brass tube which is maintained horizontally by hand. In
this way a uniform coating of carbon can be obtained on the
paper. A light chocolate colour is to be preferred to a deep black
because there is less danger of burning the paper.

A small triangular piece of photographic film or waxed paper is
fastened by a bit of wax to the end of the lever in order to serve
as a writing point. The lever is always on the right hand side of
the drum so that when the latter is rotated it pulls away from the
writing point. The lever must be at a tangent to the curved surface
of the cylinder in order to prevent the writing point leaving the
smoked surface when it rises. The writing point may be bent
inward to keep the rest of the lever free of the drum.

When ready to remove the paper from the drum, it is cut at
the overlap by a sharp knife in such a way that the knife strikes
the under paper and not the drum. A corner of the paper is
grasped against the edge of the drum with the other hand so that
it will not fall. At this stage the record may be placed on the table
and lettering or other explanatory matter inscribed. It is well
to place your name and date on each record. The record, with
smoked side up, is passed once through a rosin-alcohol solution
(120 gm. rosin to 1,000 c.c. of 95% ethyl alcohol) and then hung
over the tray to drain. When dry the parts desired are to be care-
fully cut out and preserved in the note book.

PRELIMINARY TECHNIQUE 13

Instruments for Dissection. — Each student should supply
himself with the following: Two pairs of scissors, one heavy the
other fine; two pairs forceps, one heavy the other fine; one dental
probe, one strabismus hook, one scalpel, one small serre fine and
one haemcstat. The fine scissors and fine forceps are to be used
only in delicate dissection. In order to do the best work the cutting
instruments must be kept sharp.

CHAPTER II.
VOLUNTARY MUSCLE.

One of the most striking characteristics of living things is
their power of movement. This is not so marked in plants, but
all animals, even those which lack the ability to move from place
to place, possess organs which are constantly in motion. The power
of contractility which is inherent in all protoplasm is developed
to its highest degree in muscular tissue. Thus practically every
life process which involves the movement of masses of tissues is
brought about by the action of muscles, e.g., breathing, circulation
of the blood, passage of the food down the alimentary tract, etc.

A portion of the muscle of the body is under the control of
the will. Upon this muscle we depend for locomotion, movement
of our hands, eyes and the like. Such muscle is distinguished by
the fact that it is composed of fibres with a characteristic cross
striation.

STIMULATION.

Various kinds of stimuli will cause muscle to contract, but before
deciding upon the best form to use we must know where to apply
the stimulus. Normally the muscle fibres are called into action
by impulses passing along the nerve fibres. Artificial stimuli can
be effectively applied along the same path. Thus mechanical,
thermal, chemical and electrical changes can cause a muscle to
act through its nerve. Any of these changes to be effective must
not only be of sufficient magnitude but must be abrupt. A nerve
slowly compressed may be without effect, but if quickly pinched,
a twitch of the muscle results.

Of all artificial stimuli, an electric current is not only the most
effective, but also the least injurious to nerve. Mechanical and
thermal changes great enough to have an effect injure the tissue
permanently so that these stimuli can be used only a single time.
Chemical stimulation if not harmful, is at least difficult to control.
Moderate electrical changes have none of these objections.

14

VOLUNTARY MUSCLE. 15

Although we are most interested in the physiology of mam-
malian muscle, we employ muscles from cold-blooded animals
because, while similar in function, they survive for a much longer
period when deprived of their circulation.

Experiment 2.— Nerve -Muscle Preparation. — Kill a frog in the
following manner: By bending the frog's head down, a depression
between the skull and first vertebra can be felt on sliding a
blunt-pointed seeker back along the mid-line of the top of the
skull. Make a short traverse cut in the skin at this point.
Plunge a seeker or pithing wire into this depression and turning
the instrument forward into the skull cavity destroy the brain.
Next destroy the cord by passing the seeker down into the
spinal cord. If the frog is properly pithed it immediately
becomes limp. Later the muscles may become less flaccid. A
frog thus prepared is dead although the heart and other tissues
may live for a few hours. There can be no pain whatsoever.
This method of killing a frog is called "pithing".

Slit the skin around a pithed frog just behind its fore legs
and remove the skin from the posterior half of the animal.
Lay the preparation ventral down on a clean glass plate and
keep it moistened with 0.7% salt solution. Carefully dissect
out the sciatic nerve on one side by separating the muscles on
the dorsal aspect of the thigh and freeing 'the nerve from the
surrounding tissues. Always lift the nerve with a glass hook
and avoid stretching it. Trace the nerve to its exit from the
spinal column. Split the spinal column in two at this point.
Leave parts of two or three vertebrae attached to the nerve
to serve as a convenient way of handling the nerve and cut
the remaining tissue away.

By means of the vertebrae raise the roots of the sciatic nerve
and clip beneath it until it is free down to the knee. Cut the
Tendo Achilles and separate the gastrocnemius from the other
muscles as far as the knee. Cut the other muscles and the
tibia-fibula just below the knee. Cut away all the muscles
above the knee and the femur, leaving about half an inch of bone.
In making a nerve-muscle preparation one must always
remember that slight injury to muscle or nerve may ruin it
for experimental purposes. Such injury may be caused by

16 EXPERIMENTAL PHYSIOLOGY.

stretching, pressing, drying, or chemical agents. The loss of
water may be prevented by keeping the muscle in air saturated
with water vapour or by frequently applying water to the
exposed surface. But bathing the muscle with water produces
consequences as serious, perhaps, as drying, because salts and
other substances are drawn out of the cells by osmosis. Now
if we add the same salts in the same percentage as occurs in
the tissue to the water which is used to bathe the muscle
such a solution can be used to prevent evaporation without
taking salts from the cells. NaCl dissolved in water to the
extent of 0.7% produces an isotonic solution somewhat similar
to the tissue fluid of the frog, and is therefore useful for keeping
the tissues moist without the injurious effect of pure water.
Apply this solution to both the nerve and muscle frequently.

NERVE-MUSCLE PREPARATION.

Experiment 3. — Different Kinds of Stimuli. — Fix the femur
of a nerve-muscle preparation in the flat-jawed clamp fastened
to a stand. Clamp the nerve holder to the stand just below
and then lay the whole sciatic nerve on the glass plate. Fasten
the muscle lever underneath. Attach the tendon, by means
of a hooked pin to the lever. Now adjust the after-loading
screw so that the muscle supports no weight but is fully ex-
tended when at rest. Place a ten gram weight upon the scale
pan (Fig. 2). In practically every experiment in muscle-nerve
physiology, the muscle is after-loaded in this fashion. This
after loading keeps the muscle extended and takes the slack
so that when contraction occurs there will be no sudden jerk
which might injure the muscle fibres. Arrange the lever so
that it will write on the smoked paper of the kymograph and
obtain records with the following types of stimuli :
MECHANICAL. — Pinch the nerve near its end with forceps. Can
you get a response the second time from the same place?
CHEMICAL. — Place a few salt crystals on the fresh end of the
nerve, the injured portion having been cut away. After a few
contractions of the muscle thoroughly remove the salt with
isotonic salt solution.

VOLUNTARY MUSCLE.

17

THERMAL. — Touch the fresh end of the nerve with a hot glass

rod.

ELECTRICAL. — Connect the stimulating electrodes directly to

the current supply on the table but with a simple key in the

circuit. Try the effect of make and break, beginning with 2

volts and then increasing the current.

FIG. 2. Apparatus for recording muscle contraction: (a) femur clamp; (6) nerve holder
(c) electrodes; (d) muscle lever; (e) signal magnet; (/) after-loading screw; (g) wires connected in
the primary circuit.

Next connect the electrodes to the secondary coil of the
inductorium using a primary current of 2 volts; stimulate with
make and break shocks of different intensities.

Compare the effects of make and break in direct and indirect
currents.

Minimal and Maximal Stimulation. — Beginning with a
stimulus too weak to cause contraction, then very gradually
increasing its strength, a point is reached where a slight contraction

18

EXPERIMENTAL PHYSIOLOGY.

results. This is called the minimal stimulus. In other words it is
the least stimulus which will cause a contraction. On the other
hand the maximal stimulus is the least stimulus which will produce
maximal contraction. When a series of stimuli of increasing strength,
beginning with the minimal, is sent into a nerve or muscle, one
obtains a graded response (Fig. 3). This is true no matter how
finely graded is the increase of the stimulus. (See all-or-none
principle, p. 61).

Experiment 4. — Response to Stimuli of Different Intensities.
—For this experiment employ a muscle which has been cut
away from its nerve. Instead of using ordinary stimulating
electrodes connect one terminal of the secondary coil to the

i

FIG. 3. Contractions produced by graded stimulation. Secondary coil moved 5 mm.
at each step. Both make and break shocks employed. Make contraction begins at m.

femur end of the muscle by wire and in a similar way connect
the other terminal to ihe binding post on the muscle lever.
Then a bit of fine wire is wound around the tendon and finally
fastened to the coarser wire at the binding post on the muscle
lever. In this manner a current can be passed through the
muscle.

Starting with the secondary as far as possible from the
primary, cautiously move it forward by stages, each time
stimulating the muscle with a make and break shock. When
the first perceptible contraction is produced you have reached
the threshold. Between each stimulation move the drum a

VOLUNTARY MUSCLE. 19

short distance by hand, not more than 1 mm. Now carefully
move the coil toward the primary so that at each position
there is produced a slightly greater contraction than before.
Distinguish the "make" and "break" contractions on the
record by the letters nt and b placed beneath. Also designate
the position of the secondary coil. It is possible by carefully
increasing the stimulus to obtain a graduated series of con-
tractions from both the make and break shocks (Fig. 3). When
the series is complete, indicate the minimal and maximal
stimuli.

Discuss the application of the all-or-none principle to your
records.

INDEPENDENT EXCITABILITY OF MUSCLE.

Ordinarily the question, as to whether a muscle could respond
if its nerves were absent, is of academic interest. But if we wish
to test the power of a muscle which has lost its voluntary function
by injury to the nerve, we find long after the nerve has degenerated,
that the muscle responds to direct electrical stimulation. It can
also be shown that with the nerve intact but with the connection
between the nerve and muscle paralysed, the muscle is still excit-
able. This can be done by the injection of the poison "curare"
into the circulation. After a few minutes has elapsed, stimulation
of the nerve no longer affects the muscle, while stimulation of the
muscle causes it to contract. From such experiments as these
it appears that muscle can contract without the mediation of
either nerve fibres or motor end plates.

Experiment 5. — The sciatic nerves are exposed in each leg of a
frog which has the brain destroyed. A ligature is passed under
one sciatic and around the thigh and tied tightly so as to
occlude circulation in that leg. A few drops of 1% curare
solution are injected into the back of the frog. From time to
time stimulate each sciatic nerve with induction shocks. In a
few minutes, the leg with intact circulation refuses to respond
or else responds very much less than the ligated leg. When
this stage has been reached stimulate the muscles of the un-
ligated leg directly. Their response demonstrates that the

20 EXPERIMENTAL PHYSIOLOGY.

muscle is independently irritable for the nervous impulse has
been ruled out. This has been accomplished by paralysis of
the tissue connecting nerve fibres with muscle fibres, for it
can be shown that impulses still travel along the nerve. How
does this experiment show the locus of action of the drug curare?

EXTENSIBILITY AND ELASTICITY OF MUSCLE.

In the body more or less tension affects each muscle. The
weight of various parts and the antagonistic action of opposing
muscles contribute to this tension. The result is a certain amount
of elongation. This can be shown by fastening a weight to an
isolated muscle. When the weight is removed the muscle soon
regains its former length, that is, it exhibits elasticity.

The amount of elongation increases with increase in tension,
but with each additional tension the elongation is proportionately
less. To illustrate — suppose X grams cause an elongation of 2 mm.
Then 2 X grams will not cause 4 mm. elongation but something
less, perhaps 3 mm. 3 X grams would produce 4.5 mm. elonga-
tion, etc.

Beyond a certain tension the reverse is true; the elongations
per unit increase being greater and greater with each step. Where
this reversal begins the so-called elastic limit has been passed.
Finally there comes a time when the muscle fibers are ruptured.

Needless to say a muscle should not be extended beyond its
elastic limit.

In conclusion, tension is very useful in keeping a muscle pre-
pared for contraction, for we find that a muscle will not only
respond more quickly but more energetically if it is under a certain
amount of tension. This condition holds in the body.

Muscle can be stretched and again regain its resting length
when the force producing the change is removed. It differs some-
what from a rubber band in that it does not respond by equal
extension for equal increments of weight as the total weight in-
creases nor does immediate recovery occur when the weights are
removed.
Experiment 6. — Isolate either of the two large muscles (gracilis

and semi-membranosus, Fig. 4) on the inside of the thigh of a

VOLUNTARY MUSCLE. 21

frog; cut through the tibia below its insertion and through the
femur above the knee. Remove it with the bone from which it

FIG. 4. A ventral view of frog. 1, M. Gastrocnemius.
2, M. Tibialis anticus. 3, M. Gracilis. 4, M. Adductor mag-
nus. 5. M. Sartorius. 6. M. Adductor longus. 7, M. Vastus
internus. 8. N. Vagus. 9, N. Glosso-pharyngeal. 10, N. Hypo-
glossal. 11, N. Superior laryngeal.

B. Dorsal aspect of hind leg. 12, M. Rectus anterior.
IS.M.Vastusexternas. 14,M.Semimembranosus. 15, M. Peroneus.
(After Jackson).

arises. You should have a preparation with bone at each end.
Suspend the muscles by clamping one bone in the flat-jawed
clamp. Fasten the other end to a muscle lever to which a

22 EXPERIMENTAL PHYSIOLOGY.

large scale pan is attached. Obtain a record of the comparative
length of the muscle without weight and with weights increasing
by 10 gms. at each trial. The drum should be moved by hand
between each increment. Continue until the point of rupture
is reached. Plot a curve of these results.

In a second preparation, add 10 gm. increments, but stop
before the muscle is injured. Now remove 10 gms. at a time,
securing a record in each case. Plot a curve of the recovery.
^^ In a similar manner obtain records from a rubber band,
both for extension and recovery.

Study the extensibility of a contracted muscle as compared
to the same muscle at rest. Arrange a muscle preparation as
//^. before. Obtain short records of the muscle at rest and when
stimulated by a single maximal break shock without any weight.
Continue to do this for 10 gram increments as far as possible.
What happens when a load is reached which the muscle cannot lift?
rest and when stimulated by a single maximal break shock
without any weight. Continue to do this for 10 gram incre-
ments as far as possible. What happens when a load is reached
which the muscle cannot lift?

Plot extension curves of the contracted muscle and the
resting muscle so that they may be easily compared.

THE SIMPLE CONTRACTION.

Although muscle seldom responds by a simple twitch when
called into action in the body, its behaviour can be conveniently
studied when it gives such twitches. Simple contractions can be
caused by single electrical stimuli. In order to observe the action
of a single muscle it is isolated from its neighbours and arranged as
described in the experimental work so that when contracting a
record is made upon the surface of a rapidly rotated cylinder
(Fig. 5). By means of a signal magnet and a tuning fork the time
required, for the initiation and completion of the action, can be
determined. The time elapsing between the application of the
stimulus and the first visible contraction is about 0.01 sec. for a
frog's gastrocnemius. With the more quickly responding mirror
method it is 0.0065 sec. The time consumed in the contraction

VOLUNTARY MUSCLE.

23

FIG. 5. Rapid kymograph, (a) Terminals to be connected in
primary circuit; (&) Movable ring for second stimulus; (c) Release
which starts drum.

24 EXPERIMENTAL PHYSIOLOGY.

averages 0.04 sec. while that used in relaxation is slightly more,
0.05 sec.. Roughly the total time involved in the various changes
is 0.1 sec. for the frog's gastrocnemius (Fig. 6).

The duration of the simple twitch varies in different animals.
It is less in the rabbit, 0.07 sec., and considerably less in insects,
0.003 sec.

The real LATENT PERIOD is primarily due to the time consumed
in inducing the changes in muscle leading up to contraction. Such
changes vary with the onset of fatigue, modification of temperature,
and intensity of the stimulus. Besides this, other factors are
added. If a load is attached to the muscle, the preliminary effect
of attempted contraction is a slight elongation on account of the
inertia of the load. With the ordinary apparatus friction also
slows the response.

FIG. 6. Record of an isotonic contraction of a frog's gastrocnemius muscle.
Stimulated at X. Tuning fork tracing, 1/100 sec.

Contraction Period. — A muscle requires time for the develop-
ment of the maximum contraction because each individual muscle
fibre is not stimulated in all parts simultaneously, but receives its
impulse at the motor end-plate located near the middle of the fibre.
Hence the contraction spreads outward at the rate of 3 to 4 meters
per sec. In human muscle the velocity of such a wave is 10 to
13 meters per sec. Therefore, even though each muscle fibre of a
mass may be stimulated simultaneously through a nerve trunk,
time is required for the spread of the contraction throughout the
fibres.

This can be shown experimentally in a muscle with parallel
fibres such as the sartorius. Levers are arranged resting on each
end of the muscle so that when the corresponding part of the muscle

VOLUNTARY MUSCLE. 25

contracts a record will be made on a drum. These levers are so
adjusted that they write directly one above the other. If both
parts of the muscle contracted simultaneously the levers would
move simultaneously. One end of the muscle is stimulated with
a single shock. That end contracts some time before the other,
as shown by the records upon the kymograph. By measuring the
length of muscle between the levers and determining the time
with a tuning fork on the drum, the rate in meters per sec. can be
ascertained.

Relaxation is an active process and is not entirely due to the
effect of tension upon a muscle which has ceased to contract.

If a frog's muscle is carefully isolated and dipped in olive oil
to reduce friction, it will actively relax after contraction, provided
it is floated on mercury.

Volume of a Contracting Muscle. — The contraction of a
muscle does not change its volume as the following experiment
will show.

*Experiment 7. — Two well insulated wires from the secondary
terminals of an inductorium are passed through one hole of a
rubber stopper. The bare ends of the wire are connected to
opposite ends of a frog's gastrocnemius, which is then suspended
within a wide-mouthed bottle. The hole through which the
wires passes is made water-tight. A glass tube drawn out to a
capillary is inserted in the other hole after the bottle has been
completely filled with isotonic NaCl solution. The solution
should extend a short distance into the capillary portion of the
tube. The inductorium is arranged to produce a tetanizing
current. In spite of the short-circuiting through the solution,
if a sufficiently strong current is used, the muscle can be made
to contract.

Determine whether or not any change in volume takes place
due to contraction.

Isotonic Contraction. — When a muscle lifts a light load it
shortens with little opposition, the tension remaining nearly con-
stant throughout. Such a contraction is literally one with constant
or iso-tension, hence called isotonic.

Experiment 8.— Isotonic Muscle Contraction.— The time
elements in the contraction and relaxation of a muscle can best

26 EXPERIMENTAL PHYSIOLOGY.

be studied by obtaining a record of the muscle on a rapidly
moving drum. A gastrocnemius preparation in the moist
chamber is prepared for direct stimulation by connecting each
end of the muscle to a binding post (Fig. 5). These binding
posts are connected in turn to the terminals of the secondary
coil. Moist filter paper in the chamber will prevent drying of
the muscle.

The muscle lever must be in such a position that it will
continue to write on the drum when the muscle contracts.
Weight the scale pan sufficiently to permit an appropriate
excursion of the lever. Adjust the after-loading screw so that
the resting muscle does not support this weight. The electro-
magnetic signal should be supported on the same stand as the
muscle lever and its writing point should be close to the writing
point of the muscle lever, in line vertically with it. Always
make alignment marks on the drum with all writing points
before recording a muscle contraction. Both the electromag-
netic signal and the kymograph should be introduced into the
primary circuit of the inductorium.

After determining a suitable strength for the stimulating
current, record a contraction, using the spring-driven drum.
By means of the lever at rest, record below this a base line.
Now draw an arc with the lever from the point of maximal
contraction to the base line. Obtain a tuning fork record
between the base line and signal line.

In this manner secure three or more good records. After
the paper has been removed from the drum and the surface
fixed with rosin, determine the time for the latent period,
contraction period and relaxation period.

If you have time obtain additional records in which the
after-loading screw is changed, so that the load is picked up
earlier or later than at first.

Discuss the factors which might be involved in production
of the latent period.

Experiment 8a.— Modification for Harvard Kymograph.— The
writing point of the signal magnet must be in a vertical line
with the writing point of the muscle lever. Signal magnet and
muscle lever are clamped as close together as possible on the

VOLUNTARY MUSCLE. 27

same stand. The tuning fork with writing point bent back-
ward is fastened on a separate stand at such a height that when
vibrating its record will come between that of the muscle lever
and signal-magnet. The tuning fork is placed in the opposite
direction to that of the muscle lever but as close as possible
without interference. The screw at the top of the rod support-
ing the drum is tightened so that the drum can be spun freely
by hand. Two persons are required to perform the experiment.
They should practice until they can perform teamwork without
difficulty.

The muscle and signal-magnet levers are adjusted against
the surface of the drum, then number one vibrates the tuning
fork and applies its writing point to the drum. Number two
spins the drum rapidly enough to show distinctly the individual
vibrations on the tuning fork, and as soon as the speed is
uniform number two stimulates the muscle with a break shock.
Number one must be ready to push the kymograph away from
the recording points as soon as the contraction and relaxation
are finished. Directions for the remainder of the experiment
are the same as in Experiment 8.

Isometric Contraction. — Sometimes a muscle tries to lift a
load which it cannot budge. The muscle tugs at the weight with
an increasing effort, but it cannot shorten, although the tension
increases with the effort. Such a muscle maintains a constant
length. It is said to undergo an isometric contraction.

In the preceding experiment the force which opposed the
contraction of the muscle was essentially a constant one, so that
the muscle tension remained nearly the same. In the present ex-
periment the tension is made to increase as contraction proceeds
and the change in muscle length is small. It is true that the muscle
does not maintain a constant length, but it approaches that con-
dition.

*Experiment 9. — Fasten the femur of a gastrocnemius preparation
in a flat-jawed clamp which has been fixed in the upper part of
the tripod. Connect the tendon by a hook to the spring lever,
the latter being supplied with a writing point. Connect the
secondary terminals of the inductorium with the binding posts
of the clamp and the lever. To avoid poor contacts you should

28 EXPERIMENTAL PHYSIOLOGY.

connect the binding post of the lever to the tendon by means
of a fine wire.

With the muscle under some tension secure a record of a
single contraction upon the drum of the spring-driven kymo-
graph.

Secure a second record with an increase in tension. Record
an iso tonic curve by means of the other lever on the tripod.
Stimulation signals and tuning fork records must be shown
under each record.

In order to obtain the tension or resistance overcome by the
muscle in the isometric contraction, turn the spring over and
attach the large scale-pan (weight 20 grams). Then add
sufficient weights to stretch the spring to the same extent as
occurred in the isometric contractions. Turn the drum by
hand to make short abscissae. Mark these lines with the
weights. (May also be done with Harvard kymograph as in
Exp. 8a).

Compare the latent periods, contractions and relaxations in
the isometric and isotonic records.

It is never possible to obtain a purely isotonic contraction, for
in all contractions of this kind, tension does change, though ever
so little. Nor does the purely isometric type exist, in spite of the
fact that the muscle does not lift its load and thus appears to be
free from any shortening of fibres. Watch such a muscle at work.
Some of the fibres do thicken at least in part of their course, there-
fore they must shorten. They probably do so at the expense of a
slight extension of the tendon or of other fibres.

Throughout the activity of the muscles of the body we may
have contractions predominantly isotonic or predominantly iso-
metric or these may be combined in various proportions.

EFFECT OF TEMPERATURE ON CONTRACTION.

Chemical, changes underlie muscle activity. All chemical pro-
cesses are influenced by temperature, a rise causing an increase
in the rate of the reaction, a fall causing a decrease. Within
certain limits we find this true in muscle. There is this difference
from inorganic chemical processes — the maximum reaction is soon

VOLUNTARY MUSCLE.

29

reached in muscle, increase beyond
that temperature reduces more
and more the response of the
muscle until at about 38° C. in
the frog the irritability of the
muscle is lost. Muscle is not alone
in possessing an optimum temper-
ature for its activities. All tissues
have an optimum temperature.
This optimum varies iri different
individual animals, especially cold-
blooded animals like the frog.
Winter frogs possess a lower
optimum than summer frogs.

Not only is the height of con-
traction changed by temperature
rise or fall, but the duration of its
phases is changed as well. At low
temperatures the latent period and
duration of the contraction may
be several times the corresponding
stages at higher temperatures
(Fig. 7).

Mammals differ from frogs in
maintaining a more or less
optimum temperature for their
tissues independent of external
changes.

Experiment 10.*— Fasten, by
means of thread or fine wire,
the femur of a gastrocnemius
preparation to the short arm
of the L-shaped glass rod which
has previously been fixed by
its long arm in the flat-jawed
clamp (Fig. 8). Connect the
tendon by means of a fine wire
to the pulley of the muscle
lever, suitably weighted. Con-

*This experiment may be done with the Har-
vard kymograph using the highest speed of the
clockwork.

&S,

w S

-

«-

o.2

II*

30

EXPERIMENTAL PHYSIOLOGY.

nect the secondary terminals of the inductorium with the
muscle lever and with the femur. On a rapidly moving

surface secure records of twitches immediately after the muscle
has been immersed in isotonic salt solution at the following
temperatures, each for three minutes: 4° C., 10° C., 15° C.,
25° C., 30° C., 35° C., and 40° C. Use the same

VOLUNTARY MUSCLE. 31

strength of stimulus in each case and superimpose the records,
taking care always to start at the same point on the drum.
A tuning fork record is made and the latent period is also
shown by the signal magnet.

Discuss the variations in rate and magnitude of the phases
of the single contractions at the different temperatures.

Since the temperature of cold-blooded animals depends
upon the temperature of the surrounding medium, it is much
more variable than that of the warm-blooded animal. What-
ever can be said regarding an optimum temperature for frog's
muscle can be applied in a modified form-to mammalian muscle.
The latter, however, is scarcely ever subjected to as great a range
of temperature and the optimum is higher.

If one goes beyond the temperature at which the muscle
remains irritable certain changes will take place due to irre-
versible modifications in the muscle cells. The shortening
of the muscle fibre due to coagulation of the muscle proteins
can be shown by securing a record, upon a very slowly moving
drum, of the effect of a gradual increase of temperature. De-
scribe the appearance of a muscle so treated.

Chemical changes can also be shown by comparing the
reaction to litmus paper of the cut surface of the muscle with
that of a fresh muscle.

LOAD AND WORK.

The amount of work which a muscle can do depends upon the
load which it carries. If a load is light the muscle does not accom-
plish a great deal, even though it contracts to its full extent. On
the other hand too great a load overtaxes the muscle so that the
load can scarcely be lifted and thus little work performed. There
is then for every muscle an optimum load. With this the muscle
is able to do the greatest amount of work at each contraction.
Experiment 11. — Arrange for direct stimulation of a gastro-
cnemius muscle fastened to the lever of the tripod. Adjust
the after-loading screw so that the muscle picks up the load in
the large scale pan as it starts to contract. Choose a stimulus
which is approximately maximal. The records are to be super-

32 EXPERIMENTAL PHYSIOLOGY.

imposed upon a rapid kymograph, so that after once starting
to record contractions, do not move the relative position of
the apparatus. A signal magnet is used to indicate the
latent period. One tuning fork record does for the whole set.
Using increments of ten or twenty grams, depending upon the
size of the muscle, increase the load until the muscle can no
longer lift it. Obtain a record at each increase in the load.

Compare the latent period, contraction period and relaxation
period as the load increases.

Calculate the work done in gram-millimeters at each con-
traction by multiplying the load by the vertical distance
through which the load was moved. The latter is to be deter-
mined by dividing the height of contraction as recorded on
the drum by the magnification of the lever. Now plot a curve
of work and load, with work as ordinate and load as abscissa..

What is the relation of the magnitude of the load to the work
accomplished?

INFLUENCE OF VERATRINE UPON CONTRACTION.

It is possible with certain drugs to affect one phase of the con-
traction in a muscle without materially modifying the other.
Veratrine and to some extent glycerol and nicotine produce this
effect.

*Experiment 12. — Prepare one gastrocnemius muscle of a frog
without injury to the circulation of the other gastrocnemius.
Inject a few drops of 0.1% veratrine acetate solution into the
dorsal lymph sac of the frog as soon as the first muscle has been
prepared and removed so that the other gastrocnemius may get
a sufficient amount of the drug by the time that it is wanted.
Secure a record of the contraction of the muscle first pre-
pared, using the spring-driven kymograph. Now prepare the
second gastrocnemius and obtain a contraction in a similar
fashion. From time to time take other records to discover
when the veratrine effect disappears.

Compare the contractions of the normal and veratrinized
muscles.

VOLUNTARY MUSCLE. 33

FATIGUE. —M^—

It is well known that excessive WEBSRl SBSBB
use of a muscle reduces its efficiency. j^SSSSl rW^JrltinB
In fact all phases of contraction are
affected. A careful study of the sub-
ject shows the following changes.
Repeated contraction at first causes an
increase in response, called the "stair-
case" or "treppe" effect. A maximum
is soon reached and persists for a time.
Then the contractions gradually de-
crease in magnitude until a point is
reached where the muscle fails to re-
spond (Fig. 9). In the latter stage

of fatigue the muscle may fail to relax •nnMiiiiHiiiMiiiifc •jiii
entirely between twitches so that it ^^^K'r/iIWi !K**iBim
persists in a state of "contracture."

It has been shown that certain
substances are produced during
muscular activity such as carbon-
dioxide, mono-potassium phosphate
and para-lactic acid. If these sub-
stances are injected into a fresh
muscle the first effect is to augment
the contractile response. Injection
of larger amounts reverses the effect
so that the response is reduced. In

other words it is possible to induce in MD i|BIIDS*SW^
a fresh muscle the various stages of HHf !»HlSB«iiJHsl
excitability and fatigue by means of
the waste substances produced in BBl IIISSIIB^^SB
muscle, the effect depending upon HiiilliUll
the quantity of these substances.
These observations lead one to con-
clude that the accumulation of waste IkVXV

^•k'vNt^H^SiKl ^ u

substances in a muscle is a large BBHfci T' iBSit Wl

factor in producing these conditions.
This is further substantiated by the
fact that increasing the circulation to

1 1

34 EXPERIMENTAL PHYSIOLOGY.

a muscle delays the onset of fatigue and prolongs the staircase
effect.

In frog's muscle fatigue is accompanied by a lengthening of
the time for contraction and relaxation, while in mammalian
muscle the contractions merely become shorter, with little effect
upon the duration. The using-up of materials stored in muscle
also contributes to fatigue, for in starvation fatigue is induced
more readily while the feeding of glucose helps to prevent it.

Effect of Fatigue upon Contraction. — Fatigue is partially
due to the accumulation of waste substances, therefore any re-
duction in the circulation will hasten its onset. In the ordinary
muscle preparation which has been removed from the circulation,
fatigue develops quickly and is easily studied.

Experiment 13. — Pith the brain of a frog taking care to cause as
little bleeding as possible. Make a small incision in the skin
of the heels and free the gastrocnemius tendons from the ankles.
Do not skin the legs. Fasten the frog to the frog board so that
the knees are held firmly, but the circulation to the lower legs is
not occluded. This may be done by passing a hook through the
ligaments on the convexity of the knee. Occlude the circula-
tion in the right leg by tying a mass ligature about the thigh
and connect the tendon of the right leg by means of a
thread to a lever arranged to record contractions of the
gastrocnemius. Connect one terminal of the secondary coil
to the tendon with a piece of the wire; connect the other
terminal to a wire looped about the knee. Arrange to
stimulate with maximal make and break shocks. Adjust
the after-loading screw and load the muscle with 30
grams. Because the first few contractions illustrate important
phenomena, do not stimulate the muscle until ready to record
the series of contractions resulting in fatigue.

Start the kymograph revolving slowly and stimulate as
uniformly as possible with alternating make and break shocks
at second intervals until the muscles cease to respond. Allow the
muscle to rest five minutes and record a second fatigue tracing.
The record should illustrate the phenomena of (1) Treppe,
(2) Fatigue, (3) Contracture. How do you explain Treppe and
Fatigue in terms of the All or None law? What change in the
behaviour of the muscle is responsible for contracture?

VOLUNTARY MUSCLE. 35

Repeat the experiment using the gastrocnemius of the left
leg, the circulation of which is not disturbed. Compare Treppe,
Fatigue, Contracture and Rest in the circulated and non-
circulated muscle. The circulated muscle may reach a point
beyond which fatigue fails to develop further. This is the
fatigue level. Explain why this occurs. Alter the rate of
stimulation and explain its effect on the height of the fatigue
level.

FIG. 10. Graphic record of two voluntary contractions showing their tetanic nature. Made
by pull of the index finger against a stiff spring at the end of which a writing point is attached.

TETANUS.

Voluntary muscular action is caused by a rapid succession of
impulses travelling outward along a motor nerve. The muscle
therefore does not respond by a simple twitch, but by a more or less
sustained contraction. Even the quickest voluntary movements
are a fusion of several simple contractions.

Several proofs have been obtained of the composite nature of
such contraction. First it has been found by the use of a sensitive
galvanometer that several impulses pass along a nerve which is
exciting a muscle even to brief contraction.

Second, a sound can be heard through a stethoscope applied
to a muscle contracting voluntarily. This sound is no doubt due

36 EXPERIMENTAL PHYSIOLOGY.

to vibrations produced by the rapidly contracting muscle. The
sound heard is usually an overtone because the actual vibration
rate is near the lower limit of audibility.

Third, a magnified record obtained by means of a lever, of a
muscle, under sustained voluntary contraction, is not perfectly
smooth, but contains many fine regularly occurring contractions
superimposed upon the mam contraction (Fig. 10).

FIG. 11. The effects of successive stimuli on skeletal muscle and
cardiac muscle. The vertical marks show where stimuli were introduced.
Tracings at bottom are the result of stimuli sufficiently close together to pro-
duce tetanus. Notice that the heart shows neither summation nor tetanus.
(Compiled from tracings published by T. G. Brodie and Leonard Hill).

Finally, in order to produce a sustained artificially stimulated
contraction resembling a voluntary contraction, it is necessary to
send into the muscle a rapid succession of stimuli. Such a con-
traction is called tetanus.

VOLUNTARY MUSCLE. 37

Summation. — If two stimuli are sent into a muscle in close
succession so that the muscle has not completely relaxed before
the second stimulus is effective, the latter contraction will be
higher than the first. The second contraction adds its effect to
that of the first, therefore the greatest summation will occur when
the second stimulus is effective during the greatest height of the
first contraction (Fig. 11). A certain amount of summation
results even though a maximal stimulus is used. A series of rapidly
repeated stimuli on account of the summation effects, will cause a
much higher contraction than is obtained from a single stimulus.
Analysis of Tetanus. — Voluntary muscle contraction is really
a series of twitches occurring so rapidly that relaxation is incom-
plete or fails to develop. This action can be analyzed best by a
study of two simple twitches occurring close together.
Experiment 14. — Arrange a muscle in the moist chamber for
stimulation by the inductorium. Set an electro-magnetic
signal close to and in line with the writing point of the muscle
lever. Connect the signal and the kymograph in the primary
circuit. Adjust both contacts upon the disk of the kymograph
so that two stimuli are sent into the muscle with each revolu-
tion of the drum. With the contacts far apart obtain a record
of the twitches when the drum is released. Make a series of
records in which the stimuli are closer and closer together, by
adjusting the sliding contact. Continue this series to the
point of complete summation.

In order to observe the effect of more than two successive
stimuli, replace the kymograph by the wheel interrupter in
the circuit. Make records on a slower drum, turning the inter-
rupter at different rates. With each new trial increase the rate
until complete fusion is obtained.

Next replace the wheel interrupter by the vibrating spring
on the inductorium. In this way tetanus can be produced
automatically.

Discuss summation and the development of tetanus.
In order to perform this experiment with the Harvard Kymo-
graph use the highest speed. A key is placed in the primary
circuit but one of the wires is connected to a dissecting needle.
Stimulation is obtained by quickly snapping the needle over one

38 EXPERIMENTAL PHYSIOLOGY.

edge of the binding post on the key. This can be done so quickly
that one shock is effective. Follow directions as in the original
description.

RED AND PALE MUSCLE FIBRES.

Voluntary muscle fibres may be of two kinds, one pale in aspect,
the other red. The red fibres are thinner and possess a larger
amount of sarcoplasm. They possess a greater number of nuclei
and a larger blood supply. Many of the capillaries to red fibres
are furnished with dilatations.

Red fibres contract more slowly and fatigue less readily than
do pale fibres. The former appear to be adapted for heavy work,
while the latter are for rapidity of action.

Some muscles are composed largely of red fibers, such as the
soleus in the rabbit. The gastrocnemius on the other hand con-
tains mostly pale fibres. Quite often both types of fibres are
present in a muscle, but one predominating.

HEAT PRODUCTION.

A muscle releases a considerable amount of energy in the form
of heat when it contracts. Everyone is familiar with the increased
amount of heat after vigorous exercise. Sustained contraction in a
large muscle may raise the temperature 1° C. or more, but due to
the circulation and the control mechanism, excess heat is distributed
and eventually lost.

It has been estimated that CO to 75 per cent, of the energy used
in contraction produces heat, the balance being converted into
work. Unpractised movements result in a greater proportion of
heat production. Training, therefore, is highly important in
getting the greatest work from a muscle.

There are two stages of heat production. The first is immedi-
ate or "explosive" in character and will take place in the absence
of free oxygen. For example, a muscle which contracts in Ringer's
solution from which free oxygen has been removed or in nitrogen,
releases heat of the first stage. The second stage develops slowly
and is postponed if oxygen is absent. The evolution of heat in
this stage continues for a long time after the mechanical response.
The amount of heat evolved is as great as that of the first stage.

VOLUNTARY MUSCLE. 39

RIGOR MORTIS.

Within a few hours after an animal dies, its muscles undergo a
pronounced irreversible change. They contract and lose their
extensibility. The contraction is not vigorous as can be shown by
resisting it with slight tension.

The chemical changes which seem to underlie the process, are
the production of carbon dioxide and lactic acid, which by causing
coagulation of the muscle proteins, myosinogen and paramyosingen,
bring about the contraction. Incomplete oxidation seems to favour
the development of rigor, for if plenty of oxygen is supplied neither
rigor mortis nor lactic acid is present.

Heat is also produced as rigidity comes on, which accounts for
the warmth which may be noticed sometimes hours after death.

Rigor mortis being due to chemical changes, anything which
hastens these causes an earlier development of the condition.
Thus the onset is much earlier at high temperatures; low tempera-
tures, conversely, postpone. Fatigue, by speeding up the produc-
tion of lactic acid and other substances, causes an earlier appear-
ance of rigor.

This coagulation of the muscle in rigor mortis may last for two
or three days, when it is terminated by autoly —

Coagulation of muscle proteins may be caused by heat. If a
muscle is gradually heated two stages are found to be present in
the development of rigor, the first at 39° C. in the frog and 47° C.
in the mammal, the second at 50° C. in the frog and 62° C. in the
mammal. The first is due to the muscle proteins proper and the
second to the connective tissue substances.

Heat rigor is more complete and does not disappear when auto-
lysis supervenes.

Similarly, clotting of the muscle proteins may be produced by
alcohol, chloroform or other substances.

ELECTRICAL CHANGES IN MUSCLE.

Electrical changes accompany the contraction of muscle. This
can be shown by connecting a sensitive galvanometer to a muscle
which is caused to contract voluntarily or through artificial stimu-
lation. A wave of negative variation travels from the point

40 EXPERIMENTAL PHYSIOLOGY.

stimulated, whether it be the motor end plate or one end of the
fibre in contact with stimulating electrodes. All other parts of
the muscle are positive in relation to this wave. Suppose as in
Fig. 12 the galvanometer G is connected to the muscle at 2 and
3 and the stimulating electrodes are at 1. A single contraction is
initiated at 1. This contraction is preceded by a negative electrical
condition which first reaches 2 (A), at that moment 2 will be
negative in relation to 3, causing a deflection of the galvanometer.
The wave continues to move onward, quickly passing 2 and soon
reaching 3. When 3 is reached (B) it will become negative to 2.
There will therefore be a reversal of the galvanometer. The gal-
vanometer moves first in one direction and then in the other. This
is called a diphasic variation. The electrical change attending the
contraction of muscle is designated the ACTION CURRENT.

When a muscle is cut or injured, that particular region becomes
electronegative to other parts of the muscle which are uninjured.
If connected by a conductor a slight current will be set up, called

the CURRENT OF INJURY.

FIG. 12. Diagram to show the passage of an electrical change over muscle. When
a muscle is stimulated at one end (i) a negative electrical change is started, passing
along the muscle, just preceding the contraction. A galvanometer (G) is connected
with the muscle by two leading off electrodes, 2 and 3. As the negative change reaches
2 (A) the current will pass through the galvanometer from 3 to 2. But when the
negative variation reaches 3 (B) the current is reversed. This is called the diphasic
variation.

THE NERVE-MUSCLE AS A RHEOSCOPE.

The electrical changes set up in muscle can be detected by a
vigorous nerve-muscle preparation. The nerve is permitted to
touch the muscle at a negative region and a positive region. In
a freshly cut muscle at the time of making the second contact,
the rheoscopic muscle will twitch. In a muscle stimulated through
its nerve the rheoscopic muscle will twitch at each twitch of the

VOLUNTARY MUSCLE 41

tested muscle. The explanation is that the electrical changes
in the tested muscle are strong enough to initiate impulses in the
nerve of the rheoscopic preparation. However, due to the poor
condition of the rheoscopic nerve-muscle, the experiment frequently
fails.

The sciatic-gastrocnemius preparation to be used as a rheoscope
should be taken from a very healthy frog and dissected out with
the greatest care. Abuse of the nerve may destroy its sensitiveness
so that a new preparation must be made.

Experiment 15.— Current of Injury.— One end of a gastroc-
nemius muscle is cut across transversely. This and the rheo-
scopic preparation are placed side by side on a clean glass plate.
The rheoscopic nerve, near its muscle, is placed in contact with
the uninjured surface of the muscle to be tested, the rest of the
nerve being held apart by means of a glass hook. Now the tip
of the rheoscopic nerve is allowed to touch the injured surface
of the muscle to be tested. A contraction of the rheoscopic
muscle should result at the moment of the nerve's contact with
the injured surface.

Current of Action. — Two nerve-muscle preparations are laid
side by side on a glass plate. The nerve of the one which is to
serve as the rheoscope is placed lengthwise against the muscle
to be tested and is then lifted by a glass rod at its middle so
that it is in contact with the muscle only at both ends. The
nerve of the muscle to be treated is next stimulated by single
shocks. With each twitch the rheoscopic muscle should re-
spond. If the test muscle is tetanized, the rheoscopic muscle
should be tetanized.

A secondary twitch can also be obtained by laying a freshly
prepared nerve upon a beating frog heart. The two regions of
contact for the nerves are the base and the apex, the middle
part of the nerve being held away from the contracting muscle
as before. Each beat should, produce a twitch of the gastroc-
nemius.

CHAPTER III.

SMOOTH MUSCLE AND CILIATED CELLS.
SMOOTH MUSCLE.

There are two types of muscle which are independent of volun-
tary action, cardiac muscle which is striated and smooth muscle
which lacks transverse striations. Smooth muscle fibres are
elongated and pointed at both ends. They are generally collected
into bundles. These bundles are attached at their ends to the
membranous parts where they occur.

The importance of plain or smooth muscle will be realized when
its wide distribution is considered. It is found in the lower half
of the gullet, the stomach, and intestines. In the alimentary canal
it occurs not only in the muscular coat, but as a layer in the mucous
membrane and in the villi. It is present likewise in the trachea,
bronchial tubes, bladder, ureters, uterus, glandular ducts, genital
organs, spleen, ciliary muscle and iris. The contractile element in
blood vessels consists of plain muscle. Thus it is by means of
smooth muscle that blood is shifted from one part to another and
emergencies are met. This adjustment is controlled largely through
the nervous system.

Movements of the alimentary canal, mixing of the digestive
fluids with the food and onward propulsion of the contents, are
brought about by smooth muscle. Its function in regulating blood
supply and the mechanics of digestion are alone sufficient to
demonstrate its necessity in the organism.

Extensibility and Elasticity. — -The extensibility and elas-
ticity are similar to those properties in striated muscle. However
in the stomach, bladder and uterus the extensibility is much greater.
The capacity of these organs depends upon the power of extension,
for when empty they are contracted to a small size.

Irritability. — Electrical stimulation is relatively ineffective

42

SMOOTH MUSCLE. 43

with plain muscle, strong currents being required. Induction
shocks are less effective than galvanic.

Mechanical stimulation in the form of stretching is by far the
most adequate form of stimulus, at least among artificial stimuli.

Contraction. — The LATENT PERIOD for smooth muscle response
may be as much as one hundred to five hundred times that of
striated muscle. In the frog's stomach it is from one to ten seconds;
for the cat's bladder, 0.25 sec., and for vascular muscles, 0.3 to
0.5 sec.

The CONTRACTION PERIOD may be as much as 15 to 20 seconds
for the frog's stomach. The amount and duration of the con-
traction depends upon the stimulus. In the frog's stomach a
single contraction may decrease the stomach by 45%, while tetanus
may reduce it 59%. Although the contraction may be large, it is
much gentler than that of voluntary muscle.

RELAXATION PERIOD. — -The time consumed in relaxation is much
longer than that for contraction. In the frog's stomach it is said
to be from 60 to 80 sec.

Summation and Tetanus. — Two successive stimuli properly
spaced will cause summation. A series of stimuli will cause tetanus.
To do this they need not be very frequent. A stimulus every five
seconds is sufficient in the frog's stomach.

Tone. — Smooth muscle possesses the power of remaining in a
condition of persistent shortening for long periods. Thus hollow
viscera can adapt themselves to their contents. Tone can be
varied in different ways. Cooling increases it, while heat decreases
it. The most important means of controlling tone seems to be
through the extrinsic nerve supply, e.g., stimulation of the vagus
increases stomach tone, cutting the vagus produces flabbiness in
the same organ . Throughout life the walls of the arteries resist a high
pressure, this resistance being controlled by the nervous system.

Rhythmicity. — Smooth muscle in certain parts of the body
is able to undergo rhythmical contraction. These are the alimen-
tary canal, ureter, bladder, spleen and blood vessels. In the sto-
mach and intestine rhythmical activity is induced by tension. The
rate of these contractions is somewhat as follows: stomach, 3 per
min., intestine 12 per min., spleen, 1 per min.

In this connection it is important to make it clear that these

44 EXPERIMENTAL PHYSIOLOGY.

rhythmic movements are superimposed upon whatever condition

of tone prevails.

The intrinsic nerve supply seems to preside over rhythmicity.

Experiment 16. — Contraction of Smooth Muscle. — An L-
shaped glass rod is fastened by a clamp so that it can be used
to support a piece of intestine about 2 cm. long in a small beaker
of Ringer's solution. The latter is maintained at a tempera-
ture of 37-38° C. by immersing the beaker in a large tin cup
used as a water-bath. Air is allowed to bubble slowly through
the Ringer's or Locke's solution1 by fixing a small glass tube,
connecting with the compressed air supply, so that its opening
is near the bottom of the beaker. When all is ready one end
of a piece of intestine, cut from a rabbit which has just been
killed,2 is attached to the hook of the glass rod and the other
to a heart-lever which is arranged to record on a slowly-moving
drum. Contractions of the longitudinal muscle will produce
movement of the lever.

After recording several normal contractions try the effect
of temporarily reducing the air supply. Later allow the tem-
perature of the Ringer's fluid to fall to that of the room and note
the changes in tone and rhythmic contractions. Finally study
the effect of stimulation by electricity. When ready to stimu-
late remove the Ringer's solution for a short time in order to
prevent short circuiting. Try make and break, direct and
indirect currents. Later attempt to tetanize the segment.
On account of the long latent period do not run the stimuli too
close together when single shocks are used.

Observe the direction in which a contraction travels when a
piece of intestine is pinched with forceps. In order to remember
which is the oral end a ligature should be tied around that end
when the segment is removed. Do the contractions always
move in the same direction from mechanical stimulation? Is
this type of stimulus more or less adequate than the electrical?

water with which these solutions is made up must be pure distilled. It
has been our experience in this laboratory that chlorinated lake water cannot be
used to distil from. We are compelled to use spring water from which to pre-
pare the distilled water.

2The intestine, after removal from the rabbit, is preserved in cold Locke's
solution through which oxygen or air is occasionally bubbled.

SMOOTH MUSCLE 45

CILIATED CELLS.

In addition to the muscular tissue of the body there are other

contractile elements, viz., ciliated cells.

The function of ciliated cells is to remove mucous and foreign

bodies from passages. They are therefore found in the mucous

membranes of the trachea, larynx, bronchi, nos.e, lachrymal duct,

uterus, Fallopian tubes, tubules of the epididymis, Eustachian tubes

and middle ear.

Each cell may have one or more cilia. Not only the cilia of the

same cell, but those of neighbouring cells, move in a definite

sequence, so that there is co-ordination.

The power of these cilia acting together is considerable. It is

estimated that the cilia on 1 sq. cm. of the frog's oesophagus cap

move a weight of more than 300 grams.

Warmth increases the activity of cilia; cold decreases it. Drugs

also influence their movement.

Experiment 17. — Ciliary Action. — Remove the lower jaw of a
frog and slit the oesophagus. Spread open the oesophagus with
pins. Determine the time required for a bit of cork placed
on the roof of the mouth to be moved one centimeter down the
gullet. Keep the mucous membrane moistened with salt solu-
tion. Pour iso tonic NaCl solution, warmed to 30° C. over
the membrane, drain it off, then determine the rate of move-
ment of a bit of cork. After a time blow ether vapour over the
membrane and again determine the rate of ciliary action.

CHAPTER IV.
PHYSIOLOGY OF NERVE.

All protoplasm possesses the fundamental property of irrita-
bility, i.e., it can be influenced by external stimuli and the effect
of such stimuli can be transmitted to parts at a distance from the
point of application. In animals one kind of tissue has developed
this property to a marked degree, namely nervous tissues. Con-
duction of the impulse aroused by stimulating a nerve is so rapid
that through the mediation of the nervous system, integration of

the organism is brought
about. The animal acts as
a unit not as a composite
of unrelated parts. Nerve
fibres are similar to the
~^ wires of a telegraph system

which connect the different
parts of a country with

FIG. 13. Dissection of the frog's gracilis muscle , tu pllt tup wl'rPc

to demonstrate that a nerve conducts in both direc- eacn Otner. ^UL ^ WIT
tions. Stimulation at (a) will cause contraction of j i

both(a)and(&)orru*wrjo. and communication be-

tween parts is lost. Cut

the nerves to an organ and it loses its coordination with the rest
of the body.

Just as in a wire so in a nerve fibre, conduction can take place
in both directions. This can be shown by means of a galvano-
meter. Stimulation of a fibre in the middle of its course will
cause a deflection of a galvanometer no matter at which end it is
placed. The following experiment also proves the truth of the
statement.

Experiment 18.— Kuehne's gracilis experiment. — Expose the
inner surface of a gracilis muscle in a frog. It will be seen that
the two portions of the muscle are fed by branches of the same
nerve. Separate the two parts of the muscle without injury
to the nerves. It is possible to do this so that the nerve is the
only connection between the two parts (Fig. 13).

46

NERVE 47

Stimulation of either branch of the nerve will cause con-
traction of both muscle masses. In one case the impulse must
travel to the right at the junction of the two nerves and in the
other to the left. Therefore an impulse can pass in either
direction in a nerve.

Nerve fibres are classified into two groups, depending upon the
direction in which the impulses usually travel. Those which carry
toward the brain and cord are called afferent, while those carrying
impulses in the opposite direction are designated as efferent. The
latter activate muscles and glands. Each type of fibre can conduct
in both directions, but in their normal location, conditions are so
arranged that impulses generally start only at one end — in
afferent nerves at the peripheral end, in efferent nerves at the
central end. Even though an efferent nerve be stimulated
at its peripheral end, the impulse travels only as far as the synapse
or connection with the next nerve cell or neurone. In other words
the synapse allows the passage of impulses in one direction only—
in sensory nerves, inward, i.e., towards the central nervous system;
in motor nerves, outward; i.e., towards the periphery.

When we consider the necessity of careful insulation to prevent
the spreading of an electric current in contiguous wires, it is ex-
tremely interesting to know that there is complete isolation of
impulses travelling along contiguous nerve fibres. If this were not
the case, endless confusion would result where great numbers of
both afferent and efferent fibres are closely bound up in the same
nerve trunk.

Experiment 19. — The object of this experiment is to determine
whether nerve impulses travelling over certain fibres in a nerve
trunk spread to all the fibres in that trunk or remian insulated
in fibres with which they are travelling on entering the trunk.
Pith a frog, remove the viscera and the skin of the legs. Lay
it on its back on a glass plate. The various nerves which make
up the sacral plexus may each be stimulated by unipolar induc-
tion. To do this connect the tissues of the back, by means of the
plate electrode and a wire, to the gas or water pipes leading to
the ground.

A. Attach to another wire a pithing needle and fasten the
wire to one post of the secondary coil of the inductorium. With

48 EXPERIMENTAL PHYSIOLOGY.

the inductorium arranged for minimal tetanizing currents touch
the various sacral nerves with the needle and observe whether
the same or different muscles respond in the case of stimulation
of each nerve.

With the aid of figure 4 make a table indicating which
muscles are supplied by each nerve.

B. Cut the trunk of the sciatic nerve high in the thigh and
repeat the observations.

What answer does this experiment give to the question
raised in the 1st Paragraph?

Why is it necessary to do experiment B. before this answer
can be given?

How can you explain excitation by the method of unipolar
induction?

Summation of Inadequate Stimuli.

An impulse passing along a nerve produces a change in the
nerve whether it is sufficient to cause a visible response in its
muscle or not. The nerve is left for a very brief period in a more
sensitive condition so that a stimulus which is insufficient when
applied once, may bring the end-organ into action if applied several
times in close succession.

Experiment 20. — Choose a stimulus which is just below the
threshold of that necessary to cause contraction of a muscle
through its nerve. Now apply a rapid series of these stimuli.

Velocity of the Nervous Impulse.

The time required for the passage of an impulse along a nerve
trunk can be determined by means of a sensitive galvanometer or
by the method described in the experiment below where the response
of a muscle is used.

Helmholtz, by means of a myogram of the thenar muscle when
stimulated through the median nerve at the axilla and then at
the wrist, estimated that the rate in man was 30-35 meters per
second.

Piper, by means of the string-galvanometer, has studied the
same nerve and found the rate to be as high as 125 meters per
second.

NERVE. 49

Non-medullated fibres seem to conduct more slowly, the rate

being as low as 8 meters per second.

Experiment 21. — Although the rate of transmission of the nerve
impulse is rapid, it can be determined by a high speed kymo-
graph. Prepare a gastrocnemius muscle with the nerve as
long as possible. Clamp the femur in the moist chamber, then

FIG. 14. Tracing to show rate of conduction of the nervous impulse, (i) Contraction
of the gastrocnemius when the electrodes are near the muscle; (2) contraction resulting
from stimulation at the far position. Stimulation at X.

lay the nerve across two sets of cork-held electrodes. One
pair should be close to the muscle and the other as far away as
possible. Avoid stretching the nerve. The electromagnetic
signal is placed in line with the point of the muscle lever
and alignment marks made on the smoke-drum. The signal
and kymograph are connected in the primary circuit. Adjust

50 EXPERIMENTAL PHYSIOLOGY.

the muscle lever so that only the beginning of the contraction
is recorded.

Using the base line each time, record the interval of the latent
period, first for the far position of the electrodes and then for
the near position. This can be done by changing the wires of
the secondary over to the electrode desired in each case. Esti-
mate the duration of the latent period by means of a tuning
fork tracing (Fig. 14).

The time required for the passage of the impulse along the
nerve is the difference between the latent periods resulting from
the far and near stimulations. The length of nerve traversed
may be taken as the distance between the nearer points of the
two electrodes.

Determine the velocity of the nerve impulse in meters per
second. Make four or more sets of records. Tabulate the
results. (This expt. may also be done with the Harvard drum
as in Expt. 8.)

FACTORS INFLUENCING NERVE FUNCTION.

Mechanical. — If a nerve is stretched or compressed its power
to conduct may be lost. When the compression is neither too
severe nor too prolonged conductivity may be re-established.
Compression over a broad area requires much greater force to
paralyze than when over a narrow region. The first effect of com-
pression, frequently, is to increase the excitability. This stage,
however, is quickly passed.

Thermal. — Beginning with 0° C. the rate of conduction in-
creases as the temperature is raised. Below 0° C. conduction is
suspended. At the higher limit of conduction, about 47° C. in
the mammal, due to coagulation of the proteins the nerve may
become permanently paralyzed.

Chemical. — Many drugs lower or suspend conductivity when
applied locally. Ether, chloroform, chloral, cocaine, phenol and
alcohol have this power. Conduction will return in time if the
drug does not act too long. Lack of oxygen will also destroy con-
ductivity in time.
*Experiment 22.— The Action of Carbon Dioxide, Ether, and

Chloroform upon Nerve.— A gastrocnemius muscle with its

NERVE. 51

nerve attached is fastened to a lever so that its contraction can
be recorded. The nerve is passed through the two openings
in the bottom of the gas chamber. These holes are made air-
tight with a paste made of kaolin and isotonic NaCl solution.
One pair of electrodes is set so that the nerve within the chamber
can be stimulated, another touches the nerve at the end farthest
from the muscle. Minimal break shocks are to be used for
stimulation. Carbon dioxide is passed through the chamber,
stimuli being applied from time to time at both electrodes in
order to discover changes in the response.

Wash out the carbon dioxide by passing air through the
chamber. When the nerve is again normal blow ether vapour
through until it affects the nerve, or if necessary paint the
nerve with ether. Do not prolong anaesthesia too long or it
will be difficult for the nerve to recover. In a like manner
study the influence of chloroform.

Electrical. — When a direct current is passed through a nerve
the excitability around the kathode is increased, while around the
anode it is decreased. The changed condition produced by the
direct current is called electrotonus — katelectrotonus in the first
case and anelectrotonus in the second case. Midway between the
two electrodes there is a point where the excitability of the nerve
has not been changed. This indifferent point moves toward the
kathode as the current is increased and in the opposite direction
as the current is decreased. This can be shown by stimulating
the nerve along its course during the passing of the direct current.
Experiment 23. — Electrotonus in Nerve. — When a' direct
current is to be passed through a tissue for any length of time
non-polarizable electrodes must be used as the ordinary elec-
trodes are easily polarized, thus altering the efficiency of the
current. The former consists of two boot-shaped porous
receptacles which are partly filled with a 10% ZnSO4 solution.
A piece of pure zinc attached to the source of current is inserted
into each boot. The boot is kept moist on the outside by
isotonic NaCl solution, the porous material acting as a barrier
to the passage of zinc sulphate during the time usually con-
sumed in an experiment.

The boots must be kept clean. Each time after using they

52

EXPERIMENTAL PHYSIOLOGY.

should be thoroughly washed with water and then soaked in
isotonic NaCl solution. When in use the ZnSO4 must not be
allowed to spill over as it is injurious to the nerve.

The boots are held in position by the clips on the horizontal
rod of the moist chamber base (Figs. 15 and 16).

A gastrocnemius muscle, with a long fresh nerve attached, is

NERVE.

53

arranged in a moist chamber with the nerve lying on a pair oi
electrodes in cork near the muscle and a pair of boot electrodes
farther out. Adjust the induction coil so that a break shock
just causes contraction of the muscle through the electrodes
in cork. Record the height of this contraction either on a
slow or stationary drum. Next connect the boot electrodes

FIG. 16. Top view of moist chamber in the preceding figure, showing the
arrangement of the electrodes (c) from the secondary coil in relation to the boot
electrodes. Nerve (h) is shown across the two sets of electrodes. (6) connects with
secondary, (a) connects with direct current supply.

to the current from the rheostat so that the kathode is nearer
the muscle. Try two volts at first. Determine which boot is
kathode and which is anode by dipping the two wires into
NaCl solution ; bubbles of hydrogen will issue from the kathode.
While the constant current is flowing through the nerve by
way of the boot electrodes, stimulate the nerve with the same

54 EXPERIMENTAL PHYSIOLOGY.

break induced shock as before. Then change the wires to the
boots so that the anode is nearer the muscle, and stimulate with
the same break induction shock.

Increase the direct current and test the condition at the
kathode and anode as before. A certain strength of current is
necessary to produce marked changes in the region of the
kathode and anode. It is possible to ^ompletelyj^pTock the
passage of impulses by means of a direct current in the anodal
region. The reverse effect is obtained in the kathodal region.

These electrotonic changes in the nerve only last for a short
time after which the reverse effect may occur. By reversal it is
meant that, e.g., around the kathode the stage of increased
excitability finally gives way to a condition of lowered excita-
bility. This reversal developes more quickly with a strong
current.

STIMULATION BY CONTINUOUS DIRECT CURRENT.

A direct electric current will produce stimulation of a nerve
or muscle when there is a quick change in intensity. This occurs
at the make or break, but apparently not during the passage of
the current. That there is an effect during the passage of the
current is shown in the following experiment.

Experiment 24. — Cut the skin on the ventral surface of a pithed
frog so that the sheet of abdominal muscles is disclosed. The
rectus abdominis is divided into right and left halves by a
longitudinal band of connective tissue, the linea alba. Each
half is further subdivided into bellies by five transverse tendi-
nous intersections. Place one electrode at the anterior end
of either the right or left abdominis, and the other at the
posterior end, having a reversing key in the circuit. It will
be found that at the kathode of each belly, i.e., where the
current leaves the muscle, there is a slight ridge due to con-
traction of the muscle as long as the current is passing. If the
direction of the current is reversed the contraction again occurs
at the kathode.

It can be shown that the stimulus caused by making a direct
current arises at the kathode, while that caused at the break
arises at the anode. If the kathode and anode are placed far apart

NERVE. 55

on a motor nerve the latent period for contraction of the muscle
is greater at make when the kathode is in the far position, than
when it is in the near position. Likewise at the break the latent
period is greater when the anode is in the far position than when
in the near position. Therefore the galvanic make stimulus is
kathodal and the break stimulus anodal.

Electrotonus, Kathodal and Anodal Contractions in the

Heart. — Because of the regularly occurring contractions the heart

is an admirable muscle for the study of electrotonic changes as

well as the effect of kathodal and anodal stimulation.

*Experiment 25. — Destroy the brain of a frog and expose the heart

with as little hemorrhage as possible. Two boot electrodes are

prepared by attaching a tiny strip of cotton wool to each at

the toe. The cotton is soaked with isotonic NaCl solution

and serves as a contact. The kathode is made to touch the

frog's mouth while the anode touches the ventricle. Now if the

current is closed during contraction of the ventricle, it will be

seen that at the moment of closing the latter fails to contract

in the region of the anode. This is due to the anelectrotonus

set up around the anode. Of course only a limited area will be

affected, so that very close observation will be necessary.*

Now if the current is broken during diastole the ventricle
will contract slightly in the neighbourhood of the anode.
This is the anodal opening contraction.

The condition produced in muscle by the kathode can be
studied by changing the electrodes so that the kathode touches
the ventricle and the anode the mouth. At the closing of the
circuit the ventricle contracts in the region of the kathode, if
it is in diastole. This is called the kathodal closing contrac-
tion. If the current is passing during systole there may be a
small area of heightened contraction around the kathode.
This is due to the katelectrotonus.

The response at make and break depends upon the strength
of current and upon whether the kathode or the anode is next to
the muscle.

With the KATHODE NEAR THE MUSCLE a contraction is obtained

*The local contraction or failure of contraction is revealed by a change in
color due to variations in the blood content of the heart tissue as the affected part.

56 EXPERIMENTAL PHYSIOLOGY.

at the make whether the current be weak, medium or strong.
But with this position of the kathode, contraction is obtained at
the break when only a medium current is used. The break stimulus
always being less effective than the make, its effect, when a very
weak current is used, will be below the threshold of stimulation.
On the other hand when very strong currents are used, the break
stimulus being anodal, is blocked in the nerve by the development
of electrotonic changes, so that the impulse does not reach the
muscle.

When the ANODE is NEAR THE MUSCLE, a weak current causes a
contraction at the make, but has no effect at break, not having
reached the threshold. With medium currents both make and
break are effective. In the case of strong currents, the break is
effective because it occurs at the anode, which is near the muscle
with nothing to block it. The make occurring at the kathode
has to traverse the whole region of the anode at the time when
the anode produces its greatest block to conduction, i.e., immedi-
ately after making the current.

We do not know the nature of the nerve impulse, but whatever
it is, an electrical change always accompanies it. This can be
shown by connecting a sensitive galvanometer to the nerve. For
the same reasons as in the muscle, the galvanometer will show a
DIPHASIC VARIATION, that is a deflection, first in one direction and
then in the opposite direction.

METABOLISM IN NERVE.

The metabolism in nerve is so slight that it is very difficult to
obtain indications that such changes are going on.

So far it has been impossible to demonstrate the production
of heat in nerve. Hill has used a thermo-electric couple sensitive

to ioo^boiooo °c without a posit!ve result'

Every living cell respires, i.e., it is constantly producing CO2 but
cells manifesting increased activity such as a contracting muscle
show an increase in CO2 production. Tashiro claims that the
quantity of CO2 produced by a nerve in action is more than double
that produced by a resting nerve. If this is correct it indicates
metabolic changes during the passage of the nervous impulse.

NERVE. 57

The necessity for oxygen is a further indication of increased meta-
bolism during activity. Nevertheless, nerve must use an infini-
tesimal amount of energy in its work, not only in view of the above
considerations, but because it is very difficult to fatigue.

A nerve has been stimulated continuously for ten hours and

still transmitted impulses, as could be shown by a galvanometer

or by preventing the fatigue of the muscle which is done by blocking

the contraction with curare, cold or a narcotic. Then at the end

of the time, the block being removed, the muscle contracted.

Experiment 26.— The Resistance of Nerve to Fatigue.—

Arrange two nerve muscle preparations so that they support

levers in such a position that they can write one above the

other upon the same drum. Both nerves are to lie side by side

on the same pair of electrodes. Abolish the conductivity of

one nerve by keeping it moistened with ether. Keep the nerve

moist by improvising a moist chamber from filter paper.

Stimulate the nerves by means of a tetanizing current and

obtain a record of the contraction upon a slowly moving drum.

If sufficient ether is used the muscle on that side should not

contract. As soon as this stage is reached, tetanize both

nerves until the active muscle is exhausted. When this stage

is reached, wash away the ether, with iso tonic NaCl solution,

continuing the stimulation. The muscle upon the treated side

should soon begin to contract.

It can be shown further that the fatigued muscle will
respond by direct stimulation. Therefore if nerve fibres are
still carrying impulses as is shown by response of the other
muscle, the seat of fatigue must be between nerve fibre and
muscle fibre, or at the end plate. The end plate according to this
becomes fatigued much earlier than the muscle fibre and thus
saves the fibre from excessive fatigue.

THE ALL-OR-NONE PRINCIPLE IN NERVE AND MUSCLE.

Stimulation of a muscle either directly or through its nerve pro-
duces a step-like increase in response, when successively increasing
stimuli are used. When once a step has been established further
increase of the stimulus causes no increase of contraction until a
new step is reached. The maximum contraction of the whole

58 EXPERIMENTAL PHYSIOLOGY.

muscle is frequently reached in a few definite steps. The number
of steps is never greater than the number of motor nerve fibres sup-
plying the muscle. The cutaneous dorsi nerve of the frog contains
only eight nerve fibres and each nerve fibre supplies a large num-
ber of muscle fibres, 150 to 200. Lucas found that upon gradually
increasing a stimulus to this nerve from sub-minimal to supra-
maximal that there were never more than eight steps in the height
of muscular contraction.

Experiment 27.— Adrian's Experiment.— The object of this
experiment is to determine whether a nerve impulse which is
reduced in strength by passing through an area of narcosis can
regain its strength on entering a normal part of the nerve fibre.
A large frog should be selected and from it two nerve muscle
preparations made, taking care not to injure the nerves and to
procure their entire length from their origin in the cord.

It is unnecessary to arrange recording apparatus for this
experiment. The narcotizing chamber, Fig. 16a, should be
carefully dried and placed on a dry glass plate. The glass dome
of the moist chamber should be lined with moist filter paper
so that it may be used to cover the preparation during the
experiment. Arrange inductorium so that single induced shocks
may be obtained from the electrodes. The nerves are to be
laid along the grooves of the narcotizing chamber, their proximal
ends being in vaseline at the points where the grooves open into
the circular cups, and care taken that the grooves are dry so
that when the narcotic is introduced into the cups it will not
flow back along the grooves.

When this is done regulate the strength of the stimulus so
that the muscles respond maximally to break shocks, but do not
respond to make shocks. Now introduce enough 15% alcohol
made up in Ringer's fluid into the cups of the moist chamber.
At 20 second interval stimultae the preparations, watching to
see if either of them fail, to contract. Determine how long each
one continues to respond.

When the first preparation fails to respond to a single shock
try sending in a series of shocks in as rapid succession as possible.
If a response occurs it is due to summation in conduction.
How is it explained?

NERVE

59

How do you know it is not a summation in excitation?

At the end of the experiment, if time permits, the prepara-
tions may be replaced in salt solution and the conductivity of
the nerves restored.

FIG. 16A.

The experiment may then be repeated, exchanging the
position of the two preparations so as to be sure that the result
is not due to inequalities in them.

60 EXPERIMENTAL PHYSIOLOGY.

Measure the diameter of the cups.

(a) Is the total length of narcotic through which the impulse
passed greater in the case of the two small cups or of the one
large cup?

(&) Which preparation ceased to respond first?

(c) Does the result indicate that the impulse is extinguished
immediately in a narcotized region or by progressive decrement?

(d) How do you reconcile decrement with the all or none
nature of conduction?

(e) How does the intervening normal area between the two
short regions of narcosis affect the strength of the nerve impulse?

(/) Does the energy of the impulse arise from the stimulus
or from the nerve fibre?

(g) How does this experiment prove the all or none nature
of the nerve impulse?

The passage from one step to another took place without
any contractions of intermediate size. Muscle fibres therefore
contract either not at all or to an extent which is nearly maxi-
mal. This is the all-or-none principle. According to this
principle when a muscle mass is giving a minimal or sub-maximal
•contraction only a certain number of fibres is being called into
action, the rest remaining inactive.

SECTION IIA.
CHAPTER V.

CARDIAC MUSCLE.

Cardiac muscle resembles voluntary muscle in its cross stria-
tion. The individual cells, however, are joined end to end and
many of them interconnected by fork-like projections. Anasto-
mosis is so complete that there is continuity throughout the myo-
cardium of both auricles and ventricles. Even in mammals there
is evidence of a complete syncytium in the form of muscle-fibrils
passing from cell to cell. The connection between the auricles and
ventricles consists of a narrow bundle of somewhat modified
muscular tissue, the bundle of Kent.

Cardiac muscle contracts much more rapidly than smooth
muscle, but more slowly than voluntary muscle.

The heart is involuntary in action, and undergoes rhythmic
contraction throughout life. This rhythmicity is so well estab-
lished that it is impossible to tetanize a herat by artificial stimula-
tion. Herein lies the most essential difference between cardiac
and skeletal muscles. If a stimulus is sent into the heart during
systole there is no result, but when stimulated during diastole
an extra contraction follows. The whole period of systole is
refractory to stimulation. In case an extra contraction is induced
during diastole, the relaxation which follows is prolonged to com-
pensate for the pause which was missed.

All-or-none Principle. — The all-or-none principle applies
to cardiac muscle as well as to voluntary muscle, in fact it was
first observed in the former. Due perhaps to the syncytium-like
structure of the heart muscle, all of its fibres have the same threshold
of stimulation, so if one contracts all contract. Moreover as in
voluntary muscle, those fibres which contract, do so to their full
extent if at all. Therefore the whole heart contracts to the maxi-
mum or not at all.

61

62 EXPERIMENTAL PHYSIOLOGY

Action of Cations on Cardiac Muscle.— The ions found
in the blood play a very important role in the maintenance of the
normal heart beat. Remove one of these essential ions and the
heart finally stops beating.

The SODIUM IONS not only play a large part in the maintenance
of the normal osmotic pressure but when present alone, they pro-
duce relaxation of the heart.

POTASSIUM IONS are present in much smaller quantity than
sodium and are not so essential. They tend to produce relaxation
and when increased above normal the heart rate is reduced, then
finally the heart stops in extreme diastole.

CALCIUM IONS are absolutely essential. Excess of calcium
produces calcium rigor, that is the heart finally stops in systole.
Calcium is supposed to promote contraction, while sodium and
potassium aid in relaxation.

Temperature Effects. — Although the mammalian heart is so
protected in an organism with a relatively constant temperature,
it is influenced by thermal changes if subjected to them. In fact
it acts like the heart of a frog or turtle, except that it does not
withstand the effects of a lowered temperature long. The rate
increases with increase of temperature up to a certain point.
Below 17° C. the mammalian heart does not beat, likewise above
44° C. contraction soon ceases. The heart of cold-blooded animals
responds at lower temperatures.

NERVOUS CONTROL OF THE HEART BEAT.

Inhibition. — It can be shown by stimulation that the vagus
inhibits the activity of the heart. Weak stimuli may merely
decrease the magnitude of the beat. Stronger stimuli slow or even
stop the contractions. Even with stimulation intense enough to
stop the heart, after a time the beats begin to break through or
escape. In the frog, because the vagus and sympathetic fibres run
together in the same trunk, both are stimulated. The stimulus
must be chosen so that the vagus is affected more than the accele-
rator or else there will be either no change or an increase in rate.
This is done by gradually increasing the stimulation until vagal
effects predominate.

CARDIAC MUSCLE. 63

Normally in the frog the vagus carries very few impulses to
the heart, therefore cutting the vagi has little influence upon the
rate. But in mammals the vagi continually hold the heart in
check so that cutting of these nerves permits an increase in rate.
The vagus is reflexly called into action by sensory stimulation.
A good example of this is obtained by sharply striking the abdomen
of a frog with a blunt instrument. Frequently the heart may be
stopped or slowed in this way.

An increased blood pressure will also reflexly act along the vagus
to slow the heart. This is due to the stimulation of afferent fibres
ending in the arch of the aorta. High pressures may also directly
stimulate the vagus centre.

The vagus seems to exercise a beneficial influence on the heart,
tending to oppose acceleration and overwork of the heart. The
vagus is most effective in animals which are accustomed to strenuous
muscular exercise and therefore great activity of the heart.

Acceleration. — Certain fibres, which are called accelerator
fibres, oppose the action of the vagus. They are found in the
cervical sympathetic beloAV the inferior cervical ganglion. Stimu-
lation of these fibres increases the rate of the heart and sometimes
the amplitude as well. On the other hand cutting of these fibres
permits the vagus to have full sway (in the mammal) and thus
the heart beat becomes slower. The accelerators are called into
action reflexly, although it is difficult to say just what stimuli will
affect the accelerators and what will call the vagi into action.

The arrangement of opposed fibres, i.e., accelerator and in-
hibitor, renders quick adjustment to changing requirements
easier.

Experiment 28. — Frog's Heart. — Pith a frog, destroying the
brain only. Expose the heart by slitting the upper part of the
abdomen lengthwise. Avoid injury to the anterior abdominal
vein by cutting a little to one side of the mid-line. Observe
the beating of the heart, especially the rate. Then open the
pericardium and pass a thread under the fraenum and tie.
The fraenum is a slender band of tissue which connects the
posterior surface of the heart with the pericardium. Has this
operation influenced the rate? By means of the thread raise
the heart and notice the large veins, the sinus, the white cres-

64 EXPERIMENTAL PHYSIOLOGY.

centic line at the sino-auricular junction and the auricles.
What is the sequence in the beat? Record the normal heart
beat by connecting the tip of the ventricle to a heart lever.

Prepare for dissection of the vagus by stretching the neck
tissues over a large glass rod inserted into the oesophagus.
On clearing away the connective tissue near the angle of the
jaw three nerves can be found running from below the angle
diagonally downward (Fig. 5). The upper one glosso-pharyn-
geal, soon turns and passes forward; the lower one also passes
forward in a similar manner. Lying between these nerves
before they bend upward can be found the third nerve or vago-
sympathetic lying under the edge of a fine muscle which connects
the angle of the jaw with the hyoid bone. Stimulate the vagus

FIG. 17. Effect of vagus stimulation upon the heart of the turtle. Note the increased
amplitude following the inhibition.

to make sure that the nerve is effective in modifying the heart
rate (Fig. 17). Use the tetanizing current, beginning with a
weak current and gradually increasing if there is no effect.
Bear in mind that the sympathetic is being stimulated at the
same time and therefore its stimulation may counteract the
effects of vagus stimulation. That is the reason that it is
necessary to find a current which will affect the vagus more
than the sympathetic. Obtain a record of the contracting heart
before and after vagal stimulation, using the suspension method
as shown in Fig. 18. Use the magnetic signal to mark the
time of stimulation. If you fail to obtain effects from one
vagus, try the other.

CARDIAC MUSCLE.

65

Next stimulate the white crescentic line with the electrodes.
Inhibition should be obtained.

Apply a few drops of 1% nicotine solution to the heart and
again try the effects of vagus and crescentic stimulation, from
time to time, recording the results on the kymograph. Nico-
tone will paralyze the ganglia. Try Atropine. Explain results.

FIG. 18. Suspension method for heart contraction.

Experiment 29. — Stannius Ligature. — (a) First Ligature. Ex-
pose the heart of a pithed frog in the manner already described.
Pass a thread behind the aorta. Lift up the apex and tie the
thread so that the knot lies on the crescent, thus separating
the sinus from the auricles. If properly done, the auricles and
ventricles soon cease to beat; if not, tie a second thread nearer
the auricles. If the relaxed ventricle be gently pricked it
gives a single contraction. In a short time, however, the
ventricle may begin to beat regularly but at a different rate

66 EXPERIMENTAL PHYSIOLOGY.

from that in the normal heart. A further tightening of the
ligature will again inhibit the beat. Explain results.

Certain properties of cardiac muscle can be studied with
quiescent musc?e which cannot be studied in a heart beating
normally.

Refractory Period.— Arrange to stimulate ventricle with
weakest tetanizing current that is effective. Increase the
strength of the current and record contractions. How does
cardiac muscle differ from skeletal muscle in its response to
rapidly interrupted shocks? Is there any change in the rate
of response as the current is made stronger?

All-or-none Law. — Using the same preparation, make sure
that the beat of the auricle and ventricle is inhibited by the
first Stannius ligature. Arrange the induction coil for single
induction shocks. Apply the electrodes to the ventricle and
find the weakest break shock that will cause a contraction.
Record, then turn the drum about 2 mm. After 15 sec. record
the contraction in response to a slightly increased stimulus.
Turn the drum again and increase the stimulus a little more
and so on until the heart is receiving the maximal stimulus.
Does the height of the contraction under these conditions vary
with the strength of the stimulus? Compare with skeletal
muscle. An interval of 15 sec. is used because a short interval
would give the " staircase" effect. How is this effect to be
reconciled with the "all-or-none law"?

(b) Second Ligature. Place a second ligature around the
heart exactly over the auriculo-ventricular groove. Tighten
this ligature so as to compress the heart between the ventricle
and the auricles. Note that the ventricle immediately begins
to beat, while the auricles continue in the relaxed condition.
Give a reason for this result.

Determine whether it is possible to tetanize cardiac muscle
by stimulating with the tetanizing current. Explain your
results.

Experiment 30. — Study of the Turtle Heart. — A turtle is
pithed in the same way as a frog. The plastron or ventral
portion of the shell is removed by the use of bone forceps and
scalpel. Operate so that hemorrhage is reduced to a minimum.

CARDIAC MUSCLE. 67

Study the action of the heart and arteries. Examine the
pericardium, then remove it. Where does the contraction seem
to originate? What path does it follow in passing over the
heart? Is there any delay in the passage between auricle and
ventricle?

A-V INTERVAL. — In order to observe the time relations between
the contractions of the auricle and ventricle arrange to take
simultaneous records of the two chambers one directly above
the other, using heart levers. Attach the levers to the tip of
each chamber by means of tiny hooked pins on the ends of
threads, but take care not to puncture the wall. By using a
very rapid drum the interval between the contraction of auricle
and ventricle can be determined (Fig. 19).

FIG. 19. Simultaneous tracings of the contraction of the auricle (a) and
ventricle (v) in the turtle. Time interval 5 sec. (contractions slower than normal).

STIMULATION OF THE VAGUS.— Dissect out both vagi high up
in the neck. They can be located near the carotid artery.
Pass threads under the vagi so that they can be lifted when
applying the electrodes.

Study the effect of stimulating each vagus by tetanizing
currents of different strengths. Are the right and left vagus
nerves equally efficient in stopping the heart beat? Do they
both have exactly the same effect? Explain. Is there escape
from inhibition? . Will a weak current alter the A-V interval
without stopping the heart beat? Secure records to demon-

68 EXPERIMENTAL PHYSIOLOGY.

strate the latent period, the duration of the after-effect, changes
in rate, and escapement. (Fig. 17).

INFLUENCE OF TEMPERATURE. — Continuing with the same pre-
paration obtain records with the heart at the following tempera-
tures: 4°, 10°, 20°, and 30° C. These temperatures can be
obtained by pouring Ringer's solution of the corresponding

FIG. 20. Refractory period and compensatory pause in the heart. Stimulation at X.

temperature over the heart. How does temperature influence
the rate and amplitude of the beat? What modification takes
place in the A-V interval?

EXTRA SYSTOLE. — Remove the heart by cutting widely around
the base. Drop it in Ringer's solution. After studying it
there for a short time, pin the base to a frog board and attach

CARDIAC MUSCLE. 69

the apex to a heart lever. Frequently pour Ringer's solution
over the preparation. The writing-point of a magnetic signal
in the primary circuit is exactly aligned with the writing point
of the heart lever. The secondary coil is connected by fine
wires to the base and apex of the heart. A record of the normal
heart is made upon a fairly rapid clock-work drum. Then
single break shocks are sent into the heart during different
phases of the cardiac cycle in an attempt to secure an extra
contraction (Fig. 20).

During what phase can an extra contraction be produced?
What follows an extra contraction?

When you consider that the heart obtains its rest only
between beats, what do you think of this arrangement as a
safeguard against encroachment upon the rest period?

Next, cut the ventricle across near its junction with the
auricles. If the ventricle does not stop beating, cut still closer
until the beats disappear. Now pin the cut surface of the apex
to the board and apply the wire which formerly connected the
base of the heart. Study the effects of a series of graded single
shocks. How does this agree with the all-or-none principle?
Also attempt to get the "staircase" effect. Try the effect of
two successive stimuli in order to discover whether there is a
refractory period.

Experiment 31. — Perfusion of the Turtle Heart. — The differ-
ence of different inorganic salts upon the activity of cardiac
muscle can best be shown by perfusion experiments.

In order that the experiment may have the greatest chance
of success you must understand and prepare the mechanical
arrangements before beginning the operation. A small beaker
containing Ringer's solution is placed on a shelf or stand so
that pressure is obtained for a siphon connecting the contents
with a rubber tube w^hich is long enough to be manipulated
with ease when the time comes to insert the cannula. A small
glass cannula fits into the lower end of the rubber tube, the
flow of solution being controlled by a clamp. Make sure that
the system of tube and cannula is absolutely free from air.
Expose the heart ; then carefully free a short length of the inferior
vena cava without puncturing it. Place a fine ligature around

70 EXPERIMENTAL PHYSIOLOGY.

this. Also pass ligatures around the other veins which supply the
sinus venosus. Now with small forceps pick up the lower end
of the freed portion of the vena cava. With small sharp scissors
cut obliquely downward and forward not quite halfway through
the vein. Holding the vein flap with forceps insert the cannula
and tie the ligature securely about the constriction. Air must
not be allowed in the vein or cannula. Next start a small flow
of Ringer's fluid, through the heart, immediately opening the
aorta, and then tie off all the other veins entering the sinus.
Obtain tracings of the ventricle by means of a heart lever.

Next determine the effect of the following solutions upon the
heart beat. Between each solution, Ringer's fluid must be
perfused until the heart beats normally. When changing the
siphon from one beaker to another keep air from entering by
placing the tip of the finger over the opening.

Solutions to be used are as follows:

1. 0.75% NaCl in distilled water.

2. A solution containing 0.5% NaCl and 0.25% KC1 in
distilled water.

3. 0.75% NaCl in distilled water to which a few drops of a
5% CaC^ solution has been added.

4. Distilled water.

Experiment 31a. Turtle Auricle. — Suspend the auricles of a
turtle as in Fig. 8, p. 30 of Manual, lining the beaker with
moistened filter paper. Record contractions on slow drum.
Does the heart muscle contract automatically? Does the
muscle relax to the same degree between each beat? Do these
changes occur rhythmically? Does the muscle do more work,
i.e., lift the weight of the lever through a greater distance, when
contracting after a greater relaxation or after a less relaxation?
The effect of cations on cardiac muscle is readily shown by
immersing the suspended turtle's auricle in the solutions listed
in the previous experiment.

CHAPTER VI.
BLOOD PRESSURE.

EFFECT OF CHEMICAL SUBSTANCES ON THE BLOOD
VESSELS AND HEART.

The blood stream must be maintained at a certain pressure in
order to furnish the tissues a sufficiently rapid supply of oxygen and
nutrition as well as to carry away waste products. The heart is
directly responsible for the movement of blood, so that any increase
in its output will increase the movement. Such an increase may be
obtained either by a greater rate or a greater amplitude.
Both means are used.

The blood pressure would tend to fall to zero between the con-
tractions of the heart if it were not for the elasticity of the artery
walls. With each beat of the heart the arteries are expanded.
As soon as the force of the heart is spent, the arteries contract, thus
preventing a fall to zero. Fluctuations in pressure are partly
prevented by the resistance offered by the capillaries.

The lowest pressure which is maintained is called the diastolic
pressure, because it occurs during the diastole of the heart. The
pulse pressure is the difference between the systolic and diastolic
pressures. The more elastic the blood vessels, the smaller is the
pulse pressure.

The blood supply to different regions of the body may be in-
creased by a greater output, or by a shifting of the blood from one
region to another. The latter method is made possible by the
presence of vasomotor nerves. Certain nerves cause constriction,
while others cause dilatation. Thus the supply to the muscles
can be increased by constriction of the vessels in the abdominal
organs and dilatation of those in the muscles.
Experiment 32. — The Measurement of Blood Pressure in

Man. — A sleeve containing a rubber bag, which connects with

a mercury manometer as well as with a valved rubber bulb, is

71

72

EXPERIMENTAL PHYSIOLOGY.

wrapped snugly about the arm and fastened. The sleeve
should be placed high enough to permit application of the
stethoscope over the brachial artery (Fig. 21). While palpating

FIG. 21. Sleeve in place for measuring blood pressure. It should be high enough to allow
ample room for the stethoscope.

BLOOD PRESSURE. 73

the radial artery the pressure in the rubber bag is raised above
that necessary to produce obliteration of the pulse. Now no
sound can be heard over the brachial artery at the elbow. By
gently releasing the valve the pressure is gradually lowered.
Just as soon as the pressure becomes less than systolic pressure,
the blood is forced through the occluded arteries and a distinct
thud is heard at each heart beat. At the same time or often
after the pressure has been lowered 5-10 mm. more, the pulse
is felt at the wrist. The first sound is never below the tactile
indication; if it is, some error has been made in the application
of the apparatus, usually the stethoscope. The pressure read
at the time that the first sound is heard is systolic pressure.

As the armlet pressure is further lowered, the sound be-
comes progressively louder and sometimes resembles a murmur
in character, and then, rather suddenly, it becomes feebler and
at the same time duller. At this point, the manometer indicates
the diastolic pressure. After this sudden sound, the sound may
continue to be heard for a longer or shorter time as the armlet
pressure continues to fall.

The difference between systolic and diastolic pressure is the
pulse pressure.

Determine the heart rate, systolic and diastolic pressures
under the following conditions in as many different subjects as
possible:

Supine position, sitting, standing and immediately after
vigorous exercise as running up and down stairs. How does the
pulse pressure change under these conditions?

PRINCIPLES OF HAEMODYNAMICS.

The blood propelled by the heart, circulates in a closed system
of tubes. Both the output of the heart and the response of the
vessels affect the rate of flow. Increase the output of the heart
either by a greater number of contractions per unit of time or by
increasing the amplitude; either change will produce a greater flow
of blood.

If the blood circulated in perfectly rigid vessels the pressure in
the system would rise to a maximum during the contraction of the

74

EXPERIMENTAL PHYSIOLOGY.

heart and fall to zero during the relaxation. The flow in such a
case would be spasmodic. On the other hand if the walls of the
vessels are elastic, they are stretched by each spurt of blood from
the heart, and then as the heart begins to relax the distended vessel
wall begins to contract, thus forcing the blood onward in spite of
the inactivity of the heart. The elastic wall therefore keeps the
blood pressure from falling to zero during diastole.

If the blood meets with resistance somewhere in the path the
vessels on the near side are distended more than they would be if

FIG. 22. Circulation scheme. The bulb (a) which represents the heart can
have its output varied by either a change in rate of compresssion or by sliding the
block (&) and thus changing the amplitude. The openings to the bulb are provided
with valves so that flow can take place only in one direction. The resistance which
corresponds to the capillaries is the part (c), (d). This maybe varied by means of
the clamp (c). The elasticity of the rubber tubing corresponds to the elasticity
of the arterial wall. The "venous" and "arterial" pressures are measured by mano-
meters (e) and (/) respectively.

the resistance were decreased or absent. The greater the distension
of the tubes the longer they require to contract so that a steady flow
would be kept up for a longer period. When the peripheral resist-
ance is constricted venous flow is regular ; great dilatation produces
an intermittent venous flow.

Experiment 33. — Circulation Scheme. — Many of the foregoing
principles can be demonstrated by a simple system of rubber
tubes (Fig. 22). A rubber bulb represents the heart. The

BLOOD PRESSURE.

75

rubber tubing nearest the heart takes the place of the arterial
system, while the venous system is the tubing on the other side
of the resistance.

1. With moderate capillary resistance determine a rate of bulb
compression which will cause a pronounced rise in pressure as
shown by the manometer. (If possible both arterial and venous
manometers should record in line vertically with each other; to
do this the arterial pressure zero should be higher than that for
the venous pressure.) By means of the block vary the ampli-
tude, but maintain a constant rate. What do the records of
venous and arterial pressure show? What can you say in
regard to the venous outflow?

2. With the same capillary resistance and with a moderate
amplitude vary the rate. Obtain pressure records and note
venous flow.

3. Wi,th the rate which gives the best results and with moderate
amplitude vary the capillary resistance, recording pressure
changes and noting venous outflow as before.

B

z>

//

) }

FIG. 22A.

Compare diastolic and pulse pressures under the foregoing
conditions.

What relation does capillary resistance bear to them?

How do amplitude and rate of the heart contraction affect
diastolic and pulse pressures?

Experiment 33a. — The apparatus represented in Fig. 22a may
be employed to demonstrate the part which the peripheral
resistance and the elasticity of the arterial walls play in the

76 EXPERIMENTAL PHYSIOLOGY.

conversion of an intermittent flow in the arteries to a continuous
flow in the veins.

The apparatus consists of a reservoir R, a pump P, and
two lengths of tubing, one, A, of glass and the other, B, of
rubber. Each tube is provided at its extremity with a thumb-
screw by which its calibre can be varied. A pinch-cock, D,
placed between the reservoir and the pump, allows the flow of
fluid to be cut off when the schema is not in use.

With the pinch-cock D closed, fill the reservoir with water.
Adjust the thumb-screws at the extremities of the tubes so
that the lumen of each is at its maximum calibre. Place a
basin beneath the openings of the tubes or direct them so that
the fluid issuing therefrom will flow into the sink. Open the
pinch-cock D. Compress the bulb intermittently at the rate of
about 60 per minute. What is the nature of the flow from
tubes A and B respectively? Next adjust the thumb-screws
so that the diameter of each tube is considerably reduced.
Repeat the intermittent compression of the bulb closing off
A or B by compressing the rubber tubing next the Y-piece.
What do you find with regard to the outflow from each tube?
Explain your results.

Response of Blood Vessels to Chemical Substances.

The amount of blood to any region can be increased by an in-
crease in the output of the heart or by a dilatation of the blood
vessels in that region. Dilatation may be brought about through
action of the nerves to the vessels or by chemical substances in
the blood. For example certain waste products such as lactic acid
may cause dilatation. The blood vessels therefore not only by
their elasticity help to maintain a steady blood flow, but by their
active constriction and dilatation shunt the blood to regions most
in need of it. That the blood vessels will respond to chemical
substances, can be shown by the following experiment.
Experiment 34. — Perfusion of the Frog. — Not only can the
heart be kept beating for a considerable time by perfusion, but
circulation can be maintained for some time through the blood
vessels as well. Arrange the beaker and siphon for perfusion
as used in the turtle heart. After exposing the heart of a frog

BLOOD PRESSURE 77

whose brain has been destroyed, tie off one aorta and fasten
a cannula into the other so that it will lead away from the heart.
Air must not be allowed to enter the system on any account.
Hang the frog toes down and allow the fluid to escape from
an incision in the sinus venosus. Determine the rate of flow
in cc. per minute for Ringer's solution. This can be done either
by measuring the inflow from a burette or by collecting the out-
flow in a graduated cylinder. Next perfuse with Ringer's
solution containing 0.1% NaNO2. What is the rate now?
Wash out the latter fluid with Ringer's solution, then perfuse
with Ringer's solution containing adrenalin.

Although adrenalin causes constriction of the blood vessels
in the frog and under certain conditions in mammals, small
quantities of adrenalin cause dilatation of the vessels supplying
skeletal muscle in most mammals, when injected into the general
circulation.

SECTION II B.
CHAPTER VII.

HAEMODYNAMICS

Although many fundamental facts relating to the function of
the cardiovascular system can be learned from experiments on
cold-blooded animals, it is essential for a proper understanding of
the physiology of the circulation in man that a certain number of
experiments be performed by the student on mammals. In order
that this may be done without causing any pain, the animal must
be deeply anaesthetised by an expert assistant before starting the
experiment, the anaesthesia must be maintained throughout, and
when the experiment is finished, the animal must be painlessly
killed.*

The following two experiments are selected to illustrate certain
fundamental factors governing the circulation of the blood. Other
fundamental experiments will be demonstrated before the class.

Each experiment is performed by a group containing from four
to six students. Two of these act as operators and are responsible
for the operative manipulations, one or two attend to the apparatus
(technicians), and the remainder make detailed notes of every
step in the experiment. After the experiment is completed, the
group should review the results and each member of it prepare a
report, embodying copies of the tracings secured. These reports
are to be handed in for inspection.!

*For small classes it is practicable to have these essential mammalian
experiments done on decerebrated preparations (see p. 247), but this is impossible
when large numbers have to be provided for.

fFor the experiments each student must provide himself with the following
instruments: dissecting case, including 1 pair scissors and a blunt hook (aneurism
needle or strabismus hook) ; 2 pairs, haemostats (Pean or Spencer Wells forceps) ;
2 pairs, bull dog forceps; a stethoscope.

78

HAEMODYNAMICS. 79

Experiment 35.

BLOOD PRESSURE. NERVE CONTROL OF HEART BEAT
AND PERIPHERAL RESISTANCE. HAEMORRHAGE
AND TRANSFUSION. INSPECTION OF HEART.

1. PREPARATION OF ANIMAL. — Weigh the animal. In the case of
dogs, inject morphine solution (sulphate or hydrochloride) sub-
cutaneously (0.5 c.c. of 2% solution per kilo of body weight). In
the case of cats, pass the stomach tube and pour the required amount
of ure thane solution into the stomach. (Give 1 gram of urethane
per kilogram of body weight, dissolving it in water to make a 20%
solution). These preliminary operations are to be performed by
the laboratory technician or by a demonstrator, with the students
assisting. In about one half hour the animal, now practically un-
conscious, is completely anaesthetised by administering ether. To
do this pour about 20 c.c. on a towel and hold this firmly over the
muzzle of the animal, being careful however to avoid suffocation.
The animal is then tied, back down, on the animal board, mean-
while deep anaesthesia is maintained by giving ether. Count the
pulse at the femoral artery; count the respirations and record the
rectal temperature. Note any peculiarities about the animal
(e.g., enlarged thyroid).

While the operator and anaesthetist are preparing the animal
and performing the necessary operations, the technician and
other members of the group proceed to get the manometer tubing
and cannula ready, as directed in par. 3 below.

2. INSERTION OF TRACHEAL CANNULA. Moisten the hair in front
of the neck and make a median incision down to the trachea (cut
with the edge, not with the point of the scalpel). Pass a stout
ligature under the trachea. Make a transverse incision half way
across the trachea, insert the tracheal cannula and tie it by means
of the ligature. The tracheal cannula is inserted to facilitate
artificial respiration should this be necessary. If further anaes-
thetic is required, connect the tracheal cannula with the wide-
mouthed anaesthetic bottle containing some ether.

3. INSERTION OF CAROTID CANNULA AND RECORDING OF MEAN
ARTERIAL BLOOD PRESSURE. — Pull apart the skin flaps and separate
the sterno-mastoid and sterno-thyroid muscles so as to expose the

80

EXPERIMENTAL PHYSIOLOGY.

carotid artery. With an aneurysm needle or a blunt-pointed curved
seeker, free the carotid from its sheath for about an inch and place
two ligatures under it. Tie the peripheral ligature (the one farthest

FIG. 23. Stand, manometer, pressure bottle, etc., for recording blood pressure.

from the heart). Place a bull-dog forceps on the artery central
to the central ligature.

Separate the vagus nerve from the carotid sheath on both sides
of the neck, and place ligatures, without tying them, around both

HAEMODYNAMICS. 81

nerves. Put bull-dog forceps on the ligatures to prevent these from
slipping off.

Meanwhile a cannula of suitable size is connected to the mano-
meter (Fig. 23) by rubber tubing which has been scrupulously
cleaned of any old blood. This tubing is also connected by a
T-piece with a bottle provided with a rubber bulb and containing
a 2% solution of sodium citrate to prevent clotting of the blood.
The cannula must be scrupulously clean and should be slightly oiled
before inserting. Observance of these precautions greatly dim-
inishes blood clotting. A pinchcock on the side tube which con-
nects with the bottle is cautiously opened and by gentle compres-
sion of the rubber bulb, which is connected with the bottle, the
tubing is filled with the citrate solution to the exclusion of all air
bubbles. The pinchcock on the side tube is closed and a screw
clip is placed on the tubing which connects manometer and cannula,
and closed. The writing style of the chronograph is now adjusted
so that it corresponds to that of the manometer. The line of the
time tracing therefore serves as the line of zero pressure.

The cannula, filled to the tip with the citrate solution, is now
inserted by the operator into the artery in the following way: the
wall of the artery near the peripheral ligature is picked up by a
forceps and, with a sharp scissors, a V-shaped cut is made into the
artery without cutting it clear across. The tip of the V points
peripherally. The V-shaped flap is lifted up with a fine pair of
forceps and the cannula inserted into the artery and tied by means
of the central ligature. The clip on the tubing connected with the
bottle is opened and the bulb compressed until a pressure of about
120 mm. Hg. is recorded in the manometer. The pressure in the
tubing prevents blood from entering it when the "bull-dog" for-
ceps on the artery is opened, which is now done. The writing
styles of the manometer and chronograph are adjusted to the drum,
and a tracing is taken with a slow speed drum.

4. DEMONSTRATE THE EFFECT ON THE BLOOD PRESSURE OF
VARIATIONS IN THE PUMPING ACTION OF THE HEART. STIMULATION
OF PERIPHERAL END OF VAGUS NERVE. — Expose the vagus nerve
in the neck on both sides and loosely tie the ligatures that were
previously placed around each nerve. Start the drum and record
a short piece of normal tracing, and then, without stopping the

82 EXPERIMENTAL PHYSIOLOGY.

drum, cut first one and then the other vagus between the ligatures.
Note the effect produced on the blood pressure (Fig. 27). To
signal the moment of cutting, short-circuit the time tracing for a
second or two.*

Allow several inches of tracing to be recorded after both nerves

have been cut, so as to be able to determine the after effects; then

tstop the drum and place the peripheral end of one vagus on elec-

rode s coming from a short circuiting key which is meanwhile

a b

FIG. 24. Arterial blood pressure tracing: (a) during moderate stimulation of the peri-
pheral end of vagus; (b) during stimulation of splanchnic nerve.

closed. Record a short piece of normal tracing and then by opening
the short-circuiting key, stimulate the vagus with an induced cur-
rent that is just bearable on the tip of the tongue and note the
effect produced on the blood pressure. Signal the time of stimula-
tion as above directed. Demonstrate escapement.

*By short-circuiting the time marker in this way the seconds cease to be
recorded and the break in the otherwise regular time tracing affords a useful
signal indicating the moment of cutting or stimulation.

HAEMODYNAMICS. 83

Repeat with the opposite vagus.

Repeat the experiment — using varying strengths of stimuli.

Explain the results.

5. DEMONSTRATE THE EFFECT OF VARYING THE PERIPHERAL
RESISTANCE ON THE BLOOD PRESSURE; STIMULATION OF SPLANCH-
NIC NERVE. — Make an incision along the left costal margin starting
about 1J inches from the linea alba. After stopping all hemorr-
hage,* open the peritoneum and make out the suprarenal capsule
lying over a transversely coursing vein. Just outside the suprare-
nal and above the vein expose the greater splanchnic nerve by
blunt dissection. Tie a ligature loosely around the nerve and lay
this on electrodes. While recording a normal tracing of the blood
pressure stimulate the nerve (Fig. 24). In taking this tracing all
details are to be followed as in par. 4.

Explain the result.

The splanchnic nerve is difficult to find unless the abdominal
viscera, with the exception of the left kidney, are well retracted
to the right. This is to be done by the assistant operator, who,
standing on the right of the animal, covers the viscera with cloths
wrung out with warm 0.9% NaCl solution, places the left hand
with fingers extended over the cloth, and then with the finger tips
just touching the posterior wall of the abdomen, pulls the viscera
towards the right. With the right hand this operator also pulls
down the kidney on the left side. By these procedures the suprare-
nal capsule is easily brought into view, and the operator isolates
the splanchnic nerve by blunt dissection in the region just above
the capsule.

It is often simpler to find the splanchnic nerve on the right side.
In this case, after retraction of the viscera by the procedure de-
scribed above, but towards the left side, the right kidney is exposed,
and the vein, connecting the abdominal muscles with the adrenal vein,
is ligated in two places and cut between the ligatures. The
splanchnic nerve is then brought into view by little dissection.

*The haemorrhage occurs mostly when the muscles are being cut. To arrest it
firmly sponge the wound with surgical gauze, locate the bleeding points, catch
them in the points of artery forceps (haemostats) and while an assistant gently
elevates the point of the forceps, tie a ligature around the tissue caught in them.
For venous and capillary oozing, pressure with a piece of gauze wrung out in hot
water is usually sufficient.

84 EXPERIMENTAL PHYSIOLOGY.

6. DEMONSTRATE THE EFFECTS OF HAEMORRHAGE AND OF
TRANSFUSION ON THE MEAN ARTERIAL BLOOD PRESSURE. — -Intro-
duce a cannula into the femoral artery of one side and another into
the femoral vein of the other side. The arterial cannula is intro-
duced as directed above, and connected with a piece of rubber tub-
ing through which to bleed the animal. The venous cannula is intro-
duced as follows: The vein is exposed and two ligatures placed
underneath it; a bull-dog forceps is applied central to the central
ligature, and the peripheral ligature is tied. A cannula of suitable
size, connected with about six inches of rubber tubing and filled

FIG. 25. Arterial blood pressure tracing showing the effect of a moderately slow hsemorrhag -,
followed by a rapid haemorrhage.

with 0.9% NaCl solution, held in by means of a clip on the rubber
tubing, is inserted into the vein towards the heart in exactly the
same way as in par. 3. When both cannulae are in position, the
tubing on the venous cannula is connected with a burette.

HAEMORRHAGE. — After taking an inch or two of normal tracing
of the pressure in the carotid artery, the clip on the femoral artery
is partially or completely removed and the blood which escapes
collected in a basin and defibrinated. Note the effect on the blood
pressure (a) of sudden and (b) of gradual bleeding (Fig. 25). It will
be found when the artery is fully opened that there is an immediate
fall in blood pressure due to release of peripheral resistance. If the

HAEMODYNAMICS. 85

clip is only partially opened, considerable bleeding may occur
before the blood pressure is materially affected. Continue the
bleeding until a permanent (marked) fall of blood pressure is
recorded. About 25 c.c. of blood per kg. body-weight must usually
be removed to obtain this result.

SALINE TRANSFUSION. — Now connect the venous cannula with
a burette filled with a 0.9% NaCl solution, previously warmed to
body temperature; remove the clip and allow the saline to pass
into the animal, taking great care that no air bubbles are carried
in with the solution. Meanwhile MAKE OBSERVATIONS ON THE
VENOUS BLOOD PRESSURE (which is indicated by the height in the
burette at which no more saline enters the vein); noting: (a) its
height in mm. of water and (b) whether it shows any pulsations.
Observe carefully the effect of the transfusion on the arterial blood
pressure.

The blood removed in this and in all other experiments is to
be defibrinated by beating in a clean dry basin with a bunch of
wires. When fibrin formation has ceased the whipped blood is
strained through muslin, and measured in a graduated cylinder.

BLOOD TRANSFUSION. — Again bleed from the femoral artery,
carefully noting the behaviour of blood pressure, respirations, etc.
Note any differences in the character of the blood from that ob-
tained previous to the saline transfusion. When the blood pressure
is extremely low (40 mm.) transfuse with defibrinated blood and
note the effect on the blood pressure as compared with that pro-
duced by saline solution. Note particularly with which solution
the restored pressure is best maintained.

7. OBSERVE THE HEART BEAT IN THE OPEN THORAX UNDER
ARTIFICIAL RESPIRATION. — Since natural respiration is impossible
after the thoracic cavity has been opened (explain why) it is neces-
sary to start artificial respiration by means of a pump connected
with the tracheal cannula. The air delivered from the pump is
in a continuous stream. In order to interrupt it so as to simulate
the respirations, a wide T-piece is placed on the rubber tube
between the pump and tracheal cannula, and this side tube is
opened and closed by a finger at a rate corresponding to the normal
respirations. In order to open the thorax the skin is first of all
incised down the mid line from the base of the neck to well on to

86 EXPERIMENTAL PHYSIOLOGY.

the abdomen and is quickly dissected to both sides sufficiently to
expose the rib cartilages. The latter are then separately snipped
through on both sides by a bone forceps and finally the soft tissues
are cut through by a stout scalpel from below up. Arterial haemorr-
hage must be controlled by catching the bleeding points with a
haemostat and tying, but venous oozing can be stopped by applying
cloths wrung out with hot water.

Watch the heart while the vagus nerve is being stimulated.
Place stout ligatures under the superior and inferior venae cavae
and tighten them for a few moments; note the emptying of the
heart. Place a ligature under the pulmonary artery and tighten
for a moment. Note the effect on the heart. Repeat with the
aorta.

Listen to the sounds of the heart with a stethoscope applied
directly to various parts of it and study the effect produced on
the sounds by tightening the ligatures referred to above. Stop
the artificial respiration for a few moments and note the behaviour
of the heart.

Afterwards excise the heart and open the right ventricle to
observe the contractions of the papillary muscles. Note that the
first sound is still heard in the excised heart. What conclusion do
you draw from this observation?

CHAPTER VIII.

Experiment 36.

VASO MOTOR NERVES, DEPRESSOR NERVE,
CIRCULATION TIME.

(The General Instructions for the Insertion of the Cannulae

and the Conduct of the Experiment are the same as

in Experiment 31).

1. PREPARATION OF ANIMAL. — Weigh the rabbit. Inject a 10%
solution of chloral hydrate per rectum, 5 c.c. per kg. body weight.
When the chloral effect has developed (about 30 minutes) cause the
rabbit to inhale a little ether. To fix the head make an incision
through the floor of the mouth and with an aneurysm needle pass
a stout thread through the incision and out of the mouth. Tie the
thread to the cross bar of the animal board. Tie out the limbs.

2. DEMONSTRATE VASOCONSTRICTOR FIBRES IN THE CERVICAL
SYMPATHETIC. — Clip the hair from the front of the neck and make
a median incision. Retract the skin. Separate the laryngeal
muscles from the sterno-mastoid so as to expose the carotid sheath.
Running with the carotid arteries will be seen one large and two
small nerves. Of the latter, one is the cervical sympathetic, the
other the depressor (see Fig. 26). Tie a pair of ligatures loosely
around each nerve.* Now stretch the ear on the same side as the
exposed nerve on a wire frame so as to render its vessels quite
plain. Stitch the edges of the ear to the frame. With a good light
behind it, examine the vessels of the ear, carefully noting their
size. Note also the size of the pupils. Pick up the ligatures around
one of the fine nerves, tie both ligatures and cut the nerve between
them. Look for any effect on the vessels of the ear and on the
pupils during both tying and cutting. Apply electrodes to the

*To obviate confusion the ligatures should be marked by placing bulldog
forceps on the free ends. The nerves must be frequently moistened with 0.9 per
cent, saline.

87

EXPERIMENTAL PHYSIOLOGY.

upper part of the cut nerve. Stimulate with an interrupted current
of moderate strength and watch the effect on the vessels and pupils.
If no change occurs, repeat with the other nerve. The nerve which

FIG. 26. Dissection of neck of rabbit to show relative position of nerves,
etc.: (c) carotid artery; (ft) vagus; (5) sympathetic; (d) cardiac depressor; (m)
descending branch of hypoglossal nerve. (After Jackson).

causes definite effects is the sympathetic. Explain the causes of
the results. It is often advisable to introduce the tracheal and
carotid cannulae, as directed under par. 3, before attempting to
stimulate the nerves.

VASO MOTOR NERVES. + 89

/J&

3. DEMONSTRATE THE EFFECT OF STIMULATION OF THE VAGUS
NERVE ON THE ARTERIAL BLOOD PRESSURE. — Introduce tracheal
and carotid cannulae as described in pars. 2, and 3 (Exp. 35), and
apply electrodes to the peripheral end of the vagus. Take about
an inch of normal tracing and cut the vagus on one side while a
tracing is being taken. Note any effect on the blood pressure.
Apply electrodes to the peripheral end of the cut vagus and stimu-
late with an induced current of moderate strength (just bearable to
the tip of the tongue). Repeat with stimuli of varying strengths.
Remove the electrodes to the central end of the vagus and repeat
the stimulation.

FIG. 27. Stimulation of the cardiac depressor nerve in the rabbit, showing
effect on arterial blood pressure. In the tracing to the right the vagi were intact;
in the tracing to the left they were cut. Note the slower respiratory oscillations
after cutting the nerve.

In the above experiments, the drum should be revolving slowly
while the stimuli are applied, and the moments at which this is
done should be indicated by short-circuiting the time tracing.
Explain the causes of the results.

4. DEMONSTRATE THE EFFECT OF STIMULATION OF THE DE-
PRESSOR NERVE ON THE BLOOD PRESSURE. — Pick up the ligature
tied to the central end of the depressor nerve (the small nerve
which is not the sympathetic), apply electrodes and stimulate.
A fall in blood pressure should occur (Fig. 27). Look for slowing
of the heart. Show that the fall in blood pressure, caused by
stimulation of the depressor, is not entirely due to reflex vagus

90 EXPERIMENTAL PHYSIOLOGY.

inhibition, since it persists after both vagi are severed. What con-
clusions do you draw as to the cause of the fall? Finally stimulate
the peripheral ends of the depressor and sympathetic nerves.
The sympathetic stimulation sometimes quickens the heart.

5. ESTIMATE THE CIRCULATION TIME FOR THE LESSER CIRCU-
LATION (Stewart's method). — Ligate the carotid and remove the
cannul^. On the same side of the neck insert a cannula (pointing
towards the heart) in the jugular vein (for technique see par. 6,
Exp. 31). Expose the carotid artery of the opposite side and place
under it a strip of white glazed paper resting on a piece of thin
rubber dam. Throw a strong light on the artery. Connect the venous
cannula with a burette containing a 0.2% solution of methylene
blue in physiological saline solution, at the temperature of the
body. See that there is no air in the tubing. When all is ready,
remove the bull-dog forceps and mark the exact moment that the
methylene blue enters the vein (i.e., when the methylene blue is
seen through the cannula to enter the vein). Carefully observe the
carotid artery. The moment at which the blue appears in the artery
is also noted. Repeat several times, using always the same amount
of methylene blue. To obtain accurate results no methylene blue
solution should be allowed to escape on to the wound. The results
are to be given in relationship to (1) actual periods of time and
(2) the heart-beats.

6. DETERMINE THE SEAT OF OXIDATION IN THE BODY. — Inject
an excess of a stronger solution of methylene blue until the animal
is killed by the injections, and make a careful autopsy; examining
especially sections of the muscles, kidneys and liver, also the urine,
the blood in the mesenteric vessels, etc. The methylene blue will
be found to have stained the blood but not the tissues. The tissues
have reduced it to methylene white. This is known as Ehrlich's
experiment and it shows that the tissues are the seat of reduction
in the body. Note the effect which exposure to air has on the
colour of the cut organs. On standing exposed to the air the cut
tissues will become blue because of oxidation of the methylene
white to methylene blue. What conclusions do you draw from the
experiment?

VASO MOTOR NERVES. 91

COAGULATION OF BLOOD.

Experiment 37. — A. Expose the carotid artery and jugular vein
in an anesthetized rabbot. Treat each as follows : Tie a ligature
around the vessel as close to the heart as possible. When rilled
with blood tie a second ligature 2 cm. above the first, and a
third 2 cm. above the second. Crush the vessel between the
first and second ligature by means of a hemostat without
rupturing the wall. Later, when the rest of the experiment is
completed look for an intravascular clot in each vessel " pocket."
Explain.

B. Insert a cannula into the carotid artery of the other side,
Draw 5 c.c. of blood into each of the test tubes prepared as
follows, after which, the time required for the first appearance
of a clot is determined.

1. Clean — keep at room temperature.

2. Clean— keep at 37° C. to 40° C.

3. Clean-keep at 0° C. to 5° C.

4. Containing sand with which the blood is to be shaken.

5. Thoroughly coated inside with a vaseline or paraffin oil.

6. Containing a small amount of potassium oxalate with

which the blood is to be shaken. After ascertaining
that oxalated blood does not clot add a drop or two
of 5% CaCl2 solution, increasing the quantity if
necessary.

7. Vaselined — centrifuge the blood, separating the plasma

and keeping it for observation.

C. Several glass tubes from 3 to 4 cm. long (3 to 4mm. dia.)
and tapere'd nearly to capillary dimensions at one end are
oiled inside. A short length of rubber tubing is doubled at
one end and tied with thread. This tube should fit easily but
snugly to the glass tubes and is used as a pipette bulb to suck
blood into the glass tubes.

A rabbit's ear, with hair closely clipped and washed thor-
oughly long enough before to permit drying, is oiled. When
the observer is ready to note the time, a vein is punctured by
holding the under surface of the ear against a cork and plunging
a sharp three-cornered needle through it. Four samples of

92 EXPERIMENTAL PHYSIOLOGY

blood are drawn up into the glass tubes. The first appearance
of a clot is determined by drawing out a small glass rod. Each
tube must have its own rod. From time to time (every 20 to
30 sees.) the rod is to be inserted and the first clot sought.

After having become practised in the method blood is drawn
from the human ear or finger and the clotting time determined.
In the case of the finger, it should be exercised for several
seconds, then a handkerchief wrapped tightly around it at the
second joint, the first joint being bent at a right angle. The
skin is punctured now a little below the base of the nail.

In every subject three determinations, which check within
the limits of experimental error, should be made. Obtain
observations from as many different subjects as possible.

D. Place a drop of human blood or rabbit's blood on a
slide and cover with a cover-slip. Observe the blood with a
microscope from time to time so that the fine structure and
behaviour of a clot may be seen. Between observations cover
the slide with a moist chamber cover containing wet filter paper
plastered to the inside; otherwise the blood will dry before it
clots.

Summarize the information which you have obtained form
the above experiments.

CHAPTER IX.*

POLYSPHYGMOGRAPH TRACINGS.

These experiments consist in obtaining graphic records of the
pulses of the radial and carotid arteries, of the apex beat and of the
venous pulse in the jugular vein.f

The experiments are important not only because the results
if properly interpreted throw much light on the cardio-vascular
mechanism, but because of the value of the technique in clinical
diagnosis. The tracings are often difficult to obtain, and great
care and patience must be exercised in adjusting the various re-
ceiving tambours. This is especially so for the radial pulse and
apex beat. When these two cannot be satisfactorily obtained to-
gether, the carotid should be substituted for the radial. The ob-
served person must remove his shirt and lie on a couch or table.
Sometimes it is best to lie on the side, sometimes on the back.
While cardiac tracings are being taken, the breath should be held
for a few moments.

Experiment 38.

A. The Velocity of Transmission of the Pulse Wave. (Pro-
pagation of Pulse Wave). — With the arm easily extended and
resting on the arm support, apply the button of the receiving tam-
bour over the radial artery and adjust the pressure until a maximal
movement of the recording lever is observed. Taking care that the
radial tambour does not slip, now apply the carotid tambour by
hand (apply opposite the angle of the jaw). When a maximal
pulsation of both of the levers is obtained, adjust the writing points
to a slowly moving clockwork drum. The writing points should be
as nearly as possible in the same perpendicular, and should be

*For the experiments of this chapter two sessions of three or four hours each
are necessary.

fNotes and tracings of these experiments are to be taken by every member
of the class.

93

94

EXPERIMENTAL PHYSIOLOGY

applied to the drum with a minimal amount of friction. This is
done by manipulating the adjusting screws on the tambour stands,
while the drum is moving slowly.*

When the adjustment is completed the drum is stopped for a
moment so that the two levers may draw vertical lines showing
their relative positions (alignment marks). The drum is then
allowed to revolve at a medium speed and the tracings taken.
The speed should be as great as is consistent with a definite up-
stroke of the pulse. Alignment marks should be inscribed at
frequent intervals (by stopping the drum for a moment). A time
record (0.2 sec.) is added to the tracing with the drum revolving at

FIG. 28. Simultaneous tracings of carotid and radial pulses. The rate of
transmission is determined by finding the difference between the beginning of
each pulse, after correcting for alignment of levers, and measuring off this distance
on the time tracing (l/50ths second.).

the same speed as when the pulse tracings were being taken. A
vibrating spring provided with a writing point is used for taking
the time tracing.

After the removal of the tracing from the drum, the distance of
the beginning of the upstroke is measured from its alignment mark
for each pulse curve (Fig. 28). The difference, interpreted in terms
of the time tracing, gives the rate at which the pulse wave travels
from the carotid to the radial artery. Note also the dicrotic notch

This is more satisfactory than attempting to adjust to a stationary drum
because friction is diminished. Before starting the drum, however, make certain
that the tambours are so adjusted as to give the greatest possible movements.

POLYSPHYGMOGRAMS. 95

and measure its distance from the beginning of the upstroke.
Several measurements should be taken from each member of the
group. How can the length of the pulse wave be calculated?

In the more or less arbitrary lettering of the waves observed in
pulse tracings, the main upstroke is marked 3, in the case of the
carotid and 4 in that of the radial ; the dicrotic notch of the carotid
is 5 and of the radial 6. The distances between 3 and 5 and be-
tween 4 and 6 represent respectively the time during which the
heart is pumping blood into the arteries, i.e., the semilunar valves
are open. It is called the sphygmic period ui period E.

B. The Venous or Jugular Pulse Curve. — Have the observed
person lie down with his head slightly raised by a cushion and bent
to the right side. Place the receiver (thistl" funnel) over the jugular
bulb on the right side of the neck. This lies immediately above
the inner end of the clavicle. Bring the point of the lever of the
recording tambour to write with a minimal amount of friction on
a drum. Since a venous pulse tracing cannot be interpreted with-
out a simultaneous tracing from an artery, adjust the button of a
receiving tambour over the radial artery and arrange the writing
style of its recording tambour so as to write on the drum in the
same perpendicular as the style of the venous tambour. If no
satisfactory tracing can be secured from the radial, try the carotid.
Remember to inscribe alignment marks at short intervals. While
the tracing is being taken it is usually advisable that the respiratory
movements be suspended.

To INTERPRET THE VENOUS CURVE. — Make a vertical mark
on the arterial pulse tracing corresponding to the beginning of
the pulse upstroke. If this is done on the radial pulse tracing,
measure 1/10 sec. in front of it and make another vertical mark.
(This mark is to allow for the time lost in propagation of pulse from
heart to radial. It is determined according to Exp. A).

This line 3 corresponds to the beginning of the sphygmic period
of ventricular systole, i.e., to the opening of the semilunar valves.
Measure the distance from line 3 to the nearest alignment mark.
By measuring off the same distance from the corresponding align-
ment mark of the venous tracing line 3 will be found to fall at the
beginning of a small wave which is marked c. The small wave in
front of c is marked a and is due to auricular systole. The large

96 EXPERIMENTAL PHYSIOLOGY.

wave of depression following c is marked x and is due to a fall of
pressure in the auricle. What causes this fall in pressure?

The next point to determine is the end of the sphygmic period.
This is found by measuring from the alignment mark to line 5 of
the carotid or line 6 of the radial (less 1/10 sec.) and transferring
to the jugular tracing. The line will be found to fall on a small wave
on the upstroke of the depression. This wave is marked V and is
due to the sudden opening of the tricuspid valves. Sometimes
another small wave just precedes V. It corresponds to closure of
the semilunar valves. Listen to the heart sounds while recording
a tracing in order to determine this fact.

In preparing your report of the experiment, draw in the intra-
auricular and intra-ventricular pressure curves in relationship to
the venous curve.

C. Cardiograms. — Apply the button of the special receiving
tambour to the apex beat and connect with a recording tambour.
Adjust the position and pressure of the button until a maximal
movement of the writing style of the recording tambour is obtained^
Apply the writing point to a carefully levelled drum and with this
running at moderately high speed, take a tracing. There is often
difficulty in securing a satisfactory tracing, and it may be necessary
to try another subject. Breathing should be suspended for a few
moments while the tracing is being taken.

To INTERPRET THE CURVE. — Adjust another receiving tambour
to the radial or carotid pulse with both writing styles in the same
perpendicular, and following the other directions described under
"Venous Pulse", mark on the cardiogram:
- 1. The beginning of the sphygmic period, E (line 3).

2. The end of the sphygmic period, E (line 5).

3. The auricular wave (beginning of this wave is line 1; the

end of it, line 2).

4. The beginning of ventricular systole (difference between 2

and 3 equals the presphygmic interval).

5. The opening of auriculo-ventricular valves (lowest point in

tracing; somewhat difficult to determine).

When satisfactory tracings have been secured apply a stethoscope
to the apex beat and after accustoming your ear to the sounds, mark
as accurately as possible, by free hand, the position on the cardio-

POLYSPHYGMOGRAMS. 97

gram at which they are heard. The second (sharp) sound can be
heard best over the sternal end of the second rib on the right side.

.Influence of Respiratory Movements on the Various Pulse
Tracings. — Apply the respiratory tube to the thorax and connect
with a recording tambour. Also arrange to take a tracing of one
or the other of the pulses, or of the heart beat. Record simultaneous
tracings of the respiratory movements and of the pulse. Note the
effect of inspiration and expiration respectively on the pulse curves.

How do you explain the effects?

TESTING CARDIAC EFFICIENCY.

Experiment 39. — Count the pulse arid the respirations and record
the systolic and diastolic blood pressures of a person while
sitting. Then have him stand up and raise a pair of dumbbells
from the floor to the level of the shoulders with the arms ex-
tended, at a rate of 30 times a minute, and continue the exercise
for 2 minutes. Immediately the exercise is discontinued again
repeat the above observations and continue doing so at intervals
during the recovery period, until all the observed values have
returned to normal levels. It will be necessary to repeat the
muscular exercise several times, in order to obtain a sufficient
number of observations from which to obtain a complete
picture of the after effects. It is most important that this be
done carefully since it has been *found that by such methods
the functional capacity of the heart can best be guaged (cf.
Cotton, Rapport, and Lewis, Heart, 6, 269).

Besides obtaining the data indicated above also make
careful observations on the following: (a) The precise moment
at the start of exercise at which the breathing and pulse rate
became changed, (b) The length of the period following exer-
cise during which the pulse rate and blood pressure remain
changed, (c) The con tour of the arterial pulse curve before and
after the exercise. To obtain this tracing the Dudgeon sphymo-
graph may be used. Note particularly whether a dicrotic wave
appears, (d) :The tension of CO2 in the alveolar air. The
samples must be collected during and as early after the exercise
as possible. (Since only two or three pieces of apparatus are
available for these analyses the various groups must arrange
their experiments so that this observation can be made by all.)
xThis observation is to be made only after Experiment has been performed.

SECTION III.
THE CENTRAL NERVOUS SYSTEM.

CHAPTER X.
R*EFLEX ACTION IN THE FROG.

In a nerve fibre impulses are transmitted with equal facility in
either direction (Exp. 18, p. 45), but in a reflex arc they go in one
direction only. The nerve centre is responsible for this directive
influence, and the centre may be situated either peripherally, as in
ganglia, or centrally, as in the brain and spinal cord. The exact
significance of the centres in the brain and spinal cord varies con-
siderably in different animals according to their degree of develop-
ment. In a general way the centres in the spinal cord are primarily
the local centres for each segment of the body, but the reflex
activities of other segments may be caused to cooperate so as to
bring about complex movements. The centres in the brain, on the
other hand, are to be regarded as affording higher nervous path-
ways which the reflex impulse does not necessarily traverse, but
in which, when it does so, considerable modification may occur.
The impulse may, for example, become suppressed (inhibited) or
exaggerated (augmented) by its passage through nerve centres in
which memory impressions have been stored away. If these
memory impressions indicate that reflex movements would be
harmful to the animal then the movements may be inhibited: under
the converse conditions thay may be augmented. On account of
this dominating influence of the higher over the lower centres it is
clear that a precise analysis of the physiology of reflex action
demands a simplification of the experimental conditions, by iso-
lating the spinal cord from the brain. In the lower vertebrates
this can readily be done by actually removing or destroying the
latter. In the higher vertebrates, this operation is incompatible
with life, so that the brain is merely isolated from the spinal cord
by cutting the latter below the level of exit of the nerves to the
chief respiratory muscles (diaphragm). This so-called SPINAL

9*

CENTRAL NERVOUS SYSTEM. 99

ANIMAL offers a most suitable preparation for the study of reflex
activities. Immediately following its isolation from the brain the
cord enters into a depressed condition called SPINAL SHOCK. This
is quickly recovered from in the lower animals, but may take
months to disappear in the higher.

Experiment 40. — Pith a frog, destroying the brain only and sus-
pend the animal by fixing the lower jaw to a clamp (Fig. 29).
Apply mechanical stimuli (by pinching) to the skin of various
regions and note the location and character of the response.
Commence the application of the stimuli immediately after
decerebration and observe by repeating at intervals whether
they improve with time. If they do so, what is your conclusion ?
Proceed now to demonstrate the duration of the latent period or
reflex time using as the stimulus a 0.1 per cent, solution of HoSO4.
Measure accurately the time which elapses between placing the
foot in the acid and the movement of the leg. Remove all
traces of acid from the skin by means of water. Make three
observations with the same strength of acid using the two feet
alternately and allowing some time (several minutes) to elapse
between the stimulations. Repeat the observations using 0.3
per cent. acid. Are any differences in latent period and in-
tensity of response observed, depending on the strength of
stimulus?

It is likely, with the strong stimuli employed in the above
experiments, that some spread (or irradiation) will have occurred
in the cord, so that the opposite hind limb or the fore limbs
show movements. How is this explained?

Experiment 41. — To investigate the "march" of the irradia-
tion more precisely and to demonstrate that the isolated cord
is capable of synthesising a complicated and apparently pur-
poseful group of movements saturate a piece of filter paper in
30 per cent, acetic acid, remove the excess of acid and place a
small square of this paper on one of the flanks of the frog.
After noting the character of the movements, wash away the
acid and repeat, holding the leg on the irritated side.
The Spreading of Reflexes in the cord is greatly facilitated by
strychnine, but the movements are not of the same "purposeful"
character as those just studied.

100

EXPERIMENTAL PHYSIOLOGY.

FIG. 29. Decerebrate frog suspended for studying reflex action.

CENTRAL NERVOUS SYSTEM.

101

Experiment 42. — Inject a few drops of a 0.5 per cent, solution of
strychnine sulphate into the dorsal lymph sac of the frog. Apply
weak stimuli at intervals to the skin of different parts of the
body and note the exact nature of the responses. Explain the
action of the strychnine.
Inhibition of Reflexes. — Reflex activities may be inhibited

either through afferent nerves or from the higher centres.

FIG. 30. Exposed brain of frog: (c) cerebrum; (o) optic lobes;
(d) cerebellum; (e) medulla.

Experiment 43. — Expose the sciatic nerve in the left thigh. Tie
a ligature about the nerve near the knee and cut the nerve
distal to the ligature. Immerse the right leg of the frog in
0.5% sulphuric acid and note the latent period for reflex action.
Immediately wash off the acid. After three minutes place
electrodes on the exposed sciatic nerve and as the right foot is
again immersed in the acid, stimulate the nerve with a weak
tetanizing current. Prove that the sensory endings in the skin
are still irritable by the acid. Explain the inhibition.

102 EXPERIMENTAL PHYSIOLOGY.

To show that reflexes can be inhibited by influences from
higher centres, place a crystal of salt on the optic lobes which
have been exposed in a live frog by quickly cutting off the anterior
portion of the head just behind the tympani with heavy scissors
(Fig. 30). Now attempt to elicit reflexes by the methods described
above.

THE FUNCTION OF THE SPINAL NERVE ROOTS.

Experiment 44. — Pith the brain of a large frog and fasten it ventral
side down. Slit and evert the skin over the last four vertebrae;
remove the muscles from the vertebral arches and then open
the spinal canal by means of a strong scissors, keeping them
close to the bone. Uncover three or four sets of roots. Pass a
silk ligature under a dorsal root as far out as possible. Tie and
cut distal to the ligature. Stimulate the central end of the root.
Tie a ligature under another dorsal root close to the cord and
cut the root central to it. Now stimulate distal to the ligature.
In like manner test the function of the ventral roots. In this
experiment the unipolar method of stimulation will be found
best because of the shortness of the spinal roots. The plate
electrode, attached by a wire to the secondary coil is placed
in contact with the belly of the frog. The other wire is con-
nected with a dissecting needle which then serves as the more
active electrode to stimulate the nerve roots. Note the nature
of the movements produced in the above experiments and draw
a diagram showing the functions of the roots.

THE FUNCTIONS OF THE CEREBRUM IN THE FROG.

Interesting observations can be made on a decerebrate frog,
but it must be remembered that they do not shed much light on
the functions of the cerebrum in the higher mammals because in
these the cerebral centres are relatively far more important and
participate in many reflex activities in which the spinal cord alone
is involved in lower animals.

Experiment 45.— Activities of a Decerebrate Frog.— Anaesthe-
tize a frog by placing it under a beaker together with a piece

CENTRAL NERVOUS SYSTEM 103

of absorbent cotton containing ether. Slit and retract the skin
over the skull and remove a triangular piece of the latter so as
to expose the cerebrum. The base of the triangle should be
on a line connecting the posterior border of the tympanic
membranes and should be about a centimeter wide. Avoid
injury to the brain beneath until the cerebrum is completely
exposed, then carefully remove the latter and plug the cavity
loosely with cotton. Sew the skin in place and keep the animal
where the skin will remain moist until the next day. This
can be done by placing the frog with a little water in an evaporat-
ing dish and covering all with a large funnel or something which
will prevent escape, but at the same time allow access of plenty
of air.

After 24 hours study the behaviour of the decerebrate
animal compared with a normal frog, noticing especially:
posture, ability to swim or hop, power of escaping from a vessel
of water gradually heated, ability to turn over when placed on
the back.

Again open the cranial cavity, without anaesthesia as there
can be no sense of pain, and destroy the corpora stria ta and
optic thalami. After recovery from shock observe the activities
as before. Finally destroy the rest of the brain and notice any
change in behaviour.

CHAPTER XI.

REFLEX ACTION IN MAN.

There are in general three types of reflexes elici table in man and
the higher mammals. These are: 1. The reflex movements pro-
duced by the application of hurtful or nocuous stimuli to the skin
(nociceptive reflexes). 2. The reflexes required to maintain the
joints in such a position that the animal may stand erect and move
about (postural reflexes). 3. So-called myotatic reflexes which are
contractions produced by direct mechanical stimulation of muscles
that have been brought into a hyperexcitable condition (hyper-
tonus) through reflex action.

An example of nociceptive reflexes is the flexion reflex already
studied in the spinal frog. In man this type of reflex, variously
modified according to the part of the body from which it is elici-
ted, is extensively employed in the diagnosis of nervous diseases.
The particular value of the reflex is that its presence or absence
indicates the condition of the reflex pathway at the various levels.
For the study of reflex time it is useful to employ the palpebral
reflex, but in doing so it must be remembered that there are several
fundamental differences between this and the flexion reflex, such
as the relationship between strength of stimulus and latent time as
well as intensity of response.

Experiment 46. — By means of a strip of adhesive tape attach a
thread to the upper eyelid. Pass the thread through the handle
of scissors which have been clamped to a stand to serve as a
pulley. Attach the thread to a heart lever. The head should
be held firmly in such a position that when the upper eyelid
is closed the writing point of the lever moves upward. To elicit
the reflex the subject presses a pair of electrodes against the
lower lid. The operator stimulates the subject with break
shocks only, using a current which is just strong enough to cause
the upper lid to move. The subject must not see the operator
at work. With the aid of a magnetic signal determine the reflex
time on a rapid clock-work drum. Calculate the average reflex

104

REFLEX ACTION.

105

time from a number of observations on the same subject.

Each individual of the pair should serve as subject.

There is no example of a postural reflex which can conveniently
be studied in the frog or man. (The extensor thrust and the mark-
time reflexes (see p. 240) of the spinal mammal will serve this
purpose) .

The myotatic reflexes are best illustrated in the KXEE JERK.

FIG. 31. Arrangement of apparatus for measurement of reaction time in man.

Experiment 47. — Let the subject sit in a comfortable position with
one leg crossed over the other so that the patellar ligament is
under an increased tension. The operator strikes the patellar
ligament a sharp blow with the edge of his hand or a book.
With a little practice it is easy to elicit a sudden contraction
of the quadriceps muscle giving a sudden extension of the leg.

106 EXPERIMENTAL PHYSIOLOGY.

This should be tried on a number of subjects. After observing
the extension in each subject, try the effect of REINFORCEMENT
by the following methods:

1. Clenching the hands together vigorously.

2. Strong sensory stimulation such as pulling the hair.

3. Mental effort as solving a problem.

The magnitude of the response seems to depend on the tone

of the muscle.

Tone depends on the condition of the nervous system, therefore
the knee jerk is an important means of ascertaining certain patho-
logical conditions of the central nervous system.

REACTION TIME IN MAN.

Akin to reflex latent time is the so-called REACTION TIME in man ,
that is the time which elapses between the application of a stimulus
and a prearranged voluntary reaction.

Experiment 48. — In this experiment the apparatus is set up as in
Fig. 31. With the operator's key A open, and the subject's key
B closed, the operator closes, and the subject opens B when-
ever he feels the stimulus. The signal connected with the keys
writes on a quickly revolving drum on which a time tracing in
lOOths of a second is also inscribed. Determine the average
reaction time. Now increase the stimulus to find whether the
reaction time changes. What are the additional factors in-
volved in reaction time as compared with reflex time? How do
different subjects vary?

SECTION IV.
RESPIRATION.

CHAPTER XII.
ANALYSIS OF AIR. ALVEOLAR AIR. APNOEA.

Experiment 49.—

The (Haldane) gas burette A (Fig. 32) has a capacity of 10 c.c. of which
7 c.c. are contained in the bulbar portion and 3 c.c. are on the tubular portion.

The latter is graduated to — ths of a cubic centimetre. These proportions
100

are chosen so that the oxygen in atmospheric air may be measured. The
burette is connected with a reservoir containing mercury B by rubber tubing
of sufficient length so that when B is placed on the upper hook (1) the mercury
will fill the burette. On this tubing, near the burette, are a screw clip (a)
and -a pinch cock (b). When the reservoir is hung on hook (2) the mercury
should stand at the 10 c.c. mark in the burette. At the upper end of A is
a three-way stopcock (c) opening, according to its position, either with the
outside or into a side tube which is connected by thick walled rubber tubing
(g) with thfe absorption bulb D. On the tubing above the bulb is a much
smaller bulb, E, and the lower end of the absorption bulb is attached to a
reservoir, F. A sufficient amount of 20 per cent, solution of NaOH is placed
in D and F to a mark on the tube between D and E. The gas burette is
surrounded by a water jacket and the whole apparatus is mounted on a
wooden stand.

As described the apparatus is suitable for determination of
the CO2 in alveolar air. If it is desired also to determine O2
(either in atmospheric or alveolar air) the absorption apparatus
shown in the inset of Fig. 32 must be attached at the joint G.
This will be described later.

Analysis for CO2 alone.— The first step is to test the
apparatus for leaks. Turn C in the position shown in I so that
the burette communicates directly with the outside. With
both screw clip (a) and pinch cock (b) closed, the reservoir is
then placed on hook I, after which first a and then b are partly
opened so that the mercury slowly rises in the burette and
displaces the faintly acid solution with which it was left filled.

107

108

EXPERIMENTAL PHYSIOLOGY.

When the mercury has risen to the piece of rubber tubing at
the end of tube H, b is closed and a slowly screwed down until
the mercury just fills the rubber tubing on H the last trace of
acid water being removed by means of a piece of filter paper.
The next step is to lower the reservoir B to hook 2, release the
screw clip a and cautiously open clamp b so that the mercury

FIG. 32.

slowly falls in the burette to a little below the 10 c.c. mark.
The tap C is then turned in the position III so that both
A and D communicate with the outside. The exact position
of the level of the NaOH solution in the tube between D and E
is marked on the glass (by a glass pencil or pen) and the mercury
level in the burette is adjusted, by using the screw clip a, so

ALVEOLAR AIR. 109

that the dome of the meniscus stands precisely at the 10 c.c.
mark.

The tap C is now turned so that A and D are alone in com-
munication (as shown in II) and the reservoir, B, taken off the
hook by the left hand and slightly elevated, after which a is
partly unscrewed and b held open by the right hand. The
mercury rises in A so that the air passes into D where the CO2
is absorbed from it. By cautiously raising and lowering B
the air is passed several times (five) between A and D, extreme
care being taken when lowering B to see that the NaOH solu-
tion does not rise into the narrow tubing d. If this should
happen, or a broken column of solution get into the tube d,
the clamp b should be instantly closed. Finally, the clamp
b is closed, B placed on hook 2 and the level of mercury ad-
justed by using b and a until the NaOH solution stands exactly
at the original level in d. If there has been no leakage, the
mercury should stand precisely at the 10 c.c. mark (since
atmospheric air contains too small a percentage of CO2, 0.03,
to be measurable in such a burette).

Having thus tested the reliability of the apparatus, analysis
may be made of alveolar air collected as described on p. 110.
To transfer the gas from the syringe to the burette, the former
is attached to the rubber tubing at H after this has been filled
to the top with mercury, as described above. With the reservoir
on hook 2 and the clip a partly opened clamp b is cautiously
opened and the piston of the syringe gently pressed so that
the air slowly enters the burette to just below the 10 c.c. mark.
The further steps are precisely as described for atmospheric
air. In this case, however, the volume will be found to have
become considerably reduced and the exact reading on the
burette is noted and the percentage of CO2 calculated.

At least three similarly collected samples of alveolar air
are to be analysed and the results should check to within
0.25 per cent.

When all analyses have been completed the burette is filled
with weak acid (0.5 per cent H2SO4). This is accomplished by
turning C into position I, filling the burette with mercury and
the syringe with weak acid, inserting the nozzle of the syringe

110 EXPERIMENTAL PHYSIOLOGY

at H and then allowing it slowly to fill the burette. The burette
is left filled with the weak acid.

Analysis for CO2 and O2. — For this purpose, the absorp-
tion part of the apparatus is more complicated (see Fig. 32)
and is attached at G.

The tube from the burette leads to a three-way stopcock J which accord-
ing to its position connects either with D or M; with the former for the ab-
sorption of CO2 as described above and with the latter for the absorption
of O2. This part of the apparatus consists of a spiral tube L ending below
in a bulb M which is connected with a reservoir N. M contains pyrogallic
acid dissolved in 60 per cent. KOH solution (10 grams per 100 c.c.) and N
contains mercury. These are introduced through the side tube O and the
levels are adjusted so that the jjieniscus of the pyro solution stands some-
where on the narrow tube e above the spiral. For convenience of filling and
cleaning there is a rubber joint at K surrounded by mercury so as to prevent
leaks. This is shown in detail in the small side sketch of Fig. 32. The spiral
is used to increase the surface of contact between gas and solution since
absorption of C>2 is much slower than that of CC>2.

The analytical procedure is in principle the same for O2 as
for CO2 but it is necessary that the tubing and connections be
filled with nitrogen, instead of air, before starting the analysis.
This is done by passing some air back and forth between the
burette and the spiral, until, on bringing the pyrogallate
meniscus to the original mark on e, there is no further change
in the level of the mercury in A. The tap J is then turned so
that g and d communicate, the nitrogen passed into D and
then returned to A so as to displace the air entrapped in d.
The tap J is again turned so that g and e communicate and the
last traces of O2 removed from the gas in the burette.

This preliminary filling of the apparatus with N is necessary
only when it is used for the first time after assembling; other-
wise N remains over in the tubing from the previous analysis.

In the actual analysis for CO2 and O2, the percentage of
CO2 is first of all determined by the procedure already described.
The tap J is then turned so that g and e communicate and the O2
removed, great care being taken in moving the air between
A and L that the pyrogallate solution does not rise above the
mark on e. The small bulb on e serves to prevent this by
breaking up any bubbles that form. It will be noticed that
as the O2 becomes removed the pyro solution adhering to the

ALVEOLAR AIR 111

walls of the spiral tube becomes much more transparent.
Before the final volume is read in A, the air entrapped in d
must be removed as above described1. Since it takes some
time for all O2 to be absorbed, the final volume should not be
read in A until time has been allowed for the temperature of
the gas to come to that of the water in the jacket.2

After completion of the analysis the taps J .and C are both
turned so that the tubing is completely shut off.

Calculation of the percentage of Og is shown in the example
on p. 121.

The Tension of Carbon Dioxide in Alveolar Air.— By
the gas laws each gas in the alveolar air, and also the water
vapour, will exert a partial pressure or tension which is equal
to that which it would exert were it alone present in the space
occupied by the mixture of gases. This mixture consists,
approximately, of 5.5 per cent. CO2, 16 per cent, oxygen,
79.5 per cent. Nitrogen and it is saturated with water vapour
at body temperature, 37° C. Assuming that the barometric
pressure is 760 mm. Hg., then, to find the tension of CO2 in
alveolar air we must first of all subtract from 760 the aqueous
tension at 37° C. which equals 48 mm. Hg. and multiply by
5.5
100'

xSince absorption of C>2 is relatively slow, it is advisable for routine work to
have the reservoir B raised and lowered automatically. The most convenient
way for doing this is to attach it by string to one end of a hinged metal rod
which rests on a can placed on shafting that is made to revolve at a suitable
rate by means of a small motor.

2To correct for possible changes in the temperature of the water Haldane's
dummy tube is used. This consists of a 10 c.c. tube in the water jacket, con-
nected by capillary glass tubing with the lower end of D. The capillary tubing
is furnished with a stop cock so that it can be opened to the outside. The position
of this system is shown on the dotted lines in Fig. 32. At the start of an analysis
the stop cock is connected outside and the level of the meniscus of NaOH solution
marked on the tube /. The stopcock is then turned. If this level remians un-
altered after the analysis there can have been no change of temperature. If it
has shifted, it is brought back to the original level, while g and d are in com-
munication, by raising or lowering F. By this procedure the gas in A is com-
pressed or decompressed in proportion to the extent to which it may have ex-
panded by heat or contracted by cold. Before taking the final reading the
meniscus in d is of course readjusted by manipulating the screw clip a.

112 EXPERIMENTAL PHYSIOLOGY.

Because of the free diffusibility of CO2 through the pul-
monary endothelium, the same tension must exist in the blood
as in alveolar air. If the alveolar air be collected with the least
possible disturbance in breathing, this equilibrium will be
between the arterial blood leaving the lungs but if the breath
be held it will be between the venous blood as it comes to
them. The former value, i.e., the arterial tension of CO^, is
of very great importance partly because it regulates the H-ion
concentration of the blood going to the respiratory and other

H2CO3
centres (through the equation r ) an(^ Partly because of

its specific action on these centres. The most practical method
for collecting alveolar air without serious disturbance of breath-
ing is that of Haldane.

Experiment 49a. — Haldane's method for collecting alveolar
air. — The apparatus consists of a piece of wide bore rvibber
tubing about 1 metre long provided at one end with a mouth
piece, consisting of a glass or brass tube of similar bore to the
rubber tubing, and somewhat flattened at the free end, with a
short piece of narrow-bore tubing attached at right angles.
After dipping in antiseptic solution place the mouth piece in
the mouth of the observed person and direct him to close the
lips tightly round it, and to continue breathing, by inspiring
through the nose and expiring through the tube for a minute or
two so that the person may become accustomed to the pro-
cedure. Meanwhile attach a clean all-glass 10 c.c. syringe with
the piston lightly smeared with vaseline (if there is too much
vaseline it will absorb an appreciable amount of CO2) to the
side tube by means of a short piece of rubber tubing. Now
instruct the person to make a forced expiration (really to blow
out) through the tube for a period of time equal to that of an
ordinary expiration and to close the opening of the tube in the
mouth with the tongue when he has completed the expiration'
Immediately he has done this, withdraw the piston of the
syringe (not too hurriedly so as to avoid breaking the syringe)
and when a little over 10 c.c. of air has been collected ins.truct
the person to continue breathing through the tube as before.

ALVEOLAR AIR. 113

This sample of alveolar air is discarded by being pressed back
into the breathing tube, the object of its collection being to
have the side tube and nozzle of the syringe filled with alveolar
air (instead of outside air) when the main sample is taken.
After several normal breaths have again been taken through
the tube, collect another sample (of over 10 c.c.) of alveolar
air and remove the syringe meanwhile closing its opening
by the finger, and analyse in the gas burette.

The forced expiration from which the above sample of
alveolar air was taken followed a normal inspiration but it is
clear that the percentage of CO2 which it contains will be less
than that in a forced expiration made immediately after a
normal expiration. To find the average percentage of CO2 in
the alveolar air it is therefore necessary to collect another
sample taken from a forced expiration following a normal
expiration. This is a more difficult procedure since the person
must blow out after a normal expiration for a period of time
equal to that of the normal expiration. With a little practice
it can, however, be satisfactorily accomplished. Collect and
analyse for CO2 at least two samples of alveolar air following
inspiration and expiration. If the results do not check to
within 0.2 per cent, repeat the observations until they do.
Calculate the average tension of CO2 in the alveolar air. The
chief source of error in the above method depends on the fact
that the percentage of CO2 will progressively increase in each
succeeding portion of the air of the forced expiration, because
a constant amount of CO2 is being discharged from the blood
into a diminishing volume of alveolar air. It is for this reason
that great care must be taken to have the forced expiration
of the same duration as a normal one.

Determination of the percentage of CO2 which is in equi-
librium with the tension of CO2 in the venous blood of the lungs
is much more difficult. To do it with any degree of accuracy
it is necessary to determine the percentage of CO2 in a series
of fractions of a deep expiration which immediately follows
upon a deep inspiration from a gasometer, or rubber bag, con-
taining air with about 10 per cent, of CO2. The CO2 decreases
steadily in each succeeding fraction until a level is reached

EXPERIMENTAL PHYSIOLOGY

which represents the percentage of this gas which is in equi-
librium with its tension in the pulmonary venous blood.

A rough approximate idea of the venous tension of CO2 can
be obtained by determination of its percentage in the alveolar
air at the breaking point, which means the point beyond which

. J:he breath can no longer be held.

Experiment 49b.~ After a normal inspiration hold the breath
as long as possible, then make a forced expiration through a
Haldane tube and collect a sample of the air for analysis of
COj (two if possible). Repeat this experiment after taking a
deep inspiration immediately before holding the breath. Keep
a precise record of the time during ^which the breath was held,
in both observations. Count the pulse and examine for cya-
nosis during the time the breath is held. The percentage of
CO2 in alveolar air at the breaking point is considerably above
the CO2 tension of the normal venous blood because the blood
has completed the circuit of the circulation several times during
the time the breath was held. Each time the blood returns
to the lungs it of course carries more and more CO2 which
raises the tension of this gas.

The breaking point experiment really tells us the tension
of CO2 in arterial blood beyond which the activities of the
respiratory centre can no longer be inhibited by voluntary
effort. This tension will not always be the same because it
will depend on whether or not other chemical changes are
occurring in the blood that are capable of affecting the excita-
bility of the centre. One such change is the O2 tension of the
blood. When this becomes depressed below a certain level the
respiratory centre is stimulated and it is consequently to be ex-
pected that part of the stimulus at the breaking point is due
to this cause. That this is the case can be shown by repeating
the breaking point experiment after filling the lungs with
oxygen.

Experiment 49c.— Take a deep inspiration from a rubber bag or
spirometer containing oxygen, then hold the breath to the
breaking point and analyse a specimen of alveolar air for CO2
(and for O2 if possible). Keep a record of the time during
which the breath was held.

ALVEOLAR AIR. 115

The higher tension of O2 still remaining in the arterial blood
at the breaking point is not the only cause for the decidedly
longer time during which the breath can be held in the foregoing
experiment, another cause being that the CO2-combining power
of the blood will be less because it contains relatively more
oxy haemoglobin, than in the first experiment. Oxyhaemo-
globin is more acid than reduced haemoglobin.

In both of the above experiments examine the gums and
the base of the finger nails for evidence of cyanosis (blueness)
and note the exact time at which it appears.

The Effects of Forced Breathing.— When the breathing
is made as deeply as possible, without materially altering the
rate, the alveoli become excessively ventilated with the result
that the CO2 tension falls far below the normal level and free
CO2 diffuses from the blood. The free CO2 of the blood is
'washed out' or 'blown off' with the consequence that the acid-
base equilibrium of the blood changes, leaving a relative excess
of base. The condition of alkalosis hereby established causes
alteration in various physiological functions and the following
experiments are designed to demonstrate these.

Experiment 49d. — Have a person, while sitting, respire as deeply
as possible, at the normal rate, for ten minutes. During this
time count the pulse and measure the systolic blood pressure.
Examine also for evidence of cyanosis. After the forced breath-
ing, it will be found that for a time the person does not volun-
tarily breathe — apnoea. Observe the thorax very carefully to
note the time at which the first indication of breathing returns.
Note the character of the first breath and of those which
succeed it. Make a diagram showing the character of the
breathing; also, count the pulse as frequently as possible and
look carefully for cyanosis.

Experiment 49e.— Measure the percentage of CO2 in the alveolar
air of a person then instruct him to breathe deeply for ten
minutes (as in Experiment 43d), and during the last expiration
take another sample of alveolar air. Take a third sample at
the time the breathing just returns and a fourth one when the
normal breathing is reestablished (it will be advisable to have
at least two sampling syringes for these purposes). Chart the

116 EXPERIMENTAL PHYSIOLOGY

results of the analysis along with a diagrammatic curve of the
breathing. It will be found that the CO2 tension is greatly
lowered by the forced breathing (what is the reason?) and that
normal breathing returns at a tension which is decidedly below
the normal. How do you explain this result?

Repeat this experiment with the difference that the last of the
deep inspirations is taken from a rubber bag containing oxygen.
Note the duration of the apnoea, the occurrence of cyanosis
and the alveolar CO2 tension at the return of breathing. Ex-
plain the cause for any changes from the previous results.

When the forced breathing is kept up for some time, marked
changes occur in the acid base equilibrium of the blood because
of the blowing off of CO2. A condition of alkalosis results and
the kidneys respond by excreting urine with relatively less
acid than normal. The alkalosis also causes various symptoms
the most striking of which affect the nervous system.
Experiment 49f . — Measure the alveolar CO2 and the total acidity
of the urine of a person who has not recently performed the
forced breathing experiment. For the latter purpose, shake
about 15 c.c. of urine with about 0.5 gm. of a soluble oxalate,
filter, and titrate 10 c.c. of the nitrate against 0.05 N NaOH
using phenolphthalein as indicator. Also test the knee jerks
(p. 105). Have this person breathe deeply for half an hour,
unless the process causes great discomfort when it should be
terminated earlier. At frequent intervals observe the pulse,
test the knee jerks, look for evidence of venous congestion
(cyanosis) and test the excitability of the seventh nerve by
tapping with the finger at the point of its emergence from the
skull. In a normal person this does not cause any twitching
of the face muscles but as alkalosis developes such occurs and
becomes progressively more marked. (Chvosek's sign for
tetany). Also examine carefully for the appearance of tonic
contractions of the flexor and adductor muscles of the hand and
forearm. Towards the end of the period of forced breathing
this other symptom of tetany will often be quite evident but
if not so it can be made to develop by constricting the arm by
means of a blood pressure cuff, or a rubber band. (Trousseau's
sign for tetany). After discontinuance of the forced breathing

ALVEOLAR AIR 117

observe the duration of the apnoea (and compare with that
following less prolonged hyperpnoea) , and the manner of return of
normal breathing, look also for evidence of cyanosis. Measure
the alveolar CC>2 when breathing returns. Determine for how
long hyperexcitability of the motor neurones is evident. When
normal conditions become reestablished make a record of
the subjective symptoms (the sensations experienced by the
observed person). Find out whether any visual, auditory or
peripheral skin sensations (numbness, tingling, etc.) were
experienced. Finally collect another specimen of urine and
determine its total acidity.

CHAPTER XIII.

RESPIRATORY EXCHANGE IN MAN. (TISSOT-
CARPENTER METHOD).*

The principle of the method consists in the collection of a
definite volume of expired air in an accurate spirometer and the
subsequent analysis of a mixed sample of it. It is then possible to
compute the energy metabolism (Indirect Calorimetry). After
correcting for temperature and pressure, the total CO2 output and
O2 intake, and from them R.Q., are then computed.
Experiment 50. — -The subject sits on a chair and takes the mouth-
piece A (Fig. 33) of the respiration tube in his mouth. The
mouth-piece consists of an elliptical rubber flange with a hole
in the centre connected with the respiration tube. Two rubber
lugs are provided and these are gripped between the teeth, the
flange being placed between the lips and the gums. The nose
is tightly closed by a suitable clamp.

The RESPIRATION TUBE leads to the valves, B, (Douglas' or Pearce's), the
end of the expiration tube of which leads to a large stopcock, C, connected
with the interior of the spirometer. The latter consists of a 100-litre inverted
aluminium cylinder, D, suspended from a pulley, E, in a water bath between
the double walls of an upright stationary cylinder, F. As the aluminium
cylinder rises out of the water on account of air entering it, its weight be-
comes greater. To allow for this, the wheel is made excentric, (G), so that
as E revolves, with elevation of the spirometer, a weight connected by a
thread to the circumference of the excentric exercises a progressively greater
pull on the spirometer, and so exactly counterpoises it. The top of the
cylinder is provided with holes for the insertion of a thermometer and of a
tube and stopcock for drawing off the sample of air for analysis.

*Since the main thing to be learned in this observation, apart from the methods
of analysis of the air, is the method of calculation of the respiratory quotient
and of the total respiratory exchange, the experiment can be profitably assigned
to a larger group of students than the usual. After the sample of air has been
collected, certain members of the group will proceed to analyse it and make the
necessary calculations. Since the greatest cause for delay in this experiment is in
the analysis of the oxygen in the air sample, this procedure should be practised
before starting the experiment, and a student who has already had experience
with the gas burettes should demonstrate the technique to others.

118

RESPIRATORY EXCHANGE.

119

FIG. 33. Tissot-Carpenter Spirometer, Douglas mouthpiece and Pearce's valves. The dotted
lines in the diagram indicate the outline of the aluminium cylinder while in the water seal between
the outer and inner steel cylinders, the position of the latter of which is also indicated by dotted
lines. For further references see context. The inserts show the Douglas mouthpiece and Pearce
valves, the latter of which is made as follows: Prepared casings used in the manufacture of
bologna sausage are obtained preserved in salt, and they will keep indefinitely on ice. When
needed a short piece is taken, washed free from salt by allowing water from the tap to run through it,
and softened in a weak glycerine solution. The gut becomes very soft and pliable, and does not dry
quickly. A piece of the casing about 10 cm. long is threaded through a glass tube of about 15 mm.
bore and 4 to 6 cm. long. One end of the casing is brought around the outside of the tubing and
secured by means of a thread. The lower end of the membrane is pinched off and the casing is then
cut a little more than half way across its middle, so that the opening will lie just within the free end
of the tube when the casing is drawn back through it. The loose end of the casing is slightly
twisted — an essential procedure — and is then secured by a thread on the outer side of the tube.

120 EXPERIMENTAL PHYSIOLOGY.

In conducting the observation, the spirometer is first set at zero
on the scale at the side. The subject then breathes for some time
through the valves and a side tube with the stopcock turned off.
When the breathing is comfortable, the stopcock is turned so that
the air enters the spirometer, and the side tube is closed by placing
a clamp on the rubber tubing on its free end, the exact moment
being noted. The air is allowed to collect for a definite period of
time (4 to 5 minutes for a subject at rest), at the end of which the
stopcock is again turned and the side tube opened so as to connect
with the outside. The temperature in the spirometer and the
barometric pressure are then carefully noted, as well as the volume
of air in the spirometer. The number of respirations during the
observation is also counted.

The sample of air for analysis is collected in an all-glass syringe,
the nozzle of which fits into the rubber tubing of the stopcock on
the top of the cylinder. To allow for the dead space of the tubing,
the syringe,. after filling, is removed from the tubing (after turning
off the tap again), emptied and reconnected. The piston is then
moved out and in several times, and the syringe, filled with air, is
removed to the gas analysis burette and the analysis made according
to the directions in Exp. 49.

It is particularly important to make certain that all the oxygen
is absorbed by the pyrogallate and that the air entrapped in the
tubing leading to the CCVabsorption pipette is included in the
analysis.

THE CALCULATIONS. — The following have to be determined:

1. Per cent CO2 and O2.

2. Respiratory Quotient.

3. Total vol. CO2 and O2 respired per kgm. body weight and per square
metre of body surface.

4. Calorie expenditure (indirect calorimetry).

The following report form should be filled in as indicated, the necessary calcu-
lations being made by use of the tables in the appendix.

METABOLISM SHEET.

Name Age Wt. Ht.

Surface area calculated = (Du Bois method).

Corrected Volume of Respired Air.

(1) Total volume of air respired in 15 min. = 100 Litres.

(2) Actual Temp. = 20° C.

RESPIRATORY EXCHANGE. 121

(3) Actual Bar. Press. = 747 mm. Hg.

(4) Corrected Bar. Press. ((3) -Factor in Table III) =747- (17.4+2.41) =

727.19 mm. Hg.

(5) Volume of Resp. Air at 760 mm. and 0° C. ( (1) X Factor in Table IV) 100 X
0.89062=89.062 Litres.

(6) Corrected Volume of Resp. Air

(a) per mii1ute= 5. 937 Litres.
(6) per hour = 356.248 Litres.

Calculations of 02 A bsorbed and COi Eliminated.

(7) Volume per cent. CO2 in spirometer air =4.0.

(8) Volume per cent. O2 in spirometer air = 16. 5.

(9) Volume per cent. CO2 and O2 in spirometer air (7) (8) =20.5.

(10) Volume per cent. N in spirometer air (100) - (9) =79.5.

(11) Volume per cent. N in room air taken as 79 = 79.0.

(12) Oxygen equivalent of (10) = (10X0.265) =79.5X0.265 =21.06.

(This may also be obtained from Table I).

(13) Volume per cent. O2 absorbed (12) - (8) =21.06-16.5 = 4.56.

(14) Volume percent. CO2 eliminated (7) -(.03) =4.0-0.03 = 3.97.

(15) Total O2 volume (standard conditions) absorbed per hr. (13)X6(b) =

16.24 Litres.

(16) Total CO2 volume (standard conditions) eliminated per hr. (14)X6(b) =

14.14 Litres.

The Caloric Value Calculated from the Gas Exchange.

(Indirect Calorimetry).

This can be done by using table V (Appendix) provided we know the non-
protein R.Q, which is given in the 1st column of the table. This latter is obtained
by deducting from the total CO2 eliminated, the CO2 derived from protein (found
by multiplying the urinary N by 9.35) and by deducting from the total O2 ab-
sorbed, the O2 required to oxidise protein (found by multiplying urinary N by
8.45). Suppose for example, that 14.4 gm. N is excreted in the 24 hrs. urine;
i.e., 0.6 gm. per hr. then, 9.35 X0.6 = 5.610 gm. CO2 or (since 1 gm. CO2 = 0.5087 1)
2.85 1 must be subtracted from 14.14 Litres giving 11.29 Litres, and similarly
8.45 X. 6 = 5.070 gm. O2 or (since 1 gm. O2 = 0.7 1) 3.5490 must be subtracted

from 16.24 giving 12.69. The non-protein R.Q. is therefore - - =0.889.

1 — . * K '

Referring to table V we see that at R.Q 0.889 1 litre of O2 equals 4.91 C. .'. in
Ihr. 4.91X16.24 = 79.73C were expended. As a matter of fact for many pur-
poses it is sufficiently accurate to use the uncorrected R.Q. for table V.

CHAPTER XIV.

DETERMINATION OF THE GASES OF BLOOD BY THE
PUMP METHOD.

When blood is repeatedly exposed to a vacuum all of its gases are
evolved. The evolved gases can then be transferred to a suitable
burette and their nature and relative amounts determined. The
gas cannot be completely removed by one exposure of the blood
in a vacuum, because in such a case the gas would be evolved only
until equilibrium had become established between the partial
pressure produced by the evolved gas, and that still present in
the blood. It is necessary to repeat the evacuation several times.
For this purpose a mercury pump is usually employed, but satis-
factory evacuation can also be brought about by using the simpler
apparatus described in the following experiment.

Experiment 51. — Place about 15 c.c. defibrinated ox blood in
a 500 c.c. flask, and fill the latter with alveolar air by expiring
deeply into it through a piece of wide-bore rubber tubing, or better,
through a glass tube which is connected in its course with a bottle
filled with glass beads. This condenses and removes the water from
the air. Rotate the flask, so that the blood forms a thin film on the
walls, but do not shake in such a manner as to cause the blood to
froth. While rotating, occasionally expire through the tube into
the flask so as to maintain the percentage of carbon dioxide con-
stant. Continue this procedure for about three minutes, and then
close the flask. By this means the blood absorbs oxygen to full
saturation, and carbon dioxide to the same extent as the blood in
the pulmonary capillaries.

Meanwhile the bulb of the blood receiver (Fig. 34A) is partially
evacuated by connecting it, by means of the attached piece of
rubber tubing, to a (water) vacuum pump. The screw clip (1) is
then tightened (leaving as long a piece of tubing beyond the clip

122

BLOOD GASES.

123

as possible), the side tube of the pump opened* and the blood bulb
removed. A few drops of the anti-foaming solution (caprylic
alcohol or isoamylisovalerate), is placed in the receiver. This is

FIG. 34. Apparatus for measurement of gases of blood by pump method. For
description, see Context.

*The water must never be turned off from the pump while this is connected
with a partially or completely evacuated vessel. Always allow air into the pump
before turning off the water.

124 EXPERIMENTAL PHYSIOLOGY.

accomplished by taking about 0.5 c.c. of the fluid in a glass tube
of sufficient external diameter to fit tightly the rubber tubing of
the receiver (a 1 c.c. pipette with the delivery end partially cut
off). Before inserting the pipette into the rubber tubing, the lumen
of the latter beyond the clip is filled with the anti-foaming solution,
so that no air may enter the receiver, and after inserting, the screw
clip (1) is very cautiously opened and about 0.1-0.2 c.c. of the solu-
tion allowed to run in, after which the clip is again screwed tight,
the pipette removed, and the solution still in it replaced in the
stock bottle. The receiver is reattached to the water pump and
evacuated as far as possible.

It is then attached to tube D of the blood pump and evacuation
completed by manipulating the pump as described below for the
evacuation of blood. When completely evacuated, as judged by
the inability to suck over more air, screw clip 1 is closed and the
blood receiver removed from the pump.

Ten c.c. of blood is now placed in the receiver. To accomplish
this, blood is removed from the flask by means of the special 10 c.c.
pipette, using only gentle suction and filling to the upper mark.
All air is squeezed out of the tubing on the blood bulb, and the
end of the pipette inserted in the tubing, being careful to see that
no air bubbles are present at the union. Holding the pipette and
blood bulb vertically, the clip is very cautiously unscrewed, and
the blood allowed to flow from the pipette into the bulb until the
lower mark on the former is reached. The capacity between the
two marks is 10 c.c. After tightening the screw clip, the pipette is
removed and the blood left in the tubing is squeezed out.

The blood pump must now be prepared.

This consists of a 50 c.c. all-glass (Luer) syringe (B), with vaseline between the
walls and piston, the nozzle being connected by thick-walled rubber tubing, /,
with the single tube of a three-way stopcock (C). Of the other tubes of the
stopcock, one (D) runs to connect, by narrow-bore glass tubing, with the blood
bulb and the other (£) to the gas burette (F) — a 10 c.c. graduated pipette is satis-
factory. To avoid all risk of air leakage into the syringe, this is manipulated in an
o 1-bath (O) as shown in the figure. The barrel of the syringe is clasped at its
upper end by a brass collar (L) which is held by the iron rod (R) to the upright
(£/). To the latter is also hinged the free ends of another iron rod bent on itself
(S), the bend being about 30 cm. from the free ends. Two strips of brass are
loosely attached to the two arms of the bent rod on either side of the syringe
where they cross it, and these run down to connect with the head of the piston of the

BLOOD GASES. 125

syringe. This connection will vary with the exact shape of the head of the piston, but
in any case, free play must be possible at the joint. In the syringe used in this
laboratory the head of the piston is ring shaped and is satisfactorily connected
by fitting a wooden bobbin in the ring and attaching the brass strips to the end
of the bobbin. The syringe is surrounded by a glass cylinder containing a fairly
heavy mineral oil (Mobile oil) and this cylinder is attached to the iron lever so
that it moves up and down with the piston.

The first step in preparing the blood pump is to get rid of all
the dead space in the tubing and connections. This is readily
accomplished, for E, by turning stopcock C so that E communicates
with B, raising the levelling burette (G), and simultaneously with-
drawing the piston of the syringe by depressing the lever until
about 20 mm. of mercury has collected on the top of the piston.
The stopcock is then turned so that B and D are connected and
the piston raised until all the air is expelled and mercury completely
fills tube D. Any drops of mercury falling from the open end of D
must be caught in a small beaker. The mercury left on the top of
the piston seals this completely during the subsequent manipula-
tions.

After squeezing all air out of the tubing on the blood receiver,
this is connected with D, and immersed in a jug containing water
at 45° C. Having turned C so that B communicates with D, the
piston is then depressed to about the 20 c.c. mark, and while still
depressed the screw clip (1) is opened. About this time the blood
will begin to "boil" and the gases given off from it will pass into
the vacuum above the mercury in the syringe. C is turned so that
B is closed off and the piston allowed slowly to ascend. (It must
not be allowed to ascend too rapidly, since this might break the
syringe). The gas which has collected in the syringe is then ex-
pelled into the burette (F) by turning C so that B and E communi-
cate and pressing up the piston. After all the gas is out of the
syringe, the mercury is allowed to run into E a short distance,
being careful not to allow any to get into F. This first process
obviously removes only a small fraction of the total gas in the blood,
and it must be repeated several times exactly as described above,
until no more gas can be secured. The dislodgement of the gas
from the blood is greatly accelerated by warmth and by occasionally
removing the bulb from the water-bath and shaking briskly.

It is now necessary to measure and analyse the evolved gas.

126 EXPERIMENTAL PHYSIOLOGY.

For this purpose the piston is cautiously pushed up, with E and B
in communication, until mercury stands at the zero mark on the
neck of the gas burette. The clip (4) is then screwed down and
the levelling burette (G) lowered until the menisci stand at the
same level in it and the burette. This brings the gas to atmospheric
pressure and the volume is read and noted. The reading gives the
C.C. of gas in 10 c.c. blood. The volume should be reduced to
standard temperature and pressure (for calculation see Exp. 50).
To analyze the gas a 40 per cent, solution of sodium hydroxide is
sucked from a watch glass into a 2 c.c. all-glass syringe, and the
tube of the syringe inserted in the side tube (H). All air must be
expelled from this tube. With the pinchcock (6) closed, the clip
(5) is opened, while gentle pressure is being maintained on the
piston of the small syringe so that the mercury may not run into
it. The NaOH runs up to the top of the mercury column (F), and
when it is all in, clip 5 is again screwed down. The syringe (I) is
removed and F inverted several times so that the carbon dioxide
in the gas contained in it may be thoroughly absorbed. On now
opening clip 6, the mercury will rise in F, and by adjusting the
levelling tube the shrinkage in volume due to the absorption of
CC>2 can be ascertained and the percentage of this gas determined.
The reading is taken which corresponds to the top of the NaOH
solution, a similar amount of NaOH solution being placed on the
*.op of the mercury in the levelling tube. Care must be taken to
see that all the CO2 is absorbed.

To absorb the oxygen, about a gram of pyrogallic acid is dis-
solved in 2 c.c. of water in the watch glass, the solution is introduced
into jP. and the further manipulations conducted in the same
manner as for the NaOH solution. The gas which remains when
both CO2 and O2 are absorbed is nitrogen. There should not be
more than 0.1-0.2 c.c., any larger amount being due to air leakage
into the apparatus during the manipulations.* By taking proper
precautions, however, the residual nitrogen should never be more
than 0.3-0.5 c.c. The results are to be calculated to give the amounts
of each gas in 100 c.c. of blood.

*If any considerable amount of nitrogen is left, its volume should be measured,
and after subtracting C>2, the volume of C>2 that must have been introduced, as
air, calculated and subtracted from the actually observed O2.

BLOOD GASES. 127

When the analysis is completed, the mercury is run out from the
burette by the side tube (H), after removing the stopcock (C),
and the burette thoroughly washed with water. The mercury and
alkali pyrogallate solution (which is now brown in colour) are then
washed in running water until the washings react neutral to litmus
paper. The mercury should then be transferred to a separating
funnel containing a dilute solution of sulphuric acid. The blood
bulb should also be cleaned immediately, since otherwise a sticky
precipitate which is difficult to remove adheres to the walls.

CHAPTER XV.

DETERMINATION OF BLOOD GASES BY THE
CHEMICAL METHOD

Instead of pumping the gases out of blood, they may be dis-
placed by chemical means. Each method has its own advantages.
By the pump the gases are obtained unmixed with air so that their
analysis tells us directly how much of each is contained in the blood.
By the chemical method the oxygen that is loosely combined with
haemoglobin is expelled by shaking the laked blood with potassium
ferricyanide (the oxygen liberated during the reduction of this
salt to ferrocyanide displaces the oxygen which is loosely com-
bined with the haemoglobin, and takes its place to form methaemo-
globin). The carbon dioxide is expelled by adding a non-volatile
acid (saturated solution of tartaric acid). The volume of the
expelled gases may be measured in a suitable burette, if proper
care is exercised to avoid change in volume due to variation in
temperature. The chemical method is the more practical for most
work, but since the gases become mixed with the air originally
present in the apparatus, it is not suitable for proving what these
gases may be. Logically, therefore, in studying the blood gases
the pump method should precede the chemical.

Experiment 52. — The technique is as follows: The water
jacket of the burette (Fig. 35) (G) and the water bath of the blood
bottle (D) are filled with water that has been standing for some
time in the same room, and the temperature is noted. With the
stopper removed from the bottle the fluid (solution of calcium
chloride) in the burette is adjusted to the zero mark by raising or
lowering the levelling tube (H). 20 c.c. of CCVfree weak ammonia
water* i§ then placed in the bottle (C), and 2.5 c.c. (indicated by file
mark) of a freshly prepared saturated solution of potassium ferri-

*0.5 c.c. Aq. Ammonia per 1000 c.c. of distilled water to which some barium
hydroxide has been added and then some ammonium sulphate, the resulting pre-
cipitates of BaCO3 and BaSO4 being allowed to settle.

128

CHEMICAL ANALYSIS OF BLOOD GASES.

129

FIG. 35. Haldane's apparatus for analysis of blood by chemical method. (It
more satisfactory to replace B by a glass stopcock).

130 EXPERIMENTAL PHYSIOLOGY.

cyanide in the small flat-bottomed tube (A). About 15 c.c. of
defibrinated blood is exposed in a 250 c.c. flask, to alveolar air
obtained by expiring deeply into the flask and rotating so that the
blood forms a film on the walls. The blood will become saturated
with oxygen and it will take up CC>2 until there is equilibrium be-
tween the tensions of this gas in the air and blood. (Is this all the
CC>2 with which the blood could combine?) The rotation should be
kept up for about two minutes, the air in the flask being meanwhile
repeatedly replaced by alveolar air.

Immediately it has settled to the bottom of the flask, 10 c.c.
of the blood is removed by the pipette and, after wiping the tip
with a cloth, slowly delivered under the ammonia solution in the
bottle. The bottle is then gently shaken until the blood is com-
pletely laked and a transparent red solution is obtained. (In cases
where the blood is not saturated with oxygen, as, for example, in
venous blood, it is necessary to postpone the laking process until
after the bottle has been closed and connected with the burette,
since otherwise C>2 would be absorbed from the air) . Having placed
the flat-bottomed tube (A) upright in the bottle and with the air
outlet of the burette (B) open, the stopper is firmly inserted into
the bottle, which is then immersed in the water bath, and the water
stirred. Whenever the fluid in the burette ceases to move further —
indicating that the temperature of the air in the bottle has become
the same as that of the water bath — the stopcock B is again opened
to allow the meniscus of fluid in the burette to return to the zero
mark.

To displace the oxygen, the bottle is removed from the water
bath, and while holding it in a towel, to prevent its becoming
warmed by the hand, it is tilted so that the ferri cyanide spills into
the laked blood. The bottle is shaken for about one minute without
allowing the contents to come in contact with the stopper or tubing.
The expelled oxygen depresses the fluid in the burette, and as it does
so, the levelling tube should be lowered so that there may not be
increased pressure in the apparatus, which would encourage leaks.
The bottle is returned to the water bath, and the water stirred
until the level of fluid in the burette remains constant. The reading
on the burette, taken when the levels of fluid in it and the levelling
tube are exactly the same, gives the c.c. of oxygen expelled from

CHEMICAL ANALYSIS OF BLOOD GASES. 131

10 c.c. of blood. To make certain that all the oxygen has been
expelled, the bottle is again shaken. The two readings should agree.
Great care must be taken to keep the apparatus away from drafts
or other influences that might cause the temperature of the water
to change. If this occurs, the temperature in the bath and jacket
must be brought back to the original temperature before the final
reading is taken. For this purpose a small air thermometer T, in
the shape of a U-tube (containing coloured water) connected
with a test tube, (weighted with sand) is also placed in the bath.
If the meniscus of fluid in the thermometer changes during the
observation hot or cold water must be added to the bath and this
stirred until the original temperature is regained.

The carbon dioxide is measured by a repetition of the same
technique, using tartaric acid in place of ferricyanide solution.
The steps are as follows: the stopcock B is opened, and the meniscus
of fluid in the burette brought back to zero by raising the levelling
tube, and removing the stopper of the bottle. The reagent tube is
withdrawn and washed into the bottle with as small a quantity of
CCVfree water as possible (water that has been boiled and cooled).
2.5 c.c. of the tartaric acid solution is placed in the tube, the stopper
reinserted with stopcock B opened, temperature adjustment made,
and the CC>2 displaced and measured by the same manipulations
as for oxygen. The bottle must be very thoroughly shaken since
the CO2 is difficult to dislodge from the viscid mixture of pre-
cipitated blood proteins now present in the bottle.

When the estimations are completed, the flask, pipette, bottle,
reagent tube, etc., are washed thoroughly clean, and any fluid that
may have passed into the connecting tube, cleaned out thoroughly
with water and a pipe-cleaner.

CHAPTER XVI.

THE DISSOCIATION CURVE FOR OXYGEN AND THE CO2-
COMBINING POWER OF BLOOD.

For accurate determination of the relative amounts of reduced
and oxvhemoglobin in blood exposed to atmospheres containing
varying partial pressures of oxygen no method surpasses that of
Barcroft and his coworkers. The principle of this method is to
expose a small quantity of blood in a thin film on the walls of a
relatively large cylindrical vessel (tonometer) containing a mixture
of nitrogen and oxygen gases until equilibrium has become estab-
lished between the partial pressure of the oxygen in the atmosphere
and the absorption of oxygen by the blood. Some of the blood is
transferred to a small bottle connected with a differential mano-
meter and shaken with dilute ammonia water, whereby it becomes
laked, and by taking up oxygen, causes shrinkage in the volume of
air in the bottle, the degree of which is indicated by the manometer.
The oxygen-saturated blood is then shaken with ferricyanide of
potassium which dislodges the oxygen and causes the pressure in
the bottle to rise. From the relative displacement of the fluid in
the manometer in the two observations, the percentage saturation
of the blood with oxygen is readily calculated.

For use by a class of students we have found it more expedient
to expose the blood to a partial vacuum instead of a mixture of
gases, the partial pressure of oxygen being readily calculated
from the degree to which the tonometer is evacuated as measured
by a barometer. This simplifies the technique by making it un-
necessary to analyse the gas in the manometer. For very accurate
work, however, mixtures of Q% and N are preferable. After exposure
to the partial vacuum, the blood is transferred to a differential
blood gas manometer.
Experiment 53. — The following apparatus is required:

THE TONOMETER consists of a wide glass tube (the tonometer T, Fig. 36)
of fairly stout glass, tapering down to narrow tubes at both ends. The capacity

132

DISSOCIATION CURVE.

133

should be at least 200 c.c.* The narrow tubes are connected with thick-walled
(pressure) rubber tubing which should be wired on to the glass tubes. The rubber
tubes are closed by screw clips (1 and 2). File marks are made at one of the
tapering ends of the tonometer, the distances between them corresponding
approximately to one cubic centimeter.

FIG. 36. Apparatus for the determination of the Dissocia-
tion Curve. (In place of M the manometer shown in Fig. 36a
should be used) .

THE BAROMETER consists of a vertical thick-walled glass tube about 1.25
metres long and of about 3 mm. bore bent on itself near one end, and with the
other end dipping into mercury contained in a wider flat-bottomed (specimen)
tube (the mercury reservoir) closed by a perforated cork. The barometer tube

*It would be preferable to use a tonometer twice as large since this would
diminish errors due to the addition of the gas given off from the blood.

134

EXPERIMENTAL PHYSIOLOGY.

and reservoir are firmly mounted on a stand furnished with a millimeter scale,
which is attached to the stand in such a way that it can be adjusted to bring its
zero to the surface of mercury in the reservoir, as this varies at different pressures.
The free end of the barometer tube is connected by rubber pressure tubing to a
glass T-piece (A), one limb of which is similarly connected to a stout-walled
(filtration) flask (F) joined to a good water pump (P). A capillary tube closed
by a piece of rubber tubing and a screw clip (3) also passes through the stopper
of the flask.

THE DIFFERENTIAL MANOMETER. — The manometer
shown in Fig. 36a consists of a U-tube of capillary
tubing furnished with 3-way stopcocks the side tubes
of which connect with small bottles. Suspended from
the stoppers of the bottles are small glass spoons.
The fluid in the manometer is clove oil. The mano-
meter is mounted on a board with a hook on its back
by which the board can be hung on a square glass
(museum) jar containing water. The bottles should be
immersed in the water up to the necks.

The first step is to rinse out the tono-
meter with 0.9 per cent, saline and connect
it with the side tube of the barometer T-piece
A. The pump P is turned on with screw clips
1 and 3 closed, but screw clip 2 open and the
pressure lowered until the mercury stands at
a constant level in the barometer. Screw clip
4 is closed and the mercury observed, to see
whether there is any leak. Provided there is
none, clip 3 is cautiously opened and the
mercury allowed to fall almost to the level in the reservoir (R) ;
clip 2 is tightened, the tonometer, T, removed and the pump turned
off. Defibrinated or oxalated blood (whipped ox blood is most suit-
able for large classes, but in any case blood from an etherized
animal must not be used) is now sucked into the tonometer, by
placing some of the blood in a small evaporating dish, and, with
the rubber tube dipping into it, cautiously loosening clip 1; 3 to 4
c.c. of blood should be allowed to enter the tonometer. This is
then reattached to the T-piece A of the barometer and with clips
2 and 4 open (but 1 and 3 closed) the pump is turned on and the
mercury allowed to rise as far as it will go when clip 4 is closed .
and the pump turned off. Clip 3 is now cautiously opened until

FIG. 36A.

DISSOCIATION CURVE. 135

there is a partial pressure of about 20 mm. Hg. oxygen in the
tonometer.*

When the mercury has reached this level, or one near it, clip 3
is closed and the height at which the mercury stands very accurately
noted. Clip 2 is closed, after which the mercury is allowed to fall
to zero by opening 3. The tonometer is removed and rotated so
that the blood becomes spread out as a thin film on the walls,
after which it is placed in a water-bath kept about 40° C. in which
it is constantly rotated for about 15 minutes.

On removal from the bath the pressure in the tonometer must
again be measured. For this purpose the tonometer is reattached
to A and the pump is turned on (with 3 closed) until the mercury
has risen to the level at which it previously stood. Clip 4 is closed
and 2 opened. If there has been no leak, and time has been allowed
for the tonometer to cool down, there will be practically no differ-
ence between the two readings. If a difference of more than 5 mm.
is observed it must be noted and the pressure prevailing in the
tonometer taken as the average between the two readings.

Meanwhile 3 c.c. of freshly prepared weak ammonia water con-
taining a trace of saponin (0.5 c.c. aq. ammonia in 500 c.c. water)
has been placed in the blood gas bottle. A pointed glass tube
about 30 mm. long is now attached to the rubber tubing of the tono-
meter and this is removed from the barometer and held in a vertical

*This is computed as follows: After suitable adjustment the standard baro-
meter in the room is read and from the reading is deducted the tension of aqueous
vapour at the temperature of the room (for Table, see page 277 of Appendix).
The difference gives the pressure in mm. Hg. of an atmosphere of dry air. Since
air contains 20.96 oxygen, the partial pressure of this gas in the tonometer must be

20 96
equal to — : — ths. of the difference between the height to which the mercury is

-LUU

raised in B and the corrected barometer reading. Thus, suppose the room
barometer to be 753.4 mm. and the temp. 20° C., the corrected barometer reading
is 753.4-17.4 = 736 mm.

Then--?6 X736 = 154.2 mm. 02

-LUU \

20 X 736

Suppose a tension of 20 mm. 02 is desired, then = =95.45 mm.

154 .2

That is the mercury in the barometer must be raised to 736 — 95.45 or 640.55 mm.
above the level in the reservoir (R).

136 EXPERIMENTAL PHYSIOLOGY.

position above the bottle. The screw clip 2 is opened so that the
air enters the tonometer, the clip 1 is then cautionsly loosened to
let a drop or two of blood flow out from the tip of the glass tube,*
and after closing it again the end of the tube is wiped free of blood
and placed in the bottle so that it dips under the ammonia solution.
Clip 1 is now cautiously opened and about 1 c.c. of blood allowed to
flow under the ammonia water. If this is done carefully the blood
does not mix with the ammonia water but this floats on the top as a
layer and so prevents any diffusion of oxygen between the blood
and the air. The bottle is firmly closed by its stopper, the stopcock
being meanwhile open to the outside so that the level of the clove oil
in the manometer is not disturbed. The bottle must then be sub-
merged in a water-bath containing water at about room tempera-
ture, in which it is left until, with the stopcock closed, no further
contraction of volume, due to cooling, is observed to occur.

The manometer is now removed from the bath and vigorously
shaken so that the blood becomes laked and absorbs C>2 from the
atmosphere of the bottle. On replacing the bottle in the bath and
allowing time for cooling the difference between the levels of clove
oil in the two limbs of the manometer is noted. With the stopcock
open to the outside, the stopper is removed from the bottle and
about 0.25 c.c. of a freshly prepared saturated solution of potassium
ferricyanide is placed in the glass spoon suspended from the stopper
without allowing any of the ferricyanide to mix with the laked
blood. After reinserting the stopper and cooling, the bottle is
again removed from the bath and shaken so that the ferricyanide,
by mixing with the laked blood, drives off the loosely combined
oxygen and raises the pressure, which is measured by the mano-
meter.

The relative amounts of reduced haemoglobin and oxyhaemo-
globin present in the blood are proportional to the first and second
readings of the manometer; when all is reduced haemoglobin the
diminished pressure (shrinkage) recorded in the first shaking of the
bottle is the same as the increased pressure recorded in the second.

*Enough blood should be run out to bring the meniscus of blood in the tono-
meter to the upper file mark.

DISSOCIATION CURVE. 137

The calculation of the percentage saturation of haemoglobin
with oxygen is made by subtracting the first reading from the
second, dividing by the second reading and multiplying by 100.
Suppose in the observation made at 20 mm. partial pressure of O2
the first reading is 24 mm. and the second, 108, then
108-24

77-°% Hb° and 22-3% Hb

The result must now be plotted on coordinate paper with the
percentages of HbO along the ordinates and the partial pressure of
Oxygen on the abscissae. The experiment should be repeated,
using 10 mm. and 40 mm. pressures of oxygen, and the results
similarly plotted. By joining the points, the dissociation curve for
blood is obtained. Care must be taken to see that the bottle is
sufficiently shaken so that the partly reduced blood absorbs all the
oxygen and gives it up again with ferricyanide. It is particularly
in the latter operation that care must be taken.

The Influence of Carbon Dioxide in Lowering the Dissocia-
tion Curve can be readily shown by the method. The procedure
is as follows: After the pressure has been reduced to the desired
degree in the tonometer, the latter is placed in a horizontal position
so that the blood lies along the walls, free of the ends. A CC>2
generating apparatus (Kipp's) or a bottle containing this gas is
then connected by suitable tubing with the free end of the tono-
meter, care being taken before making the connection, to fill the
tubing with CCV To accomplish this a slow stream of the gas is
maintained and the air in the tubing beyond the screw clip (1) is
squeezed out before connecting with the CC>2 generator. The most
suitable partial pressure of CC>2 to work with is 40 mm. and to
attain it the C(>2 apparatus is first of all opened and the screw clip 1
very cautiously loosened until, with clip 2 open but 3 and 4 closed,
the mercury descends about 40 mm. in the barometer. Clips 1 and
2 are then tightly screwed down, and the tonometer removed, the
further procedure being exactly as described above.

The effect of the 40 mm. of CO2 will be found in the above
example, where a partial pressure of 20 mm. C>2 was used, to reduce
the percentage of HbO from 77 to about 35.

138 EXPERIMENTAL PHYSIOLOGY.

THE C02-COMBINING POWER OF THE ALKALINE
RESERVE OF THE BLOOD.

After completing the estimations necessary for finding the per-
centage of oxy haemoglobin, in the experiments in which CC>2 is
present in the tonometer, it is of interest to determine the amount
of this gas with which the blood has combined. This will repre-
sent its ability to act as a buffer towards foreign acids. To perform
the estimation it is necessary, however, to measure accurately the
amount of blood which is removed from the tonometer to the blood
gas bottle. This can readily be done by attaching a 1 c.c. pipette
to the tubing of the tonometer (beyond clip 1), a few drops of blood
being allowed to escape from the pipette before delivering under
the ammonia solution in the bottle, and precautions being taken
not to remove any of the upper layers of blood that had been exposed
to full atmospheric pressure when the tonometer was opened.
This is ensured by removing the pipette from the tonometer before
all the blood has run out.

To dislodge the CO2 from the blood, the stopper is removed
with the usual precautions and about 0.25 c.c. of a saturated solution
of tartaric acid placed in the small test tube. After closing and
allowing for temperature changes, the acid is shaken with the
mixture of blood and ferricyanide, and the CO2 thereby evolved,
measured by multiplying the displacement of the fluid in the
manometer by a figure (the constant of the apparatus) obtained by
a preliminary experiment in which a known amount of a standard
carbonate solution is similarlv treated.

SECTION V
SPECIAL SENSES.
CHAPTER XVII.

VISION.

In order to gain accurate information through the sense of sight
about objects in the external world, their size and shape, and
their positions relative to one another, two things are necessary.
There must in the first place be a group of cells sensitive to light
waves and so connected to the central nervous system that stimu-
lation of any part of the sensitive surface gives rise to a sensation
different from that caused by stimulation of any other part.
Secondly some system of lenses is needed so that each point in
the sensitive layer does not receive rays from all directions in the
external field, but has focussed on it only those which arise from a
single part of the field. In the eye the sensitive elements are con-
tained in the retina, and the cornea and lens together make up the
focussing apparatus. In the discussion and experiments which
follow we shall first consider the way in which light waves are
brought to a focus in the eye and later the response of the retina
to them. Before taking up the complex arrangement of refracting
surfaces which exist in the eye it is well to review some of the simple
cases of the formation of images both by lenses and by mirrors.
Although the focussing in the eye is entirely done by refracting
surfaces and reflection plays no part in it, it is by reflected rays
that one examines the condition of the eye and on this account
it is necessary to have clearly in mind the laws which underlie the
reflection, as well as those of refraction.

Physiological Optics.
Reflection.

When rays of light diverging from a point are reflected by
a mirror their course is so changed that they appear to an observer

139

140

EXPERIMENTAL PHYSIOLOGY.

to come, not from the actual object itself, but from some other point
known as the IMAGE of that object. Each point on a luminous
object has a point image corresponding to it. The position of the
image of the whole object may be found by defining the position
of the image of each of its extreme or limiting points. To find these
one applies the LAWS OF REFLECTION, which state (1) that the inci-
dent ray (that is, the ray before reflection), the reflected ray, and
the perpendicular to the surface at the point of incidence, are all
in one plane, and (2) that the angle between the incident ray and

FIG. 37. To illustrate the formation of an image by a plane mirror.

the perpendicular (or the ANGLE OF INCIDENCE) is equal to that
between the perpendicular and the reflected ray (the ANGLE OF
REFLECTION). Fig. 37 shows the construction for a plane mirror.
PQ is the object, PA and PB, incident rays, and AE and BF, the
perpendiculars to the surface at the points of incidence. These,
together with the reflected rays, AC and BD, are all in the plane
of the paper, i and i' are the angles of incidence. After reflection
the rays AC and BD pass in such a direction as to make each
angle of reflection equal to the corresponding angle of incidence
(r = i, r' = i'.) These rays both appear to come from a single point

PHYSIOLOGICAL OPTICS.

141

behind the mirror P', the position of which is found by projecting
the lines of the reflected rays AC and BD back until they meet.
Since ALL rays from the point P on the object appear after re-
flection to come from one point on the image, P', the place from
which two of the rays seem to come, must be the apparent source of
all the others, in other words, P' must be the image of the point P.
The construction for Q is similar, Q' being its image. When the

FIG. 38. To illustrate the formation of an image by a concave mirror.

reflected rays only appear to come from the image, as in these
cases, and do not actually pass through it, the image is called a
VIRTUAL one. It will be noted that it is erect.

Fig. 38 shows the construction for reflection of a similar object by
a concave mirror. C is the centre of curvature of the mirror; any
line from C to the surface is perpendicular to it, since it lies on one
of the radii. One ray from P, passing through C on its way to the

142

EXPERIMENTAL PHYSIOLOGY.

surface, lies along such a line. It is reflected back along the path
on which it came (since i = o in this case, r must also =o). PA,
another incident ray from P, is so reflected (ray AB) that the angle
of incidence (i) equals the angle of reflection (r). The place (P')
where this crosses the reflected ray PC is the image of the point P,
from which these and all other reflected rays from P appear to

M

FIG. 39. To show refraction of rays passing from air into water.

come. The construction of the image, Q', of the other limiting
point, Q, is similar. The rays in this case actually pass through
the image from which they seem to come, and the image is there-
fore called a REAL one. Real images are always inverted.

Refraction.

Refraction at a Plane Surface. — When a ray of light passes
from air into a denser medium such as glass or water, the direction

PHYSIOLOGICAL OPTICS. 143

in which it travels is altered and it is bent toward the perpendicular
to the surface between the two media (Fig. 39). The angle between
the incident ray and the perpendicular is the angle of incidence
(i and i'), that between perpendicular and refracted ray, the angle
of refraction (r and r'). The extent to which this bending or
REFRACTION occurs depends on two things. One is the angle of
incidence. The greater this angle is, the more the ray is refracted,
provided of course that the medium is the same. The other factor
is the nature of the medium; the denser the medium the more it
refracts. It is found that for a given medium there is a constant
relationship between the direction of the incident ray and that of the
refracted one ; the sine of the angle of incidence divided by the sine of
the angle of refraction is always the same for the same medium,
greater when the medium is dense and less when it is rare. It is
usual to express the refracting power of a medium in this way and
to call it the REFRACTIVE INDEX, designated by ju. The example in
Fig. 39 will make this clear. AC and AI are incident rays,
CD and IJ, the rays after refraction. The angles of incidence are
i and i', the angles of refraction, r and r'.

Sine of angle of incidence

c^ r i r — r 7- — = M f°r water = 1.3;

Sine of angle of refraction

BE

BC BE GH

that is — or (since BC = CD) — =1.3. Similarly — =1.3.
L>r Dr J rv.

CD

The ray ALM, being perpendicular to the surface, is not refracted
but passes through unchanged in direction.

If the direction of the light is reversed and if it passes from the
dense to the rare medium the direction of refraction is also reversed,
the rays being bent away from the perpendicular instead of towards
it.

Refraction at a Convex Surface. — It is through a convex re-
fracting surface that the light first passes on entering the eye, as it
goes from the air into the layer of tears and the cornea.

The central point on a curved refracting surface is called its
PRINCIPAL point (P.P. Fig. 40) ; a line joining this with the centre
of curvature (N.P.) is the principal axis of the refracting surface

(P. A.). That ray from the
object which is directed
straight towards the centre
of curvature or NODAL POINT
(N.P.) of the refracting sur-
face lies of necessity on one of
the radii to the surface; it is
therefore perpendicular to it
and is not refracted (ray 2).
If a pencil of rays are parallel
to one another and also
parallel to the principal axis
(see ray 1), they are all
brought to a focus after re-
fraction at some point on the
axis, this point being known
as the SECOND PRINCIPAL
FOCUS (P.F2, Fig. 40). This
lies nearer the surface if the
curvature on the surface is
sharp and the refractive index
high, farther back if the sur-
face is part of a large circle,
or if the refractive index of
the medium is not very great.
Light to be parallel must in
theory come from objects at
an infinite distance. In actual
fact, however, rays from
points more than about ten
metres away diverge so little
that, for the eye at least,
there is no practical differ-
ence between their focus and
that of rays which arise from
further away. All parallel
rays which reach the surface
inclined at a small angle to
the principal axis are brought

PHYSIOLOGICAL OPTICS. 145

to a focus at some point in the plane of the principal focus or the
PRINCIPAL FOCAL PLANE (P. P.P.). Rays which are appreciably
divergent when they reach the surface (that is which arise from
objects less than about ten metres away) have their foci behind
the principal focal plane. The focus moves farther back the nearer
the object is brought until finally a position of the object is reached
such that the rays from it after being refracted meet only at in-
finity, that is, they are parallel to one another. This position is
known as the first principal focus (P.P. Fig. 40). If the object is
brought nearer, the rays are merely rendered less divergent after
refraction and are not brought to a focus at all.

Fig. 40 shows the way in which the position may be found of an
image formed by refraction at such a surface. From each limiting
point on the object, any two rays, the paths of which are known, are
followed until they meet. The place where they cross is the image
of the point from which they arise.

Refraction by a Convex Lens. — Rays, after they have entered

the eye, have to pass through a convex lens, the crystalline lens,

before they reach the light-sensitive cells. The refraction in this

case is similar to that which we have just considered; like a simple

convex surface, a convex lens converges rays which pass through it.

The extent of the refraction depends on how sharply the surfaces of

the lens are curved and on how much denser its substance is than

the surrounding medium. The lens has a principal axis, which

joins the centres of curvature of the two surfaces, a first and second

principal focus, and a principal focal plane. The point where the

principal axis cuts the surfaces is the principal point; rays which

pass through here are not appreciably bent if the lens is a thin one.

Experiment 54.— Set in front of the opening of the lantern the

ground glass screen, and the diaphragm with a vertical slit.

Through a convex lens throw an image of the slit on the black

wooden block. The slit is to serve as the "object". Now place

against the lens a sheet of paper perforated with two holes,

about 3 mm. in diameter, horizontally placed and less than the

diameter of the lens apart. This stops all but two pencils of

rays from the object. Note that the image does not disappear,

but becomes less bright. Cover one hole and see the further

dimming of the image.

146 EXPERIMENTAL PHYSIOLOGY.

Move the object further from the lens. There are now two
blurred images of the object. Cover the left hole and note
that one image disappears.

Place the object at a distance from the lens less than the
first. Note that in this case also a double image is formed.
Again cover the left hole and see whether the image which dis-
appears is the same one as before.

Draw diagrams of the formation of images by the lens when

the object is (a) at a distance from which the rays are focussed

on the screen, (b) at a greater distance, (c) at a less distance.

Show whether the image in each case is real or virtual.

Since the focal length of a lens varies inversely as its strength,

being shorter the stronger the lens, it is usual to express the strength

of a lens in terms of the reciprocal of its focal length. The standard

is a lens which has a focal length of one metre and this is said to

have a strength of one dioptre (D = ^r, where F = focal length,

r

D = strength in dioptres).

Refraction in the Eye.

If the rays from an object pass in their course through a succession of focussing
surfaces, a calculation of the position, size, etc., of the image which they finally
form can be made along the lines which we have indicated as long as the surfaces
are only two, or at most three, in number. When they are more than that the
problem becomes unwieldly. In such a case, however, it is often possible by
mathematical calculation to arrive at an ideal, or imaginary, single convex refract-
ing surface which will have approximately the same power as that of all the
surfaces together. For this the radii of curvature and the refractive indices of all
must be known and the surfaces must be "centred". Surfaces are said to be
centred when they all lie along the same principal axis. Light entering
the eye has to pass through a large number of refracting surfaces,
through cornea, aqueous humour, lens, and vitreous humour. Of these,
the cornea, the aqueous humour and the vitreous humour have refractive indices
which are much alike, all about equal to that of water (ju = 1.3). The lens is
denser and all parts of it have not the same composition. It is made up of more
or less concentric layers about a central core and the density increases from the
outer layers (ju = 1.40) to the core (// = !. 44). When they enter the cornea from
the air, light rays undergo the main part of the refraction which occurs in the eye,
because the densities of the two media are so different. The other con-
siderable refraction occurs on the passage of the rays through the denser substance
of the crystalline lens. In the complex system of the eye the refractive index

PHYSIOLOGICAL OPTICS. 147

and the radius of curvature of each part is known and the surfaces, although
not accurately centred, are sufficiently nearly so to make it possible to find
mathematically a single surface which represents the whole. This is used in the
construction of the SCHEMATIC EYE (Fig. 41). The single refracting surface is made
to lie a few millimetres behind the real cornea. In the unaccommodated eye
distant objects are clearly seen, which must mean that the rays from them, which
are parallel or nearly so, form sharp images on the light-sensitive surface. The
refracting surface of the schematic eye, to represent this, has its principal focal
plane on the retina. From a distant point object situated in any part of the
visual field, we know the course of one ray, that which is directed straight for the
nodal point of the simplified eye and which is not changed in its direction before
reaching the retina. By the previous argument we have shown that all rays from
such a point meet at the retina. Therefore to find the image on the retina of any
object, all we need to do is to draw straight lines from its limiting points through
the nodal point of the schematic eye.

FIG. 41. The formation of an image by the eye as represented by the schematic eye.

It will be seen that according to the foregoing construction the
retinal image is an inverted one. That we interpret this to our-
selves as an erect picture of the object may at first sight appear con-
fusing. It should be remembered, however, that we are not born
with the faculty of associating stimulation of any particular part
of the retina with the presence of some object in a particular part
of the external world. We learn this in infancy through our other
senses, mainly the sense of touch. Because a point on one side of
the retina is as a matter of fact always affected by light from some
object which we know by our other senses to be on the opposite
side of the field, we form the habit of interpreting the stimulation
in this way. We associate stimulation of the right side of the retina
with the presence of something in the left side of the visual field
and so on. To put the matter more generally, we refer stimulation
of any part of the retina to the presence of some object in the out-

148 EXPERIMENTAL PHYSIOLOGY.

side world situated along the line which joins that part with the

nodal point of the eye. That this is actually the case may be seen

from the following experiment.

Experiment 55. — Close the right eye and direct the eyes as far as
possible to the left. With the tip of the finger press lightly on
the right eye at different points near the margin of the orbit
and note the positions in the visual field of the resulting "phos-
phenes".
In this experiment although we press the right side of the retina

all the light spots which we see appear to be on our left, in the upper

field if we press below and in the lower if we press on the upper part

of the eyeball.

CHAPTER XVIII.
ERRORS IN REFRACTION.

PHYSIOLOGICAL ERRORS. ACCOMMODATION.
SPHERICAL ABERRATION.

In the description of refraction by lenses it has been stated that
a spherical lens brings all the rays from one point on the object to
a focus at the same point. As a matter of fact this is not strictly
speaking the case. The rays which pass through the outer parts
of the lenses are more refracted than those nearer the centre; they
are therefore brought to a focus a little in front of the central ones,
and, passing on, blur the image which the latter form. The differ-
ence between the foci becomes greater the nearer the object and the
more divergent the rays from it.

>j Experiment 56. — Fill the bottle with water and use it as a lens.*
Cover the opening in the lantern by the diaphragm with a
2 mm. opening, and set it about a meter away from the bottle.
Move the black screen in the pencil of light coming through the
lens until you have the light as sharply focussed on it as possible.
Note that the region on either side of the focus is dimly lighted.
Interrupt the light coming through the outer parts of the lens
and note that the focus becomes sharper. Bring the lantern
nearer the lens, set the screen at the new focus and interrupt
the outer refracted rays as before. The improvement in the
sharpness of focus is more marked.

Cover the round opening of the optical box with the clear
glass slide. Set the bottle immediately inside the opening, light
the incense in the cork and cover the box. When it has filled
with smoke place the lantern about a meter away, so that the

*The lens in this case is of course a cylindrical one. Since, however, we have
to do only with those rays which diverge horizontally, we may use the refraction
which the curved surface effects in these to represent refraction by any one plane
of a spherical lens.

149

150 EXPERIMENTAL PHYSIOLOGY.

rays pass through the bottle, and look directly down on the
refracted rays. The boundaries of the light pencil are curved,
and not plane, surfaces because the outer rays, being more
refracted than the inner, intersect the latter. Move the lantern
nearer and note that the curvature of the surfaces increases.
The refraction in the eye is corrected for spherical aberration to
some extent by the difference in the refractive indices of the differ-
ent parts of the lens. The central rays pass through the part of
the lens which is the most dense and so they are refracted more, in
comparison with the rays through the periphery, than they would
be if all the layers were of the same composition.

Chromatic Aberration. — Light which arises from objects seen
under ordinary circumstances is made up of waves of different
lengths. In passing through the refracting media each wave length
is bent to a slightly different extent and this causes another error in
the refraction of the eye. The shorter waves of the violet end of
the spectrum are brought to a focus nearest the lens, the long ones
of the red end are least refracted, while the foci of those of inter-
mediate length lie between these two extremes.*
Experiment 57. — Chromatic Aberration in Refraction by a
Convex Lens. — Set in front of the opening of the lantern the
ground glass screen and the diaphragm with a 2 mm. opening.
Place the block holding the convex lens about 15 cm. from the
opening. Using a sheet of paper as a screen move it back and
forth in the path of the refracting light until you find the focus.
Note that it is not pure white, but made up of coloured bands.
Cover the right half of the lens with a card. The light has
violet fringe. Uncover the lens and move the screen a little
nearer to it. The disc has a violent centre and red border.
Move the screen beyond the focus. The colours of the disc are
reversed.

Experiment 58. — Chromatic Aberration in the Eye. — Cover
with a card the outer half of the right pupil and, closing the
left eye, look at an electric light filament. It appears to have
a red border at the right and a violet one at the left. Draw a

*For the way in which chromatic aberration is done away with in lenses of fine
optical instruments by using layers of different dispersive power, the student
is referred to his text-books on Light. There is no such adjustment in the eyr«

ERRORS OF REFRACTION AND ACCOMMODATION. 151

diagram of the course of the red rays and of the violet ones,

using the schematic eye, and explain the apparent contradiction

between the results of this and of the preceding experiment.

When we turn our eyes to look at a certain object we habitually
place them so that the image falls on the fovea centralis. This is the
part of the retina which is capable of seeing most acutely, of making
out differences smaller than can be perceived by any other part, and
the line which connects it with the nodal point is called the VISUAL
AXIS. If this corresponded with the optical axis the refraction
would be the best that the refracting media of the eye are capable
of. In point of fact, however, there is an angle of about 5° between
the two and this is a slight additional source of error in refraction
in the eye.

ERRORS TENDING TO BE PATHOLOGICAL. — The defects in refraction which have
been described so far are found in every eye. As well as these there are several
types of faulty formation of images which are often found, any one of which if
it is at all pronounced makes the vision of the eye abnormal.

NEAR-SIGHTEDNESS OR MYOPIA. — In near-sighted eyes light from distant
objects, that is for practical purposes parallel light, is brought to a focus in front
of the retina instead of exactly on it. The rays therefore when they reach the
retina are already diverging from their focus and they form a blur on the light-
sensitive layer instead of a sharp point of light (Fig. 42 B.). This may be
because the curvature of one or all of the refracting surfaces is unusually sharp, the
length of the eyeball being normal, or it may, as is more frequently the case, be
due to an unusual length of the eyeball. There is no physiological correction for
this condition. In practice it is rectified by placing in front of the eye a ccncave
lens. Light from distant objects is thus made divergent before it reaches the eye
and its focus therefore lies on the retina, further back than the principal plane of
the shortsighted eye.

In LONG-SIGHTEDNESS OR HYPERMETROPIA the condition is reversed. The
principal focus of the refracting media lies behind the retina and the parallel rays
from distant objects have not yet arrived at their focus when they are interrupted
by it (Fig. 42, C.). An increase in the refractive power of the eye
is needed to bring the focus forward and give a clear image. An effort of contin-
ued accommodation (see below) will accomplish this, but such an effort gives rise
to various nervous symptoms, headache, irritability and the like. If convergent
glasses of suitable strength are used, the focus may be advanced the necessary
amount without any effort on the part of the patient.

ASTIGMATISM. So far as we have considered the eye as refracting all rays to
the same extent no matter in what plane they diverge from their object nor
whether they pass through the upper and lower parts, or through the right and
left sides, of the refracting surfaces. Cases are fairly common, however, in which
this does not hold good. In these eyes, known as astigmatic, all the meridians

152

EXPERIMENTAL PHYSIOLOGY.

are not curved alike. In the most common form the vertical meridian is an arc of a
smaller circle than is the horizontal and, being more sharply curved, it refracts
more than does the latter. Rays which diverge in a vertical plane from a point

T.T.

FIG. 42. The focussing of parallel rays from a distant object by (A) a
normal, (-B)e. short-sighted and (C) a 'long-sighted eye. PF in each case shows the
position of the principal focus of the combined refracting surfaces of the eye,
represented by the single surface of the schematic eye.

object are brought to a focus by such eyes in front of the focus for the horizontal
rays. The result is that the eye can never see a point image of a point object.
If the rays through one meridian are focussed on the retina then those which pass
through the meridian at right angles to the first must reach the retina as a pencil

ERRORS OF REFRACTION AND ACCOMMODATION. 153

of rays, either because they have already passed, or because they have not yet
reached their focus. The unlike meridians are not necessarily arranged in this
way, the vertical is not always that of greatest curvature, although this is the most
common form, nor are the meridians which differ most always at right angles to
one another. When they are the astigmatism is a REGULAR one; when the angle
between is not a right angle it is an IRREGULAR astigmatism. Regular astig-
matism is easy to correct by glasses ground so as either to converge the rays which
are to pass through the meridian of least refractive power or to render more
divergent those directed towards the meridian of greater refractive power.
Irregular astigmatism is more difficult to correct. In some eyes the astigmatism
is of neither of these types but consists in irregularities in the curvature of the
same meridian, a condition which cannot be made right with glasses. Even in a
normal eye there is something of this irregularity. The image of a star is not a
single point, as it would be if the refracting surfaces were of even curvature
throughout, but it has an irregular shape which is different for each individual,
because of the slight defects in the curves of the various surfaces.

Accommodation. — AVe have seen how parallel rays from a dis-
tant object are brought to a focus on the retina of a normal eye.
When however the gaze is directed to an object close at hand the
eye receives rays which diverge from one another by a considerable
angle. If no change takes place in the refractive power of either
cornea or lens these rays tend after refraction towards a focus
lying behind the retina. In consequence when they are intercepted
by the retina they form a blur and not a sharp point. To make the
image of the near object a clear one either the distance between
retina and lens must be made greater than it is for distant vision,
just as one increases the distance between lens and plate in a
camera when focussing for objects close at hand, or else the refrac-
tion by the eye media must be increased. The latter is the change
which is brought about in the mammalian eye by the act of accom-
modation. The curvature of the anterior surface of the lens is
made sharper and its refraction therefore greater.

To understand the most generally accepted explanation of the way in which
this is done, one must have clearly in mind the anatomical relationship which the
lens bears to neighbouring structures in the eye (Fig. 43). It is held in its place
by the numerous fibres which together make up the suspensory ligament of the
lens. These are more or less radially arranged about the lens, being continuous
at their inner ends with the capsule near the margin, while the outer ends of the
threads are connected to the surface of the ciliary body near its free inner edge
throughout its entire circle. The arrangement is such that those fibres which
come from the posterior surface of the lens capsule go to the anterior surface of the
ciliary body and those which arise from the anterior surface of the lens are at-

154

EXPERIMENTAL PHYSIOLOGY.

tached to the processes of the posterior surface of the ciliary body as far back as
the ora serrata of the retina. The contents of the eyeball are under a pressure
which is greater than atmospheric by about 25 cms. Hg. They therefore press
outwards on the coats of the eyeball and as a result of this the choroid coat and
the ciliary body, its forward continuation, pressed out by the vitreous humour,
pull backward as well as outward on the lens ligament. This in its turn exerts
more pull on the anterior than it does on the posterior surface of the lens capsule.
The lens itself is fluid in nature and its capsule is elastic. Its natural shape is a

Corn-*

FIG. 43. Diagram of the attachments of the lens, and of the neighbouring structures.

more or less rounded one but, being flexible, it yields to the pull of the ligament
and both its surfaces are flattened, the anterior however much the more so.
The act of accommodation is a contraction of the circular and radial muscle
fibres which are contained in the ciliary body. This makes the circle of the
free margin of the body smaller, brings it nearer to the lens, and causes the
processes of the posterior part to move forward, dragging with them the anterior
part of the choroid, with which they are continuous. The fibres of the ligament

ERRORS OF REFRACTION AND ACCOMMODATION. 155

which pass between the posterior processes of the body and the anterior surface
of the lens are slackened by these changes and the lens is allowed to bulge forward
by its own elasticity.

The extent to which the curvature can be increased by accom-
modation is limited only by the elasticity of the lens.
Experiment 59. — The Near Point. — The least distance at which
an object may be held away from the eye and still be clearly
seen is the NEAR POINT. Find how far away your own near
point lies by looking fixedly at a pin or pencil-point with one
eye closed and gradually bringing the object closer to your face.
A distance is found nearer than which if the object is held it is
seen blurred. The blurring means that your accommodation is
no longer sufficient to bring all the rays from a single point to a
focus on the retina, the focus lies behind the retina, and the
rays from each point reach the sensitive cells as a pencil, and
not as a point of light.

The near point in a young child's eye is generally about 10 or
12 cms. away; as time goes on this distance increases and as a
rule at 40 or 50 years of age the near point has receded past the
length at which it is convenient to hold a book. Convex glasses
are then used for reading and hand-work and by this means the
divergence of the rays is reduced so that objects may be comfort-
ably held and still throw sharp images on the retina. The change
in elasticity has no effect on distant vision, the shape of the lens
in the eye at rest is the same as before.

When the gaze is directed to an object near at hand two other
changes occur, associated with the act of accommodation. The
visual axes, which during rest or distant vision are parallel, are
converged so that they meet on the object at which one looks, and
make the image of it fall on the fovea centralis of each eye. This is
done by the contraction of the internal recti, which rotate the eye-
balls inward. At the same time the ACCOMMODATION REFLEX of the
pupil occurs; the circular muscle fibres which are contained within
the free margin of the iris contract and reduce the size of the open-
ing. The iris acts as a curtain or adjustable diaphragm for the
eye, limiting the size of the pencil of rays which enter it. By this
reflex contraction of the opening in near vision all rays are shut
out except those which pass through or near the centre of the

156 EXPERIMENTAL PHYSIOLOGY.

refracting surfaces. By this means spherical aberration is cut down
in near vision, in which great accuracy is usually required and in
which the aberration would otherwise be marked, because the
incoming rays are very divergent. The image is made somewhat
less bright by this device, since there are fewer rays to contribute
to it than there would be with a wider pupil, but, the object being
near, the number of rays which the eye receives from it is in any
case large and the loss is not of much importance. All three groups
of muscles which contract in near vision, the ciliary, the circular
muscle fibres of the iris, and the internal recti, are supplied by the
oculomotor nerve. The nerve supply of the radial fibres of the
iris, the contraction of which dilates the pupil, comes from the
cervical sympathetic.*

Some indication of the nature of the change in the eye media
during accommodation may be got from the following experiments.
Experiment 60. — In a dark room arrange the lens and watch glass
so that they are centred, with the lens behind the watch glass,
which is placed with its convex side forward. Hold a candle
beside your head and a little behind your eyes and look at" the
images of the flame formed by the three refracting surfaces.
These let most of the light through, but reflect some small
portion of it. The reflected rays form the images which you
see. There are three of them, one inverted and two erect.
Experiment 61. — Purkinje Images. — Look at your partner's
eye in the way in which you looked at the lens and watch glass
in the last experiment and ask the subject to gaze past you into
the distance, -along a line half way between your eye and the
candle. As before, three images may be seen but in this case
one of the erect images is larger and dimmer than the other.
Now ask the subject to focus on your finger held in his line of
vision, at about the distance of your head away. The small,
bright, erect image and the inverted one are unchanged in
position, but the larger erect one moves nearer the first and

*A stopping-down of the pupil, similar to that which occurs for near vision, is
brought about reflexly when the eye moves from a darker to a lighter place. If
the change in the intensity of the light is great the contraction is more marked at
first, becoming gradually less for some minutes afterwards as the eye becomes
'accustomed to the light". This is known as the "LIGHT REFLEX" of the pupil.

ERRORS OF REFRACTION AND ACCOMMODATION. 157

becomes smaller than before. This is the image from the
anterior surface of the lens. It is large and dim in the un-
accommodated eye because the surface which forms it is fairly
flat. During accommodation when the surface becomes more
curved the image becomes smaller and appears to move forward
because of the movement of the reflecting surface.
Experiment 62. — Schemer's Experiment. — The course of rays
which enter the eye, from objects at distances for which the
eye is not accommodated, can be traced by blocking all but
two pencils of them and finding out which regions of the retina
are affected by the two pencils of light. In a piece of heavy
paper prick two holes less than the diameter of the pupil apart.
Stick two needles upright in a strip of wood, one 30 cms. and
one 60 cms. from the end. Hold the screen with the holes
horizontal in front of one pupil and, closing the other eye,
look along the stick and accommodate for the far pin. Note
that the image of the near pin is double. Cover one hole in
the screen and see which image disappears. Repeat, with the
eye accommodated for the near pin. From your results draw
diagrams of the course of rays entering the eye (a) from an
object at the distance for which the eye is accommodated,
(b) from one at a greater distance, (c) from one at a less. Com-
pare with the diagrams from the results of Exp. 48 and explain
the differences.

CHAPTER XIX.

EXAMINATION OF THE REFRACTION OF THE EYE AND
OF THE INTERIOR OF THE EYEBALL.

THE OPHTHALMOSCOPE.

Most of the light which enters the eye passes through the
retina and is absorbed by the black pigment layer behind it but a
small proportion of the rays are reflected from the retina itself.
The rays reflected from each point are refracted again by lens and
cornea being bent away from the perpendicular in this case and they
pass out in the same general path as was taken by the incoming rays
which illuminated the point. That is, they travel back towards the
source of light. Since the eye of the observer is not under ordinary
circumstances placed at such a source it does not receive any of this
light and therefore any pupil at which one looks appears to be black.
If however one arranges to have the observer's eye at the source of
light, or a very little distance from it, some of the rays reflected from
the observed retina can be seen.

This is the principle on which the ophthalmoscope is con-
structed. It not only gives a view of the interior of the eye but
also, as we shall see, affords a method of examining its refraction.
The instrument is a slightly concave mirror with a tiny hole in the
centre of it. One holds it in front of the eye which one wishes to
examine and a light is placed beside the eye and a little behind it
(L, Fig. 44 A.). The mirror sends rays from this into the eye.
LAA' and LBB' in Fig. 44 are two such rays. There will of
course be many others, lighting up the whole area between A' and
B' and some of them falling on either side. Each point of this
illuminated area, reflecting part of the light which it receives, acts
as a source of rays, as -is shown for the point X. The observer
looks from behind through the little hole and some of the reflected
rays (as XY and XZ) returning towards the mirror pass through
the hole and afford him a view of the interior of the subject's eye.

158

THE OPHTHALMOSCOPE.

159

The subject is asked to stare ahead of him, in order to relax his
accommodation, or, what is more effective, an application of atro-
pine is made to the eye. This paralyses the endings of the third
nerve in the iris and the ciliary body and prevents accommodation,
and reflex contraction of the pupil under the stimulus of the strong
light. The rays from the retina, arising at the principal focus if
the eye is a normal one, pass out after refraction parallel to one
another. If these are to be clearly focussed on the observer's
retina his eye must be unaccommodated. He must not look there-

FIG. 44. To illustrate the use" of the ophthalmoscope. Upper diagram shows the
course of the illuminating rays and of the outgoing rays, by the direct method. Lower
diagram shows the course of outgoing rays when the indirect method is used; the
illuminating rays have been omitted.

fore at the retina which he wishes to see but must gaze through it
into the distance. If the observed eye is shortsighted, the prin-
cipal plane, the plane from which rays would have to come to be ren-
dered parallel after passing out through lens and cornea, is some-
what in front of the plane of the retina. Light rays from the
retina, therefore, since they diverge less than would rays from
the principal plane, are too much refracted by the eye media and
converge after refraction. If they are to make a sharp focus on
the unaccommodated eye of the observer they must be rendered

160 EXPERIMENTAL PHYSIOLOGY.

parallel by a divergent lens, and the strength of the lens which
one must use for this gives a measure of how much the rays differed
in direction from the parallel paths which they should have taken.
If the eye is LONG-SIGHTED the condition is reversed. The
retina is farther forward than the principal focus of the eye, the
rays diverge too sharply and after refraction they still diverge to
some extent. The observer to see clearly must in this case use a
conyergent lens and, as before, the strength of the lens gives a meas-
ure of how different the eye is from the normal. This is known as
the DIRECT METHOD of using the ophthalmoscope. The part of the
retina which one can see at one time in this way is a small one and
for that reason, and sometimes too because it is difficult for the
observer, if he is inexperienced, to relax his accommodation com-
pletely, the INDIRECT METHOD is often used (Fig. 44, B.). The
arrangement of light and mirror is on the same plan as before, but
in this case a convex lens is held immediately in front of the subject's
eye. This focusses the rays emerging parallel from the eye and
forms an inverted real image of the bright spot of the fundus, at
the principal focal plane of the lens. It is this image at which the
observer looks.

Experiment 63. — The use of the ophthalmoscope for the examina-
tion of the human eye is not suitable as a laboratory experiment
since a beginner to have success needs both time and quiet.
Each student is expected, however, at some time during this
part of the course, to procure an instrument, of which there are
several available in the supply room, and to practise with it
until he can see for himself the interior of a normal eye. Using
both the direct and indirect methods, work at the experiment
until you can see many or all of the details of the interior surface
of the eyeball described below (p. 147). Before doing this it is
wiser to practise a little in the laboratory, using the instrument
according to both methods to examine the eye of a rabbit, the
pupil of which has been dilated with atropine.
Direct Method. — Place the light and ophthalmoscope, as
shown in Fig. 44, and look from behind the mirror through
the hole. Start with it about eighteen inches away from the
observed eye and move it slowly about until the pupil is seen
as a bright red spot. Now move the mirror gradually nearer,

THE OPHTHALMOSCOPE.

161

keeping it always in the same line, until when you are about
two or three inches away from the subject's eye the bright spot
of the retina comes clearly into view. Both observer's and sub-
ject's eyes must be unaccommodated.

Indirect Method. — The arrangement is as before except that
a converging lens is held two or three inches away from the
observed eye and steadied by resting a finger of that hand

FIG. 45. Diagrams to illustrate the use of the retinoscope. No. 1 shows the path of rays
from a normal eye, No. 2 that for rays from a normal eye in front of which is held a converging lens,
or of rays from a shortsighted eye.

against the subject's forehead. Hold the ophthalmoscope
about twenty inches away and move it about and move the
lens back and forth until the image of the bright spot is clearly
seen. The observer must accommodate his eye for a point near
at hand.

THE RETINOSCOPE. Another instrument which is used to examine the
refraction in the eye is the retinoscope, a flat mirror with a little peep-hole in the
middle, which can be made to rotate through a small angle on its handle. The
principle of its use depends on the following considerations. If a spot of light is

162 EXPERIMENTAL PHYSIOLOGY.

reflected on a wall and the mirror reflecting it is rotated upward, the movement of
the spot is in the same direction as that of the mirror. If a spot of light be
reflected into a normal eye and the mirror moved, the movement of the light on
the retina appears to the observer's eye, placed at the hole in the instrument, also
to be in the same direction. This is shown in Fig. 45, No. 1 A is the first
position of the spot of light and B the second, moved by an upward movement of
the mirror. The rays from A, when after refraction they come out of the eye,
appear to come from A' at infinity. Those arising from B seem to come from B',
also at infinite distance. The mirror was moved up and the image also moves up.
If a strong converging lens is held in front of the observed eye so that the rays
coming out are made to form inverted images, at some short distance from the
lens, of the retinal points from which they arise, and if the rays are diverging from
the images again when they reach the observer's eye, the condition is reversed.
The spot of light appears to move in a direction opposite to that of the movement
of the mirror. This is shown in Fig. 45, No. 2. The rays from A, converged after
coming out of the eye, make the image at A', those from B at B'. If the observer's
eye be farther away than A' and B', the rays from the first (lower) position of the
spot of light seem to come from the upper image, those from the second place of
the spot, which was really above the first, from an image lower than the first image.
The direction of movement is thus reversed. If the observer's eye be nearer than
the focus of the lens (that is, nearer than A' or B'), the movement appears to be
in the direction of the movement of the mirror, as it was without the lens. Sup-
pose the observer's eye is always at a distance of one metre from the subject.
Then if the lens is stronger than one dioptre (i.e., has a focal length of less than
one metre) the spot will move in the opposite direction. If the lens is weaker than
this there will be no reversal, and the spot will move in the same direction as the
mirror. Now, if the subject's eye is not normal but long-sighted, the rays when
they come out diverge from each other. To bring these to a focus in front of his
eye, still at one metre distance, the observer will have to use a stronger lens than
before and the more long-sighted the eye is, the stronger the lens will have to be.
Rays coming from a short-sighted eye are already converging when they come out.
If the defect is small, a converging lens will still be needed to make the movement
of the spot appear reversed to an observer at one metre, though not so strong a one
as is needed for the normal eye. If the defect is great, the focus of the rays
without any lens at all may lie at some point between observer and subject, and
in this case the spot of light moves in the opposite direction to the mirror from
the first. The condition will be thus like that already described for Fig. 45,
No. 2, only that no lens need be imagined. A divergent lens must be used to
cause it to go to the similar direction and, as before, the strength of the lens which
must be used gives a measure of the abnormality. An examination of this kind
can be done in any plane of the eye and the method can therefore be used to
measure astigmatism. In this case the strength of lens necessary to give reversal
when one plane of movement is being tested is different from that for another
plane.

CHAPTER XX.

THE RETINA.

The view of the interior of the eyeball which one gets with an
ophthalmoscope reveals a whitish lining, the retina, over thejsurface
of which are fine branches of arteries which supply it with blood.
There are two spots on it which differ in their appearance from the
rest of the surface. One of these, situated directly opposite the
centre of the pupil, is the MACULA LUTEA. This is slightly raised,
yellowish in colour, and has a tiny depression in its centre, the
FOVEA CENTRALIS, the part of the retina capable of the most accurate
vision. The blood vessels skirt around the yellow spot and do not
pass across it. Towards the nasal side of this and some little
distance from it there is a shining white spot, the OPTIC DISC, which
marks the passage through the retina of the fibres of the optic
nerve.

In order that the student may have the structure of the retina fresh in his
mind as a basis for his experiments, he is reminded of the details of its anatomy
and histology which follow.

The fibres of the optic nerve enter the eyeball from outside arranged in a
cylindrical fashion, enclosing in the middle an artery, branches of which we have
seen ramifying over the surface of the retina. When they have pierced the outer
layers of the retina, the nerve fibres of the cylinder divide and spread out over its
whole surface, thus forming the innermost retinal layer of all. The whole retina
extends forward in the eyeball only as far as the beginning of the ciliary processes.
From here on it is represented simply by the layer of black pigment cells which
clothes the posterior surface of the ciliary body. The cells which are sensitive to
light waves, the fundamental part of the whole organ of vision, are contained in one
of the layers of the retina. The retina itself is to be regarded not simply as com-
parable to a group of sensory nerve endings, connected each by a single fibre
directly to the central nervous system, but as a part of the brain itself, from an
outgrowth of which it is formed during the development of the foetus. Accord-
ingly it contains, as well as the light-sensitive cells themselves, two layers of relay
cells through which the impulses arising in the sensitive elements must pass on
their way to the brain (Fig. 46). The elements which respond to light waves, the
rods and cones, are placed farthest away from all the incoming light. They
are the free ends of the outer cell layer, sticking out towards the choroid coat from
which they are separated only by a layer of black pigment cells. The nuclei

163

164 EXPERIMENTAL PHYSIOLOGY.

corresponding to them are nearer the centre of the eyeball and each cell sends a
connecting fibre inward into the outer nuclear layer of the retina, where it is con-
nected with the dendrites of one of the first relay cells, the bipolar cells. The rods
have slightly thickened inner, and straight slender outer, limbs. They are con-
nected with their nuclei by fine thread-like processes. The cones, which are
shorter than the rods, are also divided into inner and outer limbs. Their outer
limbs are shorter and sharper than those of the rods; the inner ones are much
thicker and connected each by a substantial process with its nucleus, which lies
nearer to it than do those of the rods. The dendrites of the bipolar cells, which are
the next units on the path of the impulse, are connected by short thick processes
with their cell bodies, the nuclei of which make up the inner nuclear layer.
The axones of these cells are short. They end in the inner molecular layer,
where they establish connection with the dendrites of the second relay cells, the
giant cells. It is the axones of the giant cells which, passing over the surface,
make the inner layer of the retina and join together at the optic disc to go out as
the optic nerve. In all parts of the retina except the fovea centralis the bipolar
cells connect each with several fibres from rod-cells or ccne-cells, and are in turn

FIG. 46. Diagram of the structures in the retina.

several of them connected with one giant cell, so that the number of nerve elements
represented in the optic nerve is a good deal less than the number of cells in the
retina, and one fibre in the nerve may carry impulses which arise from any one or
from all of a number of rods or of cones. The histology of the fovea is different
in this as in some other respects from that which we have described. It consists
only of the light-sensitive cells and the fibres which connect them with the first
relay cells. These, together with the giant cells to which they lead, are swept
aside from the fovea itself and, piled up around its margin, cause the slight
thickening of the retina which surrounds it. The sensitive cells in the fovea
are all cones, no rods are to be found, and they are each of them connected through
a different bipolar cell to a single giant cell and so to a separate fibre of the optic
nerve. The arrangement affords some histological basis for the acuteness of
vision which we have in the fovea, since each cone has a separate path to the brain.
It is found that two points in the external world may be distinguished as separate,
provided that the angle by which they are separated at the eye is 5 minutes or

THE RETINA. 165

more, and provided that they are looked at directly, that is, that their image
falls on the fovea. Construction by the schematic eye shows that an angle
of this size represents an image of about .0035 mm. on the retina, approximately
the distance from the centre of one cone to that of another. If the points are
looked at, not directly, but with 'the tail of one's eye', they must be much more
widely separated than this to appear distince from each other; the distance
necessary increases, broadly speaking, the farther they are removed towards the
periphery of the visual field.

That the cells which are most deeply placed of all should be
those which respond to the vibrations of the light waves would at
first sight appear improbable. There is abundant evidence, how-
ever, that this is the case, as the two following experiments show.
Purkinje Figures. — The retinal blood vessels, being situated
between the source of light and the receptive surface cast shadows
on the retina. These are very small and by habit are disregarded.
If, however, they can be made to fall on some part of the retina
which ordinarily does not receive them they may be seen.
Experiment 64. — Make the subject turn one eye inward and look
towards a dark wall. With a lens concentrate a good light upon
the exposed sclerotic, focussing so as to make a small, strongly
illuminated area. Now give the lens a circular or gently rocking
motion. The field appears to the subject to be reddish yellow,
with dark branching figures on it. These are the shadows of
blood vessels. In the direct line of vision a small spot free from
shadows may be seen, the macula lutea or yellow spot.
The path of the light rays through sclerotic and choroid and
the vitreous humour is shown in the diagram (Fig. 47). When
the source of light is moved there is a movement of the shadow of
the blood vessel on the retina, which is projected outward by the
subject along the line which connects the point stimulated with the
nodal point, and which is interpreted as a movement of a shadow in
the field of vision at the distance for which the eye is accommodated.
A is the first position of the light, A' the place on the retina where
it throws the shadow, A" the place on the screen at which the eye
appears to see the shadow. When the light is moved to B the
shadow moves to B' and its apparent place in the field to B".
n.p. is the nodal point of the eye, b.v. the position of the blood
vessel. Now triangles A"B" n.p. and n.p.A'B' are similar and we
know the lengths A"B", n.p.A" and n.p.A'. We can therefore find

166

EXPERIMENTAL PHYSIOLOGY.

A'B', the distance the shadow actually moved on the retina. But
triangles A'B' b.v. and bv.AB are also similar. Here we know
A'B', A.B. and (approximately) bvA., so that we can find bv.A',
the distance between light-sensitive layer and the blood vessel.
This is found to be about 0.2-0.3 mm., which is sufficiently like the
distance actually existing between the retinal blood vessels and the
layer of rods and cones to support the conclusion that these are
the elements sensitive to light.

FIG. 47. To illustrate the course of the rays w hich cast the shadows in Purkinje's
figures, and the direction along which these shadows are projected outward into the
visual field.

Blind Spot. — It may easily be shown that, although we are
unconscious of it, there is an area in the field of vision of each eye
which is blank; objects placed there are not seen. Construction
with the schematic eye shows that light rays from this area fall on
the optic disc. Here, as we have seen, there are no rods or cones,
the layers which contain them being interrupted to allow of the
passage through it of the fibres of the optic nerve.

THE RETINA. 167

Experiment 65. — Take a thin strip of paper and blacken about
1 mm. of its end. Hold a large sheet of paper with a dot in
the centre vertically in front of you. Keeping your head quite
still, close one eye and look fixedly at the dot with the other.
Move the blackened end of the paper strip slowly over the field
of vision and mark the points when it seems to disappear.
From these points map out the area of the blind spot in the
visual field. Using the schematic eye, show where it must be
in the retina.

The Response of the Retina to Light.— In addition to the
information from subjective experiments such as those which we
are about to discuss, which of course involve activity of the brain
as well as the eye, some interesting knowledge as to the nature of
the retina's reaction to light stimuli has been got from direct
experiments on the eye itself. The technique which shows the
movement of the cones, the negative variation in the retina, and
the formation and destruction of the visual purple, is unfortunately
too difficult for class experiments. The student must turn to his
text-books for information as to the nature and significance of this
work.

DARK ADAPTATION. — It is familiar that eyes at night or in a dark room can see
lights so faint that they would not be made out at all by day. This is in part due
to an actual lowering of the threshold of the light-sensitive elements, and one may
satisfy oneself by simple experiments that the main part at least of this change
in threshold occurs, not in the line of direct vision, but in the peripheral field.
The observations are entirely subjective ones and the student must make them
for himself under suitable conditions of lighting. If one looks at the sky at twi-
light searching for the first star it may appear on one side or another of the direct
line of vision, only to disappear when looked at directly. Small pieces of white
paper on a black background in a darkened room are visible as long as they are
not in the direct line of vision, but disappear, each as its image is made to fall on
the fovea. Indeed, workers in this field have not been able to find that any
lowering of the threshold at all occurs in the fovea during adaptation to the dark
and this makes it probable that the change in the threshold of the peripheral
field is a change in the rods and is not shared by the cones. The adaptation
which the rods undergo is thought to be associated with the presence in them
of visual purple, the coloured substance which is formed in them in the dark.
One of the chief reasons for this is the striking resemblance between the sen-
sitiveness of the pigment on one hand, and of the dark-adapted eye on the
other, to waves of different lengths. When several different colours are seen
under similar lighting conditions it is only a matter of judgment which of them is

168 EXPERIMENTAL PHYSIOLOGY.

the brightest, but most normal individuals if shown a spectrum in a brightly
lighted room, pick out the yellow as the brightest part of it, brighter than either
red, orange, green, blue or violet. If however all light is shut out of the room
except that coming through the prism which makes the spectrum, and if the eyes
are allowed to become adapted to the dark, the place of greatest brightness in the
spectrum appears to shift from yellow to green. This colour, it is to be noted, is
also the most effective in bleaching the visual purple. The spectrum under these
conditions also appears shortened at the red end, that is to say the long waves are
ineffective for the eyes in this state, as they are also found to be on the purple
itself.

Some further evidence which bears on the same question is got from obser-
vations on the ACHROMATIC (or colourless) THRESHOLD. If, after the eyes have
become adapted to a dark room, light is gradually introduced, all objects in the
room of any colour except red look colourless when first they become perceptible,
as if they were made up of varying shades of white and grey, and it is only on
farther illumination that they appear in their own tints. The interval during
which they seem uncoloured is the achromatic interval. Red objects have no
such interval; they appear at once in their own colour, but they require a stronger
light to make them visible than is needed to make the other objects show as
colourless grey ones. The whole phenomenon can be seen out of doors at dawn,
or, in the reverse order, at dusk. At the first daylight everything comes into
view as if in a print, all black, white or grey, and it is only as the light increases
that the leaves change from grey to green, blues, yellows and violets appear, and
red flowers turn from black objects to brightly coloured ones. These results are
interpreted as follows : in the light the cones have a threshold lower than that of the
rods and, since they are capable of distinguishing colours as well as merely bright-
ness, we see all light in which there is more of some wave lengths than of others as
coloured light, no matter how faint it may be. In the dark adapted eye the
threshold of the cones is not changed, but that of the rods is lowered, probably
because of the presence in them of the visual purple, so that they become more
sensitive than the cones. The rods are not organs of colour vision, however,
when stimulated they give rise only to sensations of brightness. Therefore faint
lights of wave lengths other than red are first seen by the rods of the dark-adapted
eye as colourless rays and only affect the cones and appear coloured when they
have become a good deal stronger. Red lights do not affect the rods at all, and
their effect does not appear until they are strong enough to stimulate the cones.

There is an abnormal type of vision known as TOTAL COLOUR BLINDNESS,
which shows a good many points or resemblance to the vision of a dark-adapted
eye. People who have this defect see, as far as one can determine, all lights as
colourless; rays of different wave-lengths differ to them only in brightness and
any two colours can be made to look exactly alike to the colour blind by varying
the brightness of one or the other. Two colours which match in their judgment
look equally bright also to the normal eye in its condition of dark adaptation, but
are different not only in colour but in brightness under ordinary conditions of
lighting. Like the dark adapted eye the colour-blind is more sensitive to the

THE RETINA. 169

shorter wave lengths; a green which appears to the colour-blind eye to match a
certain yellow seems to the normal in daylight to be not only of a different colour
from the yellow chosen, but also less bright than it. The sight of people who have
this defect is easily fatigued in bright light and is much less acute than the normal.
In the dark it far surpasses the normal in accuracy. Many colour-blind people
have a blind spot in the fovea, so that they see nothing in the direct line of vision
but only in the peripheral field. It is thought that the condition is one in which
the rods mainly function. The parallel between these eyes and the normal dark-
adapted eyes is not complete. The vision of the colour-blind is much the
more acute and is often enough so to enable the subjects to read, which cannot
be done with normal dark-adapted eyes.

CHAPTER XXI.

COLOUR VISION.

It is a familiar fact that when ordinary white light is passed
through a prism the waves of which it is composed are refracted
to a greater or less extent according to their length. As a result
the single beam which was made up of waves of all lengths is spread
out into a band of light, one end of which is formed by the shortest
of all the visible rays, the part immediately next to it by those a
little longer, and so on throughout the whole band until the other
end is reached, which contains the longest waves that the eye can
perceive as light. The physiological effect of this re-arrangement
of the waves is to make the light, which when mixed was uncoloured
or white, appear to be made up of a whole series of colours, known
as the spectrum. The colours are arranged in a definite order
according to the length of the waves of each part of the band of
light. The shortest of all look violet, those next them blue, the
waves next these again green, then yellow, then orange, and then
red, first scarlet and then, at the end of the band, crimson. Each
colour shades inperceptibly into the one next it, so that as well as
those which we have mentioned we can find in the spectrum
numbers of others, such as greenish-blue, violet-blue, greenish-
yellow, and so on.*

There are two other ways in which lights may differ from one
another. One is in BRIGHTNESS. Of two rays of the same colour
if one is more intense than the other it looks brighter.
Experiment 66. — Take two sheets of paper of the same colour.

Place one near an electric light of low, and the other near a light of

high, candle-power. The second looks brighter than the first.

*Each of the spectral colours may also be produced by the use of pigments.
The colours of these depend on the fact that, from the waves of different lengths
in a beam of white light falling on them, they absorb all except those which
belong to a very limited area of the spectrum. Blue paint, for instance, reflects
only the blue waves and a few of the green and absorbs all the others. Yellow
glass absorbs most of the wave lengths falling on it except the yellow ones, and
light which has passed through it consequently has that colour.

170

COLOUR VISION. 171

The other way in which two samples of the same colour may
differ is in the proportion of white light which they contain. A
colour which contains no white light at all is, physically speaking,
completely SATURATED or full; the more white one mixes with it
the less saturated, or the paler, it becomes.

Experiment 67. — Take two sheets of yellow tissue paper and put
one over a piece of heavy paper of the same shade and lay
the other over a sheet of white paper. They both have the same
colour but the first is fuller or more saturated than the second,
.paler one.

The evidence which we have examined so far would lead one

to think that the sensation of a given colour was entirely dependent

on the action on the retina of waves of a certain length. It would

seem that the sensation of yellow, for instance, was simply the

physiological result of the action of waves from that particular part

of the spectrum. The student's attention however is directed in

this regard to the results of the two following experiments.

Experiment 68.— The Effect of Stimulating the Retina with

Two Different Colours at the Same Time. — From the paper

discs provided choose an orange and a green, colours

situated immediately on either side of yellow in the spectrum,

the waves of one being longer and those of the other shorter

than the yellow waves. Arrange them in the rotator so that

about half of each shows and spin them rapidly until they seem

to fuse.* The colour which together they produce is a yellow

*The ideal way to mix colours on the retina is to cause the rays from those
parts of the spectrum which contain the desired colours to converge again, so that
they actually fall on the retina at the same time. A simpler means is that adopted
here, in stimulating the retina with alternate flashes of the colours following each
other in very rapid succession. Advantage is taken of the fact that the reaction
of the retina does not cease immediately the stimulus is withdrawn but lasts for a
fraction of a second after it. The result cannot be got by mixing together pig-
ments of the colours which it is desired to combine. For instance, although blue
and yellow light thrown together on the retina give white or grey, blue and yellow
pigments mixed give a green colour. The reason for this is that both blue pigment
and yellow pigment reflect, along with their particular colours, some green rays.
When they are mixed the blue absorbs yellow light which the yellow pigment
would have reflected, the yellow pigment does as much for the blue, and the only
colour left over is the green, which was completely absorbed by neither.

172 EXPERIMENTAL PHYSIOLOGY.

much like the pure spectral colour, only a little less saturated.
Change the disks, taking this time reddish-orange and yellowish-
green. These colours are also one of longer and one of shorter
wave length than yellow but they are more widely separated
from it in the spectrum than the first two were. When you spin
them you again get a yellow but this time there is a much
greater proportion of white in it, it is less saturated than before.
Replace these colours with two others still further removed
from yellow, although still lying one on either side of it; use
scarlet and blue-green. When spun together this combination
looks whitish or pure grey, the yellow is no longer produced.
Two colours such as these, which give a sensation of colourless
or white light when combined on the retina, are COMPLEMEN-
TARY COLOURS. There are innumerable such pairs, a few of
them being the following; orange with blue, bright yellow with
a more violet blue, green-yellow with violet. Try these for
yourself. It may be necessary to show in each case a little
more of one of the pair than of the other, depending on whether
or not they are equally bright. In experimenting on these
complementary colours notice that when they are arranged in
any proportion other than that which gives white, they produce
a sensation of some colour which lies between them in the
spectrum, just as those pairs did which were less widely sepa-
rated.

Now take pairs of colours too widely separated to be com-
plementary, for instance red and blue, or orange and violet,
and see that combining them gives a sensation of purple, a
colour which does not exist in the spectrum at all. It is the
complementary colour to green.

We have seen that a colour sensation, yellow for instance, can be
produced (a) by waves, acting alone, of the length of those which
make up the yellow part of the spectrum, or (b) by a great many
combinations in pairs of waves of other lengths, waves which
falling alone on the retina give colours which have no yellow in
them at all. We have also seen that two beams of different wave
lengths, as well as producing each a definite colour sensation when
it acts alone, can if combined in varying proportions give rise to
quite a large number of the other colours. ALL the colours, how-

COLOUR VISION.

173

ever, can not be got by using any one pair; to get the whole range
of possible colour sensations you must take at least three spectral
colours and vary these. There are many combinations of three
which can be taken, we have chosen those most generally used.
Experiment 69, — Effect of Simultaneous Action on the
Retina of Three Primary Colours. — Put into the rotator a
red, a green, and a violet disk. Keep on altering the propor-
tions of each until you have got (a) a colourless light, (b) each
of the colours of the spectrum and (c) a purple.
As you have seen, all the various colour sensations which we
are capable of receiving can be produced by combined action on
the retina of three simple stimuli. To put the matter differently,
the whole range of the response of the retina to light can be ex-
pressed as a function of three independent variables.

R.

ft O C^ -B V-

FIG. 48. To illustrate the Young theory of colour vision.

y

YOUNG-HELMHOLTZ THEORY OF COLOUR VISION. — 'Any theory which is to be
advanced in explanation of colour vision must be based on the fundamental facts
which we have just described. The earliest of the modern theories, and one
which has given rise to a great deal of experimental work, was first suggested by
Young and later elaborated by Helmholtz. According to this there are three
different components in the retina, whether anatomical units, or chemical sub-
stances, is not stated. Each of these is supposed to be most effectively acted
upon by light waves of a quite limited part of the spectrum, and those waves
which influence one of the substances most have very little effect on either of the
other two. For the sake of illustration, red, green and violet, were selected
as the 'primary colours.' One component is supposed to be sensitive to red
waves, less so to orange and responsive in a gradually decreasing degree to the rest
of the spectrum. The green component is affected most by green, to a less extent
by yellow or blue, and little by the colours at either end. The third component,

174 EXPERIMENTAL PHYSIOLOGY.

as well as responding to violet, is sensitive to blue, but little to the other wave
lengths (Fig. 48). If any one of these components is stimulated much more
than the others the result is a sensation of the primary colour corresponding to it.
Sensations of colours which lie intermediate to the primary colours in the spectrum
are simply the result of less unequal stimulation of two of the primary substances;
that explains why you can get such a sensation either by using a wave length (such
as blue) which affects two components (violet and green), one more than the
other, or by stimulating the retina with waves of two other colours (violet and
green) but unequal intensities, one of which stimulates mainly one, and one
mainly the other, of the same components. If any two components are stimulated
equally a sensation of white light results; complementary colours are therefore
merely those which have an equal effect on two of the components. Whether
or not a colour is more or less saturated depends on whether the components other
than the one mainly involved are responding much to the light or not.

Experiment 70. — After Images. — Look fixedly for a few seconds
at a bright object (the filament of an electric light will do) and
then transfer your gaze to a black background. The bright
image does not disappear at once but only fades gradually
away. This is a POSITIVE -AFTER-IMAGE. Repeat, after the
image has quite faded, but this time move your eyes to a brilli-
antly lighted white background. There is a persistent image
in this case also, of the same shape as the object, but whereas
the object was a bright one the image looks dark. This is a

NEGATIVE AFTER-IMAGE.

Look steadily at a tiny spot in the centre of a square of
bright red paper, well lighted. After a few minutes shift the gaze
to a white background. You see the square still, but tinged
with greenish-blue, the colour complementary to red. Repeat,
after waiting until the after-image has faded, with a yellow
square, then try with a green one. The negative after-images
are coloured with the shades complementary to those of the
objects.

The explanation of the positive after-image of a white object v
probably lies in the fact, which we have already noticed in another
connection, that the reaction of the retina does not cease imme-
diately the stimulus is removed. The positive image is merely the
after-discharge of the light-sensitive cells in the retina. The pheno-
menon of the negative after-image is comparable to the "rebound"
which occurs after a reflex, as Sherrington showed. When a reflex
arc has been for some time under the influence of one reflex, its

COLOUR VISION. 175

responsiveness to a stimulus which tends to bring about the oppo-
site reflex becomes increased. Fatigue following a reaction of one
kind makes the way easier for the opposing reaction.

In more detailed form, according to the Young theory, the reason why the
after-image of a coloured object is tinged with the complementary colour is as
follows. During the time that the gaze was fixed on the object all three com-
ponents in the cells of the retina on which the image fell were being stimulated but
the component or components which are most sensitive to the colour of the object
were being stimulated most. When the object is withdrawn and the place on
which the image falls is uniformly stimulated by white light all three com-
ponents respond. The one which was mainly affected by the former light is
fatigued however and responds less than the others. This enables the colour
complementary to it to come into prominence.

PARTIAL COLOUR BLINDNESS. — This abnormal condition of vision is inter-
esting in the discussion of the theories of colour vision as it affords a comparison
with the normal. The defect, fairly common among men, less so in women,
is physiologically quite different from total colour blindness, with which it must
not be confused. People who are partially colour blind distinguish differences
in light waves of different lengths, no matter whether their intensities are equal
or not, but they see fewer differences than normal people. If they are shown
light of different colours (the best test is light coming from lanterns with windows
of coloured glass) they select as being of similar shades the yellows and the blues
and the violets which look much alike to the normal eye, but in choosing shades
similar to a green they also take, as well as other green lights, different shades of
red, and in matching a red standard they pick out as having a colour similar to it
not only other reds but also green as well. The condition of their light- receptive
mechanism seems to be not so much different from, as simpler than, the normal;
roughly speaking, partially colour-blind vision is a reduction form of ordinary
sight. Whereas all the various shades which one sees with normal colour vision
cannot be produced with combinations of less than three given colours, two only,
if mixed in different proportions, will match all the colours seen by a subject who
is partially colour-blind.

There are two types of the defect. In the first, most common, form the eye
is relatively insensitive to red light. To match an olive green, people with this
type of the defect choose a scarlet which to the ordinary eye appears much brighter
and the visible spectrum is shorter for them at the red end than it is for other
people. Subjects whose vision belongs to the second type match a green with a
red which is about equally bright and the red end of the spectrum appears to have
the usual length.

The explanation of these facts, according to the theory of the three visual
components, is that in eyes of the first type the red component is left out and
that all wave lengths have only the other two components to act upon. In the
second type it is the green which is missing. This would express the actual con-
ditions well enough if only these abnormal types were accurately, and not merely
approximately, reduction type of the normal. More recent work however has

176 EXPERIMENTAL PHYSIOLOGY.

emphasized the fact that this is not the case and so far it has not been possible
to cover adequately the conditions of the defect with this, or indeed with any
other, theory.

HERING'S THEORY OF COLOUR VISION. — The only other theory which has
rivalled that of Young as a starting point for experimental work is the one brought
forward by Hering. This is also based on the laws of colour mixing which we have
already outlined, but it affords as well some physiological explanation for the
important fact that in our sensations light may not only be divided into different
colours, but these again appear to fall into two great classes, the 'warm' or
'bright', cheerful colours and the 'cold' or 'dull' ones. To the 'warm' class
belong reds, oranges, and yellows; they are bright of their own nature as it were,
quite apart from the brilliancy with which they are lighted. The dull ones are
green, blue and violet. There are many examples which show that we recognize
this inherent brightness or dullness of colours in everyday life. We "see things
through rosy spectacles" or we "feel blue". Decorators use yellow in north
rooms with no sunlight, but blue or green are said to be too 'cold'; children are
dressed in red in winter because it is "such a warm colour". The suggestion
which Hering made was this: there are three substances in the retina each of
which may be changed by light stimuli in two ways, being either built up or
broken down. One of them is the white-black substance; light waves of more
than a certain intensity break this down, no matter what their length, and give
as a result the sensation of white light. Darkness causes it to be built up and a
sensation of blackness arises from this. Of the other two components, one is
acted on only by red light, which breaks it down or katabolises it, or by green, the
complementary to red, which builds it up. Katabolism of the third substance
is caused by yellow light, anabolism by blue, its complementary. All other colour
sensations are mixtures of the sensations arising from the anabolism or katabolism
of the three substances. All colours which break down the components are
similar to white, which also does this; that is why they are specifically bright.
Colours which build up are like darkness and hence in themselves are dull. Com-
plementary colours give white light because, since they act equally on a single
substance, the katabolic effect of one cancels the anabolic effect of the other and
the only reaction left over is the katabolism which they both bring about in the
white-black substance.

According to this theory the reason why the after-image of, for instance, a
green object is coloured red is that the action of the green light having built up a
great deal of the red-green substance, the red waves contained in the white light
from the bright field which follows have more substance to act on than the other
waves. The after-image of a yellow object is blue because the yellow waves have
broken down the yellow-blue substance to a great extent and the anabolic reaction
to the blue waves in the succeeding light is more intense, tending to restore the
equilibrium, than it would be if the equilibrium had been undisturbed.

This theory gains some support from other observations on vision. Blackness
seems to us to be, not a negative, but a positive thing. In the ordinary resting
state of the retina we do not see blackness; our visual field is covered with vague

COLOUR VISION. 177

light-waves and is roughly speaking grey, certainly not pure black, as it ought to
be as far as the Young theory goes. According to the Hering theory the equili-
brium of the white-black substance is the resting-state, associated with grey sen-
sation, and the black sensation belongs to active katabolism.

When it comes to an explanation of partial colour-blindness this theory breaks
down. Absence of the red-green component would be the most obvious explana-
tion to advance but this does not hold in view of the two different types of the
defect. There are also difficulties arising from more detailed experiments on
after-images which stand in the way of accepting this theory as a complete
expression of the facts of colour vision. Indeed no theory has as yet provided
that, a fact not to be wondered at in view of the extreme complexity of the reac-
tions involved

Experiment 71. — Map out the extent of the colour fields on the
retina by using the perimeter. This consists of a strip of metal
bent to form an arc which can be rotated in various meridians.
The arc is graduated from the axis and it carries a small rider
to which a piece of white or tinted paper is attached. The
observed person rests his chin on the wooden pillar placed in
front of the arc, and with one eye closed he looks steadily at
a small white spot marked on the axis of the instrument. The
observer, moves the carrier from the free end to the axis
of the arc slowly, and notes the exact graduation at which the
subject first perceives it. This distance he then marks on a
chart which consists of a circle with various diameters repre-
senting the meridians each graduated to correspond to the arc,
from the centre outwards.

In transferring the readings to the chart, allowance must be
made for the crossing at the nodal point of the eye. The obser-
vation is repeated for at least twelve different meridians. Having
mapped out the visual field for white (by joining the marks on
the chart) the observation is repeated using red, yellow and
blue papers on the carrier.

CHAPTER XXII.

SIMULTANEOUS CONTRAST. VISUAL JUDGMENTS.

In the foregoing discussion it has been tacitly assumed that the
only light which has anything to do with the visual sensations
which arise from stimulation of any particular part of the retina is
the light which actually falls on that part. As a matter of fact this
is not so. Strong stimulation of one area, as well as causing the
appropriate sensation there, also tends to produce sensations of the
opposite nature in the parts nearby.
Experiment 72. — Take two pieces of the same paper, neutral grey

in colour, and lay one on a white, and one on a black back-

FlG. 49. To show simultaneous contrast.

ground. The first appears darker and the second lighter than
before because of the contrast with the surrounding field. The
effect may be increased by veiling the whole with a white tissue
paper.

Experiment 73. — Arrange a white background with a horizontal
row of candles in front of it and thrust a screen in from one
side, part way between the two (Fig. 49). One large area
of the background (GH) receives none of the candlelight and one
(AB) receives light from all the candles. Between these there
are a number of vertical strips, on the first of "which (FG) the
light from only one candle falls, on the second the light from
two (EF), and so on. One would expect each of these strips to

178

VISUAL JUDGMENTS. 179

seem of even brightness throughout; actually however, as the
figure shows, each one appears lighter at the margin which is
next the darker area and darker at the other edge, which
borders on the more brightly lighted strip.

Experiment 74. — Lay the piece of grey paper on a background of
bright red. Cover them both with white tissue paper. The
grey area looks tinged with green-blue, the colour complemen-
tary to that of the background. Change the background
several times, using various brilliant colours, and note what
influence each has on the apparent colour of the neutral grey.
The phenomenon of simultaneous contrast is seen in countless
ways in ordinary life. Shadows on sand in the sunlight look bright
blue because of their yellow back-ground. So, too, places in a
brightly lighted room which are shaded from the artificial light
but faintly illumined by daylight appear to be blue, though as a
matter of fact the light falling on them is colourless, only because
of the yellow in the light on the surrounding areas of the field.
The principal, consciously or unconsciously, is used in art, in
posters and on the stage, to make the effect of vivid colour more
vivid or to modify the shade of fainter ones. It is not clear how
the phenomenon is brought about. It does not happen in the retina,
for simultaneous contrast has been shown for the blind spot,
where there is no retina at all. The modification must take place
somewhere on that part of the brain which is concerned with sight.
In this connection it is interesting to remember that there are
other instances of the activity of one sensory area influencing the
sensitivity of another. Sherrington showed the stimulation of
one part of a receptive field lowered the threshold of the response
to a succeeding stimulation of another part of the field, for the same
or for an allied reflex (Immediate Induction).

Judgment of Space and Depth. — External objects, no matter
what their shape, can only form flat images, images of two dimen-
sions, on the retina. In spite of this we see the objects in three
dimensions, as having depth as well as width and height, and it is
of interest to analyse some of the factors which contribute to this
judgment as well as those which have to do with another closely
allied to it, the judgment of distance. It is not practicable to do
in the class room many of the experiments on the factors concerned

180 EXPERIMENTAL PHYSIOLOGY.

in visual judgments because they are so largely subjective in

nature. The student is expected to make observations for himself

along the lines indicated below. He should also use the various

charts which have been prepared to assist in this part of the work.

Experiment 75. — Some of our accuracy depends on the use of

both eyes together. Try to touch with the tip of your pencil a

black dot in the middle of a sheet of white paper which you

hold obliquely in front of you. Now close one eye and see

whether or not you can perform the movement as well as before.

MONOCULAR ELEMENTS IN JUDGMENT OF DISTANCE. — One of

these is the HAZINESS or clearness of the air between the object

and the observer. Objects at a distance are hazier, all other things

being equal, than those near by, so that we usually interpret

hazy objects as being distant. That is why objects "loom large"

in a fog. Since the things look hazy we conclude that they are

farther away than they really are. The size of the images which

they cast on the retina is not changed, however, and this together

with our idea of their distance is the only clue which we have as

to the size of the objects. If they were really as far away as we

imagine them to be, to throw images of this size on the retina they

would have to be larger than they really are, and we form our

judgments accordingly.

A second and more important factor in monocular judgment is
that known as MATHEMATICAL PERSPECTIVE. It is familiar that
lines which are really parallel to one another appear as they recede
into the distance to come closer and closer together. The rails of
a level stretch of track, for instance, appear to one standing be-
tween them to form a V, with its point beyond the horizon. We
are so familiar with this effect of distance that when we see lines
which we believe to be parallel, or nearly so, appearing much
closer together at the farther than at the nearer end of an object,
we judge the object to have a good deal of depth.

THE MUSCLE SENSE of the ciliary muscles gives important
information as to the depth of objects which are close at hand,
because of the difference in the extent of accommodation necessary
to bring into focus first their nearest and then their farthest point.
The greater depth they have, the more difference there is in the
effort of accommodation in the two cases.

VISUAL JUDGMENTS.

181

THE SIZE OF FAMILIAR OBJECTS is used to judge distances. If
the images on the retina of people or animals are small we assume
that they must be far away and other things which are evidently
near them, of which we do not so accurately know the size, are
judged also to be distant.

BINOCULAR OR STEREOSCOPIC VISION. — When we look with both
eyes at a flat object the image which is thrown on one retina is of
exactly the same shape as that made on the other. If the object
has depth, however, the two images are not exactly alike. The
right eye sees a little more of the right side and a little less of the
left than the left eye does, and vice versa. For instance, if one
looks from above at a pyramid with its top cut off, the right eye
receives an image of it of the shape of diagram A in Fig. 50,.
the left, one like B. Each of these diagrams alone looks perfectly

FIG. 50. To illustrate binocular vision.

flat. But hold them in front of your eyes about six inches away
and stare straight through them, relaxing your accommodation;
the two appear to fuse and the picture that results is one of a pyra-
mid which has depth as well as breadth and height. This is in all
probability the main factor in the better judgment of depth which
one gets from using both eyes. The principle is used in the stereo-
scope. Two photographs are taken of the same view, but the first
from a position a little to the right of that from which the second
is taken, the distance between the two positions being greater than
the distance between the eyes. These pictures are mounted on a
frame and, by an arrangement of prisms, the image of the one which
was taken from the right side is thrown on the right eye, and the
image of the left picture on the left. The result is a view in which
the relief or depth appears even greater than it really is.

CHAPTER XXIII.
HEARING.

Almost all that is known about the physiology of hearing has been got by
interpreting the structure of the ear. The subject lends itself hardly at all to
experimental investigation and on this account no attempt has been made to have
this chapter similar in treatment to the others. It is inserted merely in order to
round out for the student the outline of the physiology of the special senses.

THE PHYSICAL BASIS OF SOUND.

Preliminary to the consideration of the ear itself the student is reminded of
the following facts in the physics of sound. Sound, like light, is carried by waves,
but while light waves are strains in an intangible ether and have nothing to do
with the molecules of the medium in which they travel, sound waves are vibra-
tions of the medium which carries them and they do not pass across a vacuum.
The vibrations of the air molecules by means of which a sound wave is carried are
in the direction in which the wave itself is travelling, each particle swinging to and
fro alternately in front of and behind its mean position. The movement of the
particles close to the source of the sound is started by the vibrations of the source
itself. Those molecules in turn set those swinging which are a little farther away,
these communicate the movement to the next, and so the wave is carried. It
follows that in air in which a sound wave is travelling there must be some place
where the number of air molecules is greater than the average and somewhere
where it is less, alternate places of compression and of rarefaction (Fig. 51. No. 1).
The condition in the column of air at any one time may be shown diagrammatically
by a curve such as in Fig. 51, No. 2. Here, lengths along the abscissa represent
distances from the source along the line of the wave; lengths along the ordinate
show the number of molecules per unit area. Zero represents the average num-
ber, lengths above, numbers per unit area greater than the average (compression),
and those below, numbers less than the average (rarefaction). If the sound
is musical the wave will be a regular one and the distance from one crest to
another, or from one trough to another, always the same. Unmusical sounds, or
noises, are made by waves which have no regular shape.

Between musical sounds, which will chiefly occupy us, it will be seen that
there are several possible differences. (1) They may differ in rate, that is, in their
distance from one trough of the wave to the next, or from one crest to the succeed-
ing one. The rate of the wave determines the pitch of the note; the faster the
wave the higher the note. The note of the second wave in Figure 52, for instance,
whose period is half that of the first, is an octave higher than the first one. (2)

182

HEARING.

183

Waves may differ in intensity, in the distance from zero line to crest or trough;
the greater the intensity, the louder the note. (3) The waves may be musical,
that is, each single wave may be a repetition of the last, and still not have the
simple form shown for 1 and 2 in Fig. 52. These curves are such as would be
made by a swinging pendulum, writing on an evenly moving surface and they are
known as pendular or sine curves. The sound waves which a tuning fork gives
have this form. Waves 3 and 4 in Figure 52 on the other hand, are musical ones,
but not of this simple shape. They are each made by adding, algebraically, waves
1 and 2 at different relative phases. Curve 3 is got by adding 2 at a' to 1 at A.

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of a curve.

For curve 4, 2 at b' was added to 1 at A. Obviously, the number of such composite
curves which one might make is unlimited, since one can also choose components
of different relative rates. For instance the period of one might be to the period
of the other, as one to three, or one to four, or two to five, and so on. Further,
one might add three or four waves together instead of only two. The period of
the composite is always that of the slowest component wave which it contains.
The low note is known as the fundamental. It is the shape of the wave which

84

EXPERIMENTAL PHYSIOLOGY.

FIG. 52. Musical sound waves. Nos. 1 and 2 are simple harmonic curves; Nos.
3 and 4 are composed of a fundamental of the period of No. 1 with an overtone of
the period of No. 2 super-imposed on it in different phases.

FIG. 53. Diagram of a vibrating violin-string to show how the various overtones arise.

HEARING. 185

gives it its quality. If, for instance, one hears middle C sounded on a violin it
has a quality different to that of the same note on the piano, and this again is
different from the sound of the tuning-fork, which, as we saw, gave a simple
pendular curve. The reason for this difference is that none of the sources in
the common instruments swing with a single vibration. A violin string, for
instance, swings as a whole, the rate of vibration of the whole length determining
the pitch of the lowest or fundamental tone in the composite curve of its note.
As well as this, however, the string tends to swing as if it were subdivided into
short lengths, halves, quarters, thirds, etc. (Fig. 53). The points dividing these
lengths swing with the vibration of the whole; the parts between swing with the
rhythm of the whole and with that of the part as well. The rate of vibration of
the divisions is to the rate of the whole inversely as the length of the division is
to the length of the whole. The result of this is that the sound wave arising from
the string is a composite one, like those which we have described, made up of a sine
curve with a period like that of the vibration of the whole string, upon which a
number of other waves have been superimposed, waves with rates in simple
arithmetical ratios (1/3, 1/4, 1/6, etc.) to the first. These arise from the vibra-
tions of the parts and are known as the OVERTONES. It is the number and in-
tensity of the. overtones which determine the quality of the note and it is because
these differ on different instruments that we are able to distinguish from one
another the notes that they give.

THE TRANSMISSION OF SOUND WAVES IN THE EAR.

Of the three parts of the ear, the external and middle chambers are only
concerned with the transmission of the sound waves, the essential organ of hearing
being entirely contained in the internal ear. The waves travel in air as far as the
inner end of the EXTERNAL AUDITORY MEATUS. Here the alternation of com-
pression and rarefication of which they are made up sets in motion the TYMPANIC
MEMBRANE. This membrane is made up of two epithelial layers, enclosing
between them a layer of connective tissue which contains both radial and circular
fibres. In shape it is somewhat peculiar, being slightly vaulted, with its convex
side inward. The apex of the vault, or UMBO, which forms a centre for the
radiating and circular fibres, is not exactly at the middle of the membrane but a
little above it. Because of its peculiar shape the membrane responds to waves
of a very large range of rates, instead of having a strongly marked natural period
of its own and being set in motion mainly by waves of this period, as would be the
case if it were simply a flat drum-head.

THE MIDDLE EAR. — Sound waves travelling in the middle ear are no longer
carried by air particles but by the swinging of a chain of three small bones, one
end of which is fastened to the end of the tympanic membrane and the other to
the membrane of the fenestra ovalis, one of the openings in the bony partition
between the middle and the inner chambers. The connection with the tympanic
membrane is through a long process, the manubrium of the MALLEUS, the first of
the three bones. The body of a malleus is connected by a joint with the second

186

EXPERIMENTAL PHYSIOLOGY.

H * a d of m~

FIG. 54. Diagram of the bones of the middle ear.

FIG. 55. Diagram of a lever drawn to show the mode of action of the
bones of the middle ear.

HEARING. 187

bone, the INCUS. This in its turn is in connection, by a process running down and
in, with the third bone, the STAPES, the inner end of which is continuous with the
membrane of the fenestra ovalis. Fig. 54. shows the shape and relationship of
the three bones. The malleus is rather loosely attached by a ligament to the
upper part of the outer wall of the chamber and besides the manubrium it has a
second process, a slender one, nearly at right angles to the manubrium, running
forward to the anterior wall and firmly bound there. The bone with which the
body of the malleus is joined, the incus, has two processes, one of which, already
mentioned, extends down and in to make connection with the stapes, while the
other, directed backwards, is firmly bound to the posterior wall of the chamber.
The stapes lies almost horizontally between the tip of the process of the incus and
the fenestra ovalis. The whole chain of bones acts together as a lever, one arm of
which is the manubrium of the malleus, the other the downward process of the
incus, together with the stapes, the fulcrum being somewhere in the lower part of
the malleus-incus joint (Fig. 55). When the membrane of the tympanum swings
in, the tip of the manubrium is pressed inwards. This would tend to move
malleus and incus bodily inwards except that they are firmly bound in an antero-
posterior direction by the attachment of the processes to the anterior and posterior
walls which we have already mentioned. These act as an axis around which the
two bones swing. When the tip of the manubrium is moved in, the head of the
malleus and the body of the incus swing out. Outward movement of the body
of the incus means movement in of the process which is attached to the stapes,
and this thrusts the stapes ahead of it. It will be seen in the diagram that the
manubrium (the power arm) is about half as long again as the process in the incus
(the load arm). The lever therefore reduces the excursion of the waves which
move it, but at the same time it increases their intensity. Movements com-
municated to the fluids of the internal ear need only be very tiny indeed to
stimulate the sensitive cells, so that this cutting down of the magnitude of the
sound waves is not a disadvantage. At the same time, even these tiny colums
of fluid have an appreciable inertia and on this account the increase in power of
the wave is important. A further increase in power is got from the concentration
of the force of the wave received by the whole surface of the tympanic membrane
into the much smaller area of the fenestra ovalis.

As we have said, a movement inwards of the membrane causes the head of the
malleus to move out, locking with the body of the incus and drawing it along.
If the membrane is moved out to any unusual extent, however, as happens for
instance when the surrounding pressure falls, the inward movement of the head
of the malleus which results is not followed by the incus. As Helmholtz pointed
out, the point between them is a cog or ratchet joint and locks firmly for movements
in one direction, but slides for large movements in the other. This is probably
important as a protection to the other parts of the middle, and to the internal, ear.

There are two tiny muscles in the middle ear. One of these, the TENSOR
TYMPANI, has its origin in a groove above the Eustachian tube, that is, from the
inner wall, and is inserted into the neck of the malleus below the axis around which
the bone rotates. When this contracts it must draw the manubrium of the

188 EXPERIMENTAL PHYSIOLOGY.

malleus inward and through it exert a pull on the membrane. What the useful-
ness is of this extra tension on the membrane, and the slight increase in pressure
of the middle chamber which it causes, is not clearly understood. It has been
suggested that the action is protective, damping the response of the membrane
to excessive sounds. The suggestion has also been made that the muscle con-
tracts reflexly when the attention is directed to sounds of high pitch, or rapid
vibrations, to increase the tension of the membrane and thus to make its natural
vibration period more rapid. The second muscle in the middle chamber, the
STAPEDIUS, arises from a projection on the inner wall and is inserted into the neck
of the stapes, on which, when it contracts, it exerts a lateral pull. The conjectures
as to the use of the tensor tympani apply also to this muscle.

If the tympanic membrane is to swing freely in response to sound waves
in the air the pressure on both its surfaces must be kept equal. This is en-
sured by the opening of the EUSTACHIAN TUBE which occurs during swallow-
ing and gives a communication between the middle ear and the pharynx. When

^^

Frc. 56. Cross section of the bony labyrinth showing the spiral of the cochlea.

the mucous membrane of the tube becomes swollen and the passage closed,
as in severe colds, the gradual absorption which goes on of the air contained in
the chamber of the middle ear lowers the pressure in it, the membrane no longer
vibrates freely, and a partial deafness results.

PHYSIOLOGY OF THE ESSENTIAL ORGAN OF HEARING.

THE INTERNAL EAR. The cavity of the skull which contains this part of the
labyrinth is spiral in shape, gradually tapering off to a point (Fig. 56). It is
lined with membrane and is filled with fluid. From the part of the bone which
forms the central pillar of the spiral a ledge projects into the cavity, reaching
about half way through its width and running almost the entire length of the spiral.
This partial partition of the chamber is completed by two membranes attached
at one side to the ridge of bone and at the other to two lines a little distance apart
on the membrane of the outer wall (Fig. 57). As a result the fluid of the spiral

HEARING.

189

is divided into three columns, one on one side of the partition of bony ledge and
the first membrane, one on the other side of the bony ledge and the second mem-
brane, and the third, enclosed between the membranes, roughly triangular in
shape, its third wall being made by the membrane of the outer wall itself. Of
the two outer columns one, the SCALA VESTIBULI, is closed, at the end which is
directed towards the middle ear, by the membrane of the fenestra ovalis and the
other, the SCALA TYMPANI, by the second membrane-covered opening in the parti-
tion, the fenestra rotunda. At the top of the spiral these two columns commun-
icate over the tip of the middle triangular one, which ends blindly a little short of
the upper end. Fig. 58 shows diagrammatically the relationship of the three.
Movements of the membrane of the fenestra ovalis cause vibrations of the same

FIG. 57. One turn of the cochlear spiral, enlarged to show the organ of Corti.

rate and shape in the double column of fluid contained in the two outer chambers,
play being allowed by the membranous end of the second column at the fenestra
rotunda, which moves out when the ovalis moves in and vice versa. Because the
partitions between them are only membranes, it is to be expected that the
oscillations of the two enclosing columns must affect the contents of the third.
It is among these structures that the endings of the auditory nerve are found and
we must look here for the essential organ of hearing.

190 EXPERIMENTAL PHYSIOLOGY.

Of the two membranes which form the walls of this triangular chamber
(Fig. 57) one is histologically quite simple. The other, the BASILAR MEMBRANE
supports a double row of pillars, standing with their bases apart and their tips
leaning together and projecting into the fluid of the middle column, the endolymph.
These are known as the PILLARS OF CORTI. Above these pillars on either side,
supported by the tent-like roof which they form, are several rows of cells, among
them a large number which have hair-like projections up into the endolymph,
The whole is known as the ORGAN OF CORTI. The fibres of the auditory nerve,
which runs within the inner pillar of the spiral, pass out by the bony projection
along its entire length and end in the hair cells, which must therefore be the cells
sensitive to the waves of sound.

As we have already seen, the ear is capable of distinguishing not only pitch
and intensity but also quality, in any waves which it receives. Not only do
notes of the same pitch, one without and one with many overtones, not sound
alike but with training the hearer can distinguish what the various overtones in
the notes are, so that our theory of hearing must afford a physiological basis for
this analysis. We have no problem of simplification to deal with in the ear as we
had in the eye; in hearing the sensations are as numerous as the stimuli which
evoke them. The most generally accepted theory is one proposed by Helmholtz,
the resonance theory. If one places a number of tuning forks of different periods
near a vibrating fork or reed, giving out a simple note, among those forks which
have not been struck the one with the same natural period will begin to vibrate
and give out its note, while the others remain silent. If the vibrating source is a

f. 0 u-a, 1 1 a

FIG. 58. Diagram of the relationship of the three fluid columns of the internal ear.

violin-string, which gives a note with overtones, not only will the fork with a
period like the fundamental respond but those with periods equal to each over-
tone as well. Helmholtz suggested that some such apparatus must be contained
in the ear. He at first thought that it was the rods of Corti which, having each
a different natural period, acted as resonators to the composite vibrations of the
fluid of the outer columns. The ear however is capable of distinguishing more
notes than there are rods; besides, rods are absent from the ears of singing birds,
and this idea had to be abandoned. It was then suggested that the resonators
might be found in the basilar membrane itself. It contains a series of radial
fibres running from the bony ledge to the outer wall and the width of the mem-
brane, and therefore the length of these fibres, increases progressively from the
base of the spiral to its apex. If these fibres are the resonators the arrangement

HEARING. 191

is like that of the strings in a tiny piano, with the short treble ones at the bottom
of the cochlea and the bass ones at its top. The difficulty is to see how even the
longest of these very small fibres can have a natural period as slow as that of the
lowest note which we can hear. Of course the period of them all is increased
because of the fluid about them but, even so, the difficulty has not been com-
pletely met.

FUNCTION OF THE SEMICIRCULAR CANALS.

Experiment 76. — Place a frog on a glass plate and cover with a
funnel or beaker. Rotate about longitudinal, transverse and
vertical axes (referred to frog's body), and notice compensatory
movements.

Expose one of the white otolithic masses in an anesthetized
frog by removing the cartilage after slitting the mucous mem-
brane of the roof of the mouth medially from the Eustachian
tube. Remove one otolithic mass in one 'frog and both masses
in a second frog. After recovery from the anesthesic compare
the compensatory movements and equilibration with a normal
frog when rotated in the different axes, and when swimming.

COMPENSATORY MOVEMENTS IN MAN.

Experiment 77. — Let the subject sit in a swivel chair in which
he is rotated 10 to 20 seconds. Describe his movements after
the chair stops. Let the subject describe his sensations during
and after the rotation with and without the eyes closed.

CHAPTER XXIV.

SKIN SENSATIONS. TASTE

There are three types of skin sensations, the appreciation of
differences of temperature (temperature sense), the sense of touch,
and the sense of pain. Since our knowledge of the normal behaviour
of each of these senses depends on subjective phenomena, each
student must perform the fundamental experiments as given below,
both on himself and on others, and must accurately describe the
results in his notes.

TEMPERATURE SENSE.

It is the rate at which heat is being gained or lost by the skin,
and not the actual degree of temperature of the applied object,
that is determined by our sensations.

Experiment 78.— Place a finger of one hand in water at 2° C.
and the corresponding finger of the other hand in water at 40° C.
After no temperature sensations are felt by either finger, transfer
them simultaneously at 30° C. Note the nature of resulting
temperature sensations in the two fingers.

The temperature sensations are received by "hot" and "cold"
spots scattered over the skin, the cold spots being the more numer-
ous (Fig. 59).

Experiment 79. — Mark out an area of skin on the back of the
hand, say 20 X20 mm. square. The hand should be resting com-
fortably on a table and the subject blindfolded. With a thermo-
aesthesiometer (a test tube drawn out to a point will serve the
purpose), containing water at 40° C., proceed to explore the
selected area, systematically, in parallel lines, marking with ink
the spots at which a distinct sensation of warmth is experienced.
When all the area has been explored accurately, transfer the
spots to ruled paper, and with the aesthesiometer containing
water at a temperature of 15° C., proceed in the same way to
determine the cold spots. Note that the cold spots, besides

192

SKIN SENSATIONS.

193

being more numerous, are more sharply defined than those of
warmth; the latter are also much more readily fatigued. This
fact must be remembered in repeating the observations.
More intense heat stimuli, besides stimulating the warm spots,

also stimulate those of jgold causing a cold sensatio

Experiment 80.

Test this using a temperature of about 50° C,
Mechanical and electrical stimuli, etc., applied to hot and cold
spots, may call forth thermic sensations.

Experiment 81. — Select a few pronounced cold and hot spots.
and stimulate them by tapping lightly with a small round
pointed object, or by applying the tetanizing electric current.

IP

FIG. 59. Heat and cold spots on the skin of the palm of the left hand: (a)
heatspots; (b) cold spots. The depth of the shading in each case represents the
intensity of the sensation.

The sensation of temperature remains for some time after the
condition causing it has been removed (temperature after-sensa-
tions).
Experiment 82. — Place on the forehead a coin which has been

either cooled or warmed, and note the nature of the sensations

which are experienced after its removal.

The temperature sense is not equally acute over the body.
Many factors influence the acuity but in general it may be stated
that clothed parts are more sensitive than unclothed, with the

194 EXPERIMENTAL PHYSIOLOGY.

exception of the temples, the lower eyelids and the cheeks. The
midline of the body is relatively insensitive. The sensivity in-
creases from the far extremity of the limbs towards the trunk.
Experiment 83. — Using test tubes containing warm or cold
water, proceed to confirm the above statements.

SENSE OF TOUCH.

This is likewise mediated by spots (touch spots), which are dis-
tributed independently of those of temperature, and are most
numerous to the "windward" side of the hair follicles.

FIG. 60. Hair aesthesiometers.

Experiment 84. — Using the same area of hand as was employed
for determining the temperature spots, or a part of it, make
an exact chart indicating the position of the hairs (use a lens for
this purpose). Now cut the hairs flush with the skin, by means
of a sharp scissors, or shave the part. Take a series of hair
aesthesiometers made by mounting straight hairs of varying
thickness at right angles on wooden holders (Fig. 60). The
exact weight at which each hair bends when pressed on one scale
pan of a balance is marked on the handle. Select a hair which
provides a stimulus of such a strength that it will call forth the

SKIN SENSATIONS. 195

peculiar sensation due to a touch spot. Having made certain
that the subject who should be blindfolded, can recognize this
sensation (and does not confuse it with one of pain, which is of
longer duration), proceed, systematically, to mark the touch
spots. Transfer the results to the chart, and note the relation-
ship of the touch spots to the hair follicles. By touching a hair
the sense of touch is accentuated. Repeat these observations
on the calf of the leg, where the hair follicles are less numerous.
Experiment 85. — Select an aesthesiometer which can produce
a distinct sensation of touch when its hair is pressed to the
bending point on the end of one of the skin hairs. Apply it to a
hairless part of the skin and note that it elicits no sensation.
It is important to distinguish between the sensations of deep
pressure and touch.

Experiment 86. — -Touch the skin lightly with a blunt point; then
gradually increase the pressure and note the occurrence of the
deep pressure sensation. Note that this occurs only after there
has been decided deformation of the surface.
The threshold value for touch (touch acuity) varies in different
parts of the body. Several things, besides the presence of hairs,
determine this (thickness of skin, rate of application of stimulus,
previous friction of skin, etc.). In general, however, the order of
touch thresholds is as follows: lips, finger tips and forehead; dorsal
aspect of finger; palm, arm and thigh; forearm; while much less
sensitive are the extensor surfaces of the forearm and the loins.
Experiment 87. — Using various hairs, determine the exact thres-
hold value for touch for the above regions by finding what thickness
of hair must be used to elicit the sensation of touch at the point
at which the hair just bends. Make a table of the results*
giving the "bending weight" of the hairs for each region.
Besides touch acuity it is important to study the LOCALIZATION
of touch sensations. This may be done either by asking the
(blindfolded) person to point with his finger to the exact place
where he was touched (absolute localization) or by ascertaining
the distance by which two points must be separated in order to

*In many of these experiments on touch a fine camel's hair brush can also
be used.

196 EXPERIMENTAL PHYSIOLOGY.

bed iscriminated (touch discrimination). This "local sign", as it

is called, varies in different parts of the body.

Experiment 88. — With a pair of calipers, determine this distance
for the tip of the tongue, the palmar surface of the third phalanx
of the finger, its dorsal surface, the tip of the nose, the back and
upper arm; make a table of the results. Note that the order
of sensitivity for touch discrimination is not the same as that
for touch acuity.

SENSE OF PAIN.

Any stimulus will cause pain when it is very intense. This does
not mean, as has been thought, that pain is merely due to over-
stimulation of any receptors. There are special receptors for pain,
just as there are for touch and temperature. One of the main
proofs of this is that pain spots are more extensively and regularly
distributed than the others.

Experiment 89. — Using the same area of skin as was employed
for the location of temperature and touch spots, proceed to mark
out the pain spots by pricking with a stout horse hair or a bristle.
Note that the sensation is more lasting than that for touch and
that the pain spots are distributed in a different manner.
Another proof of the separate existence of pain spots is that they
alone are found in the cornea; on the conjunctiva they exist along
with temperature spots.

Experiment 90. — Verify the above statement, using medium
hairs.

The skin sensations are not at all influenced to the same ex-
tent when the blood supply is interfered with. This fact becomes
of interest on account of its analogy to the results which have
been observed during the regeneration of severed nerves.
Experiment 91. — By means of a blood pressure cuff and bulb
applied to the arm, determine the systolic pressure, then release
the pressure and raise the arm so as to empty it of some of its
blood, after which again apply the pressure (but not above the
systolic value). Proceed to compare the sensations of touch,
temperature and pain in exactly corresponding skin areas of the
two hands. Differences may take some time to become evident.
Note particularly for which sensations they first become so.

SKIN SENSATIONS 197

TASTE.

Closely allied to the skin sensations are those of taste and smell.
The following experiments on taste are important.
Experiment 92. — (a) Dry the tongue and apply a crystal of salt

or sugar. Note that it is not tasted.

(b) On the dry surface of the tongue apply by means of a
camel's hair brush a solution of quinine (1 to 1,000), sodium
chloride (1 to 200), cane sugar (1 to 50), and sulphuric acid
(1 to 1,000). Determine at which part of the tongue the strong-
est sensation is produced by each.

(c) Prepare a series of solutions of gradually increasing
strengths of quinine, salt, sugar, and sulphuric acid, beginning
in each case at 1 in 5,000, and apply to that portion of the tongue
most sensitive to the respective substances. Determine what
strength of solution is necessary to stimulate taste.

(d) Apply with a brush a little of the sugar solution of the
strength which just stimulated taste, and follow with a drop of
salt solution of the same taste value. Note that the salt neu-
tralizes the effect of the sugar and neither is tasted.

(e) Take a solution of strong salt water into the mouth and
wash out with distilled water. Note that the water tastes
sweet (negative after-stimulation).

PRESSURE.

Experiment 93.— Weber's Law.— With the hand outstretched on
the table and palm upward, the subject being blindfolded, a
piece of cardboard 4 cm. square is placed on the finger tips.

With 5 gms. as the standard determine the least increment
necessary to detect a difference. In all of these observations
no muscualr effort must be made; moreover the weights must
be applied as gently as possible.

Determine the increments necessary for the following
standards: 10 gm., 50 gm., 100 gm., 200 gm., 1,000 gm.
Pressure and Muscular Sense. — Repeat the above observa-
tions with the hand removed from the table, the muscles of the
hand and arm being used to their utmost in determining the
threshold of difference.

SECTION VI.

DEMONSTRATIONS.

In the foregoing chapters experiments have been described
which the student, either individually or in small groups, can
perform for himself, but there are other experiments of equal im-
portance from an educational viewpoint, in which, for obvious
reasons this is impossible and it is necessary that these be demon-
strated. The value of demonstrations of more or less complicated
experiments in physiology depends on two factors, first, that the
student is familiar with the general technique of the methods em-
ployed, and secondly that the precise application and bearing of
the results of each experiment are appreciated by him. To stage
an experiment with the sole object of demonstrating results is
valueless, however important and fundamental these results in
themselves may be ; the demonstration becomes of value only when
the student knows exactly how the result is obtained and is able
to place it in relationship with previous knowledge gained by other
methods of study.

The particular experiments which it may be deemed advanta-
geous to demonstrate will necessarily vary, not only in different
laboratories, but also from year to year in the same laboratory. It is
believed, however, that it will be useful to describe briefly the
fundamental experiments that the authors are accustomed to
demonstrate to their classes, partly to assist the students to under-
stand what is being done, and partly as a guide to others. The
experiments that have been selected may also be performed by
advanced students and are described in sufficient detail for this
purpose.

The demonstrations are grouped in chapters, each chapter
giving in practicable sequence the experiments which can most
conveniently be performed on a single animal.

198

CHAPTER XXV.

THE CIRCULATION OF THE BLOOD AND LYMPH, AND
THE RESPIRATION.

The Reflex Nerve Control of the Circulation and of the
Respiratory Movements.

In Exp. 35, p. 79, it has been demonstrated that the heart
beat is controlled largely through the vagus nerve and that the
degree of resistance to the blood flow through the splanchnic
vessels depends on impulses carried by the splanchnic nerves.
The vagus and vaso-motor nerves arise in the medulla oblongata.
The efferent nerves of the respiratory muscles (phrenic and inter-
costal nerves) arise in spinal nerve centres, which are dominated
by a higher centre. This chief respiratory centre, as it is called,
is also situated in the medulla. The present experiment is devised
to throw light on the afferent pathways through which the above
centres become stimulated.

Demonstration 1. — Simultaneous tracings are taken of the
carotid blood pressure and of the respiratory movements (p. 97),
in an etherised and tracheotomised dog. The sciatic nerve in
one leg and the vagus nerves on both sides of the neck are exposed
and loose ligatures placed around them. A hot iron is then applied
to the skin of the foot while the recording drum is revolving at
such a speed that the cardiac pulsations can readily be counted.
Observe the effects on: (1) the pulse rate (2) the mean arterial
blood pressure and (3) the rate and depth of the respirations.
What general conclusions may be drawn concerning the influence
on the centres of over-stimulation of skin nerves?

After moderately tightening the ligature, the sciatic nerve is
cut peripheral to it and the hot iron reapplied to the skin. What
conclusion do you draw from the results?

The central end of the cut sciatic is then stimulated with the
faradic current, of a strength which just produces some change in

199

200 EXPERIMENTAL PHYSIOLOGY

one or other of the tracings, the observations being repeated with
stronger stimuli. The respirations are invariably increased in
depth and rhythm (hyperpnoea) and the heart rate is often reduced
but the effect on blood pressure as a rule is not readily measurable,
because of the exaggerated respiratory oscillations which the
tracing shows. Afferent fibres to the vagus and respiratory centres
are demonstrated in this experiment, and those to the vasomotor
centre can be inferred in cases where the blood pressure rises or
remains constant, while the pulse becomes slower. Why is the
conclusion permissible?

One vagus nerve is cut after moderately tightening the ligature
around it and the central end is stimulated. Are the results similar
to those of sciatic stimulation?

The results of the foregoing experiments indicate that the
centres of respiration and of vagus and vasomotor control are
simultaneously stimulated through afferent nerves. In order to
determine the relative importance of each effect it is necessary
to eliminate one or both of the other afferent influences. With
this object in view, the remaining vagus is cut so as to remove the
heart from vagus control. (What drugs could have been employed
to effect a similar denervation of the heart? Why is there a rise
in blood pressure?) The sciatic and vagus nerves are again stimu-
lated. What difference is there in the results? Explain the cause.

Frequently, by using induced shocks that are relatively slow
and feeble, the reflex effects on arterial blood pressure are the
opposite of those observed when shocks of ordinary frequency and
strength are employed. What conclusions do you draw?

Even the elimination of reflex vagus effects may not suffice to
demonstrate the pressor or depressor influence because of the pre-
dominance of the respiratory effects. It may be necessary to
eliminate the latter, which can be done by injecting through the
femoral vein a saturated solution of curare in physiological saline.*
Under the influence of curare the respirations cease, and to keep the
animal alive it is necessary to connect the respiratory pump to the
tracheal cannula, with the anaesthetic bottle inserted in the tubing.

*The curare solution should be prepared several days previously and should be
tested on a frog before using in the above experiment. The curare at present on
the market is very unreliable.

VASCULAR AND RESPIRATORY ORGANS. 201

(Etherisation must be maintained, otherwise the animal might
suffer pain. Why?)

While maintaining the artificial respiration the sciatic and
vagus nerves are stimulated as before. What conclusions can be
drawn from the results?

The effect of asphyxia on the arterial blood pressure is then
studied, the asphyxia being induced by suspending the artificial
respiration. How do the results compare with those observed
during asphyxia with the vagi intact? After the blood pressure
has finally fallen to zero the thorax is opened and the heart ex-
amined. Extreme venous engorgement is observed. Note the
behaviour of the heart when the engorgement is relieved by punc-
turing the right ventricle. What conclusions do the observations
warrant?

CHAPTER XXVI.

The Direct Demonstration of Vasoconstrictor Fibres by
the Plethysmographic Method.

Demonstration 2. — The abdomen is opened in an etherised,
tracheotomised dog and the intestines retracted to the left side
under towels wrung out in warm saline solution. The great splanch-
nic nerve is isolated and a ligature placed around it on the right
side (see p. 83). The right kidney is freed from its bed and the folds
of peritoneum running to its upper end are cut after applying mass
ligatures. The pedicle is carefully freed by blunt dissection and
the plexus of nerves isolated from it and a ligature placed around
them. An oncometer, with cotton wool well mixed with vaseline
laid in the groove for the pedicle, is then placed under the viscus
and more cotton and vaseline placed in the groove until this is
filled (without any compression of the vein or ureter) after which the
glass lid is applied and clamped down. (There must be no strain or
kinking of the vessels during the manipulation). The side tube of
the apparatus is finally connected by moderately thick walled
tubing with a bellows recorder, the writing style of which is accur-
ately adjusted in the same perpendicular with the writing style of
a mercury manometer connected with the carotid artery. A re-
spiratory tracing should also be taken. On a moderately fast drum
tracings are taken to show the relationship of the oscillations of
the oncograph to the arterial and respiratory tracings. (What is
the relationship?)

The effect produced on the kidney volume and the arterial
blood pressure is then observed: (1) during (electrical) stimulation
of the vagus in the neck; (2) during stimulation of the great splanch-
nic nerve on the right side (Fig. 61), and (3) during stimulation of
the renal nerves. How do you interpret the results? Why is the
arterial blood pressure essential in the interpretation?

A small amount of physiological saline containing 1-10,000
adrenalin chloride is now injected into the femoral vein. What
conclusions do you draw from the result?

202

THE PLETHYSMOGRAPH.

203

111111111 infill

FIG. 61. Tracing of volume of kidney and of arterial blood pressure
during stimulation of the splanchnic nerve.

204

EXPERIMENTAL PHYSIOLOGY.

FIG. 62. Tracing of volume of kidney and of arterial blood pressure during asphyxia.
slight rise in blood pressure accompanied by marked diminution in the kidney volume.

Not

THE PLETHYMOGRAPH 205

Curare is injected into the femoral vein until the respirations
cease (see p. 200). Asphyxia is allowed to supervene by suspending
the artificial respiration (Fig. 62), but this is re-established before
the final fall in blood pressure sets in. What evidence does the result
furnish concerning the cause of the asphyxial rise in blood pressure?
(If satisfactory curare is not available, the asphyxia may be in-
duced by clamping the trachea. The results are, however, less
clear because of the respiratory convulsions).

Finally, the renal nerves are cut, and, the immediate effect on
the kidney volume having been observed and explained, several of
the foregoing experiments are repeated. How do you explain the
alterations in the results?

The anima1 is killed by haemorrhage while the various tracings
are being taken.

In one or other of these demonstrations the opportunity may
present itself of attempting the resuscitation of the animal by
intra-arterial transfusion with epinephrin. To effect this
a cannula, inserted in the central end of the carotid artery, is con-
nected with a burette containing warm isotonic saline solution,
which is allowed to flow into the artery. Artificial respiration is
also performed and after seme saline has transfused a few cubic
centimetres of adrenalin chloride is injected into the tubing which
connects the artery with the burette and the thorax is vigorously
massaged over the region of the heart. In a successful experiment
the blood pressure, recorded from the carotid on the opposite side
to that through which transfusion is being made, will be observed
to rise, at first passively because of the thoracic movements, and
later because of resumption of the heart beats. Once these have
started the blood pressure quickly recovers and the cardiac massage
may be suspended. Explain the mechanism of the resuscitation
and particularly how the epinephrin acts. Would intravenous
transfusion be of any avail in attempting the resuscitation after
complete stoppage of the heart beat?

CHAPTER XXVII.

PERFUSION OF THE EXCISED MAMMALIAN HEART.

If a solution containing certain of the inorganic constituents
of blood plasma and saturated with oxygen gas be perfused under
a certain pressure through the coronary arteries, the heart will
beat with perfect regularity for several hours outside the body.
This surviving heart preparation is particularly useful for investiga-
tions of the influence on the heart of variations in the saline con-
stituents of the nutrient fluid and of the effect of drugs. The nutrient
fluid is caused to circulate through the coronary arteries by intro-
ducing it into the aorta, where it closes the semilunar valves and
is forced into the openings of these arteries. The observation also
demonstrates most convincingly the remarkable recuperative
power of the heart, and it affords evidence that resuscitation
methods should be persistently applied after drowning accidents
and when death has occurred from other forms of suffocation.

The most convenient apparatus to use is that depicted in Fig. 63. The per-
fusion fluid (Ringer-Locke's or Tyrode's) is contained in the bottle A, and it is
kept constantly saturated with pure oxygen by bubbling the gas through it.
The fluid then passes to the cannula in the aorta through a warming device which
consists of a wide tube surrounded by a water jacket kept at the desired tempera-
ture by a thermo-siphon system, as shown in the diagram. In order that the
fluid may be thoroughly warmed it is caused to flow in a thin film on the walls of
the tube (B) by placing in this a somewhat smaller tube (C) closed at both ends.
Although it is convenient to use an apparatus in which the free end of the cannula
is ground on its outer surface so as to fit the lower end of the inner tube of the
warmer — an apparatus devised by the late T. G. Brodie — the simpler apparatus
devised by Gunn, which is also shown in the figure, is perfectly satisfactory.
In Brodie's apparatus a thermometer (T) is inserted into the aortic cannula
through a side tube; in Gunn's apparatus the thermometer is made to serve as the
inner tube of the warmer.

The heart is prepared for perfusion as follows: A rabbit is killed by a blow
on the back of the head and is immediately laid supine on the table. A longi-
tudinal incision is made through the skin of the thorax, extending for a short
distance on the abdomen. The skin is retracted to both sides of the incision
and the abdomen opened at its upper end. One blade of a stout pair of scissors
is then passed through the diaphragm under the cartilages of the ribs on one side

206

PERFUSION OF THE MAMMALIAN HEART. 207

A f

FIG. 63. Apparatus for perfusion of the mammalian (rabbit) heart. The apparatus con-
nected with the heating device is that of Brodie. Apparatus of Gunn is shown in insert.

and these are cut from below upwards, care being taken to keep the blade away
from the heart and lungs. The cartilages are similarly cut on the other side of the
sternum. The isolated piece of sternum is bent upwards and cut across at the
upper end. The exposed pericardium is picked up with a forceps and opened with

208 EXPERIMENTAL PHYSIOLOGY.

fine scissors, after which the animal is placed on the left side, so as to expose the
structures at the root of the right lung. These are cut through and the operation
is repeated on the other side. The aorta must now be made out and cut across
just proximal to where its first branch (the innominate) comes off, after which the
superior vena cava is cut a short distance from the auricle. A snip is made in the
apex of the right ventricle, using a small sharp-pointed scissors for the purpose:
this is to permit of free escape of the fluid from perfusing the heart. Finally a
thread attached to a packing needle is inserted in the inferior vena cava, and cau-
tiously guided upwards so that it emerges from the superior vena cava. The
needle is unthreaded, and the thread tied in a loop. This thread is introduced
to serve as a guide to the venae cavae into which later a tube is to be inserted.
The heart is removed, by cutting the inferior vena cava below the thread, and
placed in a dish containing cold Locke's solution. Any blood remaining in the
right auricle and ventricle is then washed out by introducing Locke's solution
from a large pipette through the superior vena cava. The special cannula is
now tied into the aorta, care being taken that no air bubbles are entrapped.
(This part of the technique is more conveniently performed with Brodie's appara-
tus than with Gunn's, but it is perfectly simple using the latter). A bent pin
hook is passed through the outer coat of the right ventricle and a thread attached
to it is carried over a pulley to a heart lever arranged to write on a drum. A
similar hook and thread may also be attached to the auricle so that simultaneous
tracings of auricle and ventricle may be taken. In order to steady the heart it is
well to pass a thread through the apex of the left ventricle and tie it below to a
glass rod fixed to a stand.

Demonstration 3. — Having secured a tracing of the heart and
noted the temperature of the perfusion fluid, the following observa-
tions may now be made:

1. The influence of changing the temperature of the perfusion
fluid. This can best be done by adding hot or cold water to the
copper vessel connected with the water jacket.

2. The influence of the local application of heat and cold in the
right auricle. To demonstrate this, pass a thin-walled glass tube
through the superior and inferior venae cavae using as a guide the
string previously placed in the vessels. The upper end of this tube
is connected by rubber tubing with a funnel, and either cold or hot
water is made to flow through it while a tracing of the beat is
being recorded. The temperature of the perfusion fluid is mean-
while kept exactly constant (cf. Sherrington).

3. The effect produced on the beat by increasing the relative
proportion of the calcium or potassium in the perfusion fluid.
This is very conveniently accomplished in the Brodie apparatus
by closing one of the side tubes on the inflow tube leading to the

PERFUSION OF THE MAMMALIAN HEART 209

upper end of the warmer with rubber tubing through the wall of
which is inserted the needle of a small hypodermic syringe con
taining the solution. For more continuous addition of greater
dilutions of the foreign solution the side tube may be connected with
a burette. In Gunn's apparatus a Y tube is provided for these
operations. By thus injecting the foreign solution at a distance
from the heart it is mixed thoroughly with the perfusion fluid and
is properly warmed before the heart is reached. Observations should
be made using first of all a few c.c. of a 1% solution of calcium
chloride. When the beat becomes very small and the ventricle is
almost in calcium rigor (see p. 70) inject a few cc. of a 1% solution of
KC1. If the beat does not become restored to the normal in a few
minutes or so, inject a little more KC1 solution. Then add still
more KC1 until the heart again almost comes to a standstill (in
diastole, p. 57), when it may again be restored by injecting CaCLj
solution.* It is, of course, evident that this observation does not
in itself prove that the two cathions are mutually antagonistic to
each other. To prove this it would be necessary to show that
without the addition of the antagonistic cathion the beat is much
more slowly restored.

4. The effect of alteration of the H-ion concentration of the
perfusion fluid. This is done by cautiously adding, by the above
procedure, 0.9 per cent, solution of NaCl containing either some
HC1 or some NaOH.

5. The effect of epinephrin. For this purpose about 1 c.c. of a
0.002 per cent, solution of adrenalin chloride solution should give
definite results.

6. The effect produced by applying a tetanising electric current
to the ventricle. Whenever the fibrillation becomes marked dis-
continue the stimulation and see whether the normal beat becomes
restored by continuing the perfusion. Observe carefully the be-
haviour of the auricles.

It is usually necessary to employ more than one heart for all
of the above observations. Sometimes, however, all can be done
on one preparation.

*It is not possible to state precisely what amount of the two solutions should
be used, because this will vary with the rate of the main perfusion. It may be
necessary to repeat the observations several times before the effects are obtained.

CHAPTER XXVIII.

MEASUREMENT OF THE BLOOD FLOW THROUGH THE
HANDS AND FEET, BY THE CALORIMETRIC
METHOD OF G. N. STEWART.

ELECTROCARDIOGRAPHY

Because of their relatively great exposed surfaces the hands,
and to a lesser degree the feet, quickly give off heat to their en-
vironment. They are excellent radiators. The muscles of these
parts when at perfect rest, contribute but a very small quota to
this heat, most of which therefore is carried to them by the blood
from other parts of the body. From these considerations it follows
that the amount of heat given out in a given time must be pro-
portional to the amount of blood flow, or expressed in precise
terms: the amount of heat given off in calories, divided by the
difference in temperature between the blood flowing to and away
from the part equals the volume flow of blood in cubic centimetres.*

If Q be the grammes of blood per minute, H the gram calories
in m minutes, T the temperature of the blood entering the hand and
T' that of the blood leaving it, S being the specific heat of blood then :

_ H 1

Q== m(T-T') T"

The calories are measured by placing the hand in a water
calorimeter; the temperature of the entering blood may be taken,
as that under the tonguef and the temperature of the leaving
blood T', as the mean temperature of the calorimeter during the
observation.

The calorimeter (see Fig. 64) consists of a tinned inner copper vessel capable
of holding somewhat over 3000 c.c. of water. This vessel is placed within a wider
vessel, the space between the two being packed with broken cork. To the open
upper end of the inner vessel is soldered a lid of copper with a hole cut in it
towards one side of the centre and large enough and shaped so as to allow the

*A small or gram calorie is the amount of heat required to raise the tempera-
ture of 1 gm. of water through 1° C.

fBecause of the liability of the mouth temperature to vary, it is more accurate
to take for T the rectal temperature less 0.5° C.

210

BLOOD FLOW IN MAN.

211

hand to pass into the vessel. About an inch beyond the edges of this hole a
strip of copper, about 1 inch wide, is soldered at right angles to the lid. Besides
the hand-hole, and on the opposite side of the centre, three small circular holes
are made in the lid each about one inch in
diameter and provided with a copper
tube of similar diameter, projecting up-
ward. A piece of insulating cork is cut so
as to form a lid large enough to fit the
outer vessel, and holes are made in it to
fit the tubes round the openings in the
copper lid of the inner vessel. After fitting
the cork cover in place, it is covered with
varnish. The three small holes are used,
one of them for a thermometer, and the
others for long feathers to serve as
stirrers. A piece of thick saddler's felt
is cut to fit the hand opening in the lid.
Several collars of the same material are
also cut, 1 inch wide, and of various sizes
to fit wrists of different sizes. The felt
lid and collars rest on the copper ledge
between the head opening and the ridge
of copper. A thermometer with a
scale which reads between 0° C. and 50°
C. divided into l/10ths is sufficiently
accurate. By using a telescope the
temperature can be read to l/50th or
I/ 100th of a degree.

Besides the calorimeter, a large bath
of 20 or 30 litres capacity (garbage can)
is necessary. It is also convenient to have
an adjustable stool (like a music stool)
on which to place the calorimeter.

Demonstration 4.— To make
the measurement, the first step
is to draw a line with a fat pencil
on the skin at the lower border of
the styloid process of the ulna of
the hand chosen for observation.
A felt collar of proper size is then
applied to the wrist with its lower

border corresponding to the pencil line, and a second line is drawn
on the skin opposite the upper border of the collar. This upper line
will serve as a mark to show that the hand is not changed in position

FIG. 64. Stewart's Calorimeter.

212 EXPERIMENTAL PHYSIOLOGY.

while in the calorimeter. The hand of the subject, who is sitting
on a high chair, is then placed in water at about 30-32° C. contained
in the large bath (the temperature of the water is taken by an
ordinary thermometer) and it is left there for ten minutes, or so, in
order that the skin and other tissues of the hand may be brought
to the same temperature as the water. By so doing, conditions
are established which are analagous to those which would exist if
the blood vessels were alone suspended in the water bath. The
opposite hand is covered with a glove, or placed in the pocket, so
that it may not become cooled and so cause reflex vaso-constric-
tion in the observed hand. While the hand is being brought
to the correct temperature, 3,000 c.c. of water is removed from
the large bath and placed in the calorimeter, the hand opening
of which is then closed by the felt lid. The water is occasionally
stirred by the feathers, the thermometer is placed in position, and
the telescope adjusted so that an exact reading of the temperature
may be taken.

When the ten minutes is up, the hand is withdrawn from the
bath, the wrist quickly wiped dry with a towel, the hand with the
fingers extended carefully passed through the hand opening into
the water in the calorimeter, and the subject instructed to keep the
fingers abducted and extended and not to move them. He must
also be warned not to touch the thermometer. The felt collar is
placed round the wrist and the height of the calorimeter is adjusted
so that the subject does not require to strain his body in order to
keep the hand in the correct position (or with the upper skin line
at the edge of the felt collar).

The observer sits on a low stool behind the subject, and having
noted the time, and stirred the water in the calorimeter, proceeds
to record the temperature. The stirring is maintained throughout
the observation, which lasts 10 minutes, and the thermometer is
read every 2 minutes. When time is up, the felt collar is removed,
the hand carefully withdrawn, and the hand opening covered by
the felt lid. It is necessary to observe the temperature in the
calorimeter for a further period of 10 minutes, with constant stirring,
so as to determine the extent to which it is losing heat to the air
(the self-cooling of the calorimeter).

The volume of the hand is determined by placing it, while still

BLOOD FLOW IN MAN. 213

wet, in water which completely fills a lipped thick-walled beaker of
suitable size. The hand displaces an equal volume of water, which
overflows into a basin in which the beaker is standing. When no
more water overflows, the hand is slowly withdrawn, and water
is added from a graduate to the beaker until it is again full; the
amount of water required gives the volume of the hand.

The temperature of the mouth or rectum is finally taken and
the temperature of the room, read from a thermometer hanging
from the chair on which the subject sits, recorded.

To apply the above formula to calculate the blood flow it is necessary to
introduce several corrections to allow for differences in the specific heats of the
hand tissues, of the metal of the calorimeter and of water. These corrections are
expressed as the water equivalents of the hand and calorimeter. The water
equivalent of the calorimeter is determined experimentally for each calorimeter,,
and for one of the above dimensions will be about 100 c.c. The water equivalent
of the hand is obtained by multiplying its volume by 0.8, this factor being the
product of the specific gravity and the specific heat of the hand.

Suppose in an experiment that the temperature of the calorimeter during an
observation lasting 10 minutes had risen from 31° C to 31.5° C and the self-cooling
of the calorimeter during the subsequent 10 minutes was 0.1° C, the mouth
temperature being 37.5° C. and the volume of the hand 450 c.c., then (applying
the above equation) :

(3000+100+360)c.c.X(0.5+0.1)°C 10*_

37.5-31.25f ( 9 =

The measurement should now be repeated under the following
conditions:

1. When the opposite hand, instead of being carefully pro-
tected from cooling, is placed in cold water. A marked curtail-
ment of blood flow, due to reflex vaso-constriction, will be observed.
This observation may be made in continuation of the previous one,
i.e., the unobserved hand kept covered for 10 minutes and then
placed in cold water for the next 10 minutes. It is particularly
important in these observations to read the calorimeter temperature
at frequent intervals, because the vaso-constriction that is im-
mediately induced gives place later to a dilatation, even while
the opposite hand is still in the cold water. The blood-flow result

*This is the reciprocal of the specific heat of blood.

fThis is the mean temperature in the calorimeter during the observation

(36 9)
of the hand ---— - -8.2

214 EXPERIMENTAL PHYSIOLOGY

for each two minutes should be plotted on coordinate paper to
show this phenomenon.

2. After the hand and arm have been used to perform work,
such as dumb bell exercise, until almost fatigued.

ELECTROCARDIOGRAPHY.

Demonstration 4a. — Electrocardiography. — The electro-
cardiograph is essentially a very sensitive galvanometer. A fibre
of finely spun glass coated with a film of silver to render it elec-
trically conductive is suspended between the poles of a powerful
electromagnet. A current passing through the fibre or string, as
it is termed, causes a deflection of the string which varies with the
direction of the current. When the base of the heart is relatively
negative to the apex the current is directed upwards, when the
apex becomes negative the current passes downwards through the
string.

By means of an arc light and a series of lenses a highly magnified
shadow of the string is cast upon a moving photographic plate
which descends at the desired speed in a carrier contained within
the camera box. In this way the movements of the string are
recorded.

The arc lamp having been lighted and the shadow of the string
focussed upon the scale situated on the front of the camera, the
observed person is seated in the chair with each hand and a foot
placed in the respective electrode basins. The key, A, should be
in the short circuit position and the key B at "off" before the
subject is connected to the electrodes. (See Fig. 64a). The magnet
should be now excited by turning on the house current and the
current of the standard cell thrown into the circuit by closing the
battery switch. The current of the standard cell serves two
purposes, to compensate for the skin current and to standardize
the excursions of the string. The electrode switch E is next adjusted
for the particular lead from which it is desired to obtain a record.
The switch B may now be turned to " patient" and the switch A
turned from the short-circuit position to 1/100. A fraction of the
current from the subject is now passing through the string and,
though the cardiac deflections, if seen at all, will be very minute,
the string moves from the zero point and takes up a new position.

BLOOD FLOW IN MAN

215

This change in the position of the string is due to skin currents and
must be compensated for. The current of the standard cell which
is in the opposite direction is used for this purpose ; the compensator
C is therefore turned until sufficient current has entered the cricuit

G O O

0!0

216 EXPERIMENTAL PHYSIOLOGY.

to bring the, string back to its original position. The switch A is
next turned to 1/10 and finally to 0, any movement of the string
from zero being compensated for each time. The skin current is
now fully compensated for and the cardiac deflections are more
pronounced.

In order that all electrocardiograms may be compared one with
another, the sensitivity of the string must be standardized so that
a given strength of current will produce a certain magnitude of
deflection. The standard used is that of Einthoven — a deflection
of 1 centimetre for a millivolt of current. The switch A is turned
to the left from 0 to 1. One millivolt derived from the standard
cell is now passing through the string and causes it to be deflected
to an extent which is dependent upon the sensitivity of the string.
With the aid of the scale on the front of the camera box the magni-
tude of the deflection is measured. If it is less than 1 c.m. the
string is not sensitive enough and should be slackened. This is
accomplished by turning the mill-headed screw above the fibre
case to the right, i.e., in the direction of the arrow marked thereon.
If the deflection is more than 1 c.m. the string is two sensitive and
should be tightened by turning the milled head to the left.

The deflections of the string are now standardized and a record
may be taken. The plate is exposed by removing its sliding cover,
the camera slit is opened and the carrier released. As the plate
descends behind the lens of the camera the string movements are
recorded. The time in 1/50 of seconds is indicated by upright
lines marked upon the plate by means of a revolving wheel placed
between the source of light and the string. Horizontal lines repre-
senting 1/10 millivolt are marked upon the plate by means of
lines engraved upon the surface of the camera lens. A record
should be taken from each of the three leads.

Each student will be provided with a record which he should
study with regard to the following points —

1. Identify the various waves upon the tracings denoting each
by its appropriate letter.

2. Measure the length of the P-R interval. What may an
examination of this interval tell you concerning the heart's action?

3. Paste the electrocardiogram in your note-book and draw
beneath it a jugular pulse tracing and an intraventricular pressure

BLOOD FLOW IN MAN. 217

curve so as to synchronize accurately the various cardiac events
represented in the three tracings.

4. Determine in millivolts the manifest value of the potential
change occurring during the production of the R wave in lead III.

5. Determine the direction of the electrical axis during the
production of the R wave.

6. Draw a diagram showing the course of the cardiac action
current and the movement of the string during the production of the
P wave.

CHAPTER XXIX.

LYMPH FORMATION.

Demonstration 5. — A large dog is given a meal containing an
excess of fat (lard) several hours prior to the experiment. After
anaesthetising with morphine and ether and inserting tracheal and
carotid cannulae the thoracic duct is exposed as it enters the left
subclavian vein at the root of the neck. This operation is rather
difficult and should be performed as follows: after extending the
incision through skin and subcutaneous tissue down to the sternal
notch, the sterno mastoid and sterno hyoid muscles are cut as low
down as possible on the left side and reflected upwards. The ex-
ternal jugular vein is then followed downward till it joins the
subclavian, which is traced inwards to its union with the internal
jugular. In the fork at the union of these two veins the thoracic
duct is sought for by very careful dissection (see Fig. 65), great
care being exercised so as not to wound the pleura which lies
immediately beneath the vein. Just before entering the vein the
thoracic duct, curving forwards and outwards, is joined by the
somewhat smaller neck lymphatic. The duct is rendered visible
by the white creamy fluid it contains, the neck lymphatic being
similarly injected to a lesser degree. (Usually it is advisable to
ligate the neck lymphatic, but when possible a cannula should be
placed in it (pointing upwards) since the lymph flow from this
lymphatic does not behave exactly like that from the thoracic
duct). To introduce the cannula into the thoracic duct two liga-
tures are placed under the latter, the one next the subclavian vein
being tied, and a slit is made with a fine pointed sharp scissors in
the duct; the edge of the slit is then caught in a fine pair of forceps
and a glass cannula (with an outside diameter of about 1/32 inch)
inserted and tied in by the second ligature. If the creamy lymph
escapes so freely from the slit in the duct that it fills the wound
its flow should be controlled by pulling gently on the free ligature.
Usually a certain amount of lymph flow is desirable since it facili-

218

LYMPH FORMATION.

219

FIG. 65. Dissection necessary for exposure of thoracic duct and neck lymphatic. Ligatures
are placed around these structures. (After Jackson).

220 EXPERIMENTAL PHYSIOLOGY.

tates insertion of the cannula. It should be remembered that the
lymph clots readily, and if this occurs in the neck of the cannula,
the clot will require to be broken up by means of a fine hair. The
cannula is connected by a short piece of flexible rubber tubing with a
bent glass tube, from the end of which the lymph is allowed to drop.

A cannula is also inserted in the femoral vein and connected
with a burette.

There are two types of experiment which may be demonstrated
on lymph flow. One of these concerns the stimulating effect of
certain chemical substances when injected into the general circula-
tion— the lymphagogues — and the other, the effect of alterations of
the circulatory conditions in the intestines and liver.

The lymphagogues are of two classes, saline and colloid. To
illustrate the former a strong solution of glucose (20 per cent.) is
injected intravenously a few c.c. at a time until an increase in the
number of drops of lymph is produced. The amount of injection is
then increased. If the neck lymphatic flow is also being observed,
particular regard should be given as to whether it behaves similarly
to that of the thoracic duct. Repeat the experiment, using a 10 %
solution of sodium chloride. Explain the results.

In the foregoing experiment it will be observed that the arterial
blood pressure either remains unaltered or rises slightly. To
illustrate the colloid lymphagogues, small quantities of a solution
of commercial peptone are injected. Note that besides affecting
the lymph flow there is a decided drop in arterial blood pressure.
(Care must be taken not to allow this to reach the danger limit).
Explain the result.

Finally it may be shown that certain drugs and hormones also
act as lymphagogues. To illustrate this, pituitrin may be used
(1 c.c. for a dog of average size, 8 kgm.).

The influence of circulatory changes in the splanchnic region
may be illustrated as follows: after opening the abdomen in the
linea alba a loose ligature is placed around the portal vein near
the hilus of the liver. When the normal rate of lymph
flow (from both ducts) has been ascertained the vein is closed by
pulling on the ligature; a decided increase in flow is observed.
If the ligature be tightened for long the lymph will become turgid
with blood. How do you explain the results?

LYMPH FOUNDATION 221

Finally the lymphatics which accompany the portal vein are
ligated. This is readily done by separating the vein by blunt dis-
section from the accompanying structures, which are then mass
ligated. A loose ligature is next placed around the inferior vena
cava just above the diaphragm, by making an opening at the
posterior end of the 6th or 7th intercostal space on the right side,
prying the ribs apart with a strong pair of retractors (meanwhile
maintaining artificial respiration) and threading the ligature
around the vein by a long aneurysm needle. After applying the
ligature, the opening in the thorax should be closed by artery for-
ceps, when it will often be found that artificial respiration can
be discontinued. Having observed the rate of lymph flow the
ligature is tightened for a few moments. Is the effect similar to
that of the previous experiment? How is it explained?

SECRETION OF URINE

Demonstration 5a.— The experiment may be performed on a
dog or cat anaesthetized, preferably with urethane (2 grams per
kilo given by stomach with an abundance of water). Insert a
canula in trachea, a canula in femoral vein for injection, and a
canula in the right carotid artery for recording blood pressure
with mercury manometer. Expose the left vagus nerve and
arrange for stimulating the peripheral end. The secretion of
urine is to be observed by counting the drops which issue from a
canula inserted into the urinary ducts. In the case of the dog
the abdominal cavity may be opened and a slender canula
inserted into one of the ureters. In the case of the cat a large
canula may be inserted into the urethra by means of an incision
made just cephalad of the pubis. The bladder will empty itself
through this canula, and if care is taken that the urethral canula
does not become blocked all urine flowing from the ureters will
pass through this canula without accumulating in the bladder.

Do not attempt to repeat the observations because the secretion
will not continue indefinitely. Observe the following:

(A) The Effect of Blood Pressure upon the Secretion of Urine.

1. Determine the normal blood pressure and rate of urine flow.

2. Inject 0.5 c.c. 1, 10,000 Adrenin into the femoral vein.

222 EXPERIMENTAL PHYSIOLOGY.

(a) What is the effect on blood pressure; on diuresis?

(b) How can these effects be correlated?

3. Stimulate the vagus.

(a) What is the effect on blood pressure; on diuresis?
(E) The Effect of the Chemical Composition of the Blood upon
the Secretion of Urine.

4. Inject 15 c.c. of a 1% solution of urea into the femoral vein.

(a) What is the effect on diuresis?

(b) Can this effect be attributed to changes in blood pressure?

5. After the diuresis returns to normal inject 50 c.c. of a warm
isotonic salt solution slowly into the femoral vein.

(a) What is the effect on diuresis?

(b) Can this effect be attributed to changes in blood pressure?

(C) The Time Interval of Urine Secretion.

Introduce 5 c.c. of a saturated solution of indigo-carmine into
the vein. Note the time which elapses until the blue colour first
appears in the urine.

(D) Asphyxiation of Kidney.

6. Close the tracheal cannula with the finger for 60 seconds.

(a) Note and explain effect upon diuresis.

(b) Can this be explained by the changes in blood pressure?
The operator may now open the body cavity and place ligatures

around the renal arteries and veins of both kidneys and around
the aorta caudad to renal arteries. Look for rhythmic movements
of the ureters.

7. Close aorta below renal arteries.

(a) State and explain effect on diuresis.

8. Close the renal veins by pulling up on the ligatures about
them. Do not check the renal circulation for more than a minute
in this way.

(a) State and explain effect on diuresis.

9. Close the renal artery by pulling up on the ligatures for three
minutes.

(a) What is the effect upon diuresis?

(b) Explain.

(c) Does the kidney recover from the asphyxiation?

CHAPTER XXX.

EXPERIMENTS TO DEMONSTRATE THE PUMPING

ACTION OF THE HEART AND THE ACTION OF

THE VALVES.

Demonstration 6. — A wide glass cannula is tied into the
superior vena cava of the excised heart of a dog or sheep and the
cannula connected by rubber tubing with a funnel. A glass tube
is also tied into the pulmonary artery, the upper end of the tube, at
a distance of about 50 cm. from the heart end, being bent double
and arranged so that the opening lies over the funnel which is
connected with the vena cava. The inferior vena cava is tied and
the preparation and tube are held by suitable clamps in a vertical
position ; water is poured into the funnel so that it distends the right
auricle and ventricle. Some of the water escapes through cut
vessels (left azygos vein) which are now tied. When the ventricle
is rhythmically compressed by the hand the water, which has mean-
while risen in the pulmonary tube to the level of the water in the
funnel, rises higher and higher with each compression, and remains
up between them, until it reaches the bend and flows back into the
funnel. This illustrates the circulation of the blood through the
heart.

It is interesting to study the effect produced by DAMAGING THE
SEMILUNAR VALVE. The fluid still rises in the tube with each
compression, but leaks back into the ventricle between the "beats".
To raise the fluid in the tube high enough so that it overflows into
the funnel it is now necessary to compress the ventricle much more
rapidly. This illustrates in a rough way how the heart may com-
pensate for a valvular inefficiency by more energetic action.

The OPERATION OF THE TRicusPiD VALVES is also readily shown
by removing the tube and cutting away most of the right auricle.
When the ventricle is filled the water flaps are floated up into
position, a narrow chink, however, remaining in the centre. This
can be temporarily closed by allowing the water to drop in the

223

224

EXPERIMENTAL PHYSIOLOGY.

FIG. 66. Gad's arrangement for observing the action of the cardiac valves. (After Tigerstedt)

HEART PREPARATION. 225

centre of the chink; following each drop the flaps come accurately
together because the waves of pressure produced by each drop
are reflected onto the under surface of the flaps from the walls of the
ventricle. What significance may this observation have in con-
nection with the mechanism of the closure of the valves in the
normal heart?

To illustrate the efficiency of the tricuspid valves, ligate the
pulmonary artery, fill the ventricle with water, and hold the ven-
tricle upside down; the water stays in.

The operation of the valves can be very clearly shown by ob-
serving them through windows inserted in the right auricle and
pulmonary artery. This is known as GAD'S HEART PREPARA-
TION.

Demonstration 7. — The general arrangement, using an ox
heart, is shown in Fig. 66. A brass tube A (40 cm. diam.)
closed by a glass window at one end and with a narrow tube soldered
into its side is tied into the right auricle, and another similar, but
narrower tube B (25 cm. diam.) into the pulmonary artery. The
side tube of A is connected by rubber tubing with the outflow of
an irrigation bottle, to the mouth of which leads a tube from B.
Through a narrow cut in the apex of the ventricle a brass tube
C connected at its free end with a rubber bulb and having a side
tube closed by a small rubber stopper, which is pierced by water-
proofed insulated wires, is inserted and tied in. The wires are
connected with a small electric lamp D which projects into the
ventricle. By rhythmically compressing the bulb E, to simulate
the ventricular contractions, water is transferred from the auricle
to the pulmonary artery, and with each pulsation the tricuspid
and semilunar valves can be very distinctly seen to open and
close in obeyance to the pressure changes.

SECTION VII.

THE DIGESTIVE SYSTEM.
CHAPTER XXXI.

THE INNERVATION OF THE SALIVARY GLANDS.

The general nature of THE NERVE CONTROL OF SECRETORY
GLANDS is most readily studied on the submaxillary gland of the
dog or cat. The responses are not exactly alike in the two animals,
but the results typify glandular innervation in general.

Demonstration 8. — A dog is anaesthetized preferably by
. urethane (p. 72) and tracheal and carotid cannulae are inserted.
An incision is made along the inner border of the ramus of the lower
jaw extending from the mouth to the angle of the jaw. The digas-
tric muscle is exposed and is strongly retracted outwards so as to
expose the mylohyoid muscle, the fibres of which run transversely
to the wound. The hypoglossal nerve is seen extending backwards
behind the posterior edge of the mylohyoid. A probe or blunt
dissector is pushed under the latter muscle and its fibres cut in the
same line as the main wound. This exposes the ducts of the sub-
maxillary (and sublingual) glands with the lingual nerve crossing
them. A ligature ,is tied on this nerveySew-- the d-uc4s and the

t.t. -^-»t<>«^^|^^»v>A^p^

nerve cut peripheral to the hgafture^after which it is traced up under
the ramus of the jaw, the small branch which arises from it and
proceeds forwards being cut. Just above where this branch comes
off another branch passes backwards and downwards to join the
ducts a short distance from the 'place of crossing of the lingual.
This is the chorda tympani nerve. It can be prepared for artificial
stimulation by tying a second ligature round the lingual nerve
above where the chorda arises and cutting above the ligature.
By holding up the piece of lingual by the two ligatures the elec-
trodes can be readily applied to the chorda. ' ' /

J ^^ **4UC4LCZ4L 9*4O'CU'v>£u4

Before attempting to insert a cannula in Wharton's duct the
chorda should be stimulated with a feeble tetanizing current after

226

SECRETION OF SALIVA. 227

placing bull dog forceps on the two ducts well forward of the point
where the lingual crosses them. This causes the ducts to distend
and into the one which does so most markedly a small cannula is
inserted by the same manipulation as for a vein (p: 8d).

The skin wound should now be carried back and the sub-
maxillary gland carefully freed by blunt dissection from the neigh-
bouring tissues, care being taken not to injure the veins. The
exposed gland should be kept moist with warm^todke's solution.

The cannula is connected by suitable rubber tubing with a bent
glass tube so arranged that the drops of secretion may fall from its
free end. Having ascertained the rate of secretion (by counting the
drops) for a normal period of one minute the chorda is stimulated
for a few moments with a feeble interrupted current. When the
secretion has returned to normal, stimulation is repeated with a
stronger current.

The relationship between the response and the strength of the
stimuli should be carefully noted. Those near the preparation can
usually see that the gland becomes flushed and apparently swollen
by the stimulation, this effect being usually most pronounced during
the first-applied stimuli. What conclusions are warranted from
the results? The observations are repeated during stimulation
of the central end of the vago-sympathetic in the neck.

The secretion pressure is now measured by connecting the
cannula, by moderately thick-walled, but yet flexible tubing, with a
mercury manometer, the tubing being filled with physiological
saline. The carotid blood pressure is also observed, and it is
advantageous to arrange the writing styles of the two manometers
so that they write in the same perpendicular on a drum. When the
chorda is stimulated with a current which gives a maximal secretion,
the pressure in the duct manometer steadily rises until it overtops
that in the artery. Explain the significance of this result.

The manometer is now removed, and, through a cannula pre-
viously inserted in the femoral vein, 10-15 mgm. of atropine sul-
phate dissolved in physiological saline is injected. When the drug
has developed its full action on the heart (how is this tested?) the
chorda is again stimulated with a maximal current, and the effect
on the secretion and the vascularity of the gland observed. What
has been the action of the atropine? Of what significance is the
experiment?

CHAPTER XXXII.

THE CONTROL OP THE PANCREATIC SECRETION.
THE SECRETION OF BILE.

As an example of THE CONTROL OF GLANDULAR FUNCTION
THROUGH HORMONES it is most convenient to take the pancreas.
The hormone for this gland is derived from the duodenal mucosa
where it is produced by the action of the acids present in the chyme
on a constituent of the epithelial cells, and it is then carried to the
pancreas by the blood.

Demonstration 9. — An anaesthetized dog that has been starv-
ed for 24 hours is prepared for registration of the arterial blood
pressure, and a cannula is inserted in the femoral vein. The abdo-
men is opened in the linea alba and the duodenum along with the
adherent pancreas pulled out of the wound. The lower (and
larger) duct of the pancreas is then exposed, and a ligature placed
under it. The position of this duct is indicated approximately by a
small lobe of pancreas, accompanied by bloodvessels, which extends
to the duodenum at a distance of about 20-25 mm. above the
point where the head of the pancreas leaves the duodenum. It
must be remembered that the duct begins branching very close to
its insertion into the duodenum so that only a small piece of it can
be dissected free. To insert a cannula into the duct the duodenum
is opened by a longitudinal incision along its free border opposite
the duct, the opening of which is then visible in the centre of a small
papilla of somewhat paler tissue than the remainder of the mucosa.
A blunt probe should be inserted into the duct and gently guided
along it so as to ascertain the exact direction of the duct and the
position of the first branch. As the probe is withdrawn a suitable
cannula with a well-marked neck is inserted and tied in position
by the ligature previously applied outside the duct. A cannula is
now placed in the common bile duct, which is readily found accom-
panying the portal vein.

228

SECRETION OF PANCREATIC JUICE AND BILE. 229

The following observations on the secretions from the two ducts
are now made.

1. Having observed the normal rate of secretion, a small piece
of cotton soaked in a weak solution of hydrochloric acid (less than
1%) is placed in the duodenum. Whenever the secretion from the
pancreatic duct becomes decidedly increased (not an invariable
result) the cotton is removed, and the duodenum washed with
physiological saline rendered faintly alkaline by sodium carbonate.
Sometimes an increased secretion of bile occurs. From what
sources may this bile be derived? What experimental steps would
you suggest in order to ascertain this?

2. The exciting influence of the acid might of course be due
to its absorption into the blood. To test this possibility inject
some of the acid into the femoral vein. A negative result is ob-
tained even when massive doses are injected.

3. Pieces (about 2 feet long) of the upper end of the jejunum and
of the lower end of the ileum are then removed, by cutting between
previously applied ligatures, and the contents washed out by tap
water. Each piece is then slit open and the mucosa scraped off by
the blunt edge of a scalpel and collected in separate watch glasses.
One half of each scraping is thoroughly ground in a small mortar
with fine quartz sand and about 30 c.c. of 0.6 per cent, hydro-
chloric acid (2 c.c. HC1 (Con) in 100 c.c. water.) The extracts are
filtered through fine muslin and nearly neutralised (but left dis-
tinctly acid towards litmus). About 5 c.c. of the extract of jejunum
is then injected into the femoral vein and observations made on
the arterial blood pressure, the secretion of pancreatic juice and the
secretion of bile. (Prior to this, however, the cystic duct should
have been tied off. Why?) The blood pressure usually falls
considerably and care must be taken so to regulate the rate (and
amount) of injection that the fall is not allowed to go too far.
What conclusions regarding the mechanism of the increased secre-
tion can be drawn from the results? To obtain satisfactory results
it may be necessary to repeat the injection using double the amount
and injecting more quickly.

The observation should then be repeated using a similarly
prepared extract of ileum. A much feebler response, if any re-

230 EXPERIMENTAL PHYSIOLOGY.

sponse at all, is observed, but the fall in blood pressure is as pro-
nounced as before. What conclusions do you draw from the result?

4. Having demonstrated the secretagoguary action of the duo-
denal extract, the question arises as to the general nature of the excit-
ing substance. The remaining portion of the acid extract of jejunum
is therefore boiled (in faintly acid reaction) and filtered through thin
filter paper. The filtrate is found to be still active when injected.
What conclusions do you draw? Has any change occurred in the
vaso-depressor effect? The secretion of bile is often observed to
respond to the injections in the same manner as the pancreatic
juice, and its behaviour therefore should be carefully watched.

5. Having established, by these experiments, that acid extracts
a pancreatic hormone from the mucosa (seeretin) the question arises
as to whether the hormone is present therein as such or as a pre-
cursor which the acid activates. To throw light on this question,
the unextracted half of the mucosa scrapings is ground in a mortar
with quartz sand, and 0.9 per cent, sodium chloride solution, and
after filtering the extract through muslin, about half of it is in-
jected intravenously. The result is negative. The remaining
portion of the extract is then rendered faintly acid with HC1,
boiled and filtered. (3n injection a slight, but yet definite increase
usually occurs in pancreatic secretion. What conclusions are per-
missible from these results?

6. Finally the influence of the intravenous injection of a solution
of bile salts on the outflow from the bile duct should be observed.

CHAPTER XXXIII.

EXPERIMENT ON THE NORMAL SECRETION OF
SALIVA AND GASTRIC JUICE.

In order to study the secretion of the digestive glands in
an unaesthetized normal animal it is necessary by surgical methods
to establish permanent openings or fistulse on the surface of the
body, through which the secretions may escape. This method has
been employed with great profit in the case of several of the glands,
the general nature of the experiments being adequately illustrated
by observation on the parotid gland (representing a gland with a
definite duct) and the stomach (representing a gland which secretes
directly on to a mucous surface). The operations necessary to
establish the fistula are performed by some competent assistant,
and the operated animals are carefully tended so that the wounds
heal without suppuration. For successful observations it is
furthermore of importance that the animal should become used to
the person who is to demonstrate the experiments.

The Normal Secretion of Saliva.

METHOD FOR MAKING FISTULA OF THE PAROTID GLAND. — A dog is anaesthetised
with morphine (morphine hydrochloride 0.01 gm. perkgm. body weight) and finally
with chloroform. The mouth is held open by means of a suitable gag and the
ductus Stenonianus located (on the mucosa of the cheek opposite the second
molar tooth). A small blunt probe is pushed into the duct and the mucous mem-
brane around it sponged with sterile surgical gauze. A circular incision is then
made through the mucosa around the duct, the area of the circle being a little less
than one sq. cm. It will probably be necessary at this stage to suspend further
operating for a minute or so in order to administer some more chloroform, and
throughout the remainder of the operation similar pauses will occasionally be
necessary. The circle of mucosa is then quickly dissected from the underlying
tissue up to the duct, after which a stab is made by a fine scalpel through the skin
of the cheek opposite the opening of the duct, the edge of the skin incision being
trimmed by a scissors so as to make the wound elliptical in shape. By means of
a surgical needle a fine silk ligature is passed through the anterior edge of the
circle of mucosa and its free ends pulled out through the skin wound. By gentle

231

232

EXPERIMENTAL PHYSIOLOGY.

traction on the ligature and by bending the free end of the probe out through the
wound the duct and encircling mucosa is brought out to the skin, care being taken
not to stretch or twist the duct. The edges of the mucous circle are now stitched
by a fine silk or chromacised catgut ligature to the edges of the skin wound. The
wound in the mouth is similarly stitched and without the application of any
dressing the animal is allowed to come out of the anaesthetic. The wound should
be bathed daily with physiological saline and a free secretion of saliva occasionally
stimulated by giving the animal some dry food (bread crumbs).

When the wound has healed, it will be necessary to accustom the dog to being
strapped into a suitable holder and it is most important for the success of the

FIG. 67. Funnel applied to skin of animal for collection of fistula juices. (Pavlov).

observations that the animal should be trained to submit to the harnessing
without fear or excitement. The knowledge that feeding is to follow soon makes
the animals eager to participate in the proceedings. To collect the secretion
Pavlov uses a funnel, made out of wax and resin* with its edges flared out so as to
form a flat surface to apply to the skin (Fig. 67). To apply the funnel the hair
around the duct is clipped close by a scissors and thoroughly dried. The funnel
is then moderately warmed and the flared edge firmly pressed on the cheek to
which it adheres, after which the narrow end is attached to a light graduated test-
tube. In removing the funnel care must be taken not to abrade the skin. Holding
a warmed metal rod near the edges helps to loosen the attachment to the skin
*50 parts, rosin ; 40 parts, ferric oxide Fe2Os and 25 parts yellow wax.

SALIVARY AND GASTRIC FISTULA.

233

Demonstration 10. — The

during the following conditions:

1 . When various foods are
fed to the animal.

2. When acid solution or
dry bread crumbs are thrown
into the mouth.

3. When the animal is
teased with a bone (which
should afterwards be given to
him).

It is also profitable to
train the animal so that he
learns to associate a certain
sound or visual impression
with the subsequent stimula-
tion of salivary secretion, as
by giving him bread crumbs.
The experiments on con-
ditioned reflexes are however
more striking when the fistula
is one involving the sub-
maxillary or sublingual ducts.
The exact nature of the
demonstrations and experi-
ments on material of this
type must naturally vary with
circumstances. Advanced
students may profitably de-
vote considerable time to this
work.

The Normal Secretion
of Gastric Juice. — Satis-
factory demonstration of the
secretory activities of the
gastric glands requires the
establishment of both gastric
and cesophageal fistulae on the same animal.

secretion should be observed

Stomach cannula. See context for

This can be under-

234

EXPERIMENTAL PHYSIOLOGY.

taken only when there is someone who can devote considerable
time every day to the proper care and attention of the animal. A
few observations may, however, be made on an animal with a
gastric fistula alone.

Method for Making the Gastric Fistula.

Through an incision 6-7 c.m. long in the linea alba just below the sternum the
stomach wall is caught by a forceps and after pulling it somewhat over to the

FIG. 69. Stand for holding animal on which fistula operation has been per-
formed. (From Tigerstedt's " Practical Physiology".)

right, the serous coat is attached to the edge of the skin wound by a few discon-
tinuous sutures. An incision is then made through all the coats of the stomach
wall, bleeding being carefully controlled by ligatures. The incision is just large
enough to admit the notched flange of the gastric cannula (Fig. 68). A cromi-
cised catgut ligature is now stitched like a purse string a short distance from the
edges of the incision and the cannula inserted and tied (not too tightly) in place
by the ligature. It is well to apply a second purse string ligature. The operation
i s completed by closing the skin wound up to the cannula with discontinuous sutures.

SALIVARY AND GASTNIC FISTULA. 235

The wound is then dressed with cotton and collodion and the outer shield of the
cannula screwed on, but not so far as to press on the wound. The tube of the
cannula is closed by the screw-in stopper.

Demonstration 11.— In about a week, when the wound will
usually have healed, the dog is placed on the observation holder (Fig.
69) and the following observations are made. A pledget of absorbent
cotton attached to a hsemostat is passed into the stomach and
gently moved over the mucosa; the nature and reaction of the
secretion which adheres to it after removal is observed. A thin-
walled rubber bag attached to a glass tube is carefully passed into
the stomach through the fistula and filled with warm water. The
tube, leading out of the fistula, is connected with a water mano-
meter, the free limb of which is attached by tubing to a sensitive
tambour arranged to inscribe its movements on a drum. Observa-
tions are now made on the hunger contractions. (It may be
necessary to wait some time before these appear). When satis-
factory records have been secured the animal is teased by tempting
it with appetising food and the effect on the contractions noted.

The rubber bag is now removed and the reaction of the secretion
adhering to it tested by means of litmus paper.

These observations on hunger contractions in a dog should be
supplemented by similar ones on man. For this purpose a thin
rubber bag is firmly tied on a narrow stomach tube and passed
down the oesophagus until the bag lies in the stomach. The bag is
then distended by air (cf . Carlson) and the outer end connected with
a water manometer and tambour. The observation should be made
on a person who has not taken food for some hours previously.

CHAPTER XXXIV.

THE MOVEMENTS OP THE OESOPHAGUS* AND
INTESTINE.

Demonstration 12. — A rabbit is narcotised by means of
urethane (p. 79) t and the oesophagus is exposed. This is accom-
plished partly by drawing the trachea to the right by a stout silk
ligature passed around it and partly by pulling the oesophagus
to the left. The vagus nerve is then followed up to where the
superior laryngeal leaves it, and this branch is cut after tying a
thread round it as near to the vagus as possible. Stimulation of the
laryngeal nerve by a Faradic current excites the act of swallowing
and it can be seen that this consists, in order, of a movement of the
floor of the mouth, of elevation of the larynx and of the passage of a
peristaltic wave along the cesophagus. A record of the peristaltic
wave can be obtained by inserting into the lumen of the cesophagus,
through a small incision, a thin-walled rubber bag (finger cot), which
is connected by tubing with a water manometer and tambour, after
distending it with warm water. J

It is now important to ascertain whether the peristaltic wave
can be more readily set up by mechanical irritation of the ceso-
phagus or of the pharynx. The stimulation can be produced by
stroking with a feather. The cesophagus is finally cut across and a
bent pin connected by a thread with a muscle lever attached to the
peripheral end of the cut tube. The superior laryngeal nerve is
again stimulated. From the results of this observation what con-
clusions can be drawn concerning the manner of transmission of the
oesophageal peristaltic wave?

*The action of the base of the tongue, etc., in swallowing may be readily
studied in the decerebrate cat, as described on p. 249.

fFor the success of the observation the anaesthesia must not be too deep.

Jit is best to tie the finger cot to a rubber catheter so that the end of the cath-
eter reaches to the end of the cot. This facilitates insertion of the cot into the
cesophagus. It is important to see that the cot becomes uniformly distended with
water before connecting with the water manometer. The edges of the incision
in the cesophagus should be stitched together so as to hold the instrument in place.

236

MOVEMENTS OF OESOPHAGUS AND INTESTINE.
THE MOVEMENTS OF THE INTESTINE.

237

Demonstration 13. — The same animal may now be used for
observations on the intestinal movements. For this purpose a
tracheal cannula is inserted and artificial respiration established.*
The thorax is then opened and the vagus and splanchnic nerves
quickly cut, this procedure being necessary in order to secure pro-

FIG. 70. Sherrington Electrodes. The nerve is pulled through the tube by means of a
thread attached to it, and is arranged so that it lies between the platinum electrodes inserted
through the side tube. The nerve musto n no i ccount be stretched when being pulled into the
electrode

nounced intestinal movements. One of the splanchnic nerves is
connected with Sherrington's electrodes (Fig. 70) the wires of which
emerge from the wound. The thoracic wound is closed by sutures,
and the abdomen opened along the linea alba. The animal is then

*It is advisable also to place a cannula filled with saline in the central end
of the jugular vein. Through this cannula drugs can subsequently be injected.

238 EXPERIMENTAL PHYSIOLOGY.

placed in a bath of physiological saline at about 33-35° C, contained
in a tank of suitable size and the intestines allowed to float out in
the saline. The pendular movements are very conspicuous and
the loops should be closely examined with the object of ascertaining
which of the muscular coats are contracting. If no peristaltic
waves are observed, they may be set up by pinching the intestine.
Observe closely the characteristics of these waves.

The effect produced on the movements by stimulation of the
peripheral end of the splanchnic nerve is now studied.

Records of the movements may be secured by the balloon
method already described for the oesophagus.

Finally, if the animal is still in suitable condition, the effect
produced on the intestinal movements by injections into the jugular
vein of epinephrin (1-10,000 adrenalin chloride) and of atropine
are observed.

SECTION VIII.
THE CENTRAL NERVOUS SYSTEM.

CHAPTER XXXV.

REFLEX ACTION IN THE MAMMALIA.

Although certain basic facts concerning the physiology of reflex
action can be learned by observations on the spinal frog (p. 90) the
variety and complexity of the reflex movements exhibited by the
preparation are too limited to enable us to understand much about
reflex action in the higher animals. Practically the only reflex
movement elicitable in the spinal frog is the flexion reflex, which is
of a type quite different from that of the reflexes that are concerned
in maintaining such animals as the dog or man in the erect posture
(postural reflexes), or in enabling movement to occur from place to
place. The functional unit of the nervous system is the reflex arc
and the ability to perform complex co-ordinated movements de-
pends on the integration of the various reflex arcs among one another
—the integrative action of the nervous system (Sherrington). In
order that we may study the principles that govern this integration
it is necessary to simplify the conditions from those obtaining in
the intact animal, which is done by breaking the connection between
the higher brain centres and those of the lower portion of the spinal
cord (the spinal animal).

Demonstration 14. — The most satisfactory preparation to
use for a study of the spinal reflexes is one in which the spinal cord
has been cut some weeks previous to the observation. Spinal shock
will have been recovered from and the following reflex movements
can readily be demonstrated :*

*These can readily be demonstrated with the animal lying on his side, but if
graphic records are desired, as for measuring latent periods, etc., it is best to sus-
pend the animal in a suitable stand with the posterior extremities hanging free.
Threads attached near the paws are carried over directing pulleys (or glass rods)

239

240 EXPERIMENTAL PHYSIOLOGY.

1. THE FLEXION REFLEX — 'by pricking the skin of the paw
with a pin or applying a moderately strong electric shock. The
flexion at knee and hip is accompanied by extension of the leg of the
opposite side, THE CROSSED EXTENSION REFLEX. By taking simul-
taneous tracings of the two legs after attaching the feet to suitable
levers by threads, the correspondence in time between the two
reflexes is demonstrated.

2. THE KNEE JERK — by giving a sharp blow with a ruler or
the handle of a scalpel to the ligamentum patellae. Note that the
leg swings limply back to its flexed position after the tap (compare
with the behaviour of the jerk in a decerebrate animal (p. 103 ).

3. THE SCRATCH REFLEX — by moving the finger or a pencil
backwards and forwards on the skin of the body. The skin area
from which the reflex can be elicited is very extensive and the paw
usually is directed approximately to the place of stimulation
('local sign'). Sometimes this property of 'local sign', however, is
very imperfect. An electrical stimulus (through the stigmatic
electrode) will also elicit the reflex, but much less satisfactorily than
the mechanical one.

4. THE EXTENSOR THRUST — by pushing the blunt end of a
pencil between the pads of the paws. The corresponding leg makes
a sudden extension movement.

The following properties of reflex action may now be studied
using one or other of the foregoing reflexes.

1. THE LATENT PERIOD OR UNCORRECTED REFLEX TIME. To
determine this it is necessary to employ electrical stimulation and to
insert a signal magnet in the primary circuit. A fast rate of drum

to be attached to the long arm of a right angled lever, from the short arm of which
a second thread runs to a straight muscle lever. The threads are adjusted so as
suitably to diminish the amplitude of movement at the writing points.

A large electrode made out of a strip of copper covered with a pad of surgica
gauze, which is moistened with salt solution, is tied on to the middle of the back
(indifferent electrode) and a much smaller electrode of the same type is tied to the
foot of one side (stimulating electrode). A third (stigmatic) electrode composed
of a stout copper wire covered at its tip by a piece of surgical gauze is also required
The wires leading from the electrodes are connected with the secondary coil of an
inductorium, the indifferent electrode with one pole and the stimulating and
stigmatic electrodes with the other. (A well insulated simple key should be
inserted in the wire leading from the attached foot electrode).

REFLEX MOVEMENTS IN MAMMALS. ' 241

must be used, the exact relationship of the signal pointer to the
recording lever being indicated by alignment marks (see p. 94).
A time tracing, preferably of 1/100 sec. must also be taken. Several
observations on the flexion reflex with stimuli of varying strength
will show the latent period to vary from about 0.03 to 0.12 sec.
The latent time of the jerk is much shorter. (The signal for the
application of the stimulus is afforded in this case by the slight
movement of the lever produced by the blow to the tendon, the
attachment of the threads to the levers being adjusted so as to make
them sensitive enough to record this).

The latent period of the scratch reflex (using the stigmatic
electrode) is relatively very long and it varies with the strength of
the stimulus. Taking these results with those already attained
on the palpebral reflex (p. 104) into consideration, what con-
clusions do you draw concerning the latency of reflex action in
general? What corrections must be made to determine the true
or reduced reflex time?

2. GRADING OF INTENSITY. — This is investigated for the flexion
and scratch reflexes by varying the strength of the electrical stimu-
lus and for the extensor thrust by applying varying degrees of
pressure to the paws. How do the results obtained from these two
types of reflex compare with those already obtained for the palpe-
bral reflex?

3. SUMMATION. Using the flexion reflex, a strength of current
is found which is just ineffectual when single shocks are applied,
a simple tapping key being inserted in the primary circuit of the
inductorium in place of the vibrating hammer. The shocks are then
applied frequently. Is the summation more pronounced than you
found it to be for isolated nerve ?

4. AFTER EFFECT. — By recording the movement of the flexion
reflex on a fairly rapid drum, the degree of after effect in relation-
ship to the strength of stimulus is observed. Both single shocks
and tetanizing shocks are employed. hi what respects do the
results differ from those obtained on a nerve?

5. REFLEX FATIGUE. — The effect of maintained stimulation is
studied in the flexion and scratch reflexes, using a slow drum. How
does reflex fatigue differ from fatigue in an isolated nerve-muscle
preparation? After the leg has been thoroughly fatigued for the

242 EXPERIMENTAL PHYSIOLOGY.

flexion reflex, the scratch reflex is elicitated by mechanical and
electrical stimuli. What do the results indicate as to the locus of
fatigue in the reflex arc?

6. IMMEDIATE INDUCTION. — The end of the handle of a scalpel
or of a small ruler is pressed against the skin of a scratch area, but
there is no response. If the same object be moved on the skin,
however, the scratch movement is likely to be set up. What is the
explanation of the result? What is the analagous experiment on
vision?

7. SUCCESSIVE INDUCTION. — Using a slow drum, record a con-
siderable number of knee jerks produced by regularly applied taps
of as nearly as possible equal intensity to the patellar tendon, and
when there is approximate equality in the heights of the contrac-
tions throw the corresponding leg for a few seconds into the flexion
reflex, meanwhile continuing the patellar taps. When the flexion
reflex subsides the tendon jerks are greatly exaggerated. Explain
why this shows successive induction. Demonstrate the same
phenomenon for the flexion reflex and extensor thrust, and for the
crossed extension reflex and flexion reflex. What is the analagous
experiment on vision?

CHAPTER XXXVI.

CEREBRAL LOCALIZATION, DECEREBRATE RIGIDITY,

RECIPROCAL INNERVATION, FUNCTIONS OF

SPINAL ROOTS IN THE MAMMAL (DOG.)

Demonstration 15.

A tracheal cannula is inserted in an etherized dog, and ligatures placed loosely
around the carotid arteries on both sides. With the animal in the prone position
the head is raised by placing a pad under it, being careful to see that there is no
kinking of the tube leading from the tracheal cannula to the ether bottle. To
serve as a landmark for the crucial sulcus of the cerebrum (which corresponds to
the Rolandic fissure on the human brain) two threads are stretched across the
head, one of them joining the outer canthi of the eyes and the other, the condyles
of the lower jaw. The crucial sulcus lies a little behind the mid-point between
the two threads. An incision is made along the mid-line of the scalp and by blunt
dissection the temporal muscle is separated from the bone far enough to make
room for two trephine holes one in front of the other and with their inner margins
at least 3 mm. from the mid line so as to avoid wounding the superior longitudinal
sinus. To manipulate the trephine, the steel point is first of all adjusted so that
it projects beyond the sawing edge, and this point is bored into the skull in order
to afford good fixation. When the sawing edge reaches the bone, care is taken it
cuts uniformly, and when a good start has been made the central point is pulled
up so that this may not puncture the brain The trephining is continued until
the inner table of the skull is cut through around most of the circle, the trephine
is then removed, and the disc of bone pried up by means of a pereosteal elevator
or a stout pair of forceps. There is apt to be considerable haemorrhage at this
stage, but it can usually be controlled by applying for a few minutes a pad of
gauze thoroughly wrung out with hot isotonic saline. When the two trephine
holes have been made they are connected together by bone forceps, taking care
not to wound the brain and controlling bleeding with hot gauze. A curved sur-
gical needle is passed through the dura and by means of it the latter is raised
sufficiently so as to be able to cut it with a sharp pointed scissors. The exposed
brain should be covered with gauze soaked in isotonic saline.

In order to stimulate the cortex a large plate (indifferent)
electrode is placed on the moistened skin of the lower dorsal region
and connected with one terminal of the secondary coil of an in-
ductorium with the other terminal of which a blunt-pointed
(stigmatic or diagnostic) electrode is connected.

Having made a rough sketch indicating the position of sulci and

243

244

EXPERIMENTAL PHYSIOLOGY.

convolutions of the exposed portion of the cerebrum, the front and
hind legs on the side opposite to the opening in the skull are loosen-
ed and the tetanizing current applied for short periods and at vary-
ing strengths until definite movements are observed to occur. For
successful results it is necessary to have the animal as lightly
anaesthetized as is consistent with the entire absence of pain.

N .

n. -..

FIG. 71. Surface of cerebrum of dog, showing on the left side approximate
positions of the various centres: n, neck; fl., forelimb; hi, hindlimb; f, face. Move-
ments of the eyes, E, accompanied by dilation of the pupil, P, are also obtained from
the portions of the cortex indicated on the right side. (After Stewart) .

When a suitable strength of stimulation has been found, the exposed
area of cerebrum is' systematically explored (Fig. 71), and the ob-
served responses noted on the sketch. The character of the move-
ments must also be carefully noted (i.e., whether there is evidence
of reciprocal innervation, etc.). The pupils and eye movements on
both sides, movements of the head and ears, changes in the respira-
tions and movements of the tail must also be looked for. The

CEREBRAL LOCALIZATION. 245

general results are indicated in the accompanying chart. Finally
the effect of the application for some time of a very strong stimulus
is studied.

The animal is finally decerebrated. For this purpose a large
aneurysm needle is passed through the posterior edge of the trephine
hole backwards until the tentorium cerebelli is felt. It is then
directed downwards and inwards so as to break across the base
of the brain stem just in front of the tentorium. Immediately this
is done the respirations are likely to cease or to become very irregular
so that artificial respiration must be started. The ether can now
be discontinued since the animal is incapable of feeling any pain.
The muscles of the extremities are carefully observed from time
to time for the onset of decerebrate rigidity and for the appearance
of reflex movements, such as the knee jerk and the flexion reflex.

For the purpose of studying recriprocal innervation* the
tendons of the rectus femoris and the semitendinosus are exposed
and isolated from adherent aponeurosis on both sides and the
tendons are cut after threads have been tied to them. The peroneal
nerves are also exposed as they lie under the skin on the inner side
of the leg or the dorsum of the foot. After attaching threads and
cutting, each nerve is placed in a pair of Sherrington's electrodes
attached to an inductorium. The threads on the tendons are con-
nected with muscle levers, by means of pulleys or angle levers,
and the writing points are arranged so that they write in
the same perpendicular on the drum, signal magnets being
inserted in the primary circuits of the two inductoria. When the
peroneal nerve on one side is stimulated it will be found that the
homolateral semitendinosus contracts,' and that the rectus femoris
simultaneously relaxes, if it be in a hypertonic condition (as a result
of decerebrate rigidity).*

If the preparation is still in suitable condition, the experiment
should be terminated by exposing the lumbar portion of the spinal
cord and studying THE FUNCTIONS OF THE SPINAL ROOTS, noting also
the effect produced on the rigidity by their section. To expose the
roots, place the animal on its belly with a thick pad or block of wood
under the lower portion of the abdomen, and make an incision in
the mid line of the back over the spines of the lumbar vertebrae.
Separate the muscles from both sides of the spinous processes and

*This experiment succeeds best on the decerebrate cat. If time is limited
the experiment should be omitted and that on the spinal roots performed.

246 EXPERIMENTAL PHYSIOLOGY.

retract strongly so that the laminae of the vertebrae are exposed.
Much of this can be done by blunt dissection, which will avoid
haemorrhage. Cut the tissues between the spinous processes of the
2nd and 3rd lumbar and also between the last lumbar and 1st
sacral and amputate at their bases, the spinous processes between
the two cuts, using a strong bone forceps. Remove the spinous
processes and after bleeding has been controlled by the application
of cloths wrung out with hot water proceed to open the spinal canal
by cutting through the laminae of the exposed vertebrae with bone
forceps, taking care that the point of this instrument does not go
deeply into the spinal canal. The spinal cord enclosed in the dura
is now exposed, but it is necessary in order to expose the roots
properly that the articular processes between neighbouring verte-
brae be picked away.* (The so-called hawk's bill bone forceps are
useful for this process).

The posterior root ganglia came into view when the articular
processes have been removed. After again stopping haemorrhage,
which is likely to be considerable at this stage, the roots are pre-
pared for stimulation.

Lift the posterior root of the 6th nerve carefully on a strabismus
hook or small aneurysm needle, and tie a ligature round it as near
to the ganglion as possible, cutting the root distal to the ligature.
Loosen the leg on the corresponding side and stimulate the central
end of the root with a tetanising current, using ordinary electrodes.
Note the character of the movements of the leg and watch for any
changes in the respirations.

Ligate and cut the 7th posterior root as near the cord as possi-
ble and stimulate the peripheral end.

What conclusion do you draw from the results of these experi-
ments?

The functions of the anterior roots are then determined by a
repetition of the same procedure as for the posterior. It is some-
what difficult to prepare these roots for stimulation, and it often
assists to pass a tape round the cord, by which it is cautiously pulled
up and to one side. Observe carefully any difference in the type
of movement which results by stimulating the anterior and posterior
roots.

Draw a diagram showing the functions of the roots.

*It will be observed that the lower (6th and 7th) lumbar roots are larger than
those higher up, the level of the 7th being about on a line with the iliac crests.

CHAPTER XXXVII.

THE DECEREBRATE CAT I.

Demonstration 16.

A cannula is inserted in the trachea in a deeply etherized cat (preferably a young
animal) and a pair of serres fines is placed on the carotid arteries on both sides of
the neck. The animal is then placed on the ' decerebrator ' with the
pelvis resting on the back platform (P), the hind legs hanging free, and the
neck on the neck block (N), care being taken to prevent any kinking of the

FIG. 72. Decerebrator. For description, see context.

tracheal cannula. The nasal septum is punctured by a scaplel and a cord is pass-
ed through to a hole on the neck block, at the lower level of the tongue guard (T).
The mouth is opened and by gentle traction on the cord and manipulation of the
head the snout is pulled over the tongue guard which lies over the tongue with its
point between the fauces and over the epiglottis. The cord is then tied to a hook
on the base of the stand and the nose piece (S) is pressed up against the snout
and fixed in position by tightening the clamp.

An incision is now made along the mid line of the skull and the skin reflected
to both sides. A distance of 30 mm. is measured back from the coronal suture

247

248 EXPERIMENTAL PHYSIOLOGY.

(which is particularly evident in young animals) and marked by a cut with a
scalpel. The preparation is now ready for decerebration. This is best done by
using a planing blade (12.5 cm. wide, 9 cm. high and 4 cm. thick) bevelled on one
edge and mounted on a wooden handle, but a good ax of about the same dimen-
sions is quite satisfactory. To perform the decerebration, the operator holding
the blade in the left hand, places the edge on the notched mark, and holds the
plane of the blade at right angles to the plane of the head. By means of a heavy
mallet (or hammer, if an ax is used) held in the right hand he gives the blade a
light blow so as to make the edge engage the skull, the blade being held accurately
transverse to the long axis of the skull and directed so that when the head is cut
through the edge will strike a mark made on the metal plate of the base of the
tongue guard. The operation is then completed by applying a couple of strong
blows and the decerebrated animal is immediately removed from the stand and
placed on its back on the table, the neck being grasped with the finger and thumb
just below the transverse process of the atlas so as to compress the vertebral
arteries. The head is kept elevated to diminish haemorrhage, and etherization is
discontinued. In many animals the respirations cease for a few moments, and
artificial respiration may be necessary. They usually return spontaneously, but
if they do not do so, the vertebral arteries should be momentarily released so as to
allow some blood to flow to the respiratory centre. The bleeding from the cut
across the head is not usually serious if the vertebrals are properly compressed
and when time has been allowed for clotting to occur these may be cautiously
released. There is no particular advantage in trying to accelerate the clotting
by applying absorbent cotton wool to the wound. It usually takes from 15 to 20
minutes before the vertebrals can be entirely released. Finally, the clamps are
removed from the carotid arteries, one at a time — any bleeding point in the cut
muscles being ligated — and the head is tied to an upright so as to keep it elevated.

The resulting preparation is suitable for many experimental
purposes (consult Sherrington's Mammalian Physiology, The
Clarendon Press, Oxford.)

The blood pressure, although it may rise considerably while
the vertebrals are being compressed (because of partial asphyxia)
soon returns to about the normal level, and the respiratory centre
responds to changes in CH of the blood much more promptly than
in anaesthetised animals. Why should this be the case? It is
particularly in the study of reflex actions, however, that the pre-
paration is of value, although certain of them are masked by the
decerebrate rigidity which develops. Since the section if properly
made goes through the mesencephalon just behind the anterior
corpora quadrigemina, reflexes involving the head end of the animal
can be demonstrated. The following are to be elicited and studied :
1. THE PINNA REFLEX. — Pinch or slightly twist the tip of the

pinna between the finger and thumb. The response consists of

THE DECEREBRATE PREPARATION. 249

a quick retraction of the pinna accompanied often by a folding
back of its free end.

2. THE ACOUSTIC REFLEX. — Pricking up of the ears when a sharp
sound is made as by clapping the hands. This reflex occurs
only when the section is forward of the anterior corpora quad-
rigemina.

3. THE DEGLUTITION REFLEX. — Swallowing movements when
water is dropped on the base of the tongue. Note the behaviour
of the respiratory movements during the swallows. Repeat the
observation by dropping the water on various parts of the tongue.
Observe the effect on swallowing produced by pressing a moist-
ened camel's hair brush on the pharynx. In order to do this,
it is necessary to slit the soft palate in the mid line. The move-
ments of the vocal cords can readily be observed by using a
laryngeal mirror.

4. THE HEAD-SHAKE REFLEX. — Rapid shaking movements in
rotary direction when air is blown into the external auditory
meatus. The reflex may also be elicited by squirting cold water
from a syringe into the ear.

5. THE FLEXION REFLEX. — By painful stimulation of the paw —
flexion at knee and hip occurs.

6. THE KNEE JERK. — Quick extension (kick) at knee produced by
tapping the patellar tendon. Note particularly that the return
of the leg after the contraction is not complete, and that if the
tendon be tapped at short intervals the knee becomes more or
less permanently extended. This result depends on a contrac-
tion remainder, which is a feature of the 'rigid' postural muscles
in decerebrate preparations. In the spinal animal there is no
postural hypertonus so that, the leg passively falls back to its
previous position after the jerk.

The scratch reflex is not present as a rule.

The most striking reflex condition produced by the decere-
bration is the rigidity, which it can readily be seen affects parti-
cularly the extensor (postural) muscles. The rigidity is not of the
same nature as the tetanic contraction produced by continuous
stimulation of the motor neurone, for the muscle yields to a slowly
applied pull and does not spring back to its old position when the
extending force is removed. This is tested by bending the knee or

250 EXPERIMENTAL PHYSIOLOGY.

elbow joints. The condition has therefore been styled ' REFLEX
POSTURAL TONUS ' and it can be shown that the weight required to
counterbalance the rigid muscle is practically the same whether
muscle is lengthened (knee in extension) or shortened (knee
in flexion).*

If the preparation is still in suitable condition, the experiment
should be terminated by exposing the lumbar portion of the spinal
cord and cutting the sensory roots of the nerves of the hind limbs
What effect has this procedure on the decerebrate rigidity? What
conclusions do you draw from the results?

*The flexor muscles acting on the joint should be paralysed by cutting their
nerves in testing these points.

CHAPTER XXXVIII.
THE DECEREBRATE CAT II.

THE DECEREBRATE PIGEON.

There are certain fundamental reflex reactions which can be
studied only after considerable operative preparation of the animal.
One of these is RECIPROCAL INHIBITION which the following experi-
ment devised by Sherrington clearly demonstrates. In it reflex
inhibition is demonstrated of the plastic tonus of the extensor
muscles of the knee in decerebrate rigidity and of the same muscles
while actively contracted as a result of reflex stimulation.

Demonstration 17.

Whenever bleeding has stopped from the neck stump of a decerebrate pre-
paration (cat) this is placed on its back on a well-heated operating table and the
hind limbs tied in an extended position. With the left leg well abducted, an
incision is made about 5 cm. long down the thigh towards the outer border, and
then curved across the thigh towards the inner border. The bluntly V-shaped
skin flap is reflected along with the subcutaneous fat towards the median line,
so as to expose the femoral artery and vein. About 8 cm. outside the artery
will be seen the psoas muscle with the femoral nerve emerging from it. There are
three branches of the nerve. Of these the outermost (a small branch) runs to the
sartorious muscle and is cut, the middle (a large branch) runs to the quadriceps
extensor and is left intact and the innermost (a small branch) is the saphenous
nerve, which also is cut. Finally, the psoas muscle is cautiously cut across piece
by piece, using small scissors for the purpose. By these operations the flexor
muscles of the hip joint are rendered incapable of acting on this joint. The
operations are repeated on the right leg.

The sciatic nerves on both sides must now be exposed, cut low
down and the central ends placed in Sherrington's electrodes, the
branches of them that run to the hamstring muscles being severed.
By this latter operation the knee joint is rendered incapable of
active flexion. To accomplish these objects an incision is made
down the mid line of the back of the thigh, starting above from a
point midway between the tuber ischii and the great trochanter and
continuing down to the outer condyle of the femur. From the

251

252 EXPERIMENTAL PHYSIOLOGY.

upper and lower ends of this incision, short incisions are made across
the limb and the skin flap is reflected forward. At the upper end
of the exposed wound the lower edge of the relatively small gluteus
maximus muscle is made out (a small vein running along its lower
border) and is cut at its lower end and reflected upward. It is now
possible by retracting the muscles between which it lies to separate
the sciatic nerve and to make out a large branch passing backwards
from it. This is the hamstring nerve and it is cut. The sciatic
nerve is next followed downwards to where the two divisions of the
trunk (peroneal and tibial) diverge. A ligature is tied around the
nerve at this point, the nerves cut distal to the ligature and their
central ends placed in Sherrington's electrodes. It will be noted
that the above nerve sections render practically all muscles in the
thigh incapable of reflex contraction except the extensor muscles.

It is now necessary to arrange for graphic records of movements
at the knee joint. For this purpose the leg is amputated near the
ankle joint. After tying a stout mass ligature (threaded by a pack
needle through the skin and muscles around the stump so as to hold
it in place) a thread is also attached to the skin of the stump, and con-
nected with a suitable muscle lever, using pulleys if necessary.
The femur must also be immobilized at its lower end so that the
knee joint may be held in a flexed position and the slightest move-
ment at this joint recorded without disturbance by any other move-
ments of the preparation. For this purpose it is necessary to
insert a threaded steel pin into the condyles of the femur on their
median aspect. To insert the drill pin, an ordinary drill, procurable
at any hardware shop, is used. The drill head is unscrewed from
the pin after this has been screwed into the bones and a brass rod,
threaded at one end so that it fits the thread of the screw pin, is
connected to the pin with its outer end held by a clamp to an up-
right stand.

These preliminary operations being completed, the preparation
is placed on its right side, and the femur of the left thigh arranged
so that it is supported by the drill pin and brass rod in a nearly
vertical position, with the hips flexed almost to a right angle.
In cases in which the decerebrate rigidity is marked, the left knee
will be nearly extended. If it is not so, it may be placed in this
position by passive extension (the shortening reaction — what does
it depend on?)

THE DECEREBRATE PREPARATION. 253

With everything ready to take a graphic record of the move-
ments of the left leg, the Sherrington electrodes are connected on
both sides, each with a separate inductorium, and the recording
drum having been started the left sciatic is stimulated momentarily
with a tetanising current of moderate strength. The leg drops into
flexion because of reflex inhibition of the postural tonus. Does the
leg go back to its extended position when the stimulus is removed?

A similar stimulus is applied to the right sciatic, when a reflex
contraction of the left extensor muscle will occur (crossed extension
reflex (see p. 240).

Finally it can be demonstrated that this reflex contraction is
inhibited through stimulation of the contralateral sciatic nerve.
To do this the right sciatic is stimulated, and whenever the tracing
shows that the left extensor muscles are decidedly contracted, the
left sciatic is simultaneously stimulated for a brief period. If the
proper strength of stimulus is employed, it will be observed that
the extensors relax. These observations should be repeated with
stimuli of varying strengths and duration. When the stimulation
of the left sciatic is discontinued while still maintaining that of the
right the muscle goes back to the contracted state. Draw a dia-
gram to show the probable arrangement of the reflex pathways in
the spinal cord.

DECEREBRATE PIGEON.

Demonstration 18.— While under the influence of ether the
skull is exposed by a transverse incision of the skin. A piece of
bone just large enough to expose the cerebrum is quickly removed
by sharp scissors. By means of a glass tube connected with a
suction pump*, the cerebrum is sucked up. Great care must be
taken to avoid injury to the cerebellum, as there is no tentorium
in the pigeon. The advantage of this method of decerebration is
that the blood is sucked away, thus allowing a clear view of the
field of operation. The cavity is plugged with absorbent cotton,
and the skin sewn up. The actual decerebration should take but a
few seconds.

*The tube should be slightly drawn out with an opening 2 or 3 mm. in diameter.
This tube is connected by rubber tubing to a filter flask so that the brain tissue
will not be drawn into the pump.

254 EXPERIMENTAL PHYSIOLOGY.

The operation should be performed at least two or three hours
before the observations are to be made in order to permit complete
recovery from the ether.

A decerebrate pigeon has no memory, therefore no sense of fear.
Tt is not conscious of painful stimulation. It sleeps unless stimu-
lated. It starves unless food is placed back in the throat so that
the swallowing reflex is started.

Stimulate the pigeon in the following ways :

1. Push it in order to get it to move.

2. Produce a loud sharp sound near one ear.

3. Allow it to smell strong ammonia.

4. Stand it on a sheet of metal and gradually heat the latter
until too warm for your own hand.

5. If the bird is not too weak, toss it into the air and it should
fly.

Make notes of the behaviour of the animal in response to these
stimuli and explain the results.

CHAPTER XXXIX.

THE SPINAL CAT (SHERRINGTON'S PREPARATION.)

In this procedure the spinal axis is cut about 4 m.m. behind the
point of the calamus scriptorius, and the spinal animal exhibits a
considerable number of complex reflex movements, although the
arterial blood pressure remains low. Spontaneous respirations, of
course, cease entirely so that it is necessary to apply artificial
respiration. The control of body temperature also disappears,
necessitating artificial warmth.

Demonstration 19. — Anaesthetise a cat deeply, insert a
tracheal cannula low down and ligate the carotid arteries on both
sides of the neck. Place the animal in the prone position and
holding the head in the left hand, make a wide transverse incision
through the skin over the occiput and retract the skin downward
so as to expose the muscles of the upper end of the neck. Feel for
the transverse processes of the atlas and cut the muscles across at
the posterior edges of the processes. Cut off the spinous process
of the axis with a bone forceps. Thread a large packing needle
(at least 15 cm. long) with stout string, and pass it close under the
body of the axis (i.e., posterior to the cesophagus) and tie it tightly
in the depths of the cross cut. This ligature compresses the
vertebral arteries as they pass between the transverse processes of
the axis and atlas.

The animal is now decapitated. For this purpose flex the head
so as to stretch the occipito-atloid membrane, and thrust the point
of a narrow (12 mm. wide) amputation knife through the membrane
moving it laterally so as to cut the cord. With the point of the
knife resting on the anterior wall of the spinal canal bend the head
forcibly to one side, and carry the edge of the knife through the
opposite occipito-atloid joint. Repeat this procedure for the other
joint and then complete the decapitation by cutting through the
remaining tissues. If there is bleeding it can be stopped by raising
the stump. When it has ceased, bring the skin flaps together over
the stump and lay the preparation on a warmed observation table.
Artificial respiration by means of a pump connected with the
tracheal cannula must of course be instituted before the cord is

255

256 EXPERIMENTAL PHYSIOLOGY.

severed, if not before. It is advisable to warm the air from the
pump, but whether or not this is done, great care must be taken to
see that the temperature of the preparation, observed by a clinical
thermometer placed in the rectum, does not fall. For some time
after the decapitation little reflex activity is shown by the prepara-
tion— why is this the case? In about one hour, however, many
complex reflex movements can readily be elicited. Of these the
following should be studied; the movements may be recorded by
tying threads to the hind limbs and connecting with reducing levers.

1. THE FLEXION REFLEX, by applying stimuli (mechanical,
electrical) to the skin of the foot or stimulating the central end of
one of the sensory nerves (peroneal) with the tetanising current.
The latent time, grading of intensity, summation, etc., may he
studied by the procedures already described on p. 240.

2. THE KNEE JERK, by passively flexing the knee joint and
tapping the patellar tendon. The prompt and limp-like return
of the leg to its original position should be contrasted with the
gradual and imperfect return observed in a decerebrate prepara-
tion (p. 249)

3. THE SCRATCH REFLEX, by stroking the skin at the side of the
neck. The scratching movement of the homolateral hind limb is
not so easily evoked as in a spinal dog that has recovered from shock,
and it may not appear until the decapitated animal has been par-
tially asphyxiated by discontinuing the artificial respiration for a
minute or so. Sometimes the preparation shows a hyperexcitable
scratch reflex, but this often depends on inadequate pulmonary
ventilation. When it occurs, the respiratory apparatus should be
examined and the tracheal cannula cleared of any mucus that may
be interfering with the free passage of air into and out of the lungs.
If the scratch reflex is marked, its inhibition may readily be demon-
strated by stimulating the central end of the peroneal nerves of
either leg.

4. STIMULATION OF THE POSTERIOR COLUMNS OF THE SPINAL
CORD, by exposing the upper end of the severed cord and stimulating
by the unipolar method. This observation is of value because it
shows that stimuli descending by the main sensory pathways of the
cord — 'because the fibres transmit in both directions (cf. p. 46) — flow
into the collaterals which are adjacent to the point of entry of the
fibres into the cord. To make the observation, place an indifferent

THE SPINAL PREPARATION. 257

electrode on one foot (well moistened with strong saline solution)
and connect its wire with one pole of the secondary coil to the other
pole of which a fine stigmatic electrode is attached. Clear away any
blood clot from the upper end of the cord (using a moistened camel's
hair brush) and while holding the neck stump in one hand, stimulate
the dorsal columns of the cord, first near the median fissure and
then as near as possible to the posterior horn of grey matter. . In
the former case it is the homolateral hind limb that flexes, in the
latter case, the homolateral fore limb. It may be necessary to
repeat the observation several times with varying strengths of
stimulation in order to secure definite results.

5. THE FUNCTIONS OF THE SPINAL ROOTS, by removing the lam-
inae and articular processes of the lumbar vertebrae. The procedure
for this operation is in general the same as that described for the dog
(p. 245) with the difference that the spinous processes of the exposed
lumbar vertebrae are not cut at their bases, but the laminae are
freely exposed by cutting away the muscles which lie over them.
The articular processes are then snipped across and while pulling
up the lowermost (7th) spinous process with a strong forceps, the
laminae are cut through beginning with the 7th lumbar and working
upwards. The ganglia of the posterior roots are brought into view
by picking away the stumps of the articular processes. The 7th
ganglion lies on a line with the iliac crests.

Finally it is important to use the decapitate preparation to
study the various conditions which control THE ARTERIAL BLOOD
PRESSURE. The technique is the same as that already described for
the anaesthetized dog (p. 79) only, of course, small cannulae must
be used and the pressure established in the tubing which connects
cannula to manometer prior to removal of the clip from the artery,
must not be more than about 50 mm. Hg.

There are certain vascular reactions which it is especially
valuable to investigate in the decapitate preparation. These are:

1. The effect of stimulation of the spinal cord on the blood
pressure.

2. The effect of varying amounts of epinephrin injected into the
femoral vein.

3. The effect of pituitary extract similarly injected.

4. The effect of asphyxia.

This last group of observations may be done by advanced
students.

SECTION IX.
GENERAL PHYSIOLOGY.

CHAPTER XL.

PROPERTIES OF LIVING PROTOPLASM.
CHEMICAL CONSTITUENTS OP PROTOPLASM.

In the broadest sense physiology is that science which deals
with the phenomena of living matter and attempts to explain these
phenomena in terms of physics and chemistry. To the majority
of people physiology means in reality "human" or at best "mam-
malian" physiology, i.e., a study of the functions of the human
body, or where this is not possible the functions of the dog or cat
are studied with the 'assumption that the physiology of these
mammals is comparable with our own. This is a perfectly natural
concept since we are primarily interested in ourselves, and are
interested in other living things only as they have some reference
to ourselves — to be cultivated if they work for our good, to be
guarded against if harmful. Recently, however, it has been
demonstrated that light may be thrown on many problems in
human and mammalian physiology by a study of the life pro-
cesses in the more lowly forms of the animal world, and of the
plant world as well. To this study of the broader, more general
aspects of the functioning of living material in any form wherever
life exists has been applied the title of General Physiology.

I. The properties of living protoplasm as shown in simple plant
animal cells.

Materials: Spirogyra, Elodea, cells of onion epidermis, Para-
mecia, Amoebae ,Stentor, stamen hairs of Tradescantia.

A. Examine the materials provided for the following points:
colour, consistency, optical nature, elasticity, viscosity and other
physical characteristics. Is protoplasm homogeneous? What is
the nature of the inclusions? What does the shape of the inclu-
sions tell one as regards the physical state of protoplasm? Crush
the cells or cut them in two. What happens to the protoplasm?

258

GENERAL PHYSIOLOGY 259

Compare the fluidity of the animal and plant cells provided. Does
the nucleus differ from the cytoplasm in optical properties? Try
the effect of acetic acid if the nucleus is not clearly distinguishable
in the living condition. What evidence can you obtain that there
is an external living membrane present? Compare the plant and
animal cells in this regard.

B. If there is movement within the cells describe this "cyclosis"
in detail, making diagrams to show the directions in which the
granules move. Does the movement follow definite lines? Com-
pute the rate in millimetres per second of the protoplasmic move-
ment in Elodea, or Tradescantia at room temperature, by means of
an ocular micrometer and stopwatch. It is a rule that the speed
of chemical reactions is doubled with each rise of 10° C. (Van't
HofFs rule). This is expressed by the equation, Qi0 = 2. For a

10

/ IT-\'-

range of less than 10° C. the following formula is used : Q

where KI = initial rate, and K2 the final rate of the reaction/ ti =
the original temperature, and t2 the final temperature. Does living
protoplasm obey this rule? Plot a heat curve containing at least
four points for (1) a plant cell exhibiting striking protoplasmic
movement, (2) contraction of the vacuoles in the paramecium, and ^
(3) the pulsations of the dorsal blood vessel of the earthworm. 'i
What is the highest temperature from which the cells will recover cS*-
after an exposure of 3 minutes? What changes in the appearance '5 '
of the protoplasm occur above this temperature? (Consult Bayliss " j*-
General Physiology pp. 41-45). j*^

C. What is the effect of various concentrations (N/10, N/100,^ t
N/1000, etc.), of acids (e.g., acetic, HC1), bases (NH4OH, NaOH), •"
salts (NaCl, CuSO4, HgCl2), anaesthetics (alcohol, ether, chloroform)
sugar, iodine, etc., on the movements of protoplasm and on the
physical state of the protoplasm? In what ways do Stentor,
Paramecia, and Amoeba each attempt a defense against a harmful
agent? Can you give an explanation of this action on purely
physical or chemical (non-anthropomorphical) grounds?

D. Plasmolysis is the shrinking of the protoplasm in a plant
cell from its cellulose wall. Is the cellulose wall living? What is

the part of the plant cell that is homologous/with the outer living
^ lyiembrane in the amoeba? Determine the/osmotic pressure of a
lant cell by rinding a concentration of(KNOj)which just fails to
lasmolyse the cell in half an hour. Determine the osmotic pres-
ure of the red corpuscles of the frog by finding the solution of
NaCl which will neither cause the cells to shrink nor to swell.
Does all protoplasm have the same osmotic pressure? How would
you expect the osmotic pressure of fresh and salt water plants to
compare? (Bayliss, Prin. of Gen. Phys., pp. 162-165).

E. What is the effect of the electric current on living protoplasm?
Use plant cells showing streaming movements, Paramecia and
Amoebae. Note particularly the effect on the cilia of the Para-
mecia. To which pole do the Paramecia move? Can you explain
the movements of the Paramecia in an electric current in non-
anthropomorphic terms? (Jennings, Behavior of the Lower
Organisms, Chap. V.).

F. Study the locomotion of the Amoeba. How are pseudopodia
formed? Does the animal move in any one direction, or are its
movements at random? Touch the amoeba with a fine glass rod at
what may be considered for the moment the anterior edge. Where
does the next pseudopodium appear? Are the pseudopodia ever
wrinkled? (Jennings, Behavior of the Lower Organisms, Chap. I).
Construct a trough by fastening a coverslip on each side of a glass
slide with Canada balsam. Observe in this trough with the micro-
scope stage vertical the movements of the Amoeba in profile.
Notice the shape of the Amoeba as it falls freely through the water.
(Dellinger, Jour. Expt. Zool., 1906).

Have you discovered in these examples of protoplasm any
evidences (1) of irritability, i.e., the capability of receiving im-
pressions from the outside world — shown so markedly in our sense
organs; (2) of conduction, i.e., the capability of transmitting effects
of stimuli to parts remote from the point of stimulation — shown in
our nerves; (3) of contractibility, as in our muscles, etc.? In other
words, can it be said that there are inherent in protoplasm itself
as a physico-chemical complex the possibilities for all the specialized
functions we find in living beings, such as the eye and brain of
man, or the swiftly moving wing of the bee? And is the explanation
of these characteristics of protoplasm beyond the realm of physical
and chemical laws? (Bayliss, Prin. of Gen. Phys., p. 3-4).

GENERAL PHYSIOLOGY O *? ? ?

, « - «

//. Chemical constituents of Protoplasm.

A. Chemical elements necessary to life and growth-
Complete water culture for seedlings contains the following:

KNO3 1 gm.

MgSO4 0.5gm.

CaSO4 0.5gm.

KH2PO4 O.ogm.

NaCl 0.5gm.

NaCl3 3-4 drops of 10% soln.

Dist. H2O 1000 c.c.— Solution to be heated to 100° C. for

5 minutes.

Certain members of the class will make up the whole solution
for control, others will omit one element. Compare the rate of
growth and general appearance of seedlings with roots placed in
the different solutions during a period of several weeks. What is
the apparent function of each element in the medium? What is
the source of carbon?

B. The more important elements in protoplasm —

1. Carbon—substance chars upon heating in dry test tube
(flour, sugar, bean, etc.).

2. Hydrogen — heat dried material as for carbon and note con-
densation of H2O in upper part of tube. (Flour, bean, etc.).

3. Oxygen— dry material; test tube closed with cork having
two holes; glass tube through one hole to end of tube and connected
with gas jet; the other connects with Bunsen burner. Pass gas
through the tube until the air is displaced. Light the burner and
heat the tube. The hydrogen is burned to H2O by O2 in the
material. (Flour).

4. Nitrogen — heat with soda-lime — NH3 is given off. Test by
its action on litmus and cone. HC1. (Flour, bean, etc.).

5. Sulphur — -boil with strong HNO3. Dilute and filter if neces-
sary— BaCl2 gives white precipitate of BaSO4. (Flour, white of
egg).

6. Phosphorus — to half of above solution add ammonium
molybdate and heat. Yellow ppt. indicates P. (Flour, etc.).

C. The chief way in which these elements are combined in
protoplasm.

262 EXPERIMENTAL PHYSIOLOGY.

1. Carbohydrates.

(a) Test cane sugar with Fehling's Soln. — Boil with dil. HC1
and repeat test.

(b) Molisch reaction. Place 5 c.c. con. H2SO4 in a test tube.
Incline this tube and pour in gently 5 c.c. of solution to be tested
to which two drops of Molisch's reagent has been added. Red to
violet zone at point of contact of the two liquids indicates a sugar.

2. Protein.

(a) Xanthoproteic Reaction. Add cone. HNO3. Heat. Cool-
should turn yellow — add NHUOH carefully in excess. Yellow
deepens to orange. Use egg white coagulated.

(b) Biuret. Add cone. KOH, mix thoroughly and add few
drops dilute CuSC>4. Purple or violet colour indicates protein. Use
egg white.

3. Fats.

Rub butter, suet, and a peanut or walnut kernel on a piece
of ordinary writing paper. A translucent spot appears. This is
not removed by heating (cf. essential oils as oil of cloves, winter-
green, etc.), nor will it wash off with water (cf. glycerine).

D. Test samples of the various foods which go to make up an
average meal, e.g., bread, meat, potatoes, a vegetable, etc., for
the three classes of foods. Write the results in the form of a table
indicating presence by +, absence by — .

"

CHAPTER XLI.
PHYSICAL CHEMISTRY OP CELLS.

Physical chemistry of cells.

A. Diffusion rates in non-living material. (1) Invert a tube of
H2O containing a few drops of phenolphthalein in a dish of dilute
KOH. Note the rate at which the liquid in the tube turns red.
(2) Repeat with a tube of solid agar to which phenolphthalein has
been added. (3Hm^4^aatA>£^Leolidified ctareh pa^tc in a oolu-??'
.tion of IKTrernhttoterraU' uf change uf mkmr. Does the physical "
character of the reagents affect the rate of diffusion? fir

B. Semi-permeable membranes in living cells. (1) Place
Spirogyra, Elodoea and Paramecia in a very dilute solution of a*xr
neutral red (one drop to a watch glass of water). Describe the
phenomena. (2) Try more concentrated solutions. (3) Place living
material stained with neutral red in N/40 NaOH, Ba(OH)2 and
NHUOH. Record the time for colour change. Return to H2O
after colour change has taken place. Is there a return to the
original condition? Have any cells been killed? To which solution

is the material most permeable? (4) Repeat with cells killed by
heat or exposure to chloroform. (5) Wash small cubes of beet in
running H2O to remove the coloured material in broken or injured
cells. Heat one half to them to boiling. Compare the rate of
diffusion of the red pigment in the boiled and raw cubes. How
do you account for this difference in permeability of living and
dead cells? (6) Place washed cubes of beet in 1.82% NaCl ( = 0.31
molar) which is isotonic with the cell contents. Note result.
Explanation? (7) Place similar washed cubes in a salt solution also
containing 0.17% CaCl2 made in the following manner: take a
3.64% solution NaCl and add an equal volume of 0.34% CaCl2.
Explain the result.

C. Artificial membranes. (1) Float minute crystals of CuSC>4
on a 2% solution of K4Fe (CN)6. Describe results fully. Is there
a suggestion of growth? (2) Try the reverse, a crystal of K4Fe

263

4*wu&»

264 EXPERIMENTAL PHYSIOLOGY.

(CN)6 in a 3% CuSO4 solution. (3) Place a lump of fused CaCl2
in a jar filled with saturated Na2CO3. Follow appearance daily
if possible. (4) Shake CHC13 with egg albumen. Examine under
microscope. The result is a collection of artificial cells each sur-
rounded by a film of protein.

D. Osmosis. Prepare collodion bags in the following manner.
Pour 5 c.c. of the dissolved collodion into an absolutely clean
short test tube. Invert the test tube over the bottle of collodion,
allowing as much of the collodion to run back into the bottle as
will do so. Now rotate the test tube in the air, still inverted, until
the collodion is no longer liquid. Withdraw the ether vapour from
the tube by suction. Finally introduce some water into the test
tube, loosen the collodion membrane from the sides of the tube
and transfer the collodion bag to water. Fill one bag with molasses
and tie to a long glass tube. Suspend in H2O and mark the height
to which the liquid rises. Note rate of rise. Do the same with egg
albumen, 2 M sugar solution and 2 M NaCl. Recall in this con-
nection experiments already performed on Spirogyra, etc. How
do you account for the difference in behaviour of sugar and salt
solution of the same molecular concentration? Calculate from the
"gas laws" the height to which a 2 M solution of cane sugar should
rise if the colloidal bag w^ere absolutely impermeable to sugar, and
freely permeable to water.

E. Surface tension. (1) Float a needle on the surface of H2O.
(2) Repeat with a soap solution. (3) What is the form of drops of
H2O falling from a pipette? Why? (4) Make a loop of wire and
attach to this a smaller loop of fine silk. Form a film on the wire
frame with soap solution. Wet a needle with soap solution and
pass through the film. Is the film ruptured? May this have any
relation to the passing of leucocytes through walls of capillaries?
Puncture the film within the silk loop. Result? Explain.
(5) Plateau's expt.— 10 cc.. 60% alcohol in test tube. Add a drop of
olive oil (which should sink; if it does not, add 70% alcohol). Add
a small amount of 50% alcohol without stirring, then a drop of
olive oil (which should not sink; if it does, add more 50% alcohol.)
Find the mixture in which olive oil drops will sink part way but
not entirely to bottom. What is the form of the drops? Try to
change that shape with a glass rod. Result? Is there any differ-

PHYSICAL CHEMISTRY OF CELLS 265

ence in this regard between large and small drops? Wet the glass
rod with NaOH and repeat the experiment. (6) Drop a small
amount of paraffin oil in 70% alcohol. Increase the surface of
the oil by pulling out projections with a glass rod. Results?
Explain. (7) Float a clean rubber band on surface of clean water.
Touch water inside the band with rod moistened with olive oil.
Result? Touch the water outside band. Explanation. (8) Place
a crystal of camphor on water. Trace the direction of the move-
/ ments. Touch the surface with a glass rod moistened in oil. Does
y this action bear any resemblance to living phenomena? (9) Place
a drop of Hg. in a watch glass. Note the form and size. Cover it
,with 2% HNO3 and place a crystal of K2Cr2O7 1 cm. away from
it. Draw changes in shape of Hg. Compare with the behaviour
^D of amoeba. (10) Prepare a cell as follows: with balsam fasten
* small glass rods to a slide at such a distance from each other that
they will support a cover glass. When dry, fill the cell with a

Xjbmi

ixture of 2 parts glycerine to one of 70% alcohol. Place on the
>ver glass. Introduce under the glass with a capillary tube one
Js small drop of clove oil. Result? (The drop should behave like
an amoeba). Now touch the cover glass with a hot wire or glass
rod. Result? (11) Into a drop of chloroform on a slide, introduce
the end of a small glass rod (1 cm. long). Result? Repeat, using
rods dipped in shellac, paraffin, gum arabic, NaCl, sugar. Does
this resemble "choice" of food in amoeba? Can you get the drop
of CHC13 seemingly to "ingest" "digest" and "excrete"? (12)
Artificial amoeba. Wet a piece of cardboard in one spot, 1 cm. in
diameter, with water. Cover this cardboard with olive oil, letting
the oil soak into the paper everywhere except, of course, at the
wet spot. Remove the surplus of oil and lay the cardboard flat
on the table. Place a drop of glycerine mixed with soot on the
cardboard at the edge of the spot where the water was. Study
the movements and draw. Can you satisfactorily explain pseudo-
podial movement as due entirely to surface tension effects?

Colloids.

(1) Effect of electrolytes on the swelling of gelatin. Soak a
sheet of gelatin in water. With a cork borer cut out discs of known
size and place them in a series of concentrations of acids, bases,

266 EXPERIMENTAL PHYSIOLOGY

and salts, N, N/10, N/20, N/30, N/40, N/50. Use discs in dis-
tilled water for control. Let them stand for several hours in these
solutions, then measure the size of each. Tabulate the results
for the whole series of all reagents. (Some members of class use
HC1, others H2SO4, NaCl, Na2SO4, NaOH, NH4OH, etc.). What
is the optimum for each electrolyte? Does valency have any
effect? Antagonistic solutions should also be tried, especially the
following: N/10 HC1 + N/10 NaCl (add equal quantities of N/5
solution of each); N/20 HC1+N/10 NaCl; N/10 NaOH + N/10
NaCl; N/20 NaOH+N/10 NaCl.

Another method, perhaps better than the disc method, is
to arrange a series of test tubes of uniform size each containing
the same quantity of powdered gelatin (0.5 — 1 gm.). 10 c.c. of
the given solution is added and when the swelling has become
constant the height of the column of gelatin is read.

(2) Brownian movement.^. Rub a piece of a water-colour
tablet in water on a slide and examine under high power both with
reflected and transmitted light (dark field). Can you trace the
path of a single particle? How would you distinguish one of these
colloidal particles from a typhoid bacillus, for example? Add a
drop of warm (liquid) gelatin to the water-colour. Result? Allow
the gelatin to cool and harden. Result on Brownian movement?

(1) Filter solution of methylene blue or congo red through bone
charcoal. The filtrate should be colourless. Now pour through
the charcoal 95% alcohol. The colour should pass through. Can
you account for this action on the supposition of changes in the
electric charges on the aggregates of molecules? (2) Place suc-
cessive drops of different dyes on the centre of a circular filter
paper. Explain results.

CHAPTER XLII.
ENZYMES AND DIGESTION. BUFFERS.

IV. Enzymes and Digestion.

1. Ptyalin from saliva, (a) Test a biscuit for sugar with
Fehling's solution. Chew a biscuit until it tastes sweet. Now test
with Fehling's. Explain result, (b) Collect a few c.c. of saliva
in a beaker (the flow of secretion may be accelerated by chewing
paraffin) ; dilute the saliva with about 5 vols. H2O. Make the
following mixtures: (1) equal vols. starch paste and saliva, (2) the
same, except using saliva that has been previously boiled, (3) same
as (1) +5 drops 10% HC1, (4) same as (1) +5 drops 10% KOH.
Divide (1) into two portions and set one outside window. Warm
all the other mixtures to 37°-40° C. (not higher) and maintain at
40° C. for 10 minutes. Test each mixture for starch (using iodine)
and for sugar (Fehling's). Discuss your results.

2. Pepsin. Prepare 12 Mett's Tubes of egg-white in the manner
demonstrated. Prepare test-tubes containing: (a) 1% pepsin in
1% Na2CO3. (b) 1% pepsin in 0.03 N HCL. (c) 1% pepsin in/
H2O. To each of these add a Mett's Tube and try effect on each ^
combination of (a) cold (out of window), (b) room temperature,^

(c) 40° C. (d) 100° C. for 10 minutes then 40° C. Make quanti- T!
tative estimations at different time intervals. Results.

3. Coagulating Enzymes. Rennin. Prepare (a) 5 c.c. fresh milk */
+ 0.03 N HCL until a ppt. formed, (b) 5 c.c. fresh milk+5 drops fi<
rennin. (c) 5 c.c. fresh milk+5 drops rennin + 5 drops 1% Na2CO3. >^e

(d) 5 c.c. fresh milk+5 drops 0.03 N HCL. Let stand for one hour A
and compare: (a) is for control to compare with (d)., Try (a) after
boiling rennin.

4. Oxidizing Enzymes. Add H2O2 to blood. Is the presence
of catalase shown? Try bits of spleen, liver, lung, kidney and
muscle of frog. Which gives greatest catalytic action? Boil each
of these tissues and repeat. Add solution of guiacum to a slice of
potato. Blue colour indicates "active" oxygen. In what part

267

268 EXPERIMENTAL PHYSIOLOGY

of the potato is this most evident? To bits of potato in an atmo-
sphere of coal gas (free from oxygen) add (without admission of
oxygen) a few drops of guiacum. Should be no colour — Why?
Now admit air. Result?

5. Is digestion in protozoa comparable to that in man? Test
alizarin in acid and alkali to determine colour changes. Feed
paramecia with a suspension of alizarin. Shortly after ingestion
the globule of alizarin should indicate digestion in an acid medium.
In what region of the animal does this take place? Later in the
course of cyclosis, an alkaline medium may be shown.

6. Write a statement concerning the characteristics of enzymes
brought out in these experiments.

V. Buffer action and determination of PH.
Titrate N/10 NaOH against N/10 HC1. Do the same with

i anS N/10 NaHCO3 and N/10 Na2CO3. How do

you account for the differences obtained? (Bayliss, Prin. Gen.
Phys., p. 203-205).

Place on a slide small bits of ciliated epithelium from each of
three regions of a frog's mouth, anterior, middle, and posterior.
Find the concentration of acetic acid which will stop the movement
of the cilia in the anterior piece in three minutes. This can be
done by diluting one drop of a stock solution of acetic acid with a
definite number of drops of distilled water. Note the time when
the ciliary motion ceases in each of the other two pieces. Place 1 c.c.
of the solution of acetic acid which stops ciliary action in the
anterior piece in three minutes in a Pyrex narrow-bore tube, add
a few drops of an appropriate indicator (brom-phenol blue) and
match the resulting colour with the appropriate colour in the colour
chart of Clark's " Determination of Hydrogen-Ions", page 40.
This will give you the pH of the solution.

Repeat using HC1 as the acid, and thymol blue as indicator.

What is the relation of acids to ciliary movement? Is there any
difference in the effectiveness of organic and inorganic acids, or is
the hydrogen-ion concentration the only factor? . How do you
account for the fact that cilia from the more anterior regions are
more susceptible to the acid?

J /*

>/ U^L^U^

/ /

CHAPTER XLIII
MOVEMENT, ENVIRONMENT.

Physiology of Movement.

1. Review the phenomena of amoeboid movement (I, F, etc.).

2. Review ciliary action (V).

3. Muscle-nerve physiology (selected experiments from Section I) .

Responses of Organisms to Environment, Regeneration.

The functional unit of the nervous system in multicellular animals
is the reflex arc. This consists in its simplest form of a receptor,
an afferent nerve, an efferent nerve, and an effector. In verte-
brates, for example, the receptor is a sense organ like the eye,
which is affected more readily by a certain stimulus than are the
other sense organs. The change wrought in this sense organ by
the stimulus affects the afferent nerve in such a way that a nervous
impulse is initiated and passes along the afferent nerve to the
central nervous system. Usually the impulse passes over an inter-
mediate neurone before it reaches the efferent neurone. From the
efferent neurone the impulse passes to some gland, or muscle, the
effector, which acts in its own characteristic fashion.

A. Experiments from Section III will be selected to illustrate
reflex arcs and the functions of the central nervous system of the
frog as an example of a vertebrate, and from Section V to illustrate
the action of receptors.

B. Invertebrates are more stereotyped in their reactions, and
their responses to external stimuli are often constant and invari-
able. To these responses has been applied the term "tropisms"
(also " taxes"). (Consult Loeb " Forced Movements, Tropisms,
and Animal Conduct").

The material to be studied is the flat worm, Planaria maculata.
Note methods of locomotion (1) on flat surface under water, (2) on
the surface film, (3) in air. Does the animal ever swim freely?
Is the entire ventral surface in contact with the substrate during

269

270 EXPERIMENTAL PHYSIOLOGY

creeping? Are any muscular movements observable during creep-
ing which may account for locomotion? (To be observed under (2)
above). Can ciliary action account for locomotion? Notice the
movements of the auricles, the triangular projections lateral to
the eyes. Is there ciliary movement? Test with powdered carmine.

With a hair or other delicate instrument explore various regions
of the body for sensitivity to touch and determine the order of
sensitivity. Touch the tip of the head with a blunt needle. If done
carefully the planarian will grip the needle and can be lifted through
the water. Response to touch is called thigmotropism, positive if
the organism moves toward the source of stimulation or remains
attached to it, negative if the organism turns or moves away. If there
is no response the organism is said to be indifferent to that stimulus.

Stereotropism. Do planarians collect in cracks with as much
of their bodies touching the dish as possible?

Chemotropism. Place a small bit of liver an inch or so away
from a shaded quiescent planarian. Describe movements of animal
in finding the liver, and during ingestion of food note action of
pharynx. Is there any circulation of ingested food?

Geotropism. Do you find any difference in response to gravity
depending on whether the animal is fed or not? Be careful to rule
out any effect of light. (Olmsted, Jour. An. Behaviour, 1917).

Galvanotropism. Are the planarians forced to travel to the
anode or cathode? Explanation?

Rheotropism. Do planarians move up or down stream in a
gentle current?

Phototropism. Place five worms in a dish one half of which is
shaded, the other half brightly illuminated. Make five trials and
note where the worms collect after five minutes. Are they nega-
tively or positively phototropic? Use coloured glass over the
illuminated half, e.g., blue, red, green, yellow. Differences in
response? Throw a narrow shadow down the middle of the dish
and start a planarian moving toward the source of light. Can you
make the animal continue to move in this direction? Describe
movements. Cut a worm's head off just behind the eyes. Describe
the responses of each piece to the light. Are the ey/^ essential
for the light response? (Consult article by Taliaferro in Jour.
Expt. Zool. 1921).

MOVEMENT, ENVIRONMENT 271

Do any parts exhibit greater permeability for dyes than other
parts? Use methylene blue. (MacArthur).

Students will be divided into groups to work out the answers
to the following questions: (1) Will transverse pieces from any part
of the body regenerate an entire new animal. (2) Will longitudinal
strips regenerate entirely? (3) Will obliquely cut pieces regenerate?
(4) On an obliquely cut piece which eye regenerates first? (5) Can
the protrusible pharynx regenerate? (6) What is the smallest
proportion of the body which will regenerate? (7) How do small
triangular pieces from the side of the body regenerate? (8) Is
regeneration more rapid in the light or in the dark? (9) What is
the effect of starvation on the size of the worm? (10) Can you get
double-headed or tailed forms by splitting the head or tail?

C. Recall the experiments in I illustrating the behaviour of
Protozoa to their environment.

D. Responses in a plant. Touch the different organs of the
Sensitive Plant, Mimosa, with varying degrees of strength. Deter-
mine the rate at which the impulse travels along a frond in cms.
per sec. Compare this with conduction in the frog and mammalian
nerve. Note the pulvinus at the base of each frond and leaflet.
Can you account for the action of the plant on the assumption of
osmotic changes?

E. The fundamental differences between plants and animals.
The fundamental difference between plants and animals does not
consist in powers of movement, since many plants are motile,
e.g., algae, while many animals, e.g., corals, are sessile, nor in
any other obvious external character. The difference lies in the
fact that plants manufacture their food from raw materials using
CO2 and H2O to form carbohydrates, nitrates as a source of protein
nitrogen, etc. Animals are unable to do this. Once the requisite
materials are made up into the proper compounds, the process by
which energy is liberated for the carrying on of life is essentially
the same in plants and animals, i.e., respiration is common to
both. Plants are unique in their powers to form materials which
store up energy. Chief of these processes is photo-synthesis, the
production of carbohydrates from CO2 and H2O by means of the
energy of sunlight.

(1) Place an equal number of Planaria of the same size in each

272

EXPERIMENTAL PHYSIOLOGY

of two pyrex glass tubes filled with water containing a few drops
of phenolsulphonephthalein. Determine the pH of the water by
comparing with known standards. Tap water is slightly alkaline.
The tubes should be tightly ctfrked with exclusion of air. Place one
tube in the dark, the other in direct sunlight. Note the change in
hydrogen-ion concentration in the two tubes after 2-3 hours or
less if the sunlight proves injurious to the planaria. If the planaria
give out CO2 the solution should become more acid, if they use
up CO2 the solution should become more alkaline.

(2) Repeat the experiment using a water plant, e.g., spirogyra,
instead of the planaria, and having the water distinctly acid to
start with by bubbling through it CO2 from your breath.

In the tube placed in sunlight bubbles may appear and a gas
collect. Determine the nature of this gas by an appropriate test.

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^ Capillary Circulation.

Pith a frog without haemorrhage. Pin web of foot over a hole in
a board. Keep rr^j06t with saline. Cover with coverglass if neces-
i sary. Focus on web with CPp. microscope.

1. Blood Vessels. Notice the movement of the blood within
the blood vessels of varying size and irregular course.

2. Velocity of flow. Is the motion equally rapid in all the
vessels? If not are the slower currents in the larger or smaller
channels? How can you distinguish an artery from a vein? Com-
pare thickness of walls and character of flow. Is the velocity
equally rapid in all parts of a vessel at a given point? Why is this

'so? Of what physiological importance?

3. Pulse. Have you seen evidence of intermittent force acting
upon the corpuscles? If so describe its influence. Determine
whether this intermittent force makes itself evident in all of the
vessels. If not, in which class of vessels is it present? Can you
detect pulsations in a small artery? Vein? Capillary?

4. Size of Vessels. Do you observe any variation in the size
of a given vessel? Carefully measure with the eyepiece micro-
meter and determine.

5. Do you see small vessels appear, apparently new, from time
to time? How do you account for this? Do you see small vessels
apparently disappear?

6. Corpuscles. In the frog the red corpuscles are larger than
the white. With high power study the behaviour of the red cor-
puscles. Do they change shape? If so, under what conditions?
Do you see evidences of flexibility and elasticity? Are the red
corpuscles nucleated? Study the white corpuscles. Where are
they the most numerous in the blood stream? How do you account
for this distribution?

7. Capillary Flow. Estimate the velocity, of a single corpuscle
by means of the micrometer eyepiece and stop watch. Of what
physiological significance is this slow rate?

273

274 EXPERIMENTAL PHYSIOLOGY

8. Process of Inflammation. Remove the cover-glass, dry the
web with filter paper and touch a point with a pin dipped into a
2 per cent, solution of croton oil and olive oil. Without replacing
cover glass, with low power, observe whether the presence of the
croton oil effects any change in the diameter of the vessels, or in
the rate of blood-flow. If there is a change in both, has one a causa-
tive relation to the other? Note and describe minutely all changes
which take place at or near the place touched with the croton oil.
If no marked changes have been produced in 10 minutes by the
croton oil, touch the point with a needle which has been dipped
in strong nitric acid. Observe with high power, using cover-glass.
Describe process of extravasation of white or red corpuscles.

^Lymph-Heart^.

Another method of pertorn
results. The tongue of a pith

are readily seen. A drop of x>

produce the necessary inflamn ation.

ng this experiment also gives good
d frog is pinned out over a hole in

a board. With the proper d( gree of stretching the bloodvessels

ol on the surface of the tongue will

1. Posterior Lymph-Hearts. Stretch the pithed frog out,
ventral side down, and watch the beating of the posterior pair of
lymph-hearts. These hearts are located (at the dorsal posterior
end of the body) one on each side of the urostyle in the triangular
spaces easily observed there between certain muscle attachments.

Cautiously turn back the skin covering the lymph-hearts being
careful not to injure the cutaneous veins and thus cause bleeding.
Moisten exposed parts with normal salt solution when necessary,
and count the number of beats per minute of each lymph-heart.
Compare. The movements of the blood-heart can be distinguished
without opening the chest. Can you detect any synchronous
relation between the rhythm of the blood-heart and that of the
lymph-hearts?

Blood-heart.

1. Blood heart-of the Frog. Expose the blood-heart. Observe
the heart "in situ" noting the two auricles; the ventricle; the
auriculo-ventricular groove; and the truncus arteriosus, which pro-
jects anteriorly from the ventricle in front of the auricles and
divides into the two aortae.

PHYSIOLOGY OF CIRCULATION 275

Tilt up the ventricle and observe the thin shred of connective
tissue — the frenum (containing a small vein) — -passing from the
pericardium to the back part of the ventricle. Tie a fine thread
around the frenum and divide it dorsal to the ligature. Lift the
heart gently by this thread and observe also the sinus venosus,
continuous with the right auricle and formed by the union of the
inferior and the two (smaller) superior venae cavae.

2. Arrange to record heart beat on slow kymograph. Rate of
heart beat and influence of temperature.

(a) Determine and record the normal rate per minute by
counting several times.

(b) Determine and record the rate per minute after bathing
the heart by means of a pipette with a few c.c. of normal saline at
20° C.;at30° C. and 5° C.

3. Movements and Sequence of Contraction.

(a) Keep the heart beat slowed down with the cold normal
saline and it will be possible to determine the sequence and details
of contraction. Determine whether the two auricles contract at
the same time. *

Carefully lift the ventricle by the ligature of the frenum and
observe the sinus venosus. Note where the wave of contraction
really begins and follow the whole cycle of contraction step by
step. Tabulate and describe the events in order through one
entire cycle.

(b) Use a pithing needle now to destroy the entire spinal cord.
What effect do you observe upon the beating of the lymph hearts?
What can you conclude concerning the nerve centres which govern
their movements? Is any change produced in the beat or appear-
ance of the blood-heart? Conclusions?

4. Effect of Temperature on the Excised Heart. Carefully
remove the entire heart from the body.

(a) Place the watch glass containing the excised heart on a
small beaker of warm water — determine the rate per second at
5° C., at 20° C. and at 35° C. Caution: Do not let the temperature
rise above 40° C. Plot beat curve and calculate Q 10.

(b) Section of the excised heart. Separate the sinus, auricles
and ventricle from each other. Does each beat independently? Do
they beat at the same rates? Note time for 10 pulsations of each.

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APPENDIX.

TABLE I.

THE PERCENTAGE OF OXYGEN WHICH IS EQUIVALENT TO THE NlTROGEN FOUND

IN THE EXPIRED AIR.

To obtain the nitrogen in the expired air, add the percentage of COz and C>2
found and subtract the sum from 100. The table gives the percentage for O2
corresponding to this figure:

%N2 78.7 78.8 78.9 79.0 79.1 79.2 79.3 79.4 79.5 79.6 79.7 79.8

79.9 80.0 80.1 80.2 80.3 80.4 80.5 80.6
%O2 20.86 20.88 20.90 20.93 20.96 20.98 21.01 21.04 21.07 21.10 21.12 21.14

21.16 21.19 21.22 21.25 21.28 21.31 21.35 21.38

TABLE II.

TENSION OF AQUEOUS VAPOUR IN MILLIMETRES OF MERCURY.
To obtain the dry barometric pressure, subtract the mm. Hg. corresponding to
the temperature of the air from the barometric pressure at the time of the experi-
ment:

Temp. 15°C. 16° 17° 18° 19° 20° 21° 22° 23° 24° 25°
Mm. 12.7 13.5 14.4 15.4 16.3 17.4 18.5 19.7 20.9 22.2 23.5

TABLE III.

TEMPERATURE CORRECTIONS TO REDUCE READINGS OF A MERCURIAL

BAROMETER WITH A BRASS SCALE TO 0° C.

Subtract the appropriate quantity as found in table from the height of the
barometer. The table is for a barometer with a brass scale, and the values are a
little lower (about .2 mm.) than for the glass scale. The corrections for inter-
mediate temperatures can be approximated.

Temp.

700

710

720

730

740

750

760

770

mm.

mm.

mm.

mm.

mm.

mm.

mm.

mm.

15°C.

1.69

1.72

1.74

1.77

1.79

1.81

1.84

1.86

20°

2.26

2.22

2.32

2.36

2.39

2.42

2.45

2.48

25°

•2.83

2.87

2.91

2.95

2.99

3.03

3.07

3.11

277

278 EXPERIMENTAL PHYSIOLOGY

TABLE IV.

TABLE FOR REDUCING GASEOUS VOLUMES TO NORMAL TEMPERATURE AND

PRESSURE.

The observed volume, when multiplied by the factor corresponding to the
temperature and pressure, will give the volume of the expired air reduced to 0°c.
and 760 mm.

Mm.

15°C.

16°

17°

18°

19°

20°

21°

22°

23°

24°

25°

720

.898

.894

.891

.888

.885

.882

.880

.877

.873

.870

.857

730

.910

.907

.904

.901

.897

.894

.891

.888

.885

.882

.879

740

.922

.919

.916

.913

.910

.907

.904

.901

.897

.894

.891

750

.935

.932

.928

.925

.922

.919

.916

.913

.910

.907

.904

760

.947

.944

.941

.938

.934

.931

.928

.925

.922

.919

.916

770 .960 .957 .953 .950 .948 .945 .940 .938 .933 .930 .927

APPENDIX

279

TABLE V.

R. Q. CALORIES FOR 1 LITER

Number

RELATIVE CALORIES CONSUMED AS

Carbohydrate Fat

per cent.

per cent.

0.707

4.686

0

100

0.71

4.690

1.4

98.6

0.72

4.702

4.8

95.2

0.73

4.714

8.2

91.8

0.74

4.727

11.8

88.4

0.75

4.739

15.0

85.0

0.76

4.752

18.4

81.6

0.77

4.764

21.8

78.2

0.78

4.776

25.2

74.8

0.79

4.789

28.6

71.4

0.80

4.801

32.0

68.0

0.81

4.813

35.4

64.6

0.82

4.825

38.8

61.2

0.83

4.838

42.2

57.8

0.84

4.850

45.6

54.4

0.85

4.863

49.0

51.0

0.86

4.875

52.4

47.6

0.87

4.887

55.8

44.2

0.88

4.900

59.2

40.8

0.89

4.912

62.6

37.4

0.90

4.924

66.0

34.0

0.91

4.936

69.4

30.6

0.92

4.948

72.8

27.2

0.93

4.960

76.2

23.8

0.94

4.973

79.6

20.4

0.95

4.685

83.0

17.0

0.96

4.997

86.4

13.6

0.97

5.010

89.8

10.2

0.98

5.022

93.2

6.8

0.99

5.034

96.6

3.4

1.00

5.047

100.0

0.0

(From Lusk.)

INDEX

Aberration, chromatic, 150

spherical, 149

Accelerator fibres of heart, 63
Acceleration of heart, 63
Accommodation, 153

reflex, 155
Acoustic reflex, 249

Action, of CO2, ether and chloroform
on nerve, 58, 50

of cations on cardiac muscle, 70, 62

current, 41

pumping, of heart, 223
Adrenalin, effect of, on blood vessels in
frog, 77

influence of, on mammalian heart,

209

Adsorption, 266
Aesthesiometer, 194
After images, 174

All-or-None principle in cardiac mus-
cle, 61

in nerve and voluntary muscle, 57
Alveolar air, collection of, 112
Analysis of gases, 107

of tetanus, 37

Anelectrotonus in nerve, 51
Anodal contractions in heart, 55
Aqueous vapor, tension of, 277
Asphyxia, influence of, on blood pres-
sure, 201

effect of, on kidney volume, 205
Astigmatism, 151

Atropine, influence of on submaxillary
secretion, 227

on heart, 65
A-V interval, turtle, 67

Basis of sound, 182

Bellows recorder, use of, 202

Bile, secretion of, 228

Binocular vision, 181

Blind spot, 166

Blood flow, measurement of, by calori-

metric method, 210
Blood gas, determination,

chemical method, 128

pump method, 122

Blood pressure, method of determining
in lower animals, 79

measurement of, in man, 71
Blood pump, 122

Blood vessels, response of, to chemical

substances, 76
Bones of middle ear, 186
Brodie's apparatus for heart per-

fusion, 207
Buffer action, 268

Calcium ions, action of, on cardiac
muscle, 70

influence of, on mammalian heart,
209

influence of, on turtle heart, 70
Calculation of respiratory quotient, 118
Calorimetric method for measurement

of blood flow, 210
Capillary circulation, 273
Carbon dioxide, action of on nerve, 50

combining power of the alkaline
reserve of blood, 138

influence of, in lowering the disso-
ciation curve, 137
Cardiac muscle, 61
Cardiograms, 96
Carotid pulse, 94

Cations, action of, on cardiac muscle,
62

influence of, on mammalian heart,
209

influences of, on turtle heart, 70
Central nervous system, 239
Cells, physical chemistry, 263
Cerebral localization, 243
Cerebrum, function of

in frog, 102

in mammal, 243

in pigeon, 253
Chemical substances, response of blood

vessels to, 71

Chloroform, action on nerve, 50
Chromatic aberration, 150
Ciliated cells, 45
Circulation, reflex control of, 199

capillary, 273

scheme, 74

time, 90
Coagulation of muscle proteins, 39

of blood, 91
Cochlea, 188
Cold spots, 192
Colloids, 263
Colour biindfless, 175

vision, 170

281

282

EXPERIMENTAL PHYSIOLOGY.

Colours, complementary, 172
Compensatory pause in heart, 69
Conduction of a nerve fibre in both

directions, 46
Contraction, isometric, 27

istonic, 25

simple, 22

Contrast, simultaneous, 173
Control of pancreatic secretion. 228
Corrections for a barometer with a

brass scale, 277
Corti, organ of, 189
Curare, effect of, 19

influence on respiration, 200
Current of injury, 40

Dark adaption of retina, 167

Decerebrate, activities of frog, 102
activities of pigeon, 253
rigidity, 249

Decerebrator, 247

Deglutition reflex, 249

Depressor nerve stimulation effect on
blood pressure, 89

Digestion, 267

Digestive system, 226

Diphasic variation, in muscle, 40
in nerve, 56

Direct current stimulation, 54

Dissociation curve for Oxygen and
CO2, combining power of blood,
132

Duodenal extract, influence of, on pan-
creatic secretion, 229

Effect of drying on muscle, 16

of temperature on contraction, 28

of temperature on contraction,
apparatus for, 29

of temperature on cardiac muscle,

62

Efferent nerve fibres, 46
Elasticity of muscle, 20, 42
Electrical changes in muscle, 39

changes, influence of, on nerve, 51

stimulation, 9, 15
Electrodes, Sherrington, 237
Electrotonus in the heart, 55

in nerve, 51
Enzymes, 267

Epinephrin, influence of, on mam-
malian heart, 209

resuscitation by transfusion with,

205

Errors in refraction, 149
Ether, action on nerve, 50
Eustachian tube, function of, 188

Extensor thrust, 240
Extra systole, 68

Factors influencing nervous conduc-
tion, mechanical, 50

Thermal, 50

Chemical, 50
Fatigue, 33

resistance of nerve to, 57

reflex, 241
Fistula, gastric, 234

of parotid gland, 231
Fixation of tracing, 12
Flexion reflex, 240, 249, 256
Fovea centralis, 163
Frog's heart, study of, 61

Gad's method for cardiac valves, 224

Gas analysis apparatus, 107

Gaseous volume reduced to normal

temperature and pressure, 278
General Physiology, 258
Gastric fistula, 234
Glucose, influence of, on lymph flow,

220

Graded stimulation, effect of, 17
Graphic record, 12
Gunn's apparatus for heart perfusion,

207

Haemodynamics, principles of, 73, 78
Haemorrhage, effect of, on blood pres-
sure, 84
Haldane's apparatus for blood gases,

129

Head-shake reflex, 249
Hearing, 182
Heart beat observed in the open

thorax, 85
contraction, suspension method

for, 63

perfusion, mammalian, 206
Hering's theory of color vision, 176
Heat production, 38
rigor, 39
spots, 193
Hunger contractions, observations of,

235
Hydrogen ions, 266

influence of, on mammalian heart,

209

Hypermetropia, 151
Hyperpnea, 200

Images, Purkinje, 156

Inadequate stimuli, summation of, 48

Independent excitability of muscle, 19

INDEX

283

Induction, immediate successive, 242
Inductorium, 11
Inflammation, 174
Inhibition, reciprocal, 251

of the heart, 62

of reflexes, 101
Innervation, reciprocal, 245

of salivary glands, 226
Intestines, movements of, 236
Irritability of smooth muscle, 42
Isometric contraction, 27
Isotonic contraction, 25

Judgment of space and depth, 178
Jugular pulse curve, 95

Katelectrotonus in nerve, 51
Kathodal contractions in the heart, 55
Kidney volume, change in due to
asphyxia, 205

change in due to splanchnic stimu-
lation, 203

Knee jerk, 105, 240, 249, 256
Kymograph, rapid, 23

slow moving, 16

Harvard. 26

Latent period of muscle contraction

voluntary muscle, 24

smooth muscle, 43
Lens, attachment of, 154
"Light reflex", 167
Load and work, 31
Localization cerebral, 243
Long-sightedness, 151
Lymph flow, influences of

circulatory changes on, 218

lymphagogues on, 220

Macula lutea, 163

Maximal and minimal stimulation, 17

Measurement of blood flow calori

metric method, 210
Mechanical stimulation, 16
Membrane, semi-permeable, 263
Metabolism in nerve, 56
Middle ear, 186
Monocular vision, 180
Movements of intestine, 236
Muscle contractions, apparatus for

recording, 16
sense, 197

Muscles of the frog's leg, 21
Musical sound waves, 184
Myopia, 151

Near point, 155

Nerve, conduction of, in both direc-
tions, 46
Nerve-muscle preparation, 14

as a rheoscope, 40
Nervous control of heart beat, 62

impulse velocity of, 48
Nicotine, effect of, on heart, 65
Nodal point, 144

Oesophagus, movements of, 236
Oncometer, use of, 202
Ophthalmoscope, use of, 158

by direct method, 160

by indirect method, 161
Optics, physiological, 139
Osmasis, 264
Overtones, 185
Oxidation in the body, 90
Oxygen, percentage of, equivalent to

nitrogen in expired air, 277

Pain, sense of, 196

Pale muscle fibres, 38

Palpebral reflex, 104

Pancreatic secretion, control of, 228

Parotid gland, fistula of, 231

Peptone, influence of, on lymph flow,

221
Perfusion of the frog, 76

of excised mammalian heart, 206

turtle heart, 69
Physical basis of sound, 182
Pigeon, decerebrate, 253
Pinna reflex, 248
Pitch, 182
Pithing, 15

Pituitrin, influence on lymph flow, 221
Plants, responses of, 271
Plasmolysis, 259
Polyshygmograph, 93
Posterior columns of spinal cord, stim-
ulation of, 256
Postural tonus, reflex, 250
Potassium ions, action of, on cardiac
muscle, 62

influence of, on mammalian heart,

209

Preliminary technique, 9
Preparation of cat and dog for experi-
mental study, 79

of rabbit, 87
Primary circuit, 10
Principal axis, 143
Propagation of pulse wave, 93
Protoplasm properties, 258

chemistry, 261

284

EXPERIMENTAL PHYSIOLOGY.

Purkinje figures, 165
images, 156

Radial pulse, 94
Rapid kymograph, 23
Reaction time in man, 106
Reciprocal inhibition, 251

innervation, 245
Red muscle fibres, 38
Reflection, laws of, 139
Reflex, accommodation, 155

acoustic, 249

action in frog, 98

action in mammal, 239

control of circulation, 199

deglutition, 249

extensor thrust, 240

fatigue, 241

flexion, 240, 249, 256

head shake, 249

palpebral, 104

pinna, 248

postural tonus, 250

control of respiration, 199

scratch, 240

summation, 241

time 240

time in frog, 99

time, in man, 106
Reflexes, spreading of, 99

inhibition of, 101
Refraction by a convex lens, 145

errors in, 149

in the eye, 146
Refractive index, 143
Refractory period in the heart, 68
Removal of paper from drum, 12
Resistance of nerve to fatigue, 57
Respiration, 107

reflex control of, 199
Respiratory exchange in man, 118

movements, influence of, on pulse
97

quotients, 279

quotients, calculation of, 120
Response of retina to light, 167
Resuscitation by transfusion with

epinephrin, 205

Retina, effects on, of stimulating with
two different colours, 171

structure of, 163

response to light, 157
Retinoscope, 161
Rheoscope, nerve muscle as a, 40
Rhythmicity of smooth muscle, 43
Rigor mortis, 39

Salivary glands, innervation of, 226
Saliva, secretion of, 231
Scheiner's experiment, 157
Schematic eye, 147

Sciatic stimulation, effect on respira-
tion, 199

Scratch reflex, 240, 256
Secondary circuit, 10
Secretin, influence of, on pancreatic

secretion, 230
Secretion of bile, 228
of gastric juice, 233
of pancreas, 228
of saliva, 231

of submaxillary gland, 226
Semilunar valve, effect of damaging,

223

Semipermeable membranes, 263
Sensations, skin, 192
Sherrington electrodes, 237
Simple contractions, 22
Simultaneous action on the retina of

three primary colours, 173
Skin sensations, 192
Smoking paper, 12
Smooth muscle, 42
Sodium chloride, influence of on lymph

flow, 221
Sodium ions, action of, on cardiac

muscle, 62

Sound waves, diagrams, 183, 184
Sodium nitrite, effect of, on blood ves-
sels of the frog, 77
Sound waves, transmission of, in the

ear, 185

Space and depth, judgments of, 178
Special senses, 139
Spherical aberration, 149
Sphygmic period, 96
Spinal cat, 255
Spinal nerve roots, function of, frog, 102

functions of mammal, 245, 250
Spirometer, Tissot-Carpenter, 118
Splanchnic nerve stimulation effect on

blood pressure, 83
stimulation, effect of, on kidney

"volume," 202
Spreading of reflexes, 99
"Staircase" effects, 33
Stannius ligature, 65
Stereoscopic vision, 181
Stewart's calorimeter method for meas-
urement of blood flow, 210
Submaxillary gland, secretion of, 226
Summation of inadequate stimuli, 48
of smooth muscle, 43
from two stimuli, 35

INDEX

285

Surface tension, 264
Suspension method for heart con-
traction, 63

Table

percentage of oxygen equivalent to

nitrogen in expired air, 277
temperature corrections to reduce
readings of a mercurial
barometer with a brass
scale, 277

tension of aqueous vapour, 277
for reducing gaseous volumes to
normal temperature and pres-
sure, 278

of respiratory quotients, 279
Taste, 197
Temperature, effect of, on contraction,

28
Temperature sense, 193

effects on cardiac muscle of frog, 62
effects on cardiac muscle of turtle,

68

Tension of aqueous vapour, 277
Tetanic nature of voluntary contrac-
tion, 36
Tetanus, 35

of smooth muscle, 44
Thermal stimulation, 15
Thoracic duct, dissection for exposure

of, 218

Tissot-Carpenter method for deter-
mining respiratory exchange, 118
Tone of smooth muscle, 43
Touch, sense of, 194
Transfusion, effect, on blood pressure, 84

Transmission of sound waves in the ear,

185

Tropisms, 269

Tricuspid valve, operation of, 223
Turtle heart, study of, 66

Vagus, influence on heart, 62

influence on respiration, 199
nerve in the frog, location of, 64
stimulation, effect on blood pres-
sure (dog), 81

stimulation, effect on blood pres-
sure (rabbit), 89
Van't Hoff rule, 259
Valves of the heart, action of, 206
Vasoconstrictor nerves, demonstration
of in cervical sympathetic, 87
fibres, demonstrated by the ple-

thysmograph, 202
Velocity of the nervous impulse, 48

of transmission of pulse wave, 93
Venous curve, interpretation of, 95
Veratrine, influence of, on contraction,

32

Vision, 123
Visual axis, 151

Volume of a contracting muscle, 25
Voltage, 9
Voluntary muscle, 14

contraction, tetanic nature of, 35

Wave of contraction, 24
Work, load and, 31
Writing point, 12

Young-Helmholtz theory of colour
vision, 176

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