General Physiology: An Outline of the Science of Life

In electrical stimulation also the depression-effects are wholly subordinate in comparison with the phenomena of excitation, and, while the latter have been investigated extensively and carefully, the former are little known. Nevertheless, there appear to be cases in which the galvanic current, especially through long influence or great intensity, is able to produce phenomena of depression. Whether the diminution of the irritability of the nerve that takes place at the anode upon the making of strong currents and at the kathode upon the breaking, and can lead to complete loss of the power of conduction at the place in question without any real destruction, is to be regarded as a phenomenon of depression is more than doubtful ; more extended experiments especially directed to this point are needed. But apparently genuine depressing effects of the galvanic current have been discovered with regard to ciliary motion by Engelmann('79, l)and Kraft ('90).

The gill-filaments of bivalve mussels are covered by a ciliated epithelium, the cilia of which on account of their length are especially well -fitted for the observation of ciliary motion. When Engelmann stimulated these filaments by means of a single, strong induction-shock, the cilia went into rigor, exactly as the cilia of Infusoria and ciliated epithelia go into heat-rigor after strong thermal stimulation. They bent into the form of a hook in the direction of the stroke, their motion ceased and they remained in this position the longer, the stronger the induction-shock had been.

Kraft made an analogous observation during the long-continued action of the constant current upon ciliated epithelia of vertebrates. At the beginning of the action the ciliary stroke was accelerated, first at the two poles, but then, by a spreading of the excitation in the tissue, in the whole intrapolar portion ; with long duration of the current the acceleration decreased gradually and gave place to 3, depression of the activity, amounting to complete standstill in the whole intrapolar portion. Hence it appears that the same relation is present here as in other, e.g., chemical, depressions, viz., the stimulus in question calls forth first a stage of excitation and then with stronger or longer action a depression. But all these relations have been too little investigated to permit a definitive interpretation.

Among the physical phenomena that are employed for the amusement of children in civilised countries, those of magnetism usually have great attraction for the childish mind. The remarkable facts, that the magnetic needle, freely suspended, under all circumstances directs one end toward the north pole of the earth, that small boats and animals provided with an iron pin and swimming in a basin of water follow the slightest motions of the magnetic needle with unfailing certainty as if conjured by a magician, that iron filings, strewn upon paper, arrange themselves in very characteristic curves over a magnet placed below them — all this has greatly fascinated us as children. Magnetic phenomena must have made the same deep impression upon the fervid fancy of the people of the Orient, who in many respects have retained childish qualities even to the present time. In the tales of the Arabian Nights there is a vivid expression of this, which still takes stronghold upon the childish heart, in the gloomy stories of the magnetic mountain and the fright of the helpless mariners who saw their ship, attracted by the invisible force, crash upon the smooth metallic rocks.

In the adult the sense of wonder and fascination connected with the magnet is largely lost because of our being accustomed to its peculiar effects. But the sensations of our childhood are again awakened when we meet with effects analogous to those that the magnet exerts upon the needle, namely, attraction and repulsion , transferred to living nature as results of a great variety of stimuli ; and when we see that these stimuli are capable of exercising upon organisms an effect that forces them under certain circumstances to turn toward or away from the source of the stimulus with the same irresistible power and the same unfailing certainty as the magnetism forces the iron. The moth returns with deadly certainty to the light ; although it has singed its wings innumerable times, it cannot resist the fascinating power and finally falls into the flame.

Since, in the higher animals, as a result of the co-operation of the nervous system, these phenomena possess a complexity that renders their examination more difficult and not rarely diminishes the certainty of the reaction, it is advantageous to consider them more especially from the standpoint of cell-physiology. It is necessary to the occurrence of the phenomena in question that differences in stimulation exist in different parts of the body. If stimuli act equally upon all sides, all the effects of stimulation described in the preceding section occur, but a directive effect is necessarily absent. Only unsymmetrical stimulation can control the direction of motion.

The word chemotaxis is applied to that property of organisms that are endowed with the capacity of active movement by which when under the influence of chemical stimuli acting unilaterally they move toward or away from the source of the stimulus. Where there is an approach to the source of the stimulus, there is positive chemotaxis, where there is a removal from the source negative chemotaxis. Unilateral stimulation with chemical stimuli is only realised when the concentration of the substance in question gradually increases from the living object in one direction.

Discovered first by Engelmann in Bacteria, observed by Stahl in Myxomycetes, studied systematically and more fully by Pfeffer, and recently investigated in leucocytes by Massart, Leber, Gabrits- 1 Although the words ' ' chemotropism " " heliotropism," etc., have been long in use, I have decided, after considerable delay, to exchange them in this edition of the book for the words " chemotaxis," " phototaxis," etc. ; my reason is that the former not only sound heavy, but suggest objections from the philological standpoint. I come to this conclusion with some reluctance because for some time I have been endeavouring to extend the former terminology from the few, earlier known "tropisms" to the corresponding phenomena associated with other stimuli, and admit new tropisms. thus indicating at once by the term used the fact that all the phenomena belong in the same class. Now, however, when the analogy of the phenomena that result from the various kinds of stimuli is fully recognised, I believe it advantageous to replace the less fortunate terms by the newer expressions, already much employed.

chevsky, Metschnikoff and others, chemotaxis has now become recognised as a phenomenon of wide distribution among free-living cells and of extraordinary significance, not only for unicellular organisms but also for life in the cell-community. Among naked protoplasmic masses chemotactic phenomena were first observed by Stahl ('84) in Myxomycetes. He allowed the yellow reticulate plasmodium of ^thalium septicum, which lives upon tan, to creep upon moist strips of filter-paper, and then hungup the strips with one end in water that was deprived of oxygen and shut off from the oxygen of the air by a layer of indifferent oil, while the other end was in contact with the air. The result was that the protoplasm of the strands that dipped into the water gradually streamed completely out and accumulated above the layer of oil upon the moist filter-paper in the air. Hence it was positively chemotactic to the oxygen of the air. That it was not the water itself which the plasmodia sought to avoid, as might perhaps be supposed, follows from the fact that the plasmodia are positively chemotactic to water and always creep from dry to moist, thus manifesting a specific hydrotaxis. The strips of filter-paper in the experiment must always be kept moist, in order that the chemotaxis toward water may not interfere with that toward oxygen. The plasmodia behave chemotactically toward other substances also, especially the tan that serves them as food. Thus, in Stahl's experiments the protoplasmic masses always crept toward pieces of tan or toward little balls of paper that were soaked with an extract of tan, and collected there, a form of positive chemotaxis which Stahl termed trophotaxis because it plays an important rdle in the habit, which is wide-spread among unicellular organisms, of searching for food. Leber ('88), Massart and Bordet ('90), Metschnikoff ('92), Buchner ('90), and others have discovered chemotactic properties in the leucocytes of vertebrates, and a relation has been found here that is of the greatest importance with respect to the behaviour of organs toward infectious diseases.

As has already been seen,1 Bacteria excrete certain metabolic products, such as the toxines, which recently have attracted greatly the attention of investigators. These products exercise upon leucocytes a very pronounced chemotactic effect, and cause them to creep in great swarms to any place in the organism where Bacteria have entered and multiplied. At the place of infection a dense accumulation of leucocytes takes place and in certain cases, as Metschnikoff has shown, they devour the bacteria and determine in part the further course of the infection (Of. Fig. 210). If the bacteria are not present in too great numbers, they may succumb in the struggle with the leucocytes, which latter in a certain sense represent the police of the body in comparison with the weaker intruders, and the infection is stopped. If the bacteria prove the

stronger, an extension of the infection throughout the organism takes place, and the course of the disease is then determined by other factors. In order to demonstrate the positively chemotactic action of bacterial products upon leucocytes, the following experiment of Massart can be performed. According to a method first devised by Pfeffer, a short capillary tube is filled with a culture of the pusforming Siaphylococciis pyogenes albus, and one end is sealed, The tube is laid in the abdominal cavity or under the skin of a rabbit, and left for some 10 to 12 hours. After this time it is found by microscopic examination of the tube that through the open end a dense swarm of leucocytes has penetrated into the interior, and has closed the opening like a thick white stopper (Fig. 208). In other words, the leucocytes are induced by the bacterial substances to creep from the tissues of the animal into the capillary tube. A critic will at once raise the objection that perhaps it is the nutrient solution in which the bacteria are cultivated, which acts chemotactically upon the leucocytes. This objection can be nullified, if, as Massart has done, there be put into the animal for

FIG. 208. — Chemotaxis of leucocytes toward pus-cocci. The leucocytes have wandered in dense crowds into u capillary tube, which contains a culture of Slaphylococcus ', they may be seen especially at the opening of the tube. purposes of control a similar capillary tube containing similar nutrient liquid but without the bacterial culture. In such a case the leucocytes do not enter. That it is not simply the bodies of the bacteria themselves, but the metabolic products excreted by them, which have the chemotactic effect, may be proved by employing for the experiment a culture liquid that has been sterilised and wholly freed from the bacterial bodies, and in which, therefore, only the dissolved metabolic products of the bacteria in question exist. The result is then the same as when the culture is employed directly : after some time the tube is filled with leucocytes that have wandered in. What is true of the culture of Staphylococcus pyogenes albus has been found also in many other pathogenic Bacteria, and there is no doubt that further investigations upon the relations between leucocytes and bacteria will make clear a whole series of points which thus far in the history of infectious diseases have been very obscure.

Moreover, leucocytes appear chemotactic, not only toward the metabolic products of Bacteria, but also, as Buchner has found, ttoward the proteids of the bodies of the Bacteria themselves, and whole series of substances of non-bacterial origin. Thus, Buchner found that broth made of wheat flour and that made of pea flour possess especially strong chemotactic power. Finally, Sicherer ('96) has recently shown that under proper conditions the leucocytes of warm-blooded animals outside the body exhibit their chemotactic properties toward very varied substances for a long time, as clearly as in the living body itself.

The chemotaxis of leucocytes plays an important role in the development of many animals. This is made clear especially by the beautiful investigations of Kowalevsky ('87) upon insects. When the fly-larva changes into the complete fly — a metamorphosis that takes place fairly rapidly — the organs of the larval body, such as the creeping muscles, become superfluous, and 209.— Leucocytes destroying the muscles in the metamorphosis of the larva of the fly. Thegranular masses are leucocytes, the striped masses bits of muscle. (After Kowalevsky.)

begin to degenerate. The substances formed at the beginning of this degeneration have a strong chemotactic action upon the leucocytes: the latter wander into the degenerating organs in great crowds, and as genuine phagocytes devour the disintegrating masses, and thus accelerate their removal (Fig. 209). It is characteristic that the phagocytes manifest their activity only in insects in which the metamorphosis takes place very rapidly ; but that in others, as in the moth, and in the degeneration of the tail of the tadpole, they have no share. Nevertheless, Metschnikoff was able to demonstrate analogous phenomena in the development of star-fishes.

Chemotaxis is wide-spread in the flagellate Bacteria, Infusoria, and swarm-spores. In Bacteria the phenomenon was first discovered by Engelmann ('81, 1 ; '94), and was at once employed practically in an ingenious manner. Engelmann observed that certain forms of Bacteria that live in decomposing infusions accumulate in great numbers in the neighbourhood of the sources of oxygen. Thus, in an exposed drop under the microscope a dense accumulation of these microbes takes place at the edges of the drop, where the oxygen of the air has the freest access. Under the cover-glass, likewise, the Bacteria congregate in the neighbourhood of the edge, and form a dense wall parallel to it. Bubbles of air, as well as plant-cells whose chlorophyll sets free oxygen in the light, act in the same way, especially when lack of oxygen is produced to a certain extent by covering the edges of the cover-glass by a layer of oil. Engelmann employed this extraordinary irritability of Bacteria toward oxygen as the basis of a method for the microscopic demonstration of very small quantities of oxygen, and this has become very important in our knowledge of the assimilatory action of various kinds of light upon the green plant-cell.1 By the external exclusion of air from a drop containing Bacteria, the place may be found where only the slightest traces of oxygen are present, and these are situated where there is a dense accumulation of the microbes. A beautiful example of such is afforded by the following observation.2 In a drop under the cover-glass a large diatom (Pinnularia) was in the field of sight, and, since in the light it gave off oxygen by reason of the activity of its chromophyll, it was closely surrounded by a wall of motionless Spirochcetoe. In the other parts of the field almost no Spirochcetce were visible. The diatom suddenly moved a little distance, and then lay still. The Bacteria, thus separated from their source of oxygen, lay quiet for a few moments ; but soon an active movement began among them, and they swam in dense crowds again to the diatom.

After one or two minutes almost all were congregated again about it in a dense mass, and motionless as before (Fig. 211, 1). Engelmann has recently figured similar observations (Fig. 211, 11 and III). The striking and systematic investigations of Pfeffer ('84/88) upon chemotaxis had their starting-point in observations on the spermatozoids of ferns, in which chemotactic relations to the egg-cell were found. It is now known that analogies to this exist in almost

FIG. 210. — Leucocyte devouring a bacterium of splenic fever. (After Metschnikoff.) all organisms, and function as the indispensable condition of the fertilisation of the egg-cell by the spermatozoon in animals as in FIG. 211. — Chemotaxis of Bacteria toward oxygen which is produced by Alga-cells in the light. 1, Diatom evolving oxygen in sunlight and surrounded by Spirilla. II, Diatom half -shaded, half-illuminated. The Bacteria have collected in the illuminated half, where oxygen is being evolved. ///, An Alga-cell surrounded by Bacteria ; A, in the dark, B, in the light. (// and III after Bngelmann.)

plants. The spermatozoon seeks the ovum, and almost everywhere in the living world is led in the right path by the chemotactic action which the metabolic products of the egg-cell exert upon the freely-moving sperm-cell (Fig. 212). The fact, which must otherwise appear very wonderful, that among the innumerable swarm of spermatozoa of the various, marine animals every species finds the proper ovum, is in a great majority of cases a direct result of chemotaxis, and is explained very simply by the further fact that every species of spermatozoon is chemotactic to the specific substances that characterise the ovum of the corresponding species. We have here an adaptive phenomenon of the simplest kind, which gives us anew an idea of how extraordinarily deeply the phenomena of chemotaxis reach into life-relations.

Pfeffer's experiment was as follows : He filled a capillary tube, sealed at one end, with a solution of c. 0'05 per cent, malic acid and placed it in a drop that contained a great number of the spermatozoids of a fern ; the malic acid gradually diffused from the opening of the tube into the drop, and thus became the source FIG. 212. — Two ova of the plant, with spermatozoids swarming about them. (After Strasburger.) of a stimulus acting unilaterally. Microscopic examination showed that the spermatozoids immediately began to steer toward the opening of the tube and to swim into it. After a half-minute some 60 spermatozoids had entered the capillary, and in some cases after five minutes approximately 600. In one experiment with 24 spermatozoids, after twelve minutes all except one, which lay at rest outside, had collected in the capillary. Hence malic acid exerts a very strong chemotactic action upon the spermatozoids of ferns ; toward all the other substances that Pfeffer tested with respect to their chemotactic power, the spermatozoids behaved with complete indifference. This suggested strongly the supposition that in the archegonium that holds the ovum it is malic acid that causes the spermatozoids to approach and enter. On account of the minuteness of the objects and the lack of micro-chemical methods, Pfeffer was not able to demonstrate malic acid in the contents of the archegonia themselves, but

he succeeded macro-chemically in establishing the presence of that acid in all parts of the plant that contain sexual products ; hence the above supposition obtains a degree of probability that borders upon certainty. The spermatozoids of mosses behaved indifferently toward malic acid, but were remarkably chemotactic toward weak solutions of cane-sugar. Later, Pfeffer extended his investigations to a large number of Bacteria and flagellate Infusoria, and obtained a series of results that are interesting in the highest degree. These show that very different substances act very differently upon different microorganisms. Substances to which one species reacts prove ineffective with another. Many substances induce only positive chemotaxis, others only negative. In the latter case the organism turns away from the source of the stimulus, and the capillary tube remains empty. The threshold of stimulation, i.e., that degree of concentration at which the substances just begin to exert their chemotactic effect, is very different for different substances and different organisms. But the most interesting fact is that many substances that induce positive chemotaxis in weak solution, induce

FIG. 213. — Scheme of chemotactic reaction. The concentration increases from the left toward the right ; at 0 the zero-point of concentration, at t the death-point. The arrows indicate the direction of movement. negative chemotaxis in the same organisms in strong solution. There exists, therefore, a stimulus-optimum toward which the organisms strive from both sides, from the weaker as well as from the stronger solution. If the solution becomes too strong, death naturally results. Hence, four important grades of concentration may be established : the zero-point, where the substance in question is wholly wanting ; the threshold of stimulation, where its concentration is such that the substance is just effective ; the optimum-point, toward which the organisms strive from all degrees of concentration above the threshold ; and the death-point, at which the concentration is too great to permit life (Fig. 213). With the same substance the optimum-point for different organisms usually exists at different grades of concentration. Massart (91) found a beautiful example of this in the different behaviour toward oxygen of a bacterium, Spirillum, and a ciliate infusorian, Anophrys. If the two species of organisms were under the cover-glass in great numbers, both congregated like a wall at the edge of the glass or around air-bubbles, but not immediately at the boundary between the air and the water ; each kept its own distance from the source of oxygen, the Anophrys nearer, the Spirillum somewhat farther.

The oxygen-optimum of each species was represented very distinctly by the distance of the individuals from the source of oxygen (Fig. 214, / and IT). Among ciliate Infusoria chemotactic phenomena have thus far become little known ; nevertheless, for a few species Massart has been able to demonstrate chemotactic properties toward various substances. The negative chemotaxis of the infusorian already mentioned, Anophrys, toward common salt may be cited ; this may be demonstrated in a very simple way. Massart laid at the edge

FIG. 214.— Chemotaxis of Bacteria (Spirillum) and Infusoria (Anophrys). I, Bubble of air under the cover-glass, surrounded by two zones, the nearer of which consists of Anophrys, the farther of Spirillum. II, Edge of the cover-glass. Anorphrys and Spirillum form similar zones. ///, Two drops of water united by a bridge of water. In the upper drop is common salt. The Infusoria (Anophrys) in this drop wander over into the drop of pure water, the more the salt dissolves. (After Massart.)

of a drop in which numerous specimens of Anophrys existed a few small crystals of common salt, and connected the opposite side of the drop by a narrow bridge of water with a similar drop of distilled water (Fig. 214, III). The result was that the Infusoria fled from the spot where the salt lay the more, the more the salt dissolved and diffused into the neighbourhood, until finally they had all passed over the narrow connection into the other drop.

Jennings l has recently made very comprehensive and systematic investigations of the chemotaxis of Paramcecium, and has employed a method that possesses in many respects great advantages. Jennings performs his experiments upon the slide under a large cover-glass which is supported by two glass rods, so that a pretty thick layer of water containing Paramceda lies between the slide and the cover. In this layer, which must be free from all admixtures, he places carefully by means of a pipette drawn out into a capillary, a drop of the solution the chemotactic action of which is to be investigated (Fig. 215). The substances in this solution diffuse at once into the surrounding liquid, in which the Paramceda are scattered uniformly and in motion. Then, according to the mode of action of the substances in question, very characteristic effects are produced. If the substances are ineffective, as, e.g., solutions of sugar, the Paramceda swim undisturbed into the drop, and after a few seconds are again spread uniformly under the cover-glass. If the drop induces negatively chemotactic properties, as, e.g., alkalies, a circle that is completely free from Paramceda, forms at the place (Fig. 215, A). But if the drop induces positive chemotaxis, as e.g., most acids, all Paramceda that are present under the cover-glass swim into it (Fig. 215, B\ If the concentration of the effective substance is above the optimum, the Infusoria accumulate about the drop in a circular zone (Fig. 215, C\ It is remarkable that Paramceda are positively chemotactic toward carbonic acid, as toward other acids. If a bubble of chemically pure carbonic acid and at the same time, for control, one of ordinary air be placed under the cover-glass, the Paramceda, leaving the air, congregate in a dense mass about the carbonic acid (Fig. 215, D). But, in proportion as the latter diffuses into the water and accumulates in a concentration above the optimum, they retreat in a closed circle from the bubble, because they are negatively chemotactic toward strong solutions of carbonic acid.

Thus very characteristic figures appear (Fig. 215, JE). Further, since Paramceda, like all other organisms, produce carbonic acid, constantly more individuals become attracted to the place where for any reason a number have assembled. We have here a very interesting case of the formation of an assemblage simply by reason of positive chemotaxis. As Fig. 215, B shows, by the transference of a drop of water from a group to another cover-glass preparation containing Paramceda a chemotactic assemblage may be produced.

Finally, chemotactic phenomena afford a means of forming an approximate conception of how extremely slight may be the stimuli that are able to exert a visible effect upon living substance. In his experiments Pfeffer found that the spermatozoids of ferns exhibited distinct chemotaxis when he employed a capillary tube containing a solution of O'OOl per cent, malic acid. If it be borne in mind that the malic acid must first diffuse into the drop in order to exercise its power, it follows that the quantity that acts upon the spermatozoids must be exceedingly small, b ut this is not all. In order to produce a chemotactic effect it is

not sufficient that a certain quantity of the substance in question be uniformly distributed in the vicinity of the organism, but a FIG. 215. — Chematoxis of Paramcecium aurelia. A, Chemotactic cover-glass preparation ; a drop of liquid that induces negatively chemotactic properties has been introduced under the coverglass by means of a capillary pipette. £, Positively chemotactic assemblage. C, The same with too strong a solution ; the Paramacia have congregated in a circle at the optimum-point of concentration. D, A bubble of carbonic acid and one of air are under the cover-glass ; the former (at the left) induces positively chemotactic properties ; the latter is indifferent. £, The same preparation a few minutes later ; the carbonic acid has diffused into the surrounding water and by its too high concentration has driven the Paramcecia to the place where they find their optimum of carbonic acid. (After Jennings.)

decrease in concentration must take place on one side. In other words, it is the amount of the difference in concentration at the two ends of the spermatozoid that determines the appearance of the chemotactic effect. Since the spermatozoid possesses only the minute length of 0'015 mm., we can form an approximate idea of how extraordinarily small must be the difference in concentration at its two poles, and therewith the amount of the stimulus that calls out a chemotactic effect. Thus, chemotactic phenomena and, as we shall see, analogous phenomena caused by other stimuli, give us a better idea than all other reactions of how excessively feeble stimuli produce a remarkable effect upon living substance. Living substance responds to extremely delicate influences. When homoeopathy affirms the effectiveness of extremely small quantities of certain medicines, its claim in this respect is fairly justified, however much superstition in other respects may attach to the homoeopathic doctrine.