_ _
PHOTOROPISMS "'I?H--tt03fdeHR<MATTtrXT(mTnnr"TTn^r5' 'AWVOLTOX.
«e.
. S tr <t*.f*-..
+<- /o • ANNIE LtAY KURD.
Subraitted 3*. partial fulfillment of the requirements for the degree of Doctor of Philosophy.
University of California
April 26, 1918.
— /-
/•
OHZBHTATIOBS ABB PHOTOTROPISH8 IB FUCU8 ABB TOLTOZ WITH HDBOCHHOJI&21C LIGHT OF XQUAL IH7HB8ZTZB8.
ABBIB
Submitted in partial fulfillment of tb§ requirements for the degree of Doctor of Philosophy.
University of California.
April 26, 1916.
/.
PHOTOTROPIST'lS WITT T'ONOCTOOMATIC LIG"T IN FUCUS Alii) VOLVOS.
The -nower of light stimuli to produce orientations an a /$r"opism3 is a phenomenon which has been widely demonstrated in both the plant ana animal kingdoms. ^ot only canfi unilateral direct movements and rrowth out in some srecies of r'-jnts, name-* ly ^quisetum, Fucus, 90t- Puccinia_,and related forms, it can establish the direction of the first cleavage plane of the ger- minating snore. Since in such cases the cell on the shaded side of the s^ore becomes the rhizoidal cell. the polarity of the
plant is determined irrespective of gravit.,- .
In any attempt to discover t e mechanics of such tropistic reactions,the first problem is to find what wave lengths of light are responsible, and to what extent they are a function of the quality of the stimulus/apart from its quantity or inten- sity. This, then, is the purpose 2$*J^ec7E££3ent' investigation.
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' >^U- *s~.0.i&-. t^CS^rdf
with monochromatic light one is struck by fehe abaanoo of quauti
ave records of -either the quality w tne interisity of the ill- umination. -It ia tfr-woll known f;^ct that "Tae ordinary li^ht filters used to obtain rnonochtomatic lig":t transmit not only
tho^e vive lengths which predo ;iiu . t.t? i i ive t .e color to the
screen, but rlso othearts of the spectrum .thariresence of can be detected only by a spectroscopic analysis. Vor example, certain results are frequently ascribed to blue light with no record of just what range of the soectrum was used nor what "ave lengths other than the predominating ones were acting.
'mother source of inaccuracy has been the neglect or over- sight of the great variation in the intensity or quantity of ra- diant energy transmitted by tjie color screens. Biological
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experimenters have fot he most part failed to take into
i
sideration "he fact that the quantity as well as the quality
of the light stimulus varies with the different colors and that the former variable must be eliminated before results can be attributed to differences in wave length alonei Quite
recently (/y/Y )such work has been aone end published with the
die
statement thst^ results! are Qualitative -eniy becau-e there is no way of comparing lights of different colors as to amounts &
,_ <»tC*tx*-^e- /*
of radiant energj . kowev-er there have been several methods de~
A vised by means of which the relative intensity of monochro-
matic lights van be measured.
The first exact work of this nature was done by itniep and Minder C^°f). They usea e blue and a red color screen and a green solution with sunlight as the source of light.
XX>t&4
The wave lengths to which each/we^e transparent "/ere known;; and the energy behind eacHi was determined ny means of a therme- pile and a, d'Arsonval galvanometer. The interference of the ±« long heat rays w.-is prevented by inserting a water layer in a parallel sided container between tne thermopile and the source pf light.
Day (/?/( }, obtained light of known wave length by mean
of a spectfcum from ."ernt Glower^s formed by e carbon bisulfide prisu and cut down by/a diaphragm witn narrow ver-
tical slits whicn could be adjusted so as to permit any de-
sired region of tne spectrum to be used. In t-iis adjustment p spectroscope w;-s used to determine the exact range of wave- lengths passing through the plit.in each of the four illumin- attions used, --- red, yellow, gfeen, and blue. He measured intensity of each with a Boys r: diomicromet, er, • nd balanced" them by varying the number of glowers employed in the lamps.
"hus there was one glower for tne red light light, two for the yellow, and three fot the green ajid biue.
('?" J Laurens in an investigation of the reactions of amphibians
used these sane metnods and the same ^ppnratus for the quanti tave analysis oi- the monochromatic light he u-ed^ ard for bnlanc- ing then with respect to their relative intensity.
Gross! Ifi2) also used these snrne methods in determining the reactions of Arthropods to raonocnromatic light.
An instrument has been devised by I'.acDougal and Gpoehr. ( ) which ner-sures the total radiant energy of any light in terras of 1 its dissociation effect on a photosensitive substance. This is measured by a galvanometer, "he advantages in the use of this "photoelectric cell" are said to be its extreme sensitiveness t% the wave lengths of the blue end of the spectrum, arid the fact thst
its action in ligi-t isHraore nesrly that of the organism than that
v
of any othor light measuring instruments available.
There have been therefore three different methods worked ot out for biological experiraentsf or the quantitoive analysis ofi light stimuli; viz., those of Kniep and Hinder, Day, and of
^ racUoupiJ and Sooehr. The interesting apparatus of Patte
add of I.oeb and Northrut) wherebya quantitave measurement of the re- action of organisms subjected to two beams of light of different intensity, is obtained. The measurement is in terms of the angular deflections from an initial path of locomotion. The same methods might be ar^olied to work with colored lights. A quantitative measur- ment of the greater effectiveness of one spectral region over another of equal intensity might be measured by the angular deviation of the rath of a motile organism from a line perpendicular to a line connect- inp the two sources.
All of these methods with the exception of those of i'nien and inder involve special a^^aratus often not easily available. It was aecessaty for fH e -rurroses of this investigation to find a simpler
of accomplishing the sa-ne results, '"his report therefore is concerned "/ith t'vo rro ultras; first, a means of making th r: intensity of monochromatic light outlined with a act of light filters equal; second, a comparative study of the tronisma obtained with differ- rant ™''ive lengths when the intensity variable in eliminated. The nar-
^henomena chosen for this study are t^ e rower of monochro- matic lights to (I) establish the rolurity of ^ucus sror el incs and :,he origin and consequent direction of the rhizoids; (2) to nro- iuce the negative rhototr -^ism of t1 n rhizoid; (3) to direct the move- icnt?? of 1rolvox.
, Ar>r«aratus and rethods for Exposure to Monochro- matic Light of Equal Intensity.
As biologocal science oec ones more exact with the tend- eouce the expression of natural -phenomena to mathematical form- it is obviously essential to define stimuli of all sorts ouant- Ltatively. Indefinaite or incomplete records of light stimuli can 10 longer bo attributed to the lack of means of measuring them be- :aune access to a spectroscore ano thermopile make it possible to inalyae any light qualitatively ana quantitatively.
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There are two methods for obtaining monochromatic light for biological experiment/S •'— . ---the projection of a snectrum upon the yijH/y/i^X organisms or <s of filtered lir-'ht passed through a colo screen. The former i .3 t"ieo etically the tHMrtr for exact ir work but technical difficulties such as the limited dispersion and low intensity make it impractical for 7iany investigations. Light filters of glass are the :r.ost convenient "leans of se- curin,- approximate! oc r ; I LC li—'t when unilateral ill-
umination is desired. Ordinary color screens transmit too wide a range of wave lengths for exact T.vork and at present there «t» are very f w iv';ose lir'it is of sufficient homogeneity .The uest ia the Wratten filter screen, made in London, <"hich consists o of a 1 ' -al-'tine film between two p/lass platesVacDougal and Sr>oehr(l9l7) have described some colored glass screens designed by them for biological work, but the range of wave lengths to which they ar-e transparent is considerably greater than for the Wratten filter.
In the experiments to be described, seven Wratten light
filters were used , each of which was fitted as a window in the
•fy
end of a dak b .x. i-lach transmitted only a narrqw range of
-ave lengths but all together they embraced the whole of the visible spectrum. The wave lengths to which each screen was transparent ^ev* determined by testing tie lipht transmitted by^ each with a direct wtsion spectroscope with a wave length scale attached. Thus the quality of the light stimulus acting in each box is accurately known.
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^-w-vox-^
is the electric ?<rc. The advantages of fts use .-; re, that all
filters aan be u^ed in each exposure, insuring identical condition? of temperature, constancy of illumination, etc. The disadvantages
are several. In the first place, the intensity is constantly chang- ing as the carbonslburn and the arc gets longer. r/orse, the lessen- ing of the intensity nay not be the sane for all the wave lengths. In the second place, fluctuations in the current cause large vari- ations in the intensity. In tjje third place, unless an ifafefefa&J- automatically adjusted arc is available, it is necessary to adjust
the carbons by hand every five to fifteen minutes^, anfl when an eight hour illumination is desired, tt$A entails considerable inconven- ience.
The dark boxes were 10x13 en. find 8 era. high, in one end ofe. which a hole was cut so that one of the light filters, 6 x 6 cm. might be fitted into it. The boxes were made light tight with
tightly fitting covers, and were painted black inside to guard
for uhe cultures
against reflections within the box. The dishes used were made of
r\
microscope slides cemented together with ainc eement so as to make shallow oblong dishes 7.5x2.5 cm. and 1 cm. deep. It was deemed necessary to use such flat sided dishes in order to prevent posrible complications from reflected and refracted light in the
curving sicleB of *» round dieMeo. In order to axrose more - - than one dish **=***& behind each screen Bo that none would be
shaded by* another, a rack was made to fit inside the box with ^
cleats projecting inward from the, ends so that three dishes c/culd -
he elipped into it one above the other.The light .entering through the screen at the end of the box , fell equally on the one exposed side of each of the three dishes. The rack containing the dishes could be easily lifted out and carried to the microscope for exam^C ation without dicing the ,^^ ^^^-^^
The spectrosccpic analysis of the light passing the screens determine, definitely the quality of light entering each box. It is at once evident that the quantity or intensity of light behind filte«**««. placed at #*£W* equal distances from the source, ^ vari33,both because the intensity of light transmitted by the differ* ent screens is dif^erent,and because the different colors are not radiated by the arc with equal intensity .This being the case , dif- erences inrf results obtained behind the screens .could not be attri- buted to differences in the ^^^^^^^^^ __ energy of *& colored lights ift^the thermopile and sensitive gal
vSnometer., The thermopile is very sensitive to tne energy of
/\
any ether vibtations whether they be the longer infra-red or so- called heat rays, or the shorter, actinic rays^of the spectrum.
The method worked out for ^eliminating the intensity variable in the use of the filters employed for thtsr investigations, consisted in finding the distance «* from the light source at which each box with its colored window should be placed in order that the intensity^
as indicated by the deflection of the galvanometer when tho opeft-
the filter Qnd Of- the thermopile was exposed to the light behind & , was
in each case.
It seems necessary, on account of the questions which have been raisedjduring .he course of this work, to stsfe here that the thermopile is equally sensitive to the energy of r the red and^he vio- let ens of .he spectrum, and is, therefore, an accurate measure
of the total amount of light acting behind each color screen.
. ie true ueeauue lEhe difference between hent and light is only a
-r- ^^
ter of wave length. -i^. ±a truo thtct (the thermopile measures jj7 in
terms of the electric current produced by a difference in temperat- ure of the exposed and unexposed junctions; but it does so by vir- tue of the fact that the energy of whatever vibrations fall upon it be thtfy long and therefore heating in their physiological ef- fect or short and therefore only perceived ad light, is converted into heat energy upon being absorbed by the exposed junction of the thermopile. In other words, the ligh- of the blue end of the spec— 1rum produces an electromotive force much less than that of the infra- red but no less measurable.
The instrumetfnta ur td in the enejjy calibration of these screens were Si ariffin^T1 ™ + "™ table thermopile with junctions of bismuth and Hwrt^SSv and a moderately sensitive galvanometer (d'Arsonval) .^BsdaP electric arc similar to the one later used in * the experiments themselves nas the source of light* The thermopile -•** with the open end screened by on,e of ;the lighjt f J.^exp, was /
exposed to the light until the galvanometer indicator reached a maximum deflection which took, ordinarily^sbout thirty second^. The number of divisions through which the spot of light re-
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fleeted from the galvanometer mirror.was noted. This was repea't" ed six times and the average deflection recorded. The other filte^rs were then used in turn to screen the thermopile t and thermopile and screen' moved to such ^ distance 'rrom the arc that the displacement^ of the galvanometer &3&& indi- ^
cator opt*'* approximately equal. This -rdi3tan.c«'was ^.al.so j^^^y for the ui'icnjroGne.d thermopile^ which rerres en red
the removal of the control from the source. For the experiment the quantity of light used can be varied for the whole set ^ of screens by multiplying or dividing these distances by the same numbervandthe intensity infall the boxes will remain equal. The actual amount o£ lisht in candlepowers can be
measured by means of a photometer. Then from the law of in-
*
verse squares, viz., that the intensity of li<*ht per unit surface varies inversely as the aquare of t^ ft. distance from -B the source , the intensity at any distance from the arc can be computed.
r
The calibration oT~nTe\screens was repeated seven times or until satisfactory checks of the distances were obtained.
some thermopiles of less rapid action than the one used here, it «^/ in impossible to get results oy waiting for the galvanometer indicator to come to a steady fftate.In such a case the deflections produced by exposure to the ligh^'""' for equal intervals of time can be compared. A series of measurements f£om five second exposures agreed very well with those obtained by the other mejjthod,
TABLE SITC'VINr,
LIGHT FILTERS^)
?ROM AN
TKAlOaTT2DARB EQUAL.
Filter No. Wave- lengths Color
70 6600-^-7000 A.U. Red
71 6200-6800 A.U. Red
Distance from light 320 cm. 275 cm.
|
72 |
5900-6200 |
Orange 23( |
|
73 |
5600-5900 " |
Yellow 25( |
|
74 |
5200-5609 " |
Green 28( |
|
75 |
4700-5200 " |
Blue 25( |
|
76 |
4000-4700 " |
Violet 25( |
|
\ |
||
|
Control |
,'JTe. 34( |
;e the same" effect it migTTE oe tne resort or a general inten- iy gradiant; if only the blue ligh<; is effective, the problem one of a chemical effect; if only the red it might be assumed
in. TROPIRTIC REACTIONS TO MONOCHROMATIC LIGHT IN FUCUS
SPORELINGS.
The power of external factors to determine the polarity of a germinating spore is, without doubt, the power to orient the spin- dle of the first dividing nucleus, if that polarity is established by the direction of tne first cleavage plane. The work on such
orientations is very, limitedjaml has often resulted in negative resuWs
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^kjjito<-JT~-\ ^2-c*u) » woril^U**^ o» frgflS'9' eggs
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-omoryo QJ subjecting *aasQ.o pressure. The spindle parallel to the flattened sides of the egg. This is consistent with Hertwig's theory that the spindle. should form pareifcel to the longer aocis ofl a iding cell. It is conceivable that the power of light to orient <*- tfe* spindle might result from its power to cause more rapid growth
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along one axis, with tho rtfeult thayche elongated celljin turn * orients the spindle® No such effect of light has been demonstrated
with animal eg,^s? but in certain pi an ts . vi z . , Equi se turn , Fucus .Pi c-
¥o^v^f( C/i<-ocxxw<xOc »*
fystoseira,, it has been found that one-sided 'A
illumination with daylight causes the first cleavage plane to be formed perpendicular to the direction of the incident light and the cell on the darker side of tne spore to become the rhizoidal cell. Equal illumination on all sides retards or prevents germination. Gravity add contact cannot establish the polarity of these spores.
All these experiments were conducted witr natural light as the source of illumination. The first purpose of the present investi- gation ?/as to determine tne ower of monochromatic light to es- tablish the the polarity of germinating spores of Fucus evanescens.
It was hoped that the results o4 such experiments might give a clue
Y> W
to the mechanics of aZuA orientations. If all wave lengths should pre-
s\
duce the same effect it might be the result of a general inten- sity gradient; if only the blue ligho is effective, the problem is one of a chemical effect; if only the red it might DC assumed th*"t~
the orientation is due to a metabolism gradiant. The second pur- pose qC&^a study of the photro*ropism of the young rhizoids in monochromatic light. It has been shown (^T^jft that they are neg- atively heliotropicjbut the apparatus designed for the polarity experiments made it very easy to answer^, several questions concerning
tnyfr phenomenon wore definitely. These questions are:(j) What wave- lengchs are responsible for the turning away from a source of white
lightt (2) What is the role of intensity of the light or is it a matter of quality alone or both? (3) Do all lights which have any efiect at all produce the same negr tive tropism profluced by white ^ light?
The f irat work establishing the nov: km I fact that one- -
sided illumination will cause the first division plane of a germinating
J
spore to be perpendicular to the direction of the incident light s done by Stahl(/f? $"*) on Equi fee turn. He found th?<t the first wall
is formed perpendicular to •£& light rays striking the spores on one side only, and that if ? 11 sides ore illuminated by rotating "
spore? on a clinostat , the formation of ine vrall is retarded or prevented. The cell on tne shaded side of tne s -ore become? the rhi - zoidalfcell. in darkness the formation of the first v/all follov/s no rule and the rhizoids extend 4n every di ection. Stahl refers to
enrlier v/ork on liar s i 1 ia and n ara. whicn indicates /I that gravity
a is ton. controlling factor in the otientation of the first division
A
plane
, spore regardless of the light direction. SK ^Je conclude^ that no$T
A only light but a difference in the concentration of oxygen on the
two Fides of the spores could determine their pol; rity. He says that as a result of their respiration tne water in the center of the groups of scores is lesn rich in oxygenjand as a consequence the rhizoids^p. are formed on th?>t side. Inlsupport of this theory is the fact th» ':. although light can determine the polnrity of all
Rosenvinge(1889) showed that in Fucus spiralia there is no
relation between gravity and the first division plane nor did con-
. tacf with a solid body have any effect. He got tne same orientation
- to light in Ascophylium and ./ucrus' that Stahl did with Bfruisetum.
but with puzzling exceptions. There the spores were in groups the
^k_ /T_x£jv«-^W «-— *^
cell/ toward the -4e&r, interior of the Gr°up became; and0 in the lov/er part of hanging drops the rhizoids appeaWon the upper side of the
/f
tne specie^- led excet> Vucua serratus ;viz. .Agcojohyllum nodosum. Fucus vesciculosus, J<\ soirniis, n/.d Pexvetia comiii culrit.p , their sensibility to light differs and the oxygen factor or internal caue~
es t&tjS&Bfe?/ produce frequent exceptions in all but Pelvetia. The rhizoids of the latter species are alwa/y sf oraed on the darker
side of the spore, and this is the one species in which tne egg is surrounded by an oogonial wall which might prevent any of he
effects of varying oxygen concentrntion^which can net more potent- ly thsn light on the spores of the other species. Rosenvinge quotes
Kny as finding that neither light, gravity, ijor contract c?t( influ- ence the point of origin of the pollen tube from pollen grains,
but that in tne neighborhood of other grains tne tubelwill be sent out from t,-.'.e side away from them, on which side the supply of
oitygen or nutritive elements would be greater.
-iS^SHSV, farmer and Will iaias ( iS'fv ) state/ that if Fucus spores
/
are illuminated on all sides they tend to remain spherical in%tead of producing B rhizoid bu t<:e elongation of one of the two cells. Again(l898) they experimented with one-sided illumination with the usual result that most of .he rhizoids originated on trie shaded side of the spore and the others *fere turned that Iway. The fact that some graw out at an angle to tne incident light «4s attributed to "the character of the egg itself".
¥inkler(#//1900) found the some orienting effect of light on tne spores of Cy s t o s e i r a __b ar b a ta but failed to find any dffect of
a difference in the oxygen content of the -vater. He too
A
that gravity and contact are not factors in the establishment of polarity of the sporelings. r.e found that the direction is determin- ed before the firs', division takes place. Since t is polarity is established during the firstjfour hours of illumination and connot changed afterwards by any change in the direction of the incident light, he concludes that light can orient the spore only during fertilization.
- •
Randolph and I:^rce(l905) performed *m£ one-sided illumination experiments on Sicbyota. JJictyooteris. Lanrencia, and Gystoseira, and poinl^out the certainty of the j^/diium-jy action of other factor
because rhizoids are formed in .he dark and in all-sided illumin- •i
ation. They say that although Y/inkler (1900) suggest^ tne possibility of stopping germination toy changing the direction of lign-; every three hours, i ; could not be done with Sictyopteris. They emphasized^ theroossibility of influences preceding the illumination affecting tr:e polarity.
The work of Fronrne'Rev. 1914) on the urediniospores of Puccinia
rhamnif shauid oa men-. ioned horo/. He *&* that in darkness the
^ U
germ tube grew from any side^oT the spore, but that in inilateral
e tubes almost always issued from the darker side of the a* spore.
of.
the first cleavage icus scores. germinated in u**Tlateral have failed so far
™^L^' _^^ A
All attempts to solve thi groblera as to wh^tr"wave lengths of
;ht are responsible lor the orientation
\ plane o
^—^ ^
and apparently -•aTnvalsucceed^fith the • pparatus described
in the first port of th^>»^pape>>*41owever it seems worth while to report t-e metlpyrfs used and why theyhmte failed. T7ork v/ill be continued on^fenis investigation at the earliest o*pXjrtunity, PS it is mereiar a matter of obtaining the right conditions of "gmjwth
bejtffnd the light filters, together with a sufficiently strong
illumination.
The fruiting plants of Fucus eyanescens were collected at Sausilito at low tide of one day and kept over night in damp news1*'
papers. The next mornibg they were dried slightly by exposing them to the air for about half an hour or less. The fruiting tips were then cut off a'^ submerged in *ta- v/ater in the culture dishes. After fifteen minutes many eggsjand sperms have settled to the bottom of the dish or can be scraped 6ff into the water. The piece of plant is
then removedVnd the dish is placed in the rack which fits into a dark box behind a filter screen. Each box with its cultures
is placed at the proper distance from the light so that each is il luminated with equal intensity. These distances for each
the raethod by which they -'ere obtfii- ed^
by five as the case might be
distance given tnei? however was divided by four or
; / *\
in order to increase the intensity of the illumination equally
for all. The illumination with the electric arc was continued
V— ^
for eight in an otherwise darkened room, the carbons being ad-
justed every five minutes. This time was judged more than sufficient
M*4
to produce **»jseffect since it was found in Cystoseira (
that the polarity of the egg is so firmly established by four hours
exposure to daylight that it cannot be changed.
/ The f<ct that Ku cuy^evan es c ens is a monoecious specita makes it impossible to tell the exact time of fertilization, £• but it occurs soon after the eggs escape from the oogonial sac into the water. The sperms at this time can ba seen escaping fro-n the antheridia and swimming rapio.i./ around the eggs, then scattering as, presumably, one of than succeeds in entering. The first oross-wp.li can "03 seen very plainly twenty- four hours after the cultures are started. The mucilage accompanying the eggs caused them to aether e so firmly to the distil that it is not necessary to use f sol^d media to ke p the sporelings from being displaced when the
cultures are moved to the microscope sjage for examination.
However «U-Hha eggs germinated behind the color screens^
and illuminated for the first eight hours after fertilization.
-
showed 4«r orientation with respect to the direction of the- light. They grew 'corually as if in darkness with the direction of the clear - age plane following no rule and the rhizoids extending in every dir- ection. Evidently the intensity of the light was too low to have any effect, or the time or duration of the stimulation was not right.
Attempts to increase the intensity by bringing the cultures closer
that is. within about seventy-five centimeters »
to the wrcTfre suited invar iably in the death of the spores Before any
A placed (5"cm. from the naked arc »
development occured. A cooling device^ consisting of a layer of water
A
(p cm. thick between two glass plates, reduced the temperature from 29 to v*s*^y-+*^=^fa^^\\\\\'r^,^f^iFrjrtp^/ but al so Ar educed che intensity of "#Te light that no orientation was obtained. Attempt* to use direct sunlight as the source of light also caused the rapid death of the eggs, presumably on account of ',he high temperature accompanying it.
Future work on this puoblem auafr therefore involve the use
of stronger sources of illumination ajdn cooling devices to
"-a an ^
/r
keep :;he temperature below the death point.
interesting results were obtained in an experiment to dis-
£#6^4+ cover the weakest stimulus whicu would produce the charact4ristic
A
orientations of natural illumination. Cultures were placed at in- tervals of 15 cm. froia an east window a^n exposed 8 hours on a cloudy day> Upon examination a few days later every culture was-te
found to show strikingly the strong orienting effect of unilateral in
daylight. Even the one fartherest away, 350 cm. from the window^ were practically all oriented
c^~«4«j,* "l^more'dfr lets^/ r._ivery Culture js nowing this orientaion has.f requent ex-
ceptions to the rule. Every worker on this problem has reported such exfceptiens and they toave been ex*~plained by the assumption of the existence of an inherent polarity which as a rule i§ over- come by the stronger light stimulus. The fact that in absolute -fr darkness, germination and normal growth are as rapid or more so than in light, also points 60 an inherent polarity. But the tend- ancy varies enormously in individual spores. Therefore it is easy to understand that slight variations in the quantity and quality of illumination might easily produce'-b*^ changes in the ] sensitiveness of the spore toward light, which ;eans failure of experiments with monochromatic light until the right combination of other external £ factors in produced.
It was^oon discovered in the course of this investigation that although the WJLtf intensity of the electric are. used was too low to orient the cleavage plane of the egg and too low to cause the cell away from the ligh , to become the rhizoidal cell, it was strong enough to produce a conspicuous neeative heliotropism of the rhizoids of these same spores if the illumination was resumed after the rhizoids fead develop d- OKly seven hour exposures were used to
obtain this effect. On th^day after s-ich illumination examin-
ation showed that the rhizoids of th_; cultures behind t
sharply blue and violet filters were all turned away from the light,
tff/ltr
behind -3d*- t.1'.^ filters continued in the direction in which the£
jtAAJSs™ £*SQ -4^£~~tt—
had started, .anqpoaring no different thair-&aa control in dark-
*
ness. Therefore the wave IS-ngths responsible for the neg-
ative phototroplsm ffi////jiin white lighyare those of the blue end of the spectrum. It should be remembered that -Hi there is no question of a difference in intensity entering here because each screen was placed at such a distance from the arc that the quantity of light in each box qas the same. These re- sults are summarized in the following table
.. KELIOTROPISM
Filter^ COLOR TAV^ L^CGTH? DISTANCE FROM , LIGHT CULTUPES NO. I + J
70 red 6600-7JOQ A.U. T& <~™ .
71 orange 62^0-6800 ' bff - orange 5900-6200 -rt -
yellow 5600-5900 fe>2. -
green 5200-5600 70
blue 4800-5200 (* %• "
violet 4000-4700 •* " + -f
white tfo<Jo -7000 -/-
^L.
T Ke s e
A Dlmilar experiment was triedVrth sunligTit; as" the source
of illumination. The young plants were exposed behind the fil- ters all day in a south window. The same results were obtained as when the arc was used. Then the experiment w :s repeatedwith diffused light. The boxes were placed in an east window fir eight
hours_ --- 10 A.M. to 6 P.M. --- on March 28. Again the rhizoids
/i^txz^-d-vt/e-*—- in the blue and violet light showed the frrMffiisrii, but in addition
a considerable but much smaller portion ware affected in the same
jP • J // fJui. c+Ts*s*h£<^^> f£ <*s<-*-<^££ -*
behind the green filter. &******«[< , £->J"Jr<7L /' /• L ^JLtZ^^eu^^*^ •+*&* **^J^*
waj
These experiments 'ndicate that the light of wave length
*
4ooo-520CWImgstrom yUnitaVf^ responsible for the phenomenon,
but that some rhizoids, more sensitive, will respond to those of
5200-5600. It seems very possible in view of the results on
irolvox reported in the third part of this parer that with a^greater ^ intensity, the rays of the red end of the spectrum might caWse the sane negative phototropism. •*&?
Only the growing tips of the Jifrhizofds are sensitive to light^, mhis results in a sharp angular turn if the direction of illumination is changed through 90 or 180 degrees, or if the plants be brought from darkness into unilateral light. As pointed out by Loeb and others tropismJla uncftubljunliy due to the di ference in the speed of the
chemical reactions going in the two dides of the growing tip.
The first protuberance of the germinating spore is not affected by light striking it from th'i side; and if it is so illuminated during the early stage of elongation of this cell, the first bend occurs at the cross wall separating it from the next rhizoidal cell. In
s~*
other words, the firs t rhizoidal /^elongation continues in the orig- inal direction established by the first cleavage plane. and the wall separating it from the next cell of the growing ehizoid appears as the axis of the tropism.
In every culture of Pucus evanescens whether germinated in darkness or in strong unilateral light a most striking orientation of the first cross-wall with reference to adjacent spores appears, ^her- e-teer a group of spores are lying within about 0.2 mm. of each other, the first cleavage plane is perpendicular to the direction of the cen-
t^er of the group. THe cell toward the interior invariably becomes
C )
the rhizcidal cell. This phenomenon was reported by Fosenvinge in
--"
other species of Fucus and in Ascophyllum.
^
A study of this phenomenon ^pfs madeM'to determine the strength ,
compared to *^'**^'
of the stimulus producing this effect, *«*- that of light. in
A
A
orientation lu lh ) lUnaaliou of itc rayo.lt was at once very
evident that for most spores the formal? provaiirs when the spores are within a short distance of eachf other --- .2 mm. or often more --- and beyond this distance, the chemical^) stimulus becomes too weak and only the light is able to determine the polarity of the plant. Only the comparatively isolated spores therefore ever show the orient- ation to light ;i-rrj )i I'JL I'H-iml » with the^ource^ of illumination uned
here.
The phenomenon is V3ry conspicuous ia groups o"' 2, 3, or 4
eggs as well as in masses of fifty or a hundred.. In these 1 ,rge ,-roups it is made evident by the invariable rtile that no ;%fl/^/rhizoid ever
extends outwards from a group. Then two spores are within the distance
which through the stimulus is effective, the first cleavage planes of the two
are often parallel and the rhizoids grow towards each other and often meet tip to tip. The groups of five or six often make symmetrical star-like designs when the rhizoids have grown and p/oject beyond the group.
""he spores are more rarely affected in thas way when the distano^ between them is over*>.3 mm^but the phenomenon is sometimes observed in spores as much as O.5 mm. apart. Tithin a distance ofo.2 mir.. there are practically no exceptions.
^he relative sensitiveness of a sj ore towards light and towards this cherr.ical( ? ) stimulus varies greatly for different spores. "'hen cultutes were placed in the window to get as strong a light stimulus as possible in order to determine at what distance from each other the eggs had to be not to show a greater sensitive- ness towards/the chemical stimulus than towards the light, it was found that^this distance followed no rule, the scores showing the greatest in- dividual differences. Of two spores lying within 0.3 mm. of each other one might be ^entirely oriented by the adjacent sporephile the otherj apparently like it, Would show only the action of the light stimulus.
In many canes two such spores wouljl see- to show a rss-iltant effect of the two stimuli so that both -YOU Id be half turned towards each other with fe£*VhlzOidal ceil« showing a^tcTtake -a direction away from the -
•££oK- iv*~-* • —
light at » resultant angle.
T osenvinge ascribes this group orientation to a difference
d>n the concentration of oxygen or of nutritive substances on the two
forms on th > side sides of the spore. He thinks the rhizoid^i'/ 4,/V/1/ toward
of a group or towards another egg because the water on that side is less rich" than on the outer side of the spores as a result o,' their
/\ £**u^A.
metabolism, '"inkier (^ ) working with Cystoaeira barbata^that a dif- ference in oxygen concentration ha? no such effect. Apparently the phe- nomenon does not occur naturally in)thisjspecies since Wwr figure given shows nothing but the effect of light. I have nevor seen a culture, of Fucus evanescenjlth spOres germinating so near each other, which s! owed only light orientation and ::ot the group orientation. aAl- raost invariably when the spores- of this species germinate in such close proximity, lipht appears to have no power to establish the po- larity of the plant.
The possibility that the group orientation is due to a pol- arity established by the position of the egg in the oogoniura is
tut*** suggested by finding many groups of lying just as they escaped fror-
the oogonial sac and conspicuously oriented with respe?t to each other. The fact that groups of ten or of two are as regularly oriented, would refute thT suggestion; but in order to prove that the phenomenon is the resuljr of a stimulus acting after the eggs leave the oogonium, a group of them were transferred to a watch crys- tal and mixed with the point of a needle until their relative posi-fc-
"/ions were entirely changed. But when they germinated the character- istic orientation with respect to each other was found to be with- out an exception.
mhe only apparent explanation of the group orientation is '.hat of a diffusion gradiant of some substance emanati^ng from a growing spore, or of some substance being used up by it. A continuation of the inve/stigati ^n of this problem will b3 an attempt to dis- cover a substance or substances which can so effect the dividing nucleus of the egg cell that its unequal distribution on *M sides of the cell will orient the qxis of the spindle, The effect of bub- bling carbon dioxide or_ oxygen through one end of a culture will be tried as being the most probable factor^ involved.
A striking phenomenon which seems to be further evidence of the power of a chemical stimulus to orient the spores was found in three different cultures. ?.mall pieces of a foreign substance, apparently sand, caused the orientation of the spores adjacent to •astey'ith the result that the rhizoids all extended inward towards t: e substance. This material was present in too small amounts for chemical analysis. A search for some difficultly soluble substance, or an insoluble substance which might be sligBiJfly soluble in the weak carbonic acid produced by the growing spores, which, when added to the cultures , might orient the eggs adjacent to it, has failed to brin* any positive results. They do show ^however, that the phenom- enon is not due to a contact stimulus. "Ehe^//^ substances tried W3repcheelite/ Ca^o^Oj; 'Magnetite/ FQj0J, Ba ite/Ba^So), Casser-ite
n oAnornblend, tfubellite, beach sand, granite, and silve~r filings. The substance or condition originating in the activity ofacfjacenf
spores which has so powerful an effect in orientating the first plane and in determining which cell shall become the
A
zoidal cell has no power to cause any chemotropism of the rhizoids
after they are started. No rhizoid has been found to have its direct- Ti on modified by the presence of other spores adjacent to it. In the absence of any light s$imulus)the rhizoids continue in the direc - tion that they take priginally from the spore.
-i-Ct^Yrt/ma/W
TO summarize briefl" '. .is work on light and chemical stimu li as they affect the polarity of ^ucus e .d the direction of growth of the rhizoida:
(1) Green, blue, 8$rt violet "-ight (4000-5609 A.n.) of an inten- sity great enough to produce negative phototropisms of the growing rhizoids is without effect on the direction of the first cleavage f plane of the egg.
3
?N > J
chemical stimulus which orients the direction of the
first cleavage plane and determines which cell shall become ths
rhizoidal cell
alli
diet
vhemi
' ~-' hrig no power to •^s&r,. c cf
rhizoids .
(£)Th laxative photctropism or the rhizoicr : . *i .^o-
.«. ~ c ' _ . '. .' , . "^ " .a_. ^ 13
since, with equal intensity of illumin; ." ;ht
• : peel . Lthoti ^
j -
—244.
V. -- Pho tot rop isms ci Volvox in Mono chro 'na tic Light of Equal Intensity.
Oltraanns ( /?<fR ) , Holmes ( J<p3 ) and Mast have reported in detail the reactions of Volvox to li^t stimuli. No one, however, has studi- ed its response to monocnroraatic illumination. The purpcs of the present study is a comparison of its phototropisrne in different spectral re -ions for which the intensity is made equal.
The recent article by Mast (l?'7) suns up in a table the litera- ture on the reactions of plants and ini^als to colored light so cc - pletely and so concisely that there is no necessity for a repetition here. ite summarizes briefly results jiven in the table as follows:
"For seedlings <• f gr 5 »n plants, plumules and radicles, the region in the spectrum of maximum stimulating effect ia in the blue or violet. For the fungi it is somewhat nearer the red. For Bac- terium pho tour? tri cum it is in the infra-red and the orange. For Oscillaria and Piraiaeoium bursaria is it questionable, activity and aggregation being probably determined by cuemioal changes in the solu- tion associated with the colors. For Chlp.mydomonas it is in the green; for all r ther unicellular forms tested it is in the blue, as it is also for the co^l^nterates and vermes and for a few of the molluscs an.-"1 Arthropods. >ut for most of the molluscs and arthropods it apcears to be in the ,^re ^n or yellow."
T -)t tb ? -:if<st effective wave lengths -ir*3 not the erii.ie for even related forms in the low«r organisms iB furt)i«r borne cut by lias Us ( l^l"[) results. ue found that the maximum efficiency is near 483 for 3uglena, Tra^relomonas, Phacus, Gonium, Arenicola and Lumbric.s;
—25--
near 524 /y for Pandorina, Eudorina and Spondylonio ruin; and near 503u./^- for Chlamydoinonas and olcwfly larvae. The wor? of Schaelferf^ ) on amaebae 3 i iteresting as his results indicate that the prw»r cf a beam of red light to cause the organisms to move towards it is as great as a beam cf white light, and hae mere attraction than the blu»- Most investigators of phc to tropisms find either that the red is less effective than the blue or that it has nc effect. A few find that in equal brii'Jitness, tha effect is nearly the same for all colors, e Volvox colonies upon which the following experiments w~r made wre collected by Dr. N. L. Gardner in a small la>e in Golden Gate Park on April 2, 1918. They we*e so abundant that the water was quite :Tr?«n vrith them.
On April 5 they were exposed to direct nunlifiht behind the seven Vfrat '.en litfit filters used in the work on iucus. The same rectangular, flat-aided dishes made of microscope slides cut and ce- mented together were used. The disfcss, filled with a Vclvox -us- pension were planed in the dark boxes in one end of erioh of wliioh a color sere ?n waa fitted. The series of eight boxes were tnen ex- posed to direct sunli.-jit. A: intervals the covers were raised to de- termine roughly the ^r^ntage of org'inisns reipondinst as shown by the completeness of thei c aggregation. Ths results of Uiis series are surnra-'xrized below, the plus ^i ,hs i •:! opting an obvious positive hello tro pi am with Uie org nip-in coll e -ted against the ^lass on the side toward the sun, a 2<=>ro indicnti".;r no ^vidence of any eftect of the light.
--26 —
Table No. 1.
Volvox Reactions to Sunlight behind Wratten Colored Filters.
Filter
70 71 72 73 74 75 76
Control. Table No. 2. 70 71 72 73 74 75 76
Control.
Wave Length in Angs t r o m un its.
- 7000
3~c[ OO — (o^OQ 5<o oo - y<?o°
4-700 -
4-066 47OO
H- o o o —
QO -
- (c^OO 5(0 00 —
s'Zoo - ^6,00
y-ooj
Lj-Odi) — JOOO
Color
y£ )
Sxposure
|
y ' |
10' |
%o |
|
T IT |
r IE |
r IT |
|
o.o |
-1- o |
f- ° |
|
o o |
0 0 |
-f o |
|
0 0 |
0 0 |
0 CP |
|
+ t |
4- -f- |
r t |
|
f- + |
4- -f |
t -f- |
|
i- t |
t + |
t t |
|
+ t |
t + |
4- t- |
|
+ t |
-h i |
t f |
t
|
3" ' |
jo |
i sT |
^ |
_ JO |
|
O |
o |
O |
o |
' |
|
0 |
<^ |
0 |
•f |
t |
|
O |
o |
o |
f |
t |
|
* |
* |
t- |
f- |
-f- |
|
t |
,h |
f |
t |
t |
|
-h |
f |
f- |
1- |
-j- |
|
-f- |
* |
^ |
t |
-h |
|
•f |
± |
f |
f- |
t- . |
Table 1 shows that wave lengths from 4000 to 4700 A. are de- cidedly more powerful in directing the movements of Volvox than are those of the remainder of the visible spectrum, 4700 to 7000 A. U. There is an indication, however, that some individuals respond to the long waves.
Table 2 shows that the stimulating efficiency of the difierent
—27--
is dependent on the duration of t;:e exposure. At the -^nd of only three minutes, the organisms of the gre«n, blue and violet were all over to the brighter side of the dish. At the end of 15 mi-iutes the red, orange and yellow had still oroduced no visible effect. But, after 20 minutes, all but the longest waves, 6600 to 7000 had caused a definite ag regation;and after thirty uinutes,' the effect was se^n benind this r^d screen.
As striding a? tnis relation of the time factor to the power of light stiumli to produce the phc totropisrn was the difference in the response behind the 'four filters of the green, blue, and violet. After only 3 minutes of exposure the aggregation effect was most marked in the violet, less in the blue and still less in the green. The difference in the appearance of the four disnes was due to the difference in the number of individuals that had responded positively and the completeness of their response, i.e. in 4-0 o-j — i+j o practically every colony -va« igainst the glass on the outer side of the dish; in A =. *f 9o - frffctf ^- A0 there were more scattered about in the water. In 5200 - 560^tf still more and in 560 - 590^
there w»re just barely enough affected to make it evident that some were positively «[z?«o%4'(l; After 10 minutes this gradiant was still visible. After about 15 minutes, the dishes all looked alike. Tnis again shows the importance of the time factor, since the less sensitive organisms which respond less rapidly are after a longer time afteoted to the sane decree.
In all the succeeding exposures with sunlight this rule held good, that the organisms behind 76snd75 respond most rapidly and most
--2; --
crrnpletely , followed "by those "behind 74 and then 73. It WT.S < f oen quite impos ible to detect • ny dit'terenoe in the response behind 75 and 76.
It was noticeable in every series exposed that in the -vhite light of the control complete aggregation of the colonies as the re- sult of their phototropism was no*- so rapid as in A=*tof-i^o /*-P- Repeatedly the observation was made that the control locked more like the culture in fr*1- VR» /+yt^ • T:.e expl.-unticn in prob-ibly
either that the intensity was fso raiich greater that some individuals were caused to respond negatively, or were neutral; or that the great- er reflection from the back of the dish ma e all parts of the wat»r more nearly ali>-e in intensity of illumination and so tended to equalize the Rtiraulus acting on the different sides cf the orginism.
When a series of cultures was »xpooed in the same way with the sun hidden by clouds no respcns -> could be obtained behind the red, cringe ^nd yellow filters.
Theae experiments srow the greater stimulating eiticiency of th e blue end nf the apectrura since comparison of the order of efficiency of the screens in produciri/-; the positive response is not in agreement with the order of the intensity of the light behind them, (see table 1 ) In fact the stimuli of least energy produce the most rapid response. They also show that with a longer exposure or more intense stimulus the less effective wave lengths may produce the s-ime degre "• of r spons e* In none of the exposures to sunlight were there any negative reactions or reversals of the response.
On April 6, the experiments ^«re repeated -vith the electric arc as the source of li ht, and with the dark boxes at such distances from it that the intensity behind all was the same. The results are
--2S--
as follows: Filter 70 71 72 73 74 75 76
Wave -^-'n.-
(.(.oa ~ "7 °0 &
b 1(1 O - & % 00
ac, - *>
Color
Distance
osure
AJL
CJ
+•
H y oo - 5 '
^ 0 6 0 ~ Lf -J u0
- -i000
ft.
r
r t
t
f-
t
1-
The ri.pid reBpcne-- in. t>m re'. v;s aurpriging sinoe the energy must have be-n leea '.han in sunlight. It took 20 minutes to get a noticeable reaction in -Jie orange and yellow in sunliriht the day befor Here with the arc as a source of light it took hut thr°e minutes. It toc-y 30 minutes to get the resporv e in the red that was produoed by the arc in 5 minutes. The explanation rni^ht be that the colonies were niore sensitive to light nn April 6. It was unfortu- nate that tjiis day vrae oloudy so that the experiment -*ith sunlight o uld not be repeated. That the ability of the colonies to re- spond is dependent to a large extent en their own condition as aflected by external factors of their invironment was shown striking- ly by the fact that organisims kept over night in a small mouth bottle -vould not respond to -my lirjit stimulus, while o th-TS cf the same collection ?ept in shallow pans reacted rapidly. Mast ( found considerable difficulty in ,;etting consistent responses on account of the gre t change in the reactions of . is organisms with Change in their con di lion due supposedly to the li, ht and temp ^r-s oonditons.
—30--
ism, Also it, J.B prob-'.ble in vi-^w-of jLoab^s -vor>- (1^6 ) that the phototropio reactions of all organisms change with their age.
Another explanation which was suggested by the fact that white light was often slower in producing a response than the b lue and violet, is that the greater energy of the red in sunlight might have tended to reverse the positive reaction or neutralize it; or it
ight h-iv* be<?n so reflected within the dish as to act more nearly with the optimum intensity on -<11 sides of the organism. •"» that as it mri.y, the fact remains that repeated trials prove Conclusively that at the end of a fiv- minute exposure, all wave lengths of the visible centrum hnve produced equal eft^ots when the source of li;Tht ic the electric arc. This is further proof of Mast's Wf ) contention tir.it: "There ia nc evidenc-- i-.-Mc ting that stimula- tion i-, n.:r; of the • 3 studied is independent of lurranous in-
tensity, for if the light in the spectrum on Cither side of the maxi- mum be made sufficiently intense it becomes iaore efi active than at the maximum, M
On April 20, another collection of Volvox was made. It was ho r;ed that a rough approximation of applicability of the Roscoe-
Buns>?n Law in the reactions of th is form U> li :ht. This law states
•ing that o'J<-r conditions "be/eqial, the product of the intensity and
the duration of the exposure Is .'? co .stant. Owing, however, to the short time required for response and the variations in the sensitive- ness of individuals making it impossible to determine the end points exactly no thin- c ,n be concluded o.s to the truth of this law for Volvox u Li] . nr? RQ urate ra^ans of ..leosuring the b 'ginning and end of the reactions is obtained. To show the diilic^lty produced by
—51 —
th« i :}- responses of the colonies, -..he tira" taken for most
individuals to ^rosa the dish, i. e. ^' .;a».. , in white li.'ht bO om. from the source in a straight line towards the aro was about 1 minute. ?n this distance was doubled, the time was still about
1 minute. When the c Iture was re::.cved 200 om. many c lonies still requii-ed rrily 1 rainu^e to get icroes Uie dish.
This effect of Changing the distance was repeated with the No. 70 filter, since as it transmits the longest wav~ lengths the time cf the reaction nhculd be longer. However, they were still too n.^.rt to obviate tho lar.je errors due to the indefinite end points. At a distance cf 25 cm. Lhere was a noticeable collection on tiie li(shter si 3 of the dish. After 13 minutes nearly all had moved over. At a distance nf 50 cm., ICO cm., :»nc 200 cm., the same positive reoulte v/ re c^ 'ith t>ie five minute exposure. Al-
though failing to si:ow the applicability of the Rosooe-Bunsen L w this exp^riniont r^.cws th<; potency of the long red rays to oause the same pro to tropisms produced by the blue waves when they aot with the right intensity.
~:ry exposure througiiouc, tli^-^e experiraents was made with a fresh culture BC t.-at in no inst nee was tiiere a ch.ince of the sensitiveness being ..ifior^nt for the succeaaive exposures because of the of -fact I -ceedint:; illumination.
• j (^9o3) reports that tiie positive p}:ototropism in
medium li'jht is reversed by strong light. Att-=>mpts w^ro made t): ere- f re to find the turning point for each screen by .lessening its is- tan*5e fro^i the aro. *t the control behind clear glass was the
only one in which any negative pho to t ro ;: i 3in could be obtained nnd the response M -re varied at different times. In one experiment, the
—32--
orgaii^ms reacted positively at 40 cm. from the aro, negatively at 20 cm. In another, the neutral point seamed to be 50 cm. away. Quite often Tit this distance there were nearly as raany colonies swimming away from as towards light. The response wis always more definitely positive at 100 om. than at 50 era. Sometimes, however, the response was mrstly positive as close as 10 om. , so that the re- versal is dependent to a large extent en the -cndition of the org'-n- isms as for the other screens, the greatest intensity obtainable be- hind any was too low to produce any negative renuonse.
An entire carbon aro run was projected on one dish of
organisms. By looVing down it could be seen immediately that while those cr£T.nir»"3 fin din selves in the yellow, ^reen, blue and
violet lip;ht swam LowirdH the litf-.t, these of the red waved not to- wards the front of the dish but in a direction por.^ii^i to it, i.e. towards the yellow. The yellow apr.eared to the eye the Brightest part of the dish. The organisms in it were dcubtless decidedly photo vronic boc-'i-se of the relatively greater intensity, while those of the blue were so in almost equal degree because cf the greater ef f ectiveneflr* of these wave Tr-«n;-:tr.8 even ^ t low intensity. In the red the intensify wna not gre t en< ugh U) overcome the ineffectiveness of the lon/3 r;iys, 30 tiie organisms responded either to an intensity gradiant whi oh led them towards the yellow, or to •> wave length gradiant which would also lead tnen toward the yellow. Or the r - suit might have be-=>n due to a diffusion of the more ei-:iri«nt rays into the red. The raraa red rayp in th<5 naVed aro exposure th«re was no -iefl -ntion of the pat!"! leading dire^t-ly toward? the source of light.
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It is interesting in this connection to note Holt and Lee's ) conclusions as to the vole of the intensity factor in phcto- tropis . They ntate that, "T'n^re is no evidence t/iat
•
respond to any other property cf li^ht than its intensity. All phototatio response in ^xplainea by the ii tensity of the 11,-^rt, and the direction fro;a wai ^Ji it -lornes.11 But my experiments 'Vith Volvox show oonolu'9ively t/iac for Any Driven intensity the blu3 and violet wave lengths tire --test stim luting effioienoy, but tliat by
increasin-; th-j duration of the exoosure the same effect is produced
Ln d tlie otli«r BCte^naJ and "by , aity of the
other colors they oan be made to produce an equally rapid re ponse. This is in in or a i . 1 -.c-'t's ( '^'7 ) Conclusions that photo-
tropio resp'-na-jB are net entirely in ependent of intensity.
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NON-CIRCULATING BOOK
783071
UNIVERSITY OF CALIFORNIA LIBRARY