—————— ne — —— i ——— as — SS SSeS —— a —— an I ——— oS a ————— -— —— eee ns — — oe — cette —= a ee ee — — —— _ a — a —— —= ————— ee SS SSSSSSSasassqm—n—nq—sSsSSSSSS ee —— — _ —— = — en a een ear —— ee S —————— ——— _ a ————— rere etennsg grrr ee OS nn ee —— ee = ——— = ee er pe ea = —+ oo —————————— = ——— = — oe Se 5 agen —— ee ———— ———— —————— ————— —— ee SS ee ee ——— ———— oe re —— = =— ; SS rn eS ——— ee ————— ———— ——— — oe — — : a rr ee ee —<- > a A ES A ——— —= - ~ — a -_——— ——— _ = a ene a) ————————— ————————— ——_— = -— ——————— es ie —————eee — ee — ———— — — == ——————— ne SS SSeeeesssasasasassSsSS5S5 5353535 —— ———————— ——————— ——————— —— ———————— oe ——— oe —————_— — = — eee —- ——— ee ee —— ————————— ——— —< — — a —— ———— ———— SS ———== oo ————————— ——— a ————— = ——= SSS ———— er : —————— re —— a ne ——————— ———— ——= ————————— ———— Se ———_ ———— ——-———— —— ———- ————— ————— So ———— = ee ————— —_—_— = ————— SS —————— SS SaaS ————— ee —————— —~ — —————————— —————— — eee —————— ; — = ——— i —— ——— SSS —— ———— Se ——— —— nn ae ——— ————— SS SS ——— ———————— ——— — —————— —————— ———— —————— SS ——-_ ———— —— ———— - ee a a ——-— ———— ——————— = —— —- — ———————— —————— SS ————. ————————————————— ————— —-. oo ——————————————oooeeeSS ———— eo —— oo SSS ——— : EE ———— ——— — ——— ——————— ————— eee ——— ————— or —— ————— — ——— ee ——— ———— ees 2SS55 = ———— ————— = = ————— See —— _s ——=— > A TS _- — ————— —_— SS — eo as ——— ————— ——— ——= ——————— = ———— oS ee ——— eee ————— —~ -— —_ a SSS ———aaeaeaeaes~=q—s— SSS ————— — — — — — — _— = - _ A ps — =. ——_-——— ee ————— so = — ——— - oye 257 Bud of syncytium from intervillous space ..=».....«+s+s «8 264 Chorionic villus and decidua serotina .!..... :../5....).¢3 eee 264 Section of chorionic villus at the fiitth week .. 2... 2. 2 9m 265 Section from sixth week pregnant uterus . .°.°- 4). . 6 sa) 268 Decidua serotina and chorionic villi at the sixth week. ...... 268 Sections of full-term chorionic villi. 2. jes. Wa eee 269 INTRODUCTION : first public lecture under the auspices of the Harvey Society was held at the New York Academy of Medicine on October 7th, 1905. The President of the Academy, Dr. Dana, addressed the audience, and was followed by President Graham Lusk, who spoke for the Harvey Society as follows: ‘‘Gentlemen of the Medical Profession of New York: On the behalf of the Harvey Society, I beg to thank President Dana for the kind words he has spoken concerning our under- taking, and through him to thank the Academy of Medicine for its patronage, so helpful to the success of our newly-formed society. ‘“‘Tt is perhaps fitting to describe the reasons which have led to the formation of the Harvey Society. ‘‘There exist to-day in New York two classes of medical societies. The first class is devoted to clinical work, and has existed since the foundation of the town. The other class is employed in research along the lines of experimental medicine. The societies of the latter class are of recent origin and' may need a word of introduction. Seven years ago the Society of Physiological Chemists was established and still affords oppor- tunity for the discussion of problems in an ever widening field. The Harvey Society is indebted to this sister association for a goodly subsecription—the total amount of its surplus funds. The Society for Experimental Biology and Medicine was founded three years ago through the influence of Dr. S. J. Meltzer. Members of this Society must be workers along experimental lines. It is evident that the societies having clini- cal discussions for their aim must of necessity be of different character from those whose objects are exclusively experimental or theoretical. But it is also true that knowledge of the latter sort, if presented in a broad and liberal spirit, may be made of value to the members of all classes of the medical fraternity. 9 10 INTRODUCTION ‘‘Having in mind this idea of creating a common meeting ground, some of the laboratory workers in New York came together last spring and decided upon a new experiment. ‘This experiment is the Harvey Society, founded for the diffusion of knowledge of the medical sciences through the medium of public lectures by men who are workers in the subjects pre- sented. Dr. Theobald Smith has expressed our thought as regards the character of these lectures. He writes, ‘If I under- stand the scope of the society rightly, such a lecture should be prepared with great care, from the broadest and most advanced outlook, while being at the same time purely instructive.’ ‘‘In Berlin and in Paris men of science give public lectures with the wish of serving those busy with the art and practice of their profession. The Harvey Society is of this spirit. We would welcome the support of the notable audience of the medical profession of New York.’’ THE THEORY OF NARCOSIS * HANS MEYER, M.D., Professor of Pharmacology, University of Vienna. ESTEEM it a high honor as well as a great pleasure to address you on this occasion and to have been asked to open the first course of lectures of the Harvey Society. The estab- lishment of these lectures is additional evidence, if such be required, of the great interest displayed by American physicians in theoretical conceptions and in scientific research, and, in addition, of an earnest effort to encourage and diffuse such knowledge. I look, however, on the invitation extended to me as evidence of your friendly and fraternal sentiments toward the whole body of German scientists, and I may be permitted on their behalf to offer you an expression of their cordial appreciation. Following the suggestion of your president, I have selected as a theme for this evening’s lecture a subject which, for a long time, has repeatedly attracted and interested not only pharma- ecologists, but biologists as well, namely, the relationship between the pharmacologic action of a drug and its recognized chemical or physical properties. The solution of this problem presents great difficulties. Even with a knowledge of the chemical and physical properties of an active substance it is yet impossible, without further knowledge, to determine which of these prop- erties is responsible for the specific action on the animal organ- ism. And this the more so, since we do not know the chemical point of attack in the organism, and hence can not know the nature of the chemical reactions which occur between poison and protoplasm. Only in one way can we reach a conclusion that admits of probability. If we find a large series of different * Lecture delivered October 7th, 1905. 11 12 HARVEY SOCIETY substances with different chemical and physical properties, which all possess identical or else very similar pharmacologic actions, then we can fix on the chemical and physical properties common to all and on which the common pharmacologic action naturally depends. So, for example, out of a mass of keys different yet all able to open the same lock we can determine which part of each key is the essential one, what form common to all fits the lock. And it is hardly necessary to point out that from this we can obtain an insight into the construction of the lock itself. In our case this means an insight into the chemical organization of protoplasm. The first experiment in this direction was made by two English investigators—Crum-Brown and Fraser. They discov- ered the notable fact that practically all the ammonium bases, that is, organic bases in which the pentavalent nitrogen is connected with four valencies to carbon, exercise the same pharmacologic action, regardless of other differences in their constitution and nature; the action in this case being the same as that of curare, a paralysis of motor nerves. This definite relationship has been confirmed by many investigators, but the explanation for it is still lacking. It appears to me possible that the strongly basic character of all these ammonium bases is the chief factor. They are much more strongly basic than alkaloids and even sodium and potassium, and the few excep- tions, such as betain, antipyrin and others which do not possess this strongly basic character, are also without the curare-like action. Another series of experiments along this same direction, in connection with the action of the neutral alkali salts, has been conducted by Hofmeister. He has shown that all the effects of these salts, including their laxative and diuretic actions, may be explained by their physical properties, their diffusibility and osmotic strength. This brings us to a third especially large group of substances whose actions are all identical in principle. I refer to those substances which are commonly designated anesthetics. To this group belong bodies quite distinct from each other chemically ; THE THEORY OF NARCOSIS 13 aleohols, aldehydes, ketones, esters, ethers and numberless others. They all possess the common action of depressing the cen- tral nervous system. Wherein is the relationship? On which of their common properties is their narcotizing action dependent ? The chemical composition of the nervous system itself gives the first clue for an understanding. It differs from all other tissues in its richness in fat-like constituents, and on the basis of this peculiarity Bibra and Harles attempted many years ago to explain the action of anesthetics. They found that the anes- theties dissolved ordinary fat, and, as a result of a quantitative estimation of the fat content of the brain of a normal and nareotized animal, assumed that the anesthetics directly removed fat-like substances from the brain. But these con- clusions could not be confirmed. From another standpoint, however, Hermann arrived at a similar opinion of the action of narcotics. Hermann had discovered the presence of lecithin in the red blood cells, and since the anesthetics, such as ether, chloroform, ete., dissolve the blood cells, he explained this by their power of dissolving lecithin, and pointed out the paral- lelism between this process and the narcosis of the central nervous system. In both of these hypotheses there appeared to me to be an element of truth, and in order to establish this and define its character I myself instituted a series of experiments. I started with the following assumption: If fat-solubility is indeed a necessary condition for narcotic action it is to be expected that all indifferent, fat-soluble substances must act as narcotics if they can enter the cells, and that, on the other hand, if by any circumstance they lose their fat-soluble property, then they must also become inactive. I have tested this assumption by investigating a series of substances made by combining components which in themselves had no narcotic action, but whose combinations were soluble in fat. As examples may be mentioned the amides of organic acids. These amides are neutral compounds which are soluble in fat and which all possess the typical narcotic action, with, however, one single exception, carbamide, and this particular one is insoluble in 14 HARVEY SOCIETY fat. Another group (comparable to the amides) is composed of condensation products of glycerin; the chlorhydrins, acetins and glycerin-ether. These also are soluble in fat and act as narcotics. All these substances, however, are very readily split up by hydrolysis into components insoluble in fat and on such decomposition lose immediately their narcotic power.* Now, if from these and similar observations the conclusion can be drawn that the solubility of an anesthetic in fat is cer- tainly one of the conditions for narcosis, the further question presents itself whether this condition is an essential one and whether it can be utilized as a measure of narcotic power. Were this the case, a quantitative relationship between narcotic power and solubility in fat must exist. But it is obvious that other factors must influence the action of narcotics in the animal body, for their affinity for the watery components of the body, as well as that for the fat-like constituents, must be considered. According to their relative solubility in the fat-like and non- fat-like constituents of the body they will distribute themselves between those constituents. So, for example, such substances as are very little soluble in water will dissolve for the most part in the fat-like constituents. Richet had previously made the observation that the anes- thetics which are little soluble in water possess a marked narcotic action, and he regarded this relationship as a general law. Asa matter of fact, however, from this single relationship no general law can be deduced, for substances such as alcohol and chloral, which are both equally soluble in water, possess very different narcotic powers. The proper expression of the law must, therefore, be that the distribution relationship, the so-called distribution coefficient, of the narcotics between fatty and watery solutions, is the determining factor of narcotic action. To test the correctness of this hypothetical law I have determined the strength of action of a large number of different narcotic poisons by estimating the smallest molecular concen- 1The amides yield fatty acids and ammonia salts, the glycerin derivatives, glycerin and acetic or hydrochloric acids. THE THEORY OF NARCOSIS 15 tration of their solutions which was sufficient to induce narcosis of small fish and tadpoles placed in them. A second series of experiments was then carried out with the same substances for the purpose of determining their distribution coefficient between water and fat. A mixture of water and oil was used to estimate the relative quantities passing into these two substances. The comparison between the distribution coefficient so obtained and the narcotie strength of the narcotics did, as a matter of fact, yield the expected result, as a glance at the following table reveals: TABLE SHOWING THE RELATIONSHIP BETWEEN THE DISTRIBUTION Co- EFFICIENTS F/W AND THE CONCENTRATIONS, EXPRESSED IN GRAM MOLECULES, OF SOLUTIONS ExEerRTING EquaL Narcotic ACTION. F/W Concentration RE Oaeo.c. Prac dsb. hada o o caouote'a 4.4 0.0013 NRE, oobi 4s oid aie. G4 ee do ee wee 4.0 0.0018 Se ee eer mre ca 1.6 0.002 EE ate a oc. nd: 9 6 Pete has cA 0.006? ONIN bc aha wid kc va ae pis see oe 0.7 0.002 ES CN FO gh ae ee ee ge Py 0.6 0.002 MRC Gre BEG or tt he og as a ah 0.3 0.010 OS Se Bete eo ee Cree er 0.23 0.015 SMR MAAEM 5c toa sc bigs Xo six oie 9 2 0.22 0.025 ny RCN ha Se Fite vps Sees ss 0.14 0.025 Nn eRe Sears eae oy ee wer as 0.06? 0.02? Ee ee ae 0.04 0.4 ST UNND. So's aie thee Ga oo s ue 0.03 0.5 With an increase in the distribution coefficient there occurs an almost parallel increase in the narcotic strength, that is, decrease in the molecular concentration necessary for narcosis. The few departures from the general rule which occur in the table can be explained by the naturally inexact method of estimating the narcotic power. A further proof of the correctness of the view described may be offered. It is known that the solubility of most sub- stances in water and fat changes in a different way with variations in temperature. The distribution coefficient is also variable according to temperature. It must be expected, then, that the narcotic strength as well will vary with changes in 16 HARVEY SOCIETY temperature. And this is the fact. I examined six substances, of which with higher temperature three gave higher and three lower distribution coefficients. And it was found that in exact accordance with the rise or fall of the distribution coefficient so the narcotic strength rose or fell, so that tadpoles which were just narcotized by a certain chloralhydrate solution at 30 degrees C. were aroused and quite active on cooling to 3 degrees, and on subsequently warming to from 25 to 30 degrees again passed into narcosis. From this the direct dependence of narcosis on the physical relationship of the narcotie to the fat-like substances, the lipoids of the body, and the watery constituents seems to be definitely proved. As a result of all these studies we arrive at the following explanation. of narcosis: The narcotizing substance enters into a loose physico-chemical combination with the vitally important lpoids of the cell, perhaps with the lecithin, and in so doing changes their normal relationship to the other cell constituents, through which an inhibition of the entire cell chemism results. It also becomes evident that the narcosis immediately disap- pears as soon as the loose, reversible combination, which is dependent on the solution tension, breaks up. It follows further that substances chemically absolutely indifferent, as the volatile saturated hydrocarbons, can act as narcotics. Quite in opposition to this idea, it has been frequently put forward and accepted that the breaking up of the narcotics, with a chemical action of definite atomic groups thus set free, as the ethyl group, for instance, is responsible for the narcosis. But even in the case of sulphonal and its related sulphones, from which this idea originates, it can be shown that the action is induced by the entire unchanged molecule, and that the lack of activity of certain sulphones is due not, as is generally believed, to their not being broken up, but to a low distribution coefficient. This simple theory also explains the fact that all structures capable of stimulation, not only the cells of the nervous system, but all others, and all plant cells as well, are depressed by the narcotic members of this series, for in all living cells lecithin, THE THEORY OF NARCOSIS 17 a lipoid body, is to be found. And, indeed, the establishment of the fact that the effect on the lipoids by narcotics, such as ether and chloroform, is such as to immediately inhibit the vital processes of the cell, shows us that these lipoids are among the constituents essential to the life of the cell. Moreover, by establishing this fact it seems to me that the general biologic significance of the theory becomes apparent. That many narcotics induce not pure narcosis alone, but often show other distinct actions, as, for example, the occurrence of convulsions, which quite overshadow any narcosis present, is easily to be understood when one remembers that the narcotics may possess an affinity not only for the cell lipoids but for other cell constituents as well, and through some union with these, concomitant effects quite different from narcosis may be induced. This occurs, for instance, in the case of the phenols, whose narcotic action is thrown into the background by the appearance of clonic spinal convulsions. No attempt is made to explain every type of narcosis by means of the theory presented here. It is very probable that some other disturbances in chemical equilibrium ean occur in the cell and inhibit the performance of its function and that substances such as morphin are narcotic through their relation- ship to other points of attack than the “‘ alcohol lipoids ’’; and most probably the same can be said concerning the very remarkable nareosis from magnesium salts, lately discovered by Meltzer. I desire to add in conclusion that shortly after I had pub- lished my theory of alcohol narcosis the physiologist Overton published experiments which, carried out independently of mine and from a different point of view, in fact with somewhat different methods, brought him to an identical conclusion, 2.e., to a similar theory of narcosis, so that he has confirmed my work and accepted the formulation of my theory literally. I take this as a strong and gratifying argument for the correctness of our assumption. MODERN PROBLEMS OF METABO- LISM* CARL VON NOORDEN, M.D., Professor of Medicine, University of Vienna. OUR president has conferred on me a great honor in asking me to deliver one of the opening lectures of the new Harvey Medical Society of this city. It is your aim that from this well-established center waves of scientific stimulation for research work may arise and reach not only the circles of the professional workers of this city, but even those of the whole country. At this moment, when a society, promoted under such favorable auspices, opens its career by a course of lectures, I think it is opportune, not only to recount the results of investigations already completed, but principally to consider those problems which still await solution. IT am perfectly aware that in doing so I must renounce giving to my hearers the harmonious impression which a well-worked scheme ealls forth; for I am to touch manifold subjects and points of view which stand wide apart and in no organic relation to each other. Even in confining myself to a very small sphere of the problems of metabolism, a complete and exhaustive representa- tion of such will be impossible. Only a small selection can be made, and even this will savor of arbitrariness. IT may touch, perhaps, on several subjects which to you appear quite unimportant, and, on the other hand, I may omit many points which are of recognized importance. I expressly remark, therefore, that I shall mostly confine myself to questions which enter into my own program for future investigations on the problems of metabolism. If, as a result of my communications, *Lecture delivered October 14, 1905. 18 MODERN PROBLEMS OF METABOLISM 19 you gain the impression that details of the problems are thrown together by arbitrariness or by chance of selection, I hope that, on the other hand, the personal factor will be the joining link for compensating such disadvantages. A short retrospect of the history of several problems of metab- olism may form a useful preface. All the first investigations, decades ago, were directed toward the recognition of the quality of the chemical changes in the body. The substances which resulted from the breaking up of the tissues of the body and of the ingested food were the earliest to be demonstrated. The end-products of animal metabolism were determined. Most important rules were discovered concerning the production of CO,, urea, uric acid, kreatinin, indican and hippuric acid, ete. Among the normal end-products, many substances were found which appeared only under certain conditions, and were regarded as characteristic for particular diseases. As examples of such substances I may mention sugar, the various types of albumins, peptone, leucin, tyrosin, lactic acid, cystin, ete. Fol- lowing this period, in which the names of Wohler and von Liebig stand out prominently, came the second era, viz., that of pointing out the quantitative changes of metabolism. First introduced by Bischoff, the work in this branch of investigation was carried on and thoroughly established by Carl von Voit and von Pettenkofer and their pupils. Originally confined to the physiologic circumstances in animals and men, this ‘‘quan- titative study of metabolism’’ has since obtained new triumphs in its application to clinical medicine and to the study of pathologie processes. It is scarcely twenty years since these investigations commenced, and already, both in the physiologic and in the clinical laboratory, these quantitative estimations are being placed in the background, while attention is being directed to the newer field of the intermediary processes of metabolism. The finest and best work of late years relates to these questions. Hence to-day the investigations on metabolism approach again in character to those of the first period; but what then appeared impossible is now being attacked from all sides. Then one had to be satisfied with a knowledge of the 20 HARVEY SOCIETY end-products only ; to-day one endeavors, through the prominent discoveries in chemistry, to make clear the intermediate stages, through which the metabolites pass to their final conditions. An infinite number of new questions is thus presented by the recent advance in physiologic and pathologic chemistry. A number of important questions, which are of interest to the physiologist and pathologist alike, however, were left unsolved during the earlier periods of quantitative estimations, and it is only now that—thanks to the better technic of recent times—exact measuring methods are available for their investigation. First of all, there is the question of the metabolism of energy. Since the time of Voit and Rubner it has been customary to express and to measure body ‘‘ energy ’’ in terms of calories. In part through the relation of the body weight to the necessary intake of food, and in part from the amount of oxygen con- sumed and of CO, expired, certain average figures have been determined. When an adult man is in a condition of complete muscular rest, from 22 to 24 calories per kilo of body weight are necessary during each twenty-four hours; with usual light work, from 32 to 36 calories are required. The daily food must have these calorific values if the weight of the body shall neither increase nor diminish. With the increase of muscular work, the amount of energy consumed increases in certain proportions, and these latter have been sufficiently ascertained. We know also that children require a relatively high, and old people a relatively low, exchange of energy. Still, all these are only average numbers and they require the further support of numerous careful and exact observations. Even the most trustworthy figures, obtained by the use of methods of whose accuracy there is not the slightest shadow of doubt, showed that under exactly the same conditions a difference of from 20 to 25 per cent. arose between single individuals; this can only depend on the so-called individual factors. In future, however, this difference may not be slurred over by the use of the mystic word ‘‘ individuality ’’; we must endeavor to make clear the reasons for the rise above the average MODERN PROBLEMS OF METABOLISM 21 in oxidative processes in one person, and the fall below the average in another. Such information would provide us with a clear—I might even say a mathematical—insight into the condition which we now designate by the term ‘‘ individuality.’’ An important by-question which arises in regard to the physiology of nutrition, is the problem of the influence exerted on the consumption of energy by the respective constituents of the food. Certain experiments which Max Rubner and Ed. Pflueger have carried out on animals, tend to show that when the food contains an excessive quantity of proteids the energy-exchange rises considerably above the average. The energy production appeared to rise higher than was necessary for the muscular work done and for the maintenance of the body warmth. These results remind one of the old theory known by the name of ‘* Luxus-consumption,’’ if it does not even entirely compass it. They are too few and insufficient to revive the old hypothesis, which we have long known to be erroneous. As, however, one of the bases of the new science of nutrition is touched by it, the point should be thoroughly cleared up by new and better experiments on human subjects. If the excess of proteid intake really exerts a marked influence on the oxidative processes of the human organism, then we must change many of our views and explain differently a number of former experiments in metabolism. Up to now we trust that not the kind and the amount of food but only the internal and external bodily work rules the extent of the oxidation. The question is not a theo- retical one only. Recently manifold endeavors have been made to shake the old standard numbers for the albumin intake of healthy men settled by the school of Voit. It would be water on their mill could the supporters of vegetarianism from whom these endeavors originate, prove their contention, that large amounts of albumin raise the consumption of energy to an unseemly, that is to say, to an unnecessary and prodigal, extent. The theory of vegetarianism would also receive a specially strong support were it possible to confirm the oft-spoken asser- tion that the prodigal expenditure of energy only follows an 22 HARVEY SOCIETY excessive intake of animal albumins and does not result from a similar quantity of vegetable albumins. A few experiments we made lately turned against the theories of Rubner and Pfiueger. Of greatest interest and importance are, of course, those alterations of the exchanges of energy which occur in various diseased conditions. Single and occasional former investiga- tions excluded, we first commenced only about ten or fifteen years ago to busy ourselves with these matters. One single important fact is thoroughly established, viz.: the increase of the energy exchange which follows the administration of thyroid gland substance. This observation, which was made in my clinic by my former assistant, Prof. A. Magnus-Levy, was suggested by the practical experiences of Yorke-Davies and Leichtenstern on the influence of thyroid gland tablets on obesity. Later, Magnus-Levy discovered a similar increase in the transformation of energy in exophthalmic goiter and a decrease in myxedema. But these are the only diseases in which, up to now, spontaneous changes in the output of energy are known to occur. Thus the studies—I might call them pre- liminary—which have hitherto been made on the extent of the processes of oxidation and the amount of nutriment necessary in diseased conditions, afford sufficient reason for the use of our improved methods in further investigations in this field. Many of these problems are of great practical importance for bedside treatment. Next, there is the old question of how great the metabolism energy is in people who are run down by chronic disease or by insufficient nourishment. Do these persons require the same amount of food as do healthy individuals, reckoned per kilo of body weight, or do their bodies diminish the extent of exchange on some self-regulated plan? It is cer- tain that the albumin metabolism is diminished. It has even been asserted that the total production of energy also is dimin- ished, but on this we are as yet without definite proof. My preliminary observations point to the contrary, but the question has not been investigated with scientific exactitude. . The extremely painstaking and brilliant work of Neumann in Kiel, MODERN PROBLEMS OF METABOLISM 23 and of Chittenden in America, which has demonstrated the surprising extent to which the food of an adult man may be diminished without affecting the capacity for work and without altering the nitrogenous equilibrium of the body, leaves untouched this particular question. Obesity is quite the contrary. For a very long time it has been asserted that there are two forms of obesity. One type is said to result from an excessive intake of food or from insuf- ficient muscular exercise; the other is said to arise from an endogenous retardation of metabolic exchanges. The question is of great theoretical interest, but, as every one must admit, it is also of marked practical therapeutic importance. Since I first approached the matter, some twelve years ago, by investi- gations on the respiratory exchanges, the question has been constantly discussed. Some differences exist between the results of clinical observation and of laboratory experiments. Clinical reports indicate the occurrence of cases in which the obesity is due only to abnormal lowering of the oxidation, that is, to a diseased state of the protoplasm. Scientifically exact experiments, however, have failed to discover such relations. The results of some work done in the clinic at Basel seemed to point to abnormal low oxidative changes during muscular work and during the digestive processes of obese persons, but they must be discounted by the fact that the methods of estimation employed were not free from objections; correct deductions from them are therefore impossible. I am convinced, however, that with the advent of more satisfactory methods the views of the practitioners will be confirmed by laboratory experiments. Since the earliest days of investigations on metabolism, the question as to the energy exchanges in fever has received atten- tion. That the albumin exchanges are increased is quite cer- tain ; toxic influences are the reason. But why does the patient waste during the periods of fever? Why does he also lose so much of his body fat? As a matter of fact, in every case of long-continued fever, we observe an enormous loss of weight, even if we endeavor to avoid this loss by the administration of rich and nutritious foods. Does the cause lie in the fact that 24 HARVEY SOCIETY in spite of all our care an individual can not ingest the normal average calories of the food, since the digestive organs during fever are unable to take in or to digest the necessary amount? Or do the oxidative processes in the fever periods rise markedly above the normal? If this is the case, the food requirements of the fever patient will not be satisfied by ordinary quantities ; the amount of food sufficient for a healthy individual would not prevent the patient wasting during the stages of fever. The practitioner of earlier times did not doubt that fever was always accompanied by a substantial increase in all the processes of oxidation. The exact investigations on metabolic changes which have been made during the last decades do not, however, con- firm these ideas. These consist, in particular, of the works of Senator and some investigations by F. Kraus and by the pupils of Zuntz. If we thoroughly and critically read through these works, we find that they are full of contradictions and by no means permit of any final conclusions being made. The technic of to-day promises, however, a satisfactory and objection-free solution of this old problem. Still, the working out of the matter is naturally dependent on clinical material, and, unfor- tunately, the majority of hospitals to-day are not equipped with the necessary apparatus. Among other diseases in which the energy exchanges should be further investigated, I may mention diabetes mellitus. In slight cases, the relations are simple and undisputed. Such eases do not exert any influence on the energy exchanges. For a long time, however, it has been supposed—and lately the assertion has been revived on many sides—that in severe cases of diabetes the production of energy, and consequently the food requirements, are distinctly diminished. It has been calculated that in these patients the daily energy needs are satisfied with from 18 to 20 calories per kilo of body weight, while the healthy person requires from 34 to 36 calories under parallel conditions. The question is of great practical importance, because a clear conception would be of real assistance to us in the difficult dietetic treatment of diabetes mellitus. I do not admit that the just-mentioned figures, concerning the diminished production MODERN PROBLEMS OF METABOLISM 25 of energy in severe cases of diabetes, are quite correct; and I am of the opinion that the few previous exact observations on the production of CO, and the consumption of oxygen, are quite sufficient to prove this. Anyone who possesses a large respiratory apparatus can definitely settle the entire question in a few days. We leave now those questions which are intimately connected with the transformation of energy, and turn to another very interesting and important problem, relating to the metabolism of albumin. Earlier experiments on animals and recent inves- tigations on human subjects have taught us that an excessive amount of food compels a retention of nitrogenous substances in the body. The usual nitrogenous equilibrium is disturbed; a smaller quantity of nitrogen appears in the excreta than was present in the food. This retention of nitrogen may be attained by the administration of large amounts of albumin, but much more thoroughly and surely by a simultaneous excess of fat or especially of carbohydrates. The albumin-sparing proper- ties of the two latter substances, of course, are well known. The ultimate effect of such over-nutrition is always an increase in the total quantity of fat. We apply this knowledge thera- peutically in our ‘‘ feeding cure,’’ ete. But regarding the nitrogen there was until a short time ago the opinion that in spite of such an excessive nutrition, the nitrogen retention was only slight in quantity and short in duration—at least so far as well-nourished adults are concerned. It was taught that the body always endeavors to maintain a nitrogenous equilibrium so that, in the case of over-nutrition while the excess storing of fat may continue for a long time, a similar storing of proteids is soon stopped. In certain cases, however, the storing of body proteids seemed to be both extensive and long continued, as for instance, during the period of body growth, or after chronic exhaustive diseases, or after periods of lowered nutrition—it is always during the new growth of tissues. The occurrence of considerable nitrogen retention has recently been noted, apart from the conditions just mentioned. In a case of my own, I found that in two months not less than 370 grams of nitrogen 26 HARVEY SOCIETY were retained. Expressed in terms of meat this is more than 11 kg. of flesh. Is this retained nitrogen really built up into pure albumins and protoplasmic substance? Our general knowledge tends to indicate otherwise. We know that excessive feeding produces obese, but never athletic, individuals. A priori, it is very improbable that the nitrogen retained during excessive nutrition indicates the formation of pure albumins or a new formation of tissue substance. Perhaps the nitrogen only exists in the form of nitrogen-containing fragments of the large molecules of albumin, which are held for a time and _ are then cast off at a later period. In favor of this supposition there is the fact, that when the period of excessive nutrition is stopped, it is usual for an enormous quantity of nitrogen to appear in the urine. . The exact form in which the nitrogen is retained within the body is still, however, entirely unknown. This question is important, because a knowledge of it would throw light on the changes which the molecules of albumin undergo in the body. This problem leads to a consideration of the intermediate stages of metabolism, the special field of modern physiologic chemistry. Naturally, most questions of the ‘‘intermediary’’ metabolism concern themselves directly or indirectly with the fate of the albumin molecules; with their disintegration as well as with their synthesis. It seems that the synthesis of albumin in the body may originate from much simpler molecules than we could conceive of until lately. By intense and long-continued tryptic digestion of albumin, the latter has been broken up until the solution no longer yields the biuret reaction. In spite of this, the administration of the products of such diges- tion to animals serves for the substitution of pure albumins and for the maintenance of nitrogenous equilibrium. In close theoretical relation to this brilliant and important experiment of Otto Loewi, stand those considerations which are bound up with the discovery of erepsin in the walls of the alimentary canal. This ferment splits up the albumoses and peptones into simpler substances and, in particular, splits MODERN PROBLEMS OF METABOLISM 27 off the amino-acids. Hence, it has been assumed that this action represents the regular arrangement of processes, that the organism normally lives on the amino-acid mixtures, and that from these basal substances are formed the albumins which ultimately circulate in the blood stream. Such a sweeping conclusion, however, is a little too previous, for it has been shown recently that erepsin occurs in all the organs of the body and thus is not specific for the alimentary tract. The ferment, which in vitro is able to split up the albumoses, may syntheti- eally form albumoses from amino-acids when acting in the intes- tine. Such reversibility of ferments is already known. ‘These considerations appear to indicate, therefore, that the real func- tion of the intestinal mucosa is not to break up the albumoses into amino-acids, but, on the contrary, to build up albumoses and similar substances out of the amino-acids, which pass into the intestinal wall from the lumen. The question is certainly an available one for further experimental investigations. First of all, the living and surviving intestinal mucosa must be allowed to act on a mixture of amino-acids. This experiment has not yet been made. It is one, of course, which is very important for our ideas concerning the assimilation of albu- minous substances. If the investigation yielded positive results, there would be a remarkable analogy between this process and that of the synthesis of fat by the intestinal cells. Concerning the fate of fats, we know (1) that a fat-splitting ferment (lipase) is present in the intestinal wall; (2) that by the aid of this ferment there also occurs in the intestinal wall a syn- thesis of fat from fatty acids and glycerin; (3) that in vitro this synthetic process can be reproduced by the aid of lipase. For these brilliant and important investigations we are indebted to your own countryman, Dr. Loevenhart. These questions are of great significance in practical dietetics, since they bring into greater application the until now almost entirely neglected amino-acids. In particular, rectal feeding would receive a new impetus. Among the amino-acids there are many substances which are less irritable to the mucosa of the large intestine and are more easily absorbed than the 28 HARVEY SOCIETY usually prescribed albumoses and peptones. We have already commenced investigations on this point. Associated with the amino-acids, which represent the chief nitrogen-containing group of the albumin molecules, are many other questions, only a few of which can be touched on here. The chief of my clinical laboratory, Dr. G. Embden, has lately made an investigation on the amino-acids under physiologic and pathologic conditions. What I have to say on this point is due chiefly to the important theoretical and analytical work of Dr. Embden. First, I have to mention that glycocoll probably can be split off from all the amino-acids, the chains of the higher constituted amino-acids being broken up between a and B C-atom. Leucin CH; CH, ~~ a CH NH, + H_ CH, NH, COOH ~ (COOH (Glycocoll.) This process, it would appear, plays a great part in the organism. Recent investigations in my laboratory have shown that glycocoll is normally present in all urines, and in such quantities as to approach up to 1 per cent. of the total nitro- gen output. This well-grounded observation is striking, because we have for a long time known that glycocoll introduced into the stomach is very easily assimilated and reappears as urea in the urine. The formation of glycocoll within the body must be very large, if the kidney is able to catch and to eliminate some portion of it. This production of glycocoll from the higher amino-acids may also explain how it is that the body always has glycocoll at its disposal for coupling or combination pur- poses. I recall to mind the instances of hippuric and glycocolie acids. Further experiments must be performed, however, in order to determine whether or not the administration of large quantities of the higher amino-acids is followed by the appear- ance of a supernormal amount of glycocoll in the urine. If the formation of glycocoll from the higher amino-acids MODERN PROBLEMS OF METABOLISM 29 takes place in the manner which our preliminary investigations suggest as being probable, then a new light will be shed on the question of the formation of sugar from albumins. Since it has been established that higher amino-acids, such as alanin, form a definite source for sugar (G. Embden and H. Salomon), G. Embden, working in my laboratory, has shown that after the removal of the pancreas in dogs, sugar is formed from glycocoll equally as it is from alanin and other higher amino- acids. Glycocoll seems to be one of the most prolific sources of sugar that we know of. This fact, taken in connection with the previously mentioned conditions of glycocoll formation out of other amino-acids, explains at one stroke the question of sugar formation from albumins, and effectively removes those objections which have been heard during the most recent years. Glycocoll, however, does not seem to be the only amino-acid from which sugar can be rapidly formed. Our attention has also been directed specially to leucin, for the reason that of all the amino-acids leucin occurs most largely in the majority of the proteids of food. We are quite certain that lactic acid can be produced from alanin, and that in fact this procedure takes place within the body. A similar possibility obtains for the formation of lactic acid out of leucin. Theoretically, on the addition of water and oxidation, leucin breaks up into acetone and lactic acid, while at the same time amides are split off. Only just recently attention has been drawn to how often this process occurs in the chemistry of animal tissues and how im- portant it is. In this case the chain of C-atoms is broken between the 8 and y atom. You see, there are different possi- bilities of disintegration of the same molecule. CH, CH, CH, OH, ba pene | acetone CH = CO CHNH, Se | Lactic Acid + NH, COOH +0+H,0 COOH) In confirmation of these theoretical possibilities, we have just proved that the ‘‘ surviving ’’ liver always excretes some acetone 80 HARVEY SOCIETY into perfused blood, and that the amount of acetone consid- erably increases when leucin is added to the inflowing blood. At the same time, as we have determined with certainty the formation of lactic acid from leucin, we are met with a new problem, associated with the as yet unknown changes during the passage of carbohydrates through the body. We certainly _ know the end-products of carbohydrate disintegration—carbonic acid and water—but in regard to the intermediary metabolism of carbohydrates and the manner in which those end-products are produced we are still in the realm of theories. Entirely disconnected facts are alone our guides. One of the earliest theories related to the formation of lactic acid from carbohy- drates; but until now no satisfactory proof of this was given to us, at all events so far as muscular tissues are concerned. Many physiologists consider the lactic-acid formation in muscle to be due to post-mortem changes. The modern technics, which have advanced perfusion methods to a remarkable extent, will make possible a definite solution of the problem; until now, it is only solved so far as the liver is concerned. In my labora- tory, Embden and Almaggia have thoroughly demonstrated that lactic acid ensues in fact from disintegration of carbo- hydrates by the liver. This result arises from the action of a ferment and, as all our experiences with organic ferments indicate that the action of these ferments is reversible, so this procedure may also take place in the reverse way. As a matter of fact, we know that very often the administration of lactic acid to individuals affected with severe diabetes, and more especially to dogs after removal of the pancreas, is followed by an increase in the glycosuria. We also consider lactic acid as a rich souree of glycogen. These few available facts lead to the following hypothesis: A part of the sugar which is broken up in the muscle circu- lates in the blood as lactic acid; the lactic acid passes to the liver and is there rebuilt up to carbohydrate and eventually reaches anew the muscles in the form of sugar. With this conception of the intermediary stages and circulation of the carbohydrates in the form of lactic acid, some well-known facts MODERN PROBLEMS OF METABOLISM $1 are in full agreement. After extirpation of the liver, sugar disappears from the blood stream and lactic acid makes its appearance. Another remarkable fact may also be explained on this hypothesis. "When the pancreas is removed from birds, glycosuria does not result. In these animals, lactic acid is not bound to be regenerated into sugar, but with the addition of ammonia, it ean form uric acid. If this view be a correct one, then the uric acid of the bird is partly a derivative of sugar. I advance this theory, of course, only in the form of an hypoth- esis; it has, in any case, the advantage of promoting further investigations on the intermediary stages of carbohydrate me- tabolism and of providing a new aim and a definite proposition for further proof. I have already mentioned that, theoretically, acetone may be produced from leucin, and that we have been able to demon- strate this procedure by experiments on animals. This result is very remarkable, since the opinions of to-day designate the fatty acids alone, and the lower fatty acids in particular, as the source of the acetone bodies, and because until now we have always accepted the oxybutyrie and diacetic acids as the neces- sary precedents to acetone. This latter view thus requires correction, although our experiments in no way show that in the formation of acetone leucin plays an important figure in respect to quantity. At all events, it indicates that the acetone question can not yet enter into a condition of rest. Also the problems of the formation of acetone from fat and the hindrance to the production of acetone through the simultaneous oxidation of carbohydrate, are still sufficiently enigmatical and can not be solved until we know much more about the intermediary disintegration of fats and of carbohydrates than we do up to this day. With this I wish to conclude my survey of modern problems of metabolism. As TI stated at the commencement of the lecture, it has been necessary to roam over a large amount of ground and to consider subjects that are but slightly related to each other. You will observe that to-day we are busying ourselves in a much more intimate manner with the details of metabolic 32 HARVEY SOCIETY processes than in not very remote periods was deemed either necessary or possible. Already the little that has been men- tioned here is more than the working powers of one single man can master; but on all sides we see new young energy pouring into this interesting and important branch of medical investi- gation, in order to harvest this inexhaustible field. We greet it with joy and with satisfaction. The results will not be long in coming. We are all convinced that these marked steps into the wonder- land of animal metabolism will not only advance the theoretical science, but, as we have always seen, that every advance in physiologic and pathologic chemistry has been followed by improvement of our bedside treatment. The achievements of the dietetic treatment of diseases have gone hand in hand with the advances in theoretical investigations. If we compare the progress in dietetics that has been made during the last decade with the wonderful successes of the surgeon, the medical clinician no longer need feel either shame or envy. In the same period a vast amount of work has been done by the internist in regard to therapeutic matters. The close relations which have been maintained between the progress in clinical bedside treatment on the one hand and physiologic and patho- logic chemistry on the other, have been very fruitful indeed, and still fruitful will remain. Great problems still await solution and rich outside help is necessary thereto. With confident expectation, medical science looks to this country, in which in recent times numerous ardent and honest research-loving young workers have entered into the service of problems of metabolism, and in which the riches and the munificence of its inhabitants more than elsewhere have provided that external aid which has made more easy the prosecution of great and far-reaching investigations. I close with the prophetic words of our Goethe: “ Amerika, Du hast es besser Als unser Continent der Alte.” ON TRYPANOSOMES* FREDERICK G. NOVY, Sc.D., M.D., Professor of Bacteriology, University of Michigan, Ann Arbor, Michigan. NTIL very recently the bacteria or plant organisms have been given a preponderating and almost exclusive role in the production of infectious diseases. The studies of the past few years, however, have brought to light another group of organisms which play an exceedingly important part in the causation of diseases peculiar to the warm countries. Unicellular forms of animal life are to-day the recognized causes of a large number of diseases whereas but a few years ago they claimed but scanty attention. The pathogenic protozoa, in a remarkably short time, have risen from an obscure to a com- manding position by the side of the pathogenic bacteria. Under the head of protozoa are classed: First, the try- panosomes which are met with free in the blood plasma; second, the hemocytozoa which find their habitat within the blood cells and are represented by the malarial organisms in man and by related forms in the lower animals; also by the piroplasmata found in Texas fever and allied affections. And, third, the amebe which are found in the intestine in dysentery. Many other forms of pathogenic protozoa are known but these are of relatively little interest compared with those mentioned above. Although the first of these, the trypanosomes, have acquired special importance during the past decade, it is nevertheless an * Lecture delivered November 4, 1905. The lecture was illustrated by a large number of lantern slides of which only a few can be reproduced in connection with this paper. The author wishes to acknowledge the courtesy of the Journal of the American Medical Association for the loan of the cuts of the illustrations which are given in the text. 3 33 34 HARVEY SOCIETY interesting fact that the first representatives of this group were described more than sixty years ago. The credit of the dis- covery of this group belongs to Valentin who, in 1841, found the first trypanosomes in the blood of salmon. In the following year a similar organism was found in frog’s blood and it was to this that Gruby gave the generic name trypanosoma, implying a screw- or auger-like body, the name being suggested by the peculiar motion of the parasite. It will be seen from this that the discovery of these organisms antedates that of anthrax (1849) and of the Spirillum Obermeieri (1873). Up to 1850, trypanosomes had been found in the blood of several mammals, notably field mice, moles and rats, but these observations were eventually lost sight of, and it was not until 1877 that they were rediscovered by Surgeon-major Lewis at Calcutta. It was the interest aroused by the findings of Lewis that led to the discovery by Evans in 1880 of a trypanosome in the blood of animals afflicted with a disease known in India as surra. This organism, now designated as Trypanosoma Evansi, is consequently the first known really pathogenic try- panosome, inasmuch as the organisms met with prior to that time are commonly looked upon as harmless parasites. During the following fifteen years, although many observa- tions were made upon the trypanosomes in rats, fish, frogs and birds, they attracted but little attention, largely because of the all-absorbing interest in the study of bacterial diseases. The past decade, however, has witnessed a truly remarkable progress in our knowledge of the diseases due to protozoal organisms and the credit of bringing about a just recognition of the etiological réle of trypanosomes belongs to Colonel David Bruce, of the British Army Medical Service. In 1894 he began the classical investigation of the terrible tsetse-fly dis- ease or nagana of Zululand, the results of which study were published in 1895, 1897 and 1903. He showed that this dis- ease was due to the presence in the blood of a trypanosome now known as 7'r. Brucei, similar to that studied in surra by Evans and Lingard; that the disease was transmitted from the infected to healthy animals by the bite of the tsetse-fly, Glossina ON TRYPANOSOMES 35 morsitans; and that the persistence of the disease was due to the presence of the parasite in the large game which conse- quently acted as a reservoir for the virus. The trypanosome which he discovered was transported to England by Dr. Wag- horn in 1896 and it is this virus which has been utilized in most of the researches carried out in Europe and in this country. During his travels in East Africa, in 1898, Koch presumably encountered this same disease which he considered to be identi- eal with the Indian surra. He pointed out at the time the morphological differences which exist between this trypanosome and that of rats, and by means of a simple animal experiment he was able to differentiate sharply between the two organisms. Thus, in the blood of the rat the two organisms develop side by side and are quite easily distinguished by the aid of the microscope. If, however, the blood of such a rat is injected into a dog the nagana trypanosome alone appears in the latter and eventually causes death. This work of Koch served as the immediate incentive for an exhaustive study of the rat trypanosome which was made in 1899 by Rabinowitsch and Kempner. To these workers much credit is due for introducing the staining methods which have thrown so much light upon the structure of these organisms. They were able to demonstrate many facts bearing upon the mode of multiplication, and in addition they showed that active immunity to 7'r. Lewisi could be produced in rats. The study of the rat trypanosomes begun in 1900 by Laveran and Mesnil has led to the splendid series of researches on the pathogenic trypanosomes which have come from the Pasteur Institute since that time. The Tr. Lewisi as found in the common rat is an excellent type of the whole group, and owing to the almost universal distribution of its host it has been found in all parts of the world. There is probably no city in which the rats are wholly free from this infection. This fact is of interest because it enables any one who is desirous of studying this organism to readily procure the needed material. It should be borne in 86 HARVEY SOCIETY mind, however, that the infection is not necessarily evenly distributed throughout a community, but that it may be and is often localized in one or more places. Thus, at Ann Arbor, of the first 107 rats caught at different places and examined, five were found infected, and all of these came from the same barn. During the past six years we have repeatedly secured rats from this particular barn and invariably one or more in a catch have been found to harbor the parasite, whereas rats taken in other parts of the city have been relatively free from the infection. It is an interesting fact that at times the white rat will be found to be spontaneously infected with Tr. Lewisi. The first observation on this point was made by Laveran and Mesnil in Paris, and later by Terry in Chicago. We have also repeatedly met with infected white rats purchased from dealers in the large cities. Before taking up the more strictly pathogenic forms it will be desirable to consider briefly the structure and mode of multi- plication of the rat trypanosome. In general, the facts ascer- tained by the study of this parasite hold true for the other forms. In the fresh blood film, when examined by the means of a medium objective, such as Leitz No. 7, the trypanosome readily attracts attention by the sudden commotion of the blood corpuscles in its immediate neighborhood. These are pushed aside or lashed about as the organism moves from place to place. The motion is fairly rapid, more so than is usually the case with the other blood trypanosomes. It will be seen that the body of the parasite is somewhat spindle-shaped and that one end terminates in a single long free whip or flagellum. The presence of this organ is an important characteristic and it 1s because of it that the trypanosomes are classed among the Flagellata. On careful observation it will be seen that the trypanosome usually moves with its flagellum foremost, picking ~ its way among the corpuscles. The flagellar end is conse- quently spoken of as the anterior end. The opposite extremity or posterior end is rather sharply pointed, a fact which distin- guishes it readily from the other trypanosomes. Fic. 1.—Trypanosoma Lewisi in blood of rat. Note the sharp posterior end opposite the free flagellum; also nu- cleus and micro-nucleus. Magnification 1,500 times. Fic, 2.—A multiplication rosette of Tr. Lewisi in blood of rat, showing division into eight cells and remnant with flagellum of original parent cell, The elongated micro-nucleus in each young cell shows two flagella, a long and a short one, indicating that further division is about to take place. Magnification 3,000 times. ON TRYPANOSOMES 37 On further inspection it may be possible to note the presence of an undulating membrane—a fin-like structure which extends from a point near the posterior end of the body to the base of the flagellum. This undulating membrane constitutes the real locomotive organ, the free flagellum probably having very little to do with the actual movement of the cell. This is seen in the fact that at least one blood trypanosome (7'r. dimorphon) is devoid or nearly so of a free flagellum. Moreover, in arti- ficial culture many trypanosomes, although provided with a very long free flagellum, show scarcely any motion, owing to the rudimentary character of the undulating membrane. The body of the trypanosome in the living condition is color- less or nearly so and the contents are nearly homogeneous; at most a fine granulation may be observed. Further details as to structure can only be made out by the application of suitable staining methods. For this purpose the Romanowsky method in some of its numerous modifications is quite generally employed. No other process of staining brings out so well the structural character- istics of the trypanosomes and for that matter of all protozoa. The principle of the method consists in the use of a ‘‘ripened’’ or polychrome methylene blue which, with eosin, stains the nuclei and other nuclear structures a deep rose-red, whereas the plasma of the cell shows a pale to deep blue. In this way the details of the structure are brought out in sharp relief. The altered methylene blue owes its peculiar staining properties to the presence of at least two decomposition products, methylene azure (— . atl 7 q v2" <) the ‘se-fly ‘ d of rat. One of the or Mal du coit) in blood of mouse. — One of tl Tsetse-fly disease) in bloo Magnification cells in process of division. Magnification 1,500 times. : | Fic. 5.—Trypanosoma Brucei (Nagana or Fic. 6.—Trypanosoma equiperdum (Dourine cells is in process of division. 1,500 times. Fic. 7.—Trypanosoma equinum (Caderas) in Fig. 8.—Trypanosoma dimorphon (Gambian blood of mouse. Note the apparent absence of! horse disease) in blood of mouse. One cell is blepharoplasts or micro-nueclei. The double dividing. Note the absence of a free flagellum. flagellum indicates divisional changes. Magni- Magnification 1,500 times. fication 1,500 times. ON TRYPANOSOMES 53 remain susceptible to infection with the other trypanosomes (as has been shown by Laveran and Mesnil, also by Nocard and Vallée), and consequently this reaction may be interpreted as demonstrating that nagana is specifically different from surra and the other trypanosomiases. In the large animals the affection is characterized by fever, anemia, edemas of the extremities and abdomen, and extreme emaciation. The animal may waste to a mere skeleton and often becomes blind. Its appetite may last until the end. In the blood of the diseased animals Bruce discovered in 1894 the trypanosome which now bears his name. As has been pointed out heretofore, this organism is scarcely to be distin- guished either morphologically or biologically from the T7’r. Evansi (Fig. 5). The immunity experiments referred to above have served to show that the diseases are distinct entities. The Tr. Brucei was the first pathogenic trypanosome success- fully cultivated (Novy and MacNeal). Some difficulty is experienced in securing the first generation, but after that there is none. It has been possible to carry such a culture through nearly 100 generations at room temperature and at 25°. When once thoroughly adapted to the blood agar medium a very rich growth can be obtained in from three to four days. Even after cultivation for two years such cultures are capable of infecting guinea-pigs. It has not been possible as yet to obtain non- virulent cultures which could be used for the purpose of vaccinating animals against the virulent type. The cultural characteristics are very marked and serve to differentiate it from Tr. Lewisi on the one hand and Tr. Evansi on the other. In view of the fact that nagana of some parts of Africa differs to a greater or less extent from that of Zululand, which fact has suggested the existence of a number of varieties or even a group of allied diseases, it is desirable that in such cases the cultural characteristics of the organism be ascertained. TRANSMISSION. The general impression regarding the réle of the tsetse-flies in the transmission of nagana was fully established by the 54 HARVEY SOCIETY splendid investigation of Bruce. This was executed in Zulu- land, on the top of Ubombo, at an elevation of about 1900 feet. Although this hill was in the midst of the fly-country, Ubombo itself remained free from tsetses from the beginning of the investigation in 1894 until 1897, at which time nagana for some reason spread beyond its former boundaries, and even reached the top of the hills. Bruce early noted the remarkable fact that no cases of the spontaneous disease occurred on Ubombo hill, notwithstanding the constant and close association of healthy horses, cattle and dogs with those suffering from the disease and the presence of several species of blood-sucking flies other than tsetses. This observation was confirmed later by Martini who found two infected Togo ponies in the Berlin Zoological Garden, and yet in spite of the presence of ordinary biting flies the infection did not spread among the other animals. These facts go to show that nagana is not spread by ordinary biting insects such as Stomoxys, mosquitoes and fleas. Bruce further showed that horses when taken for a few hours into the fly country—even though they were not allowed to eat or drink while there—contracted the disease, thus showing that the latter was not conveyed by the food or drink but in all probability by the bites of the flies. It was further proved that tsetses brought to Ubombo and kept for several days could then bite dogs without producing the disease. Infection, however, did occur if they were allowed to feed on dogs immediately after feeding on an infected animal. This result was also obtained if the flies were allowed to bite at 12, 24 and 48, but not at 72 hours after the infective feed. Bruce examined the proboscis of the flies at varying hourly intervals after they had fed and found rarely more than a single trypanosome. In the stomachs of such flies the trypano- somes were usually present as long as the blood remained. Up to 55 hours they were always present, but none were found on the sixth day. Owing to the presence of the trypanosomes in the stomachs it might be expected that such flies, if minced up and injected into a dog, would produce the disease. This, however, Bruce was unable to accomplish except in one case ON 'TRYPANOSOMES 55 where the fly had fed but half an hour before on an infected animal. This result is difficult to explain, especially since we have been able to infect mice by injecting mosquitoes which had an infective feed 24 to 36 hours before. Within the past year it has been shown by Koch that several species of tsetse-flies contain trypanosomes in their stomachs, at the time of capture even when there is no evidence of the presence of blood to indicate that the flies had recently fed on an animal. It was supposed by him that the flagellates found in the stomachs of the flies represented developmental forms of Tr. Brucei, although he was unable to infect flies by feeding on infected animals, and moreover failed to infect rats by injecting the contents of the stomachs of such flies. As will be seen similar observations have been made by Gray and Tulloch with reference to the trypanosome of sleeping sickness. The facts noted, however, are all open to an entirely different interpretation, and it is more than likely that the organisms seen in the flies represent, as we have shown elsewhere, a harm- less intestinal parasite much the same as that found in mosquitoes and other insects. Notwithstanding the above, the fact remains that the tsetse- flies can and do convey the disease provided that they bite within a few hours or at most a day or two after they have fed on a diseased animal. The fly, therefore, acts as a mere carrier of the trypanosome, and that being the case it was necessary to establish the natural source of the disease. It had been shown previously by Lingard in India and confirmed later by Musgrave in Manila that rats may harbor the surra trypanosome. Bruce was led, therefore, to believe that a similar source might be found in the big game of the fly-country, especially as the general impression pre- vailed that where there was no wild game there was no nagana. Accordingly, he examined the blood of various animals shot in the fly-country, but could find no trypanosomes in their blood. Subsequently, however, he did find the adult organism in three wild animals. Acting on the supposition that the parasites might be present in such small numbers as to escape micro- 56 HARVEY SOCIETY scopical detection, he injected the blood into dogs, with the result that in the first series nine out of thirty-five dogs thus inoculated developed nagana. In brief, it has been shown.that the large game harbors the parasite without any injurious effect in much the same way that the common rat carries 7'r. Lewisi. In such animals the disease is probably chronic and non-fatal and hence they serve as a reservoir for the virus which they supply to the tsetses. It has since been shown that with the extermination of the large game in some parts of South Africa, due to the introduction of rinderpest, the fly disease has become a negligible quantity. Although considerable effort has been expended to discover a preventive and curative treatment for this infection, the results thus far have not been very satisfactory. The admuinis- tration of arsenic is of no preventive value but in infected animals serves to prolong life. When given in sufficient dose to the small animals it causes the trypanosomes to disappear from the blood, but only temporarily. After a few days they reappear, and on repeating the injection of arsenic they vanish as before. By repeating the treatment it has been possible to keep rats alive for 79 days, whereas the controls die in about 5 days. Laveran and Mesnil were unable to effect any cure with arsenite of sodium employed in this manner. Thomas has obtained much better results with a compound of arsenic and anilin known as atoxyl. With this he has been able to cure infected rats, guinea-pigs and a rabbit. The trypan-red introduced by Ehrlich and Shiga likewise causes a temporary disappearance of the trypanosomes, but it only exceptionally is able to do more than prolong the duration of the disease. A number of other anilin dyes have been used, some of which have given very encouraging results. The recent studies of Mesnil and Nicolle on the ‘‘benzidine colors’’ are very promising, and it is to be hoped that an efficient ‘‘chromo- therapy’’ will be discovered. The alternate treatment with trypan-red and atoxyl has given fair results, and it is quite possible that some modification of this procedure may prove serviceable in practice. ON TRYPANOSOMES 57 The action of human serum as a curative agent is of especial interest. It may be assumed as established that man is not subject to infection with Tr. Brucei. At all events, though man is attacked in the fly-country by the tsetses, no ill effects have been noticed from such bites. The normal sera of diverse animals have been tested by Laveran as to their effect on the trypanosome and of such only the human serum was found to be efficacious. It not only can prolong the duration of the disease but is capable of curing mice. An alternate treatment with arsenic and serum has given still better results. From the brief outline given above it will be seen that arsenic, certain anilin dyes and human serum are the only agents known at present which can influence the course of the infection with Tr. Brucei. A practical preventive and curative treatment, however, has not yet been discovered, and as it is obviously impossible to reach and destroy all infected animals the preven- tion of nagana remains a serious problem. DOURINE. This disease, unlike surra and nagana, is not limited to the tropical countries, for in the past it has been met with in many parts of Europe and it is even said to occur in the United States. Such reports, however, are based purely upon circum- stantial and clinical evidence, for up to the present time no one has demonstrated in this country the presence of the specific organism in such eases. The disease it should be stated is especially prevalent along the Mediterranean littoral. The trypanosome of Dourine, T'r. equiperdum, was discovered in 1894, the same year as T'r. Brucet, by Rouget, a French army veterinarian. By successive passages through rabbits he was able to keep the organism, derived from a stallion in Algeria, for over two years. Since then other observers have confirmed and extended his work. One notable feature of the disease is that it is not spread, so far as is known, by flying insects. In this respect it differs therefore from surra and nagana and the other trypanosomiases. 58 HARVEY SOCIETY The evidence on hand goes to show that it is spread exclusively by sexual contact, hence the designation Mal du coit. It is restricted to breeding equines, and on account of the peculiar skin lesions it is sometimes spoken of as horse syphilis. The usual chronic form of the disease is characterized by edema of the genitals, a moderate fever and slow wasting. In from one to two months, plaques of varying size appear on different parts of the skin. The wasting becomes more pro- nounced and paralysis of the hind quarters becomes manifest. In the last stages of the disease the anemia is profound, the animal becomes paralyzed and is unable to rise. The infected animals may die in from two to ten months, exceptionally they live for as long as two years. Experimentally, the disease is readily reproduced in the horse, ass, dog and rabbit. Rats, mice and guinea-pigs are apparently refractory to the fresh virus, but after repeated passage the latter will prove infective. It is an interesting fact, first established by Rouget, that a lesion of the mucous membrane is not necessary in order that infection shall result. The direct application of the virus to the conjunctiva or to the vagina is sufficient to cause an infection. ‘The disease has not been produced by ingestion of the parasite. Although very searce, the trypanosome can be detected in the blood of the infected animal and especially in the freshly formed plaques. Morphologically, it can hardly be distinguished from the other pathogenic trypanosomes (Fig. 6). The most important difference according to Laveran and Mesnil consists in the absence of the protoplasmic granules such as are present in Tr. Brucei. Thomas and Breinl were able to keep the organism alive on blood agar for 17 days, at which time it was still infective, but no sub-cultures could be obtained. Animals such as dogs which have recovered from the disease possess an active immunity. But when inoculated with caderas (Ligniéres), or nagana (Nocard), they promptly succumb to these diseases, thus showing that dourine is specifically different. The treatment of dourine is on the whole as unsatisfactory as that of the other trypanosomiases. Arsenic, trypan-red and ON TRYPANOSOMES 59 human serum have an action similar to that noted in connection with nagana. CADERAS. The disease known as Mal de caderas is widely prevalent in many parts of South America from the Amazon on the north to Bolivia on the south. It is the only trypanosomatic disease which has been recognized on the American Continent. It has been the subject of repeated studies, but its nature was not recognized until 1901 when Elmassian at Assumption discov- ered the trypanosome which Voges designated as Tr. equinum. The name given above refers to the characteristic symptom of the disease, the paralysis of the hip or hind quarters. Caderas occurs almost exclusively among horses, although instances of spontaneous infection of dogs are known. The ass and mule are more resistant than the horse. The disease may be of short duration lasting from a few weeks to one or two months, or it may be of a very chronic form which persists for many months. It is attended with a marked remitting fever; the animal rapidly loses weight, although the appetite continues. LEvent- ually the hind quarters begin to drag and finally complete paralysis results. Anemia and albuminuria are common, and hematuria may be present. A notable feature is the almost complete absence of edemas which are almost always present in nagana, surra, ete. The trypanosomes are not very numerous in the blood and often cannot be detected microscopically. The inoculation of such blood into susceptible animals will result in infection. Rats and mice are particularly susceptible, and in these the trypanosomes become extremely rich and death occurs in from one to two weeks after inoculation. Most of the smaller mam- mals are likewise susceptible, but the disease is of longer dura- tion. Goats, sheep, cattle and hogs develop a very mild infec- tion without any visible manifestation. The organism is present in the blood in such very small numbers that it can 60 HARVEY SOCIETY searcely be detected except by inoculation of the blood into mice. By this means it has been shown that the blood may harbor the trypanosome for from two to six months. With these animals recovery is the rule, and they become immune. The Tr. equinum, although it is of the same form and size as the other pathogenic trypanosomes, is nevertheless readily dif- ferentiated from these by one important characteristic and that is the apparent absence of the micro-nucleus or blepharo- plast. This structure is so inconspicuous that its existence has been denied by some (Fig. 7). Attempts at cultivation have been made by Rabinowitsch and Kempner, and Laveran and Mesnil but without any definite suecess. Thomas and Breinl observed an apparent multiph- eation in a blood agar tube, 29 days old, and with this success- fully infected a rat. Sub-cultures were not obtained. As to the mode of transmission of caderas very little is known at present. The conveyance by insects is by no means estab- lished, although it has been possible to infect horses by Sto- moxys which had previously fed on infected animals. On the other hand, healthy and infected animals have been kept together or at most separated by a fence without the infection spreading. The results of treatment in caderas have on the whole been more favorable than in the case of the other trypanosomiases. Thomas and Breinl obtained with atoxyl cures in rabbit, guinea- pig and rat. Ehrlich and Shiga were able to cure mice by means of their trypan-red. Moreover, previous inoculations with the dye served to prevent infection. Several other dyes are now known which in a single injection will cure infected mice (Mesnil and Nicolle). Human serum has been-shown to be as active against caderas as against nagana. As with the latter, the disease is prolonged in the infected mice and excep- tionally a cure is effected. GAMBIAN HORSE DISEASE. This disease was first recognized by Dutton and Todd, in 1902, among the horses of Senegambia. Of 36 examined 10 ON TRYPANOSOMES 61 were found to have the trypanosome in their blood. As far as known no other domestic animal is subject to the disease, although most mammals, including sheep, goats and cattle, can be infected. The natural disease is very chronic in character and differs from nagana by the absence of edemas. In the latter respect it agrees with caderas, but as will be seen the trypanosomes of these two diseases are easily differentiated. The duration of the disease is not known, though it probably lasts for a few months to more than a year. In an experimental infection of a horse, Laveran and Mesnil noted the formation of an edematous patch, but otherwise the animal did not appear to be ill. There was an occasional rise in temperature and try- panosomes were present at first in the blood, but later were recognized only by inoculation of rats and mice. In this way they were found to be present as late as the one-hundred-and- eighty-third day. The disease is presumably transmitted by biting flies, although no positive evidence on this point has been obtained. The examination of living and stained preparations shows the presence of two forms, hence the name of the organism, Tr. dimorphon. The short form is about 124 while the long one is 20 to 254. A similar occurrence of long and short forms has been noted in galziekte and in bird infections. Unlike as in Tr. Brucei, the undulating membrane is not conspicuous but by far the most important characteristic is the absence of a free flagellum. This condition is due to the prolongation of the protoplasm of the cell along the flagellum to the very tip. This feature serves to identify the organism in the same way that the minute blepharoplast characterizes the trypanosome of caderas. The artificial culture of this organism was attempted by Laveran and Mesnil, but although they succeeded in keeping it alive on artificial media for over a month they were unable to secure sub-cultures. Thomas and Breinl] were more success- ful, for they maintained it on blood agar for 78 days and were able to infect animals as late as the twenty-third day. 62 HARVEY SOCIETY The structural peculiarity of the trypanosome serves at once to differentiate the infection from all other flagellate diseases. As further evidence of its individuality it may be mentioned that animals immunized to the other trypanosomes remain susceptible to inoculation with Tr. dimorphon. Thus, goats which have been vaccinated against surra, nagana and ecaderas, are very sensitive to this parasite. The treatment of the experimental infections has not been as good as with nagana and the other trypanosomiases. Thus, arsenic, either in the form of arsenite of sodium or atoxyl, causes the organisms to disappear temporarily and the duration has been prolonged, but no cure has been effected. Trypan-red has a similar action on the trypanosomes, but neither Laveran and Mesnil nor Thomas and Brein] have noted any definite curative powers. Human serum in large enough dose may also cause the disappearance of the trypanosomes for a varying length of time. The action, however, is more feeble than in the case of nagana. GALZIEKTE. A trypanosomiasis of cattle wholly distinct from nagana or surra appears to exist throughout South Africa and probably it occurs elsewhere. Thus, a similar if not identical infection has been observed in Hast Africa (Sander, Panse), in West Africa (Schilling, Ziemann), in the Trans-Caucasus (Ziemann), and in India (Lingard). The disease itself has been known for many years and is known by a variety of names, such as gall-sickness or galziekte, malaria, jaundice or bilious fever of cattle. Compared with the other diseases of domestic animals in Africa this is of but slight importance. The disease is marked by a light fever, which lasts several days, and a severe anemia, which may be either acute or chronic. The mortality as given by Theiler is but 12.5 per cent. In the blood of the infected cattle in 1902 Theiler discovered an unusually large trypanosome which Laveran and Bruce, independently, designated as Tr. Theilerr. It is the largest ON TRYPANOSOMES 63 of the pathogenic trypanosomes and is about the size of the large form of Jr. avium as met with in robins and blue-jays. Like the Tr. avium and Tr. dimorphon it occurs in the blood in two forms, one short and the other long. The former are 25 to 30u in length by 2 to 3 in width; while the latter may be 60 to 70% long and 4 to 5, wide. It is very actively motile and has a prominent undulating membrane and a long free flagellum. Experimentally the disease can be readily transmitted to cattle by injection of the infected blood. The trypanosomes may be very numerous or on the other hand quite scarce. Like the trypanosome of the rat the Tr. Theileri appears to be limited to a single host, since all attempts to inoculate other animals have failed. After recovery the cattle are immune. The disease appears to be transmitted by the bite of a fly, Hippo- bosca rufipes. HUMAN TRYPANOSOMIASIS. (Sleeping Sickness. ) For a long time it was supposed that man was not subject to trypanosomatic infection. This appears to be true for surra and nagana as well as for the other diseases discussed above. Cer- tain it is that the bites of insect carriers are without effect in -man, and even accidental inoculations have occurred without any observable result. In a way, this immunity of man to the animal infections is largely if not wholly due to the peculiar action of human serum on these trypanosomes. The first authentic case of human infection with trypano- somes was observed at Bathurst, Gambia, in 1901, by Forde, who, however, did not recognize the nature of the organisms which he described as ‘‘small, worm-like, extremely active bodies.’’ On subsequent examination of the patient, who was an Englishman in the government service, in December, 1901, by Dutton, the organism was at once recognized as a trypano- some and was named by him Tr. gambiense. On his return to England, Dutton examined 115 blood films obtained from native 64 HARVEY SOCIETY children in Gambia and one of these preparations showed a double infection with malarial parasites and trypanosomes. The result of these discoveries was of far-reaching importance. The Liverpool School of Tropical Medicine at once sent out an expedition consisting of Dutton and Todd to Senegambia with the result that these observers recognized seven cases out of 1043 persons examined. About the same time (1902) Manson diagnosed the disease in a woman, the wife of a Congo mis- sionary. Other cases were soon reported by Broden at Leopold- ville, Brumpt at Boumba, and by Kermorgant. The existence of a human trypanosomatic fever was thus established but its relation to the terrible disease known as Sleeping Sickness was not suspected. Sleeping sickness itself has been known to exist on the West Coast of Africa for more than a century. From the time of Winterbottom, who described it among the slaves of Benin in 1803, it has repeatedly been studied by English and French physicians and missionaries on the Gold Coast and at Sierra Leone. Although undoubtedly hundreds of slaves infected with the disease were transported to the West Indies, there is no reason to believe that new cases ever developed on this side of the Atlantic. The disease is stated to exist on the West Coast of Africa from Senegal on the north to Benguella in Angola on the south. In certain localities of this vast territory it has proved particu- larly destructive, notably along the Lower Congo. Since the establishment of the Congo Free State in 1885 the disease has been carried to the Upper Congo in the first place by traders, and secondly by military expeditions (1892-1896) against the Arab raiders. In 1901 its presence was reported for the first time in Uganda, where already it had caused an enormous destruction of life. This serious outbreak is generally supposed to be due to the return of the remnants of Emin Pasha’s army which were brought from the regions west of Albert Nyanza, during the years 1892-95, and established on Victoria Nyanza in Busoga. Whatever its origin, the disease has since its intro- duction spread along the entire north shore of Lake Victoria ON TRYPANOSOMES 65 Nyanza and has even passed down the Victoria Nile as far as Wadelai (Greig). The British and Portuguese governments, recognizing the need of definite information regarding the cause and spread of sleeping sickness, appointed commissions to investigate the dis- ease. Considerable attention was given at first to the supposed bacterial cause, and it was while engaged in this study that Castellani in Uganda noted the presence of trypanosomes (Noyv., 1902), in the cerebro-spinal fluid of five cases of sleeping sick- ness. At the time he did not consider that this trypanosome had any causal relationship to the disease, but later, on the suggestion and with the aid of Colonel Bruce and others, he examined additional cases of the disease and was able to report the presence of trypanosomes in 70 per cent. of the cases (April, 1903). Subsequent studies by Bruce, Nabarro, Greig, and others have demonstrated the constant occurrence of the Tr. ugandense (Castellani) in either the blood or cerebro-spinal fluid of sleeping sickness eases. The trypanosome found in sleeping sickness was at first supposed to be distinct from the Tr. gambiense of Dutton, but subsequent researches have shown that in all probability the two organisms are identical and that the trypanosomatic fever is but the first, while sleeping sickness is the last, stage of the human disease. Still more recently the important fact has been brought out that glandular enlargements are a constant feature of early eases of human trypanosomiasis; in other words, that sleeping sickness during the early stage is a specific polyadenitis caused by the Tr. gambiense (Greig and Gray). It has been shown that the trypanosome could practically be always found in such enlarged glands, and Dutton and Todd have pointed out that cervical gland palpation is a simple and very accurate method of detecting cases of trypanosomiases in which clinical signs are wanting. The recognition of the existence of such cases explains the ease with which the disease has been carried into uninfected districts by the migration of apparently healthy persons. And furthermore, being a simple means of diagnosis 5 66 HARVEY SOCIETY of the earliest stage of the infection, it enables putting into effect measures for the prevention of the disease. This means the exclusion and removal of persons having glandular enlarge- - ments from uninfected territory and their segregation as far as possible. The necessity of adopting every possible means of arresting the progress of the disease is seen in the fact that in many villages Dutton and Todd found from 30 to 50 per cent. of the population infected; which means, since the disease so far as known is invariably fatal, that at least a third of the people in such districts will probably die of trypanosomiasis. That this is far from exaggerating the conditions of things is evidenced by the history of the spread of the disease in Uganda, where in a few years hundreds of thousands have died of the infection and whole regions have been depopulated. It has been shown conclusively that sleeping sickness is con- veyed by the bite of a tsetse-fly, Glossina palpalis. This species is different from that which carries the nagana of South Africa. Whether other species of this genus can convey the disease has not been established. In all probability, as in nagana, the fly is a mere vector, a mere mechanical means of carrying the trypanosome from the sick to the healthy persons. The pres- ence of multiplication forms of trypanosomes in the stomachs of such flies (Gray and Tulloch, Koch) has been taken to show that the fly is not a passive carrier of the organism. ‘The tsetse trypanosome, however, has not been shown to be identical with the human parasite—in fact, there is reason to believe that they are in no wise related, and that the former (Tr. Grayi, Novy) is a harmless parasite peculiar to the fly. Human trypanosomiasis is characterized by two stages. In the first the trypanosomes exist in the blood but always in small numbers. An irregular remitting fever is the chief symptom of this stage. The pulse and respiration are accel- erated. Slight edemas and erythemas are at times met with and in addition enlarged glands and spleen. Owing to the mildness of these symptoms the disease passes unnoticed among the natives. The second stage follows after the lapse of a variable length of time. It is this stage which is known as Fic. 9.—Trypanosoma gambiense (human _ try panosomi- asis or sleeping sickness) in blood of a rat. Two types are shown; the broad pale form (female ?) isdividing. Magni- fication 1,500 times. MacNeal’s stain. Fic, 10.—Trypanosoma gambiense from same preparation as preceding, showing the usual form, some of the cells in process of division. Magnification 1,500 times. ON TRYPANOSOMES 67 sleeping sickness. The fever is marked especially toward even- ing. The patients become dull and apathetic and complain of intense headache. Weakness of the arms and legs develops, speech becomes difficult, and emaciation sets in. Somnolence increases and a comatose condition supervenes with death. The Tr. gambiense is present in but small numbers in the blood of man. Most of the experimental animals are subject to infection and in the blood of such it may become very numerous. The course of the disease in monkeys is at times very, suggestive of that in man. The baboon has been supposed to be refrac- tory but Thomas and Breinl have succeeded in infecting four of these animals. The strain isolated from one of these proved to be highly virulent. The macaques are quite susceptible, and die in from one to two months. In the horse and donkey the infection is very chronic, with very few parasites in the blood, and recovery seems to occur. The cow is even more refractory than the horse. Sheep contract also a mild infection and recover. Goats are apparently more susceptible and death may result (Thomas and Breinl). Dogs are easily susceptible but may survive for six or even nine months, though death may occur in from five to six weeks. Cats are likewise subject to infection and especially kittens, in which the parasites appear in large numbers and cause death in from three to seven weeks. In rabbits and guinea-pigs the trypanosomes are very scanty, especially in rabbits, and the infection is very chronic, lasting for several months. In mice the infection is also very slight and recovery may take place. On the other hand white rats are quite easily infected. As a result of intraperitoneal injec- tion we have seen the parasites appear in the blood within three days, though the period of incubation is usually given as about fifteen days. The parasites may increase enormously in numbers for a while and then almost completely disappear from the circulation (Figs. 9 and 10). Death occurs in from one to three months. Morphologically the Tr. gambiense resembles very closely Tr. Brucei and Tr. Evansi. The cultivation of the organism has been attempted by Laveran and Mesnil and also by Thomas and 68 HARVEY SOCIETY Breinl. The former were able to keep it alive on blood agar for nineteen days but were unable to obtain sub-cultures or to infect rats with such material. The latter succeeded in main- taining it for sixty-eight days but they also failed to obtain actual sub-cultures. Normal human serum which possesses a pronounced action on the trypanosomes of caderas and nagana is without effect on Tr. gambiense. According to Thiroux the serum of cases of sleeping sickness in which the blood is free from trypano- somes possesses a slight protective action with respect to mice. The treatment of experimental animals with arsenic (atoxyl), trypan-red and other anilin dyes (Mesnil and Nicolle) has given very encouraging results. With the aid of an alternate treatment with arsenic and trypan-red Laveran was able to cure monkeys. With the exception of one case in a woman, reported by Dutton and Todd, the treatment of the human eases has thus far been ineffectual. KALA-AZAR. A brief consideration must be given at this place to a peculiar organism which is present in Kala-azar and the cachexial fever of India, especially since recent studies go to show that it is a flagellate and closely related to the trypanosomes. In 1903 Leishman found certain bodies in the spleen of a fatal case of the disease and surmised that they were degenerated forms of trypanosomes. Very shortly after Donovan at Madras con- firmed this finding but he was unable to get any trace of try- panosomes. lLaveran, to whom specimens were submitted, pro- nounced the parasite to be a Piroplasma and gave to it the name Piroplasma Donovan. Ross regarded it as representing a new genus and for that reason he called it Leishmania Donovan. The investigations of Rogers, which have since been con- firmed by others, have thrown much light upon the nature of this organism. He found that when the fresh blood, obtained by spleen puncture, was transferred to test-tubes containing a few drops of 2 to 5 per cent. citrate of soda in normal salt solution, the parasites remained alive for many days, and after ON TRYPANOSOMES 69 about three days some of them developed into elongated flagel- lated bodies which he took to be trypanosomes, although no undulating membrane could be detected. Rogers has since found that the flagellation took place more uniformly and regularly if the citrated spleen blood was faintly acidified with citric acid. The flagellated forms develop best at about 22°, the same as in the case of the cultural trypanosomes. ‘The Leishman-Donovan bodies it may be said resemble greatly rounded up forms of trypanosomes. They show a nucleus and a micro-nucleus. In the citrated blood these forms increase in size, elongate, and give off a flagellum. The latter starts from the blepharoplast which lies close to the anterior end of the cell. It is perhaps on account of this close proximity to the end that, as is the case of the mosquito trypanosomes, no undu- Jating membrane can be made out. At all events, on account of the absence of this structure, Rogers has recently come to the conclusion that the organism belongs to the herpetomonas group and not to the trypanosomes, and he has designated it as the Herpetomonas? of Kala-azar. The fact that we have in this case an undoubted flagellate developing from the Leishman-Donovan bodies goes to establish a certain relationship between this disease and those which have been heretofore considered. The exact position of the organism can only be determined by further study. It certainly presents some of the cultural characteristics of the bird trypanosomes and more especially of the mosquito flagellates. It may be added that, while ordinarily the mosquito herpetomonas fails to show an undulating membrane, in some cultures evidence of such can be observed, and hence the apparent absence of this structure does not necessarily exclude the organism from the group of trypanosomes. As to the transmission of this fatal disease nothing definite can be stated. Rogers is of the belief that the common bed- bug or possibly mosquitoes are the most likely hosts. By allow- 1 Through typographical error this genus is given as Hepatomo- nas in Rogers’ paper. 70 HARVEY SOCIETY ing mosquitoes to bite a patient Patton has been able to find in their stomachs flagellates, herpetomonas and crithidia, such as we have described in these insects, and consequently these forms cannot be considered as stages of this parasite. From a private communication it appears that Patton has succeeded in finding developmental forms of the parasite in the common bed-bug. TRYPANOSOMES OF OTHER ANIMALS. In the foregoing an effort was made to give a brief résumé of the mammalian trypanosomes, particularly of the pathogenic species. These organisms, however, are by no means limited to the mammals, but on the contrary, they may be found in almost all forms of life down to the insects. Some of the earliest observations made upon trypanosomes were on those of the frog. At the present time a number of species are known to occur in these animals. They are nearly all characterized by their great bulky size, although long and slender as well as short forms are known. Williams and Lewis obtained a successful initial culture of the 7'r. rotatoriwm, while more recently Bouet has given a detailed study of the cultural forms. Not only batrachians, such as frogs, but also reptiles, as turtles and snakes, have been shown to harbor trypanosomes in their blood. Of particular interest perhaps are the flagellates present in the blood of fish either from fresh or from salt water. Of these two genera have been recognized,—Trypanosoma and Trypano- plasma. The latter genus was created by Laveran and Mesnil and includes forms which differ from the true trypanosomes in having a posterior as well as an anterior free whip. The trypanoplasmata have been studied, particularly by Laveran and Mesnil, Brumpt, and by Keysselitz. The latter has observed a double infection with the two genera in fourteen species of fish. Although a considerable number of species of fish trypanosomes and trypanoplasmata have been described, Keysselitz regards the latter in the fish studied by him as rep- resenting but one species, T'rypanoplasma Borreli. The percent- ON 'TRYPANOSOMES 71 age of naturally infected fish cannot be readily given, but there is reason to believe that it is very large. As in the case of birds, the flagellates may be present in but very small numbers and hence escape detection. The infection is undoubtedly spread among fish through the agency of blood-sucking parasites and more especially the leeches. The studies of Brumpt and of Keysselitz have shown that a large percentage of the leeches contain variable numbers of flagellates in their intestinal canal. Owing to the many diffi- culties attending such investigations it has not been possible as yet to prove definitely that the flagellates observed in the leech are derived from those in the blood of fish or that con- versely the fish flagellates develop from those multiplying in the gut of the leech. Nevertheless, it has been assumed and it is quite generally accepted, that the leeches are the intermediate hosts of the parasites. Keysselitz, and especially Brumpt, has succeeded in infecting fish by placing on them infected leeches. It is an interesting fact that flagellates are present in the gut of many insects, irrespective as to whether these feed on blood or otherwise. The herpetomonas of the house-fly is an example of infection of a non-biting insect. Many similar observations could be given to show that intestinal parasitism by flagellates is a common occurrence. We have shown that 15 per cent. of the wild mosquitoes may be infected with herpe- tomonas and crithidia, while Ross, Legér and others have shown similar parasites in the larval and pupal stages. These two organisms when grown on blood agar retain the same form as observed in the insect, thus demonstrating that they are cultures m vivo and, as such, that they are really multiplication forms of trypanosomes and not distinct genera. It follows from the above that much caution must be used in drawing conclusions as to the relation of flagellates found in insects to the blood trypanosomes or to the intracellular para- sites found in vertebrates. As has been pointed out, the try- panosomes of tsetse-flies are not to be regarded as multiplication forms of Tr. Brucei and Tr. gambiense, and the same con- clusion holds for Schaudinn’s views regarding Tr. noctue and 72 HARVEY SOCIETY Spirocheta Ziemann, which two forms were considered as flagellate stages of the halteridium and leucocytozoon of the owl. The foregoing summary of the trypanosomatic infections would be incomplete without a brief reference to the flagellates of birds. These were first studied by Danilewsky, who described a large and a small form of Z7'r. avium (1885). Since then these forms have been found by a number of other invyes- tigators, and for the details of this work the reader is referred to the monograph on ‘‘Bird Trypanosomes’’ by Novy and MacNeal. It has been shown by us that flagellate infection of birds is exceedingly widespread and that it can be recognized best by the cultivation method. A number of species have been shown to exist in our common birds. These can be readily cultivated and the characteristics presented by the cultures readily permit the differentiation of species. Since then Thiroux has been able to grow the Zr. paddw and Cerqueira has been equally successful with the trypanosome of the Nicticorax of Brazil. The most important result of these studies on bird trypanosomes has been the demonstration that these flagellates are in no wise related to the intracellular parasites. AUTOLYSIS* P. A. LEVENE, M.D., Rockefeller Institute for Medical Research. TISSUE DISINTEGRATION. HE most constant property of living matter is disinte- gration. This precedes every other manifestation of life in the simplest and in the most complex animated organism. The development of a fertilized egg begins with disintegration of its original structure. When the egg is placed under con- ditions which make the initial disintegration impossible, life remains suspended. Disintegration also is the most lasting property of living matter, for when all other functions are extinct this property is still in evidence, and, if conditions are favorable, the disintegration proceeds until all evidence of organization or of structure disappears, giving place to a mix- ture of organic and inorganic substances. Disintegration of organic matter present in the cells and tis- sues is also the primary source of that form of vital energy which controls most functions of the living organism. Con- traction of muscle, secretion of glands, peristaltic movements of the gastro-intestinal tract, growth and reproduction of an organ- ism, are possible only so long as the breaking-down process continues. This has been realized by physiologists at all times. However, the mechanism and the exact nature of the chemical reactions associated with animal functions have never been very clear, and even to-day the question remains a topic of consid- erable controversy. Lavoisier was the first to emphasize strongly the similarity between the chemical reactions in the living organism and those in the process of combustion. Ever since that time physiolo- gists have adopted the term combustion in order to signify * Lecture delivered November 18, 1905. 73 74 HARVEY SOCIETY the chemical reactions in the organism which result in the production of animal energy. Physiologists speak of burning or non-burning in the human or in the animal body of proteid, fat or sugar, of burning of the body tissues and of body cells. Indeed, the ultimate products of the reactions in the body are very similar to those from the burning of carbonaceous material. Carbon dioxid, water and heat are formed in both instances. However, there has always existed an utter lack of information regarding the agents causing the powerful oxidation of organic material in the animal body. The simplest way out of the difficulty seemed to ascribe the power of combustion to a peculiar property of the living eell. In recent years there has accumulated a great number of observations tending to show that various functions previously regarded as the result of life, as the result of cell assimilation and disintegration by the animal tissues, are actually occasioned by substances which ean be isolated from the living cell. An instance illustrative of this statement is found in the work on alcoholic fermentation. The formation of alcohol from grape sugar by the yeast cell was regarded as a chemical reaction brought about by the activity of the cell. Alcohol thus was considered a catabolic product of cell metab- olism. Mme. Manassein, and, more convincingly, Buchner, have demonstrated that one substance from the cell can be obtained which is capable of accomplishing the alcohohe fer- mentation of sugar in the same manner as the living cell. This startling discovery marks a radical change in our conception of the process of life. For centuries it was thought that the activity of a complex organism was needed to manufacture spirits out of grape sugar, and the foregoing work demonstrated that the cell may be crushed, its life may be extinct, and one substance soluble in water may be extracted which possesses the power to accomplish the work of the entire cell. True, the substance is of a very subtle nature and needs to be handled with great care. It does not resist the action of heat and other strong chemical and physical agents, but while intact it is capable of inducing chemical reactions into which it apparently AUTOLYSIS 75 does not enter itself, and thus is capable of performing work out of proportion to its own mass. Substances endowed with this power are designated enzymes or ferments. The work of Buchner caused a very intense interest to be directed toward the older and unsystematic work on the enzymotic processes in the cells of the simple and complex organisms. As already stated, the source of all vital and animal energy hes in tissue disintegration, and the prevailing conception has been that the disintegration was brought about by the power of the living cell to burn its own components. It is the great merit of Salkowski to have shown that a cell or tissue in which all visible signs of life have disappeared still retains the power of self-dissolution, of self-disintegration, of autolysis. True, the phenomenon had not escaped the observation of earler workers, and in 1871 Hoppe-Seyler wrote:+ ‘‘ All organs suffer- ing death within the organism, in the absence of oxygen, undergo softening and dissolution in a manner resembling that of putrefaction. In the course of that process, albuminous matter gives rise to leucin and tyrosin, fat to free acids and soaps. This maceration, identical with the pathologic concep- tion of softening, is accomplished without giving rise to ill odor, and is a process similar to the one resulting from the action of water, acids and digestive enzymes.’’ In 1874 the French chemist, Schutzenberger 2 observed similar changes in yeast which had been allowed to remain for from 12 to 15 hours in water suspension at a temperature of from 35° to 40° C. However, before going into the details of chemical analyses, attention may be called to the structural, morphologic changes which cells and tissues undergo when they are placed in con- ditions which do not permit continuation of life. It is well known that animal tissues and organs are readily invaded by micro-organisms, causing putrefaction. The application of 1 Tiibinger Med. Chem. Untersuchungen, 1871, p. 499. 2 Compt. rend., vol. xxviii, and Bull. de Soe. Chemique, vol. xxi, p. 204 76 HARVEY SOCIETY the recent methods of aseptic surgery allows the removal of organs from the animal body and the preserving of them free from all contamination with micro-organisms. Hauser, as well as Rindfleisch and Meissner previous to him, succeeded in preserving tissues for months and years free from infection with any bacteria. In organs kept in an absolutely sterile con- dition Hauser observed general softening, and microscopically he noted the destruction of the most typical structural part of the cell, the nuclear material, and decay of the mass of the cell, made apparent by the development of changes which are designated by pathologists as fatty degeneration. Thus the term softening is not merely a figure of speech, but applies to an actual occurrence. This is made evident particularly through the work of Schutzenberger and the work- ers who followed him. Normal fresh organs on extraction with boiling water give off only a small fraction of their constituents, while those that have undergone the process of softening allow a very considerable part of their substance to pass into the boiling water. Thus, fresh yeast on boiling with hot water leaves a residue consisting of from 20 to 21 per cent. of its original weight, while the residue of yeast kept in water for from 12 to 15 hours does not exceed 13 per cent. However, in the experiment of Hauser, although the tissues were placed in conditions unfavorable for continuation of life, death set in slowly, and the possibility is not excluded that the softening was accomplished by the vital force not yet completely extinct. In the experiments of Schutzenberger also this possibility was not excluded; besides, yeast always contains bacteria and it is difficult to separate the part of the changes wrought by the action of micro-organisms from that induced by the surviving yeast cell. Salkowski was the first to preserve the material employed in his experiments under conditions which checked all fune- tions but that of dissolution, bacterial growth being impossible. This was achieved by the use of chloroform water instead of the 3 Arch. f. exper. Path. u. Pharm., vol. xx, p. 162, 1886. AUTOLYSIS 77 pure. Salkowski repeated the experiments of Schutzenberger on yeast and arrived at the same conclusions as the first observer. He extended the work to animal tissues, using the liver and muscle. ‘The results are best seen in the following table: From 1000 grams of liver Autolyzed Control Difference were extracted by hot water organ Organic substance ..... 45.97 gms. 33.73 gms. 12.24 gms. PE PA bye Dae Maa ck ef 7.95 gms. 7.21 gms. 0.74 gms. Phosphoric acid ....... 1.957 gms. 1.359gms. 0.598 gms. Nitrogen in form of nitrogenous substances 6.239 gms. 3.152 gms. 3.087 gms. In the main experiment the finely-divided organ was mixed with three times its weight of chloroform water and allowed to stand. At given intervals analyses were made. In the con- trol experiment the organ was heated and then further treated in the same manner as in the principal experiment. The table clearly shows that, on standing, substances soluble in hot water have developed in the organ. Very similar changes occur in tissues subjected to the influence of digestive enzymes, either in the digestive tract or outside of the body. Because of this analogy Salkowski introduced the term ‘‘self-digestion’’ in order to designate the process occurring in tissues allowed to stand under antiseptic conditions. For reasons which will be made clear in the course of the discussion, the process later was named by Hofmeister ‘‘ autolysis.”’ Thus the researches of Salkowski have established the fact that tissues, placed in conditions which do not allow contamina- tion with living matter, undergo changes resembling those occurring during the course of digestion; but they offered no information regarding the réle played by the process in the economy of the organism, in those transformations of matter which create and maintain life. It was still undecided whether or not the capacity of self-digestion was a universal property of all tissues. The probability was not excluded that the autolysis of an organ was brought about by the action of enzymes ab- sorbed from the gastro-intestinal tract and transported to the various organs. The researches following those of Salkowski 78 HARVEY SOCIETY endeavored to give an answer to these questions. The solution of the first problem was comparatively an easy matter. It was necessary only to repeat his experiments on various other organs. This was accomplished most successfully by the efforts of Hedin and Rowland.* It may be noted here that the last two investigators employed in their experiments not the entire tissue nor the tissue extracts, but the plasma of the organs. In this manner they made certain that no cellular elements were playing any part in their experiments, and that the reactions were caused by a soluble substance present in the plasma. Previously Schwiening,°® a pupil of Salkowski, had established the same fact by employing filtered tissue extracts. The work was further ex- tended by Martin Jacoby ® and by Stookey and myself.* As a result of all the work it may be regarded as established that the power of self-digestion is shared equally by all organs. The solution of the second problem, namely, of the origin of the autolysing power, required more ingenuity and perhaps more work. Attempts were made to obtain the desired information in various ways. If the digesting power present in the organs be due to a substance derived from the pancreas, the autolysis of organs must be influenced by the same factors and in the same manner as pancreatic digestion; further, if that assump- tion be correct, one would expect to find among the products of autolysis those substances which arise on tryptic digestion. The chemical composition of animal organs is very complex, but the pancreatic gland is capable of disintegrating all the principal tissue constituents, although it resorts to a different mechanism, perhaps to a different substance, for the digestion of the individual substances. The principal components of tissues are albuminous material, carbohydrates and fats. In the course of self-digestion all these components are disinte- erated, and it is a matter of convenience to discuss separately 4 Zeitsch. f. phys. Chem., vol. xxxii, 1902. 5 Virchow’s Archiv., exxxvi, 1894. 6 Zeitsch. f. phys. Chem., vol. xxx, 1900. 7 Jour. Med. Research, vol. x, 1903. AUTOLYSIS 79 the change which each of the components undergoes in the course of autolysis. Of all enzymotie processes, that resulting from the breaking up of the proteid molecule has been studied in the greatest detail. For this reason the study of the proteoly- tic action of organs was employed for the investigations into the origin of the autolytic power of tissues. Two proteolytic enzymes of distinct individuality have always been known—pepsin, elaborated by the glandular apparatus of the stomach, and trypsin, formed in the pancreatic gland. The principal point of distinction between the two substances is that one requires for its action the presence of acid, while the other is most active in the presence of alkali. Further, it has generally been accepted that pepsin is incapable of pro- ducing the same degree of cleavage as trypsin. The formation of erystalline products of amino-acids has been noted only on tryptic digestion. Most typical for the cleavage by the ferment of the pancreatic gland is considered the appearance of a substance giving a peculiar color test with bromin, named tryp- tophan. In the course of digestion by either of the two en- zymes, albumoses and peptones are formed. Biondi, a student of Salkowski, has noted that the pro- teolytic action of the liver is facilitated by the presence of acids. This difference in intensity of digestion under the two different conditions is made very conspicuous by the following table : Out of 1000 grams of liver Experiment 1 Experiment 2 passed into solution with 0.28% HCl without HCl Organic substances ........... 100.10 gms. 59.0 gms. OS ats et REGa Eres tao A ee 26.90 gms. 11.12 gms. N. in nitrogenous substances... 11.76 gms. 7. gms. PEETIOEE: bien iste aie e's cidsce baw a Trace. Trace. BOGE: -o Meer oes ck es ete > None. None. The conditions influencing the intensity of autolysis were studied in greater detail by Hedin and Rowland,* whose in- vestigations were made on tissue plasma obtained by Buchner’s method. It was established by these writers that the self-di- gestion of the majority of organs is facilitated by the presence 80 HARVEY SOCIETY of 0.25 per cent. of acetic acid and is depressed by the presence of alkalies, by calcium carbonate and magnesium oxid. The only deviation from this, according to Hedin and Rowland, is in muscle tissue, where the intensity of digestion is not affected by the presence of alkali or acid. On the other hand, cardiac muscle is subject to the general rule of autolysis. The autolysis of nerve tissue, and of the testes also, is facilitated by the presence of acid, as was demonstrated by Stookey and myself. These observations are important, for the reason that they make very improbable the supposition that self-digestion of tissues is caused by trypsin deposited in the organs by the blood supply. On the other hand, Salkowski, in his early work on autolysis, has noted the appearance of leucin and tyrosin, and in this respect the proteolytic action of animal tissues resembles tryptic digestion. Contradictory to this seemed the observations of Biondi.* This author could not detect tryptophan in the experiments in which the absence of bacterial growth was made certain. Another peculiarity of the autolytic cleavage noted by Biondi is the comparatively insignificant formation of al- bumose and of peptone. Jacoby also, in his very exhaustive study on autolysis, invites special attention to the foregomg difference between tryptic and autolytic digestion. On the other hand, Jacoby demonstrated tryptophan among the prod- ucts of self-digestion of tissues. Thus the chemical process of autolysis bears some resemblance to either form of digestion, peptic and tryptic, and yet is different from each of them. This alone makes it very probable that animal tissues do not borrow their power of disintegration from either gastric or pancreatie gland, and that self-digestion is one of the general properties of living or, rather, surviving organs. Additional evidence in support of these assumptions was brought forward by Matthes.* It is well known that urine of normal individuals contains a proteolytic enzyme resembling 8 Virchow’s Archiv., vol. exliv, 1906. ® Archiv. f. exp. Path. u. Pharm., vol. li, 1904. AUTOLYSIS 81 pepsin. Matthes demonstrated that after the removal of the stomach of dogs the enzyme ceases to be eliminated by the urine. It was natural on the basis of this experiment to view the stomach as the source of the urinary pepsin. ‘The same method of investigation was applied by Matthes to the study of the origin of the self-digesting power of organs and tissues. Dogs were deprived of their pancreas and allowed to recover from the operation. The organs were then examined for their proteolytic power. No difference could be detected between the organs of the normal and those of the operated animal. Thus, all evidence seemed unanimously to support the view that self-digestion is a constant property of surviving tissue. However, for the interpretation of the rédle of this function in the economy of the living organism, it still remained to be established whether or not the process of self-disintegration takes place also in life. Jacoby ° was the first to give experimen- tal trial of the question. For this purpose he performed on dogs the following operations: The hepatic artery and the portal vein were ligated and, after several hours, the liver was extirpated and analyzed for amino-acids. lLeucin and tyrosin were found to be present. Further, he obtained the same results on ligating a part of the liver. These substances were also obtained by Jacoby from organs extirpated asepti- eally and kept under conditions in which contamination was impossible. It may be remarked that all these methods are open to some objections. More convincing seems to me the analysis of the developing organism. It has been known for some time, through the work of Schulze and his pupils, that in the course of germination and growth of plants, substances appear which arise also on proteolytic digestion of the seeds. I have made a similar observation on the developing egg of fish and of fowl.t° In the course of development of the egg one can notice the breakdown of the albuminous matter and the appearance of products of the nature of nitrogenous acids. 10 Zeitsch. f. phys. Chem., vol. xxxv, 1902. 6 82 HARVEY SOCIETY So, at the present time, there is sufficient evidence for the assumption that disintegration or self-digestion is a constant occurrence in living as well as in surviving tissues. However, there is still a lack of information regarding the role of this function in the mechanism of life. In the animal tissue, organ or cell, one has to distinguish two different parts, one representing the organized mechanism controlling its function, the other consisting of various organic substances stored up or deposited in the organs, as a supply of fuel material. Blood plasma and lymph, which envelop every part of the organ, are not integral parts of its tissue. They only furnish the material which the organ may or may not use. White of an egg and the greatest part of its yolk are only building material for the developing organism. In physiology there are two views regarding the production of animal energy. One is that a substance can not be utilized by a living cell unless it has been assimilated and transferred into organized cell substance. Liebig was the author of this theory and Pfliiger most vigorously defended it. On the other hand, Carl Voit claimed that in higher organisms the principal supply of fuel material is furnished to the organs by the blood. The albuminous matter carried to the organs was named by Voit ‘‘ circulating proteid.’’ Opinions on the subject are still divided and it is possible that in a way both views are correct. Since there was some foundation for the view that the pro- cess of autolysis is the one which controls tissue disintegration, it seemed important to make clear whether or not the mecha- nism is capable of breaking down albuminous matter derived from other sources than that of its own body substance. The first observation in this direction was made by Theobald Smith, who noted that fresh tissues removed from the organism under aseptic conditions were capable of digesting gelatin. On the other hand, Martin Jacoby “ noted that during the process of liver autolysis, of the proteids only the globulins suffered a 11 Zeitsch. f. phys. Chem., vol. xxx, 1900, also vol. xxxui, 1903. AUTOLYSIS 83 disintegration; and in a later work he observed that the self- digesting liver was completely incapable of digesting lung tissue. Thus, on the basis of this work, one would be led to the view that the process of autolysis is incapable of causing the digestion of circulating proteid, and that the two processes are totally independent of each other. However, Hedin * has shown that the spleen possesses the power to digest not only its own proteid material, but also the proteids of the blood. Thus the question still remains an open one. PRODUCTS OF TISSUE DISINTEGRATION. The work thus far reviewed possessed primarily theoretical interest only. It aimed to elucidate the mechanism controlling the disintegration of tissue components in the living and in the surviving organs. Nevertheless a detailed knowledge of the products of tissue autolysis is of importance from the stand- point of practical medicine. In the human organism, as well as in that of many animals, all substances which are consumed as food and nourishment, no matter how greatly they differ in their chemical composition, are finally broken down into a few very simple bodies, which are rejected by the organism through the kidneys, bile and other excretory mechanisms. Urea and carbonic acid are the two substances into which nearly all food- stuff is transformed. In a complex organism the metamorphosis is a gradual process. Before a nitrogenous substance is trans- formed into urea it undergoes numerous degradations. Before sugar is oxidized to carbonic acid it suffers numerous changes. Further, it is not improbable that in a very complex organism individual organs are concerned only in one definite phase of the transformation, leaving the other organs to continue and to complete the work. In his recent address on the subject, Professor v. Noorden ?? pointed out that the information re- garding the nature of intermediate products of metabolism, as well as the seat of their formation, is lacking. Attention of investigators has turned to the study of the products of autoly- 12 This volume, page 18. 54 HARVEY SOCIETY sis of various organs in the hope of filling in the gap in our knowledge of the mechanism of nutrition and of self-preserva- tion of the organism. However, the study of the substances arising in the course of autolysis was preceded by very active work on the normal composition of tissues and tissue components. Indeed, it was to be expected that within the body, tissue constituents would break down into their component parts. Recent years are marked by astonishing progress in the knowledge of the chemi- eal nature of tissues. It was owing to this progress that the study of autolysis was made a comparatively easy matter. -As already stated, the principal tissue components are albuminous matter, sugars and fat. The changes which each one of these components undergoes in the course of self-digestion have been the subject of special investigation. Under the term proteid is generally understood the sub- stance which represents the most important and most character- istic part of living matter. It is colloidal in nature and is composed of various nitrogenous acids. On heating proteid with strong acids or alkalies, the original substance disappears, giving rise to the nitrogenous acids. Of those already known are the following: Glycocoll. Lysin. Alanin. Arginin. Aminovalerianie acid. Histidin. Leucin. Prolin. Glutamic acid. Tryptophan. Phenylalanin. Cystein. Tyrosin. Of the proteids, one group attracts special attention. Its members are present in greatest quantity in the nuclei of all cells, and it has been assumed that the function of the nucleus is closely associated with the presence of these substances. They are named nucleins, nucleoproteids, nucleoalbumins, ete. They are more complex than ordinary proteids, containing in their molecule, besides the usual constituents, a body termed nucleic acid. This acid is composed of substances to which a consider- able role in the pathogenesis of disease has been attributed. AUTOLYSIS 85 Its components are as follows: Phosphoric acid, carbohydrate, thymin, uracil, eytosin, adenin, guanin, hypoxanthin. Normally, components of simple and complex proteids occur as such in tissues in very insignificant quantities. But it is found that in the course of self-digestion an organ may underge such deep changes that nothing remains of its original structure. in its place the following substances appearing: & : q = o n ee ie Ate cy 4 aD Kd & SOM nannies ae6'n, 0.8 wide alas — — — — — PAT oe dm lala i Sesceaia es -L -t- ate ae ate Aminobutyriec acid ........ ++ <= ao a - Aminovalerianie acid ....... ? 2 + 2 + PIN Lg a seas S aca 8 ate es a 1. ot -E + OFIGATMNC BOM oe ess ce ewe e's -|- 4. ate ~- a RAPHIC ACID o iilicic cei’ oes — = a al. + Pyrrolidin earbonie acid ..... 2 2 + a= -— RE Ae i a winx oie 9. 6a s ws + + 4- = —: SCC ao i ae ~ + + + + A glance at the table shows clearly that the action of the autolytic process in organs is as powerful as that of strong acids combined with high temperature. Nearly all the products which are obtained on prolonged boiling of proteids with strong mineral acids arise also in the course of autolysis. However, there are noted some differences in the two processes. If it be allowed to name substances appearing on cleavage with mineral acid as primary cleavage products, the distinction may be made that on autolysis the primary products undergo further trans- formation. It is a matter of convenience to discuss the points of difference according to the three principal groups of sub- stances in which they occur, namely: 1. The nitrogenous acids containing only one nitrogen in their molecule, monoamino- acids. 2. Acids with more than one nitrogen in the molecule (The substances of this group arising from proteid cleavage were named by Kossel hexon bases. They generally possess basic properties). 3. Substances resulting from the nuclear degradation, nuclein derivatives or nuclein bases. The most 86 HARVEY SOCIETY appropriate method for investigation was: first, to study the products obtainable on boiling organs with strong acids; second, to study those arising on autolysis of the same organs and, finally, to analyze the substances appearing on boiling with strong acids of organs previously subjected to self-digestion. On acid cleavage all the amino-acids are obtained which are known to appear on the breaking down of proteid material. Among the end-products of self-digestion of the pancreas, Emerson ** discovered oxyphenylethylamin, which is not known to be present in the proteid molecule, and which may be re- garded as a secondary product derived from tyrosin. Further, on autolysis of various organs the formation of glycocoll was not observed, and prolin could be demonstrated only in a few experiments. It should be remarked that the present methods of analysis of amino-acids are not fully satisfactory, and too much weight should not be attached to the results thus far obtained. However, the results of the analysis of the amino-acids obtained from the fresh and from the self-digested glands seem to indicate that in the course of the latter process some destruction of the substances takes place. This may be seen from a table showing the results of experiments not yet published, although completed: Fresh Spleen Autolyzed Spleen 5 pounds 5 pounds Glycogolll sce tg eee ae 0.700 0.700 Adana 5 oer. chee meet eet 8.6 Lit Aminobutyric and aminovalerianic ACIGS. Vranas cata se ee Recs 5.25 5.00 PieU eo Sea as Os ete aC 14.75 12.0 Aspartie eld (c.f ahe trae ore 2.24 0.8 Glutaniie Sate! This author accepts the presence in the tissues of two specific enzymes, one capable of acting on guanin and the other on adenin, and further of an oxidizing enzyme, the function of which it is to complete the transformation of nuclear material. The deductions of Schenek** are similar to those of Jones, and Schittenhelm ”° has corroborated in a gen- eral way the same conclusions. However, he does not support the assumption of the existence of more than one enzyme which ean transform the amino-purins into the corresponding oxy- derivatives. According to Schittenhelm, the entire nuclear destruction is accomplished by three enzymes, one breaking up the nucleic acid into its components, the second splitting off the nitrogen from the nitrogenous constituents, and the third com- pleting the oxidation of the purin derivative. It must be admitted that more detailed information concerning the intermediary products of nuclein metabolism is still wanting. With this I wish to conclude the review of the products arising on autolysis of surviving organs. Reference should be made to the products of autolysis of sugar and fat, but thus far the investigations in that direction are few in number and the results obtained from them not very significant. This also concludes the review of autolytic action in normal organs. It remains to discuss these results in connection with the original problems which led to all these numerous investigations. DISCUSSION OF RESULTS. It has been stated already that the principal object of the work was: 1. To elucidate the nature and the mechanism of those chemical reactions which make the functions of the body possible. 2. To interpret the role of individual organs in the 26 Zeitsch, f. phys. Chem., vol. xliu, 1904, 92 HARVEY SOCIETY animal metabolism. 3. To study the intermediate products of metabolism, since there is a general agreement that by the accumulation in the organism of these substances many diseased conditions are occasioned. The foregoing review leads one to the conclusion that the knowledge of intermediate metabolism has been furthered considerably. On the other hand, a comparative study of the products of disintegration of various organs fails to bring out marked differences among them, although, during life at least, some organs are known to be the seat of special chemical reac- tions. This leads one to the assumption that in the animal body the process of self-digestion does not control all chemical reactions occurring in organs, perhaps even not all the processes of disintegration. A most conspicuous instance illustrating this statement is found in the work in which Eck’s fistula was employed. By this name is designated a fistula between the vena cava and the portal vein. The aim of the fistula is to exclude the liver from the portal circulation. The organism of the dog possesses a very intense power of burning uric acid; the acid is present in the urine of this animal only in traces, even after injection of two to three grams of the substance. Dr. Sweet and I have demonstrated that animals kept for weeks on a diet free from all precursors of uric acid excrete consider- able quantities of it in the urine as soon as an Eck fistula is performed on them. The output is especially increased after the administration of the substance itself. Evidently under these conditions the organism fails to dis- integrate urie acid, although the process of self-digestion is not depressed in the tissues. Thus the mechanism of ‘‘burn- ing’’ uric acid in the living organism is not known yet. Pro- teid combustion also in the normal living organism apparently is different from the proteid disintegration in the course of autolysis. The great mass of products in the process of self- digestion remain in the stage of nitrogenous acids. A small part of them lose their nitrogen and a still smaller part give rise to carbon dioxid. In the living organism the splitting off of nitrogen from proteid material is a very rapid process, and AUTOLYSIS 93 the transformation of all carbonaceous material to carbon dioxid also occurs with much greater rapidity than it possibly could take place in the course of self-digestion. But it has been stated that disintegration by the process of autolysis does occur during life. This and the foregoing are not contradictory to each other. Every tissue consists of cells of different age, of dif- ferent states of nutrition and of different resistance. Work on hemolysis has brought out most clearly that individual blood cells vary in their vulnerability. Cells in a state of defective nutrition succumb to the process of self-digestion. In the course of that process enzymes are liberated which are capable of digesting extraneous material also. It is difficult to demonstrate the correctness of this view on a complex organism, but it is made very clear from observations on the yeast cell. It is the function of that organism to convert grape sugar into alcohol. So long as conditions for this func- tion are favorable there is little evidence of the process of self- digestion in a colony of yeast cells. But as soon as conditions are so altered as to make the normal life and the alcoholic fer- mentation impossible, a very active proteid-splitting enzyme is developed by the yeast cell which causes digestion of the cell proteid and of other proteid material. In the living organism the two forms of metabolism undoubtedly coexist. One is the result of the function of the organs, the other of their disinte- gration. The supply of energy required for the maintenance of life is furnished possibly by the first process. It has already been stated that by means of autolysis proteid is converted prin- cipally into amino-acids. By this conversion proteid could not furnish the organism with its full calorific requirement. On the other hand, autolytic enzymes may act on cell proteid and on the surrounding proteid in a manner similar to that of the enzymes of digestive glands, namely, rendering them a more suitable material for rapid combustion. It is marvelous that, notwithstanding the presence of destruc- tive agents in all tissues, organs succeed in guarding their integrity. A most clever investigation of recent years throws light on the mechanism by which this is accomplished. The 94 HARVEY SOCIETY integrity of the gastric wall, the function of which is to elab- orate digestive enzymes, has been the cause of much speculation. Weinland ** demonstrated that this was due to the presence of an antiferment in the digestive glands. In the blood also were found antitryptic substances by Hahn,?* Landsteiner,”® Glaessner *° and Catheart.*t The same property was noted in tissue extracts by Dr. Stookey and myself.’ Furthermore, we have demonstrated that tissue extracts exercise an action antagonistic to that of autolytic enzymes. However, in health the two tendencies are so regulated that the tissue disintegra- tion is sufficient to permit the organs to perform their function, while excessive wear is avoided. But as soon as the normal nutrition of the organism is disturbed the autolytic power of tissues increases. The mere fasting of an animal suffices to occasion in the tissues an exaggerated tendency for self-destruc- tion. This was demonstrated by the experiments of Lane- Claypore and Schryver. It has been known for years that in cases of starvation animal organs lose in weight and that the loss varies in different organs. It is not improbable that products formed by disintegration of some organs serve to sup- port the integrity of other more important organs. More marked is the high destructive power of tissues in diseases of a grave nature. Thus, in diseases of the respiratory system and of the heart, an intense self-digesting tendency of the tissues was noted by Schlesinger. In infectious diseases a similar observation was made by Flexner.*? The work of this author preceded that of Schlesinger, and is very important for the reason that it furnished an interpretation for some old observa- tions of pathologists. Flexner demonstrated an unusually high rate of self-digestion in organs removed from individuals who succumbed to typhoid 27 Zeitsch. f. Biol., vol. xliv, 1903. 28 Miinch. med. Wochft., 1903. 29 Cent. f. Bacteriol., vol. xxvii, 1900. 30 Hofmeister’s Beitrige, vol. iv, 1903. 31 Jour. of Physiol., vol. xxxi, 1904. 32 Univ. of Penn. Bull., July, 1903. AUTOLYSIS 95 fever and other infectious diseases. ‘The observation on typhoid is of special interest, since the exaggerated autolysis in the course of the disease can not be ascribed to the action of the micro-organism, for it is known that the proteolytic power of that germ is very slight. That this high rate of self-digestion is not merely a post-mortem phenomenon may be concluded from _ the old clinical observation, that products of proteid digestion are eliminated by the kidneys in the course of infectious dis- eases. ‘The occurrence of peptonuria in these pathologic forms is not infrequent. Thus, in the light of the new investigation, this symptom acquires a special significance. So long as the nutrition is in a sufficiently good condition to prevent wasting of the tissues of the body, peptone is not present in the urine. However, the softening and the wasting of tissues is most striking when these are under the influence of protoplasmic poisons. For this reason the study of phosphorus poisoning has attracted great attention. Martin Jacoby ® first pointed out the high rate of autolysis of the organs removed from animals killed by phosphorus poisoning. In this condition self-digestion takes place not only in surviv- ing organs, but also during life. This may be concluded from the work of Abderhalden ** and his co-workers, who recently have demonstrated the presence of crystalline components of the proteid molecule in the urine of animals poisoned with phosphorus. The importance attached to the study of phosphorus poison- ing is largely due to the resemblance which the clinical symp- toms of this condition bear to that of a spontaneous pathologic form known as yellow atrophy of the liver. While the morpho- logic changes in the liver in the two conditions are not abso- lutely identical, still they present many points of similarity. The most striking are the disintegration of cellular elements and the so-called fatty degeneration of the organ. Through the work of many investigators, and particularly through that of Wakeman and Waldvogel, it has become evident that the changes which the liver undergoes in these pathologic conditions are identical with those which the organ suffers in the course 96 HARVEY SOCIETY of autolysis. Indeed, the fact that the liver of persons who have succumbed to yellow atrophy contains products of proteid digestion was demonstrated by Salkowski more than twenty years ago, and was recently corroborated by Alonzo Taylor.** However, in neither of the two forms are the changes limited to one organ. Jacoby has demonstrated that the blood in phos- phorus poisoning presents a striking loss of coagulability. Still more striking is the power it possesses of liquefying coagulated blood. This peculiarity was interpreted as being due to the presence of a proteolytic enzyme in the blood. In the course of yellow atrophy, products of a proteid cleavage have been found in the blood by Frerichs. This has been corroborated by many investigators, and very recently Neuberg and Richter ** have shown that leucin, tyrosin and lysin may be present in the blood in quantities which clearly show that their origin could not be limited to the liver alone. Thus, in the foregoing forms all tissues are apparently affected in the same manner. Marked autolysis in them may be consid- ered a symptom of decline in general health and nutrition. However, there are conditions in which self-digestion is located in one organ only ; thus the atrophy of the thymus, the involution of a puerperal uterus, are accomplished by a process of auto- lysis. The softening of tumors is brought about by the same mechanism. This was made clear through the work of Petry,” who demonstrated that freshly removed tumors contain products of proteid cleavage. The same author further demonstrated that the rate of self-digestion of the new growth is higher than that of a normal tissue. An attempt was also made to study the toxicity of the products resulting from this process. How- ever, neither from a chemical nor a pathologic point of view could a difference between the end-products of autolysis of tumors and those of normal tissues be detected. Indeed, the intensity of self-digestion is high in all organs composed of cellular elements endowed with rapid growth. 33 Jour. of Med. Research, vol. viii, 1902. 34 Deutsch. med. Wochft., No. 16, 1904. AUTOLYSIS 97 The occurrence of local autolysis is not, as a rule, productive of a lowering in general health. On the contrary, it tends to restore normal conditions when these have been disturbed. There are other conditions in which the process of autolysis is of aid to the organism in the efforts to maintain its integrity. It has been stated already that in the course of infectious diseases tissues possess a high power of autolysis. Investi- gations of Blum,'* Conradi* and Levaditi** have shown that autolysis may be one of the means to which the organism resorts in order to elaborate protective substanees. Substances of two distinct groups are formed in the organism as the result of infection. Those of one group aim to destroy the micro- organism and are designated bactericidal; the purpose of the other is to neutralize the toxin elaborated by the micro-organ- ism—these are commonly named antitoxin. Normal tissues in course of autolysis may give rise to substances of either group. Blum has shown that the products of autolysis of lymph glands possess the power to neutralize tetanus and diphtheria toxins and cobra venom. ‘The mechanism of this action is not based merely on the physical properties of the autolyzed gland, for it is possible to save animals from death by injecting the products of autolysis subsequent to the injection of toxin. Further, Conradi has tested the bactericidal power of the products of self-digestion of various organs. This author noted that the last, added to a suspension of bacteria in broth, pre- vented their growth. The intensity of bactericidal power varied in different organs, as presented in the following table: SMI eh eats peat bt a Oise kare CH v5 ow ee ak ak Strong PemCMINET EMAC), Sao sie hee ee Cea wees Uae sides Strong ee Bi enh tebe adr giao Pex euute ule SB ots Strong NR eae ne a fe Sst aha ast elavnrs aishaieie G0; kowele bie Strong asia 2s 0 Mei ay ie 28 Ta ga le Marked SMITE RC Lan hie oa Sees dee She tereelild UD Marked STOUR DA lg ar SS Brat Rnd IAS) by Gd |s, Wid GNA B-aintle a's wd Sas Slight ke vila bi ainck oie,0 bin, Ral 8) None APS UN SARS oe ity, SEE eS Ona ee en None 35 Hofmeister’s Beitriige, vol. i, 1902; vol. v, 1904. 36 Ann. de l’Inst. Pasteur, vol. xvii, p. 186. 7 98 HARVEY SOCIETY Glancing over the table, one is struck by the fact that organs rich in leucocytes are most efficient in elaborating protective substances against infection. And one is naturally led to the analysis of the role played by the white blood cells in the effort of the animal body to maintain its integrity. A review of all the information gained concerning the action of leucocytes on the tissues shows that this is similar to the action of the digestive tissue enzymes. a 16,300,948 11,798 73.7 TRAST 16,526,135 13,247 75.1 1890 cise 16,526,135 11,059 66.9 101 kare 17,689,146 12,389 70.0 TAOS Seine 18,330,737 14,200 77.5 S03 SoS Goo 18,467,970 15,726 80.4 RESULTS OF SERUM THERAPY 125 Year Population Deaths Rate Diphtheria and Croup per 100,000 OE pte 19,033,902 15,125 79.9 re Ne aes 19,143,188 10,657 55.6 MN nw acd but the striking peculiarity of the heart muscle, namely, the refractory period, is still retained. It is difficult to understand how Rohde could have reached a different conclusion, unless indeed he was misled by an inac- eurate method of registration. The objection that I have made to the general application of Carlson’s results on the heart of the limulus retains, therefore, its full significance and prevents us from accepting this work as giving a final solution to the problem. Some further facts which tend to support the myogenic view may now be considered. The Reversal of the Beat—Under various conditions the beat of the heart may be reversed, that is, the wave of contraction may begin in the ventricles, proceed thence to the auricles, and finally to the sinus venosus. As Gaskell, Engelmann and others *See Amer. Journ. of Physiol. 25 In this connection see Porter: Amer. Jour. of Physiol., 1905, VO. XV, p..1. 21 322 HARVEY SOCIETY have pointed out, this reversal, while easily understood on the myogenic theory, is opposed to the usual form of the neurogenic hypothesis. A nervous mechanism, consisting of a principal motor center in the region of the sinus and subordinate motor centers in auricle and ventricle, can not, according to our experience with such mechanisms in other parts of the body, work in both directions; a system of connecting neurons is a mechanism that conducts and co-ordinates only in one direction. To account for this phenomenon the neurogenists are obliged to assume that the nervous apparatus in the heart forms a peculiar interconnecting network, the like of which is not found in the other automatic nervous mechanisms of the body, not even in those of the intestines. For when any portion of the intestine in a condition of rest is stimulated at a given point the wave of contraction or of contraction and inhibition pro- ceeds onward in normal fashion; the movement is a peristalsis and not an antiperistalsis. In the heart, on the contrary, when at rest, any adequate stimulus applied to the ventricles will set up a reversed rhythm. The necessity forced on the neurog- enists to make a new and unproved assumption to meet this case, does not, of course, strengthen their side of the argument. So, too, the well known zigzag experiment by Engelmann fits well into the myogenic theory, but is difficult of explanation in terms of the neurogenic hypothesis without recourse to the unknown properties of a nerve network. In this experiment it was shown that when a ventricle is so cut as to form an irregular piece, with intervening narrow bridges, a stimulus applied at either end arouses a wave of contraction that spreads in orderly sequence over the whole piece. The Auriculoventricular Bundle.—It was formerly held that the myogenic theory is inapplicable to the mammalian heart be- cause no muscular connection exists between auricles and ven- tricles. This objection has been completely removed in recent years. On the anatomical side the work of Kent, W. His, Jr., Retzer, Braeunig, Humblet and Tawara has shown beyond doubt that a small muscular slip passes from the auricle into the ventricular THE CAUSE OF THE HEART BEAT 323 septum. In man, acording to Retzer, this bridge is about 1.5 mm. in thickness, 2.5 mm. in width and 18 mm. long. On the physiological side the experiments of His, Hering, Humblet, Fredericq and especially of Erlanger, have proved with equal certainty that it is along this narrow bundle that the wave of excitation is conveyed from auricle to ventricle. The last named observer has shown in a series of brilliant experiments that if this bridge be compressed by a specially devised clamp, the sequence of the ventricular on the auricular contractions may be removed either completely or partially. In the former ease there is complete heart block, and the ven- tricle, after a preliminary pause, beats with a slow rhythm entirely independent of that of the auricle. In the latter case the block is partial, and the ventricular beats exhibit a 1 to 2 or 1 to 3 rhythm, as compared with those of the auricle. Erlanger has succeeded in showing that in man under certain pathological conditions an exactly similar condition prevails, forming the important feature of the Stokes-Adams syndrome.”® Autopsies, indeed, have shown that in some of these cases a demonstrable lesion exists in the region of the bundle. No one ean deny the importance of this bundle as the physiological link connecting auricles and ventricles. If it so happened that the tissue composing it was entirely devoid of nerve fibers the myogenic hypothesis would be practically demonstrated. Ac- cording to Tawara, however, the bundle is provided with a nerve network similar to that found enveloping the muscular tissue of the rest of the heart, and naturally the neurogenists attribute to this network the functions that the myogenists would assign to the muscular bundle itself. A definite answer to our problem is, therefore, again post- poned. The myogenist may, however, urge with justice that the probabilities here are once more in favor of his view. This bundle constitutes the only known muscular connection between auricles and ventricles, while nerve connections between the two 26 Erlanger: The Jour. of Exper. Medicine, 1905, vol. vii, No. 6; ibid., 1906, vol. viii, No. 1. 324 HARVEY SOCIETY chambers exist freely in other parts of the auriculoventricular ring *‘; yet severance of this small muscular bridge is all that is necessary in order to interrupt completely the physiological connection between auricle and ventricle. Investigation of this interesting structure has but just begun. We may hope that future work will develop facts of fundamental significance for the physiology and pathology of the heart. Already one sug- gestion has arisen out of the work, which indicates the possi- bility of a new point of view regarding the cause and sequence of the heart beat. Tawara maintains that the cells composing the bundle are not ordinary heart muscle, but that variety of cardiac muscle which has been designated as Purkinje fibers or cells. He believes that the bundle after entering the ventricle spreads out to constitute the Purkinje cells that are known to form a layer beneath the endocardium, and one may conceive that this layer has a still further distribution within the mass of the heart musculature. There is thus presented the possi- bility of a widespread occurrence of a specially modified type of contractile tissue which may be intimately connected with the phenomenon of automatic rhythmicity as well as conduction. We must, however, await further investigation before attempt- ing to speculate on this modification of the myogenic theory. The Action of the Accelerator Nerves—lIt has been known that the hearts of both warm-blooded and cold-blooded animals may be kept alive for hours after excision from the body, provided they are supplied with an artificial circulation. Recently the possibilities of thus maintaining an isolated heart, or of reviving its activity after death, have been devel- oped in a remarkable way. Kuliabko was able to restore the beat of the heart in animals that had been dead three or four days, by the simple process of supplying the coronary vessels with a Ringer’s solution. He obtained also similar successful results on human hearts as late as twenty hours after death. In an experiment made on the heart of a man who had been dead eleven hours, Hering was able to restore its beat for a *7 See Lomakina: Zeits. f. Biol., 1900, vol. xxxix, p. 377. THE CAUSE OF THE HEART BEAT 325 period of several hours. Hering has made use of this possi- bility to study the source of the heart’s automatic rhythm.*® He found in the course of experiments on the isolated mammal- ian heart that the inhibitory and accelerator nerves continued to give their respective effects for a long period of time. In one case in a monkey the vagus retained its inhibitory action for six hours after death, and the accelerator for more than fifty-three hours. On the other hand, comparative experiments made on rabbits and dogs showed that sympathetic nerve ganglia, such as the superior cervical or the ciliary, lose their irritability very quickly after death, even when supplied with an artificial circulation of Ringer’s solution. It seems probable from these experiments that the maintenance of an automatic rhythm in the heart so long after excision or after somatic death can not be due to the activity of intrinsic ganglion cells. Since, moreover, the accelerator fibers retained their irritability in these experiments for very long periods after death it would seem probable that they do not end in the ganglia of the heart but are distributed rather directly to the heart muscles. Such a conclusion implies that the rhythm of the heart beat originates in automatic processes within the muscular tissue itself. THE MYOGENIC THEORY OR THE NEUROGENIC THEORY ? From consideration of this brief review of the current litera- ture and discussions on this subject it appears to me that every impartial observer will be forced to come to the same conclusion as that reached by Hofmann in his excellent critical paper published in 1898, namely, that the myogenic theory is the most probable of any that have been proposed so far. The theory in its most general form assumes that contraction waves or excitation waves arise in the sinus region and are conducted by the muscular tissue over the whole heart, the visible effect at each point being dependent on the condition of the muscu- lature at that moment. At the passage from auricle to ventricle there is a slowing of the conduction due to the small size and 28 Hering: Pfliiger’s Archiv., 1903, vol. xcix, p. 253. 326 HARVEY SOCIETY special properties of the narrow bundle connecting the two chambers. Moreover the condition of the musculature at any point may be influenced in opposite directions by nervous influences, inhibitory and acceleratory, which, however, have nothing to do directly with the origination or conduction of the initial motor impulse. This theory gives in general an adequate explanation of the phenomena of the normal heart beat, and of those variations that occur under pathological and experimental conditions, but in order to apply it in detail we need to know more of the processes that lead to contraction and relaxation. In fact, the phenomenon of the co-ordination of the beat is not sufficiently accounted for, whether we adopt the myogenie or the neurog- enie theory. The difficulty is apparent if we stop to consider those conditions which lead to meo-ordinated contractions, such as are exhibited in the peculiar fibrillated movements of auricle or ventricle. In this condition the mass of the musculature of the ventricle, instead of contracting simultaneously or in a rapid wave running from one end to the other, exhibits feeble local contractions and dilatations which involve only small areas, and give the entire ventricle the appearance of a flutter- ing, trembling mass. One of the most remarkable means of thus throwing the co- ordinated contraction of the ventricles into ineo-ordinated fibrillary movements is the heart puncture as described by Kronecker and Schmey.”® — - = - + —<-- ~ _ a nnams tentang naeeehoaraoninmnaqeanetytgacverent yok ePL RO SELATAN Ane adn er as - oon — ean cbeeerereemensetn anna an enhao nae ga eae Oe a a ae ae —_ =i ———— —_ a =r nS Sarre ta sven yoestunene@gneoconarencask o-recelb teen tnone on eNOS Ae nena Cas SAG Int OS ATID SLO a ee ae A A « ee nee reaons sonanetamecenpenasnciaensventctnowaiarghaaastaneueav=-ces-ausanaens finan Vento Oreeae erent fees sas Sen SS che ovenenrmp ee nine _ ~y Sh sane aosoannahenennsesnaysonessatusnener—eemcnanaret cae a Stirs aC aaa SA SMe a O Cae nn ts ———————— — a a a a aE © we. ane ae mae earn erga cceeseuameednenact{hhteer-ahreaieneernna seen he Anse sas aien ee ee a ne Se eee ai ange amen te Ont Pye ar ae ———— Sieneianna en cnateanaanetnnaraenatiadll A EA TS AS SET oe - Aras) fig cccossoteasenclamere sowmnonnne—sonsat-onseesndnasrssene sanwssnanresea easuty O-aaitwesioxsuetcsnonresnatne ent stiiio~ Sci ries Mere Swoon S ob erenenssane ten em a — —_—— aE a a Sa eee ae nen aeeener yrererunyeseear - scans; sj asoudatesadveptactensenewsratncasnemanayevumeatcng§ osmspeanen we frenwnsisuahee sun atemeananrvaneesanseenrmeery cease, a eee hee eens, reel a a a = = > = Sern are eevee arr a yO La ee eee Dotan gece — ——————— - ~ - — pea as arnt ot SE ne cana aye eicaonarsoms eaienanarean ate oe ae egar emanate nea Teen OAD Oat NN ene ~ “4 > ar ite spencer rae poornomcnopeceng rt natnfetrerberaee senate est nant fee owe ees ae a age dens anteater eee est a a A TC A SA ee - wee 4s) nae aiasamnnnncanneen annage'eomenawwmnen Beieten ence nat4eestnscnanabasnae—-oabeeestinenre samyeoomsshabea Ante nteniGinennn srs A Re Anas Nt tS —_ oC — Se a eae rrrenrecgenenn nooo pmenounecionreerererosenseanenyseavener ed; nseev ow mncentwarrOHssaatnr ae sesshee Neen oss nesacoeeeen aap eh A Li SS ee ao ee se voctumepaweetenssenays sei Seenssee-® gr-ahersuangioene ase anny nOnorecs (seman teiwnewnesgieseres eRe a = eee > —— a ae Pes « ee he ee Tierronses: Yom qumietenssouns-aioastestey toeaceanyeywaet ust Si SeNneeSOgNeaner AAAS PSMA Aes airs ON ak neta ee ——<———— a a ee et an ene nb nn en =- —— red seers veseseuoas atennoareq/saaun Geevdssessebrenrdaese=-aRIeaa sens ea OOEe Ape me seas CE7E PORN Omen ee an a —— — cen AA A ASA ne aa Spann aah ener nate - ees fae rane qreeveig baba tenn sonsaranieesoremn caved —erwnn tnaerw —tebanuih ter tAnmaAgoestirwar as sasy hry a an ee nc —_ ———— a ca aa a a LY PNT SSN cae car Ceaenasee a mat ps eran © Pe —— nh ppeoeneren apo aT PALM 7 eee NT TP Oa nT 7 ner engeneg trereoeahootrean ar oread AE EROS ean aT OTT att IT TE nt a enna ——-— Sean reer AE TTT OTS DCN Sore eeken eee ener ans SAT BE ere —— ns a Oe see caareaios a ts tresnsesnmecton-ocsansodaerenossharungere.invenasts ahsustareeesiencstarebiGSGWt?*tsGPe Seems ine are Seite Tene an en aN att cenerensagvanee i A a oa! ree oe ran ae eee a a TT AD NTT CANIS ASE neat empresa eesten ef == co eases eoaayeynenenageeuncntesossntanadusyeonanan tor -eatatenwnn narvorennssefpvtmenmaren smestssit snttn Sn — ae a a TO ea RT PTT D ca eee eres ——— ae rooms teeen "sue oe #0. sag -shbneege = Hane wenemen et Sanat renames: ores ne an a a a A TDG ALOR A AAT AE . aosetehenenparesnaelanynp anes se—seelbeene Ser Tre 0 ae a me wtralaterosemaune. Syeseecnmandn 1aagwonta nate ingest sar yosOeE? A ade EaBenremann satin —eemaenreeas Smee ot ee ne eno a ge a cence a eee a nee A a OTST NL ASA OPTRA ASS EE oo eS meena tasadesotennanesenehanaheanonensustnresaaete Setters Sia AGES ALA Ae NaS OTST oS NN - ——— A I yO an a A ET oa. eee aT aan Teena Meudreusig ntvnssehandes ius va sorends wert viner-mknmamnsed tsar euinsummsuteter orvewhksnt Pement eae ee nnn —————— an a NT Te aS ey ope aera rae een Te ear ape TS panera nan pet Dane meee ——— meres oe eames wants aogracartecanulsectqeqeunnssabpgyhrtenajreeneisgirrrunim nae sonnt wemeyt -agne-anis puarerm matches moe =F ata — — — = en a a a a Ee Nt Ay Se ra ananeannnssannes ta netanaactepatamnenee Siee OP CCAA LAAT COTO ATTA SI AE, © =" moor — an own - ae — - a = ve - mney _- = = ~ ~ ~ ‘ ana hchna 7 ~ _ — or