Protoplasmic Action and Nervous Action

General physiology has been defined by Verworn^ as '^ cellular physiology," in accordance with the general conception of the cell theory that the ultimate living units of any organism are the cells. According to this conception the cells are the simplest units capable of independent life; hence general physiology, aiming at the anatysis and characterization of life-processes, should be equivalent to cell physiology. There appears, however, to be a certain arbitrariness in this idea. The cell is already a complex system with a definite organization, usually containing a nucleus and exhibiting other special structural differentiations. The question of the physiological significance of the cellular organization constitutes a special problem in itself. While it is remarkable that all higher organisms show this type of organization, it seems hardly justifiable to regard all organisms as consisting of cells and products of cells. Such a conception regards the simplest living unit as having a certain definite type of structural organization; i.e., it is essentially a morphological conception. A chemical characterization seems to meet the requirements of the case more completely. Many organisms are known which do not show the chief structural feature of the cell, differentiation into nucleus and cytoplasm; e.g., bacteria and blue-green algae. Usually bacteria are regarded as plant cells of a special kind; it is question-

able, however, if micrococci, and especially the organisms in filterable viruses, can be considered as cells in the true sense. The case of the ultra-microscopic organisms present in the filterable viruses is of special interest. These organisms can be demonstrated only by the effects which they produce (infection); they prove themselves to be living by their power of automatic growth, shown by multiplication in the body of the host or in culturemedia, and also by exhibiting other properties characteristic of protoplasm in general, such as thermolability and susceptibility to toxic agents of the disinfectant class. They may be described as complex and chemically active (metabolizing) material in a fine state of subdivision (like that of colloidal material), possessing in addition to the other properties of matter in this state the special vital properties of assimilation, growth, and multiplication. As already pointed out, this ability to transform environmental material into its own specifically organized and active substance is the distinctive criterion of living as distinguished from non-living matter.

Our conceptions of the nature of living organisms must be broad enough to include the ultra-microscopic forms. Cells, as found in higher organisms, are units of a relatively complex and highly differentiated kind, representing a comparatively advanced stage of evolution. They are by no means to be regarded as the only systems in nature exhibiting the characteristics of life. In higher organisms, however, we have definite experimental evidence that the smallest unit capable of continued independent life is the nucleated cell. The

protozoa remain as single cells throughout life. The higher animals and plants are single cells only at the beginning of their development — in the germ-cell stage; in later developmental stages and as adults they consist of large, closely associated aggregates or colonies of cells, which, together with the intercellular fluid media serving for transport (blood) and various other products of cellular activity (skeletal and other structures), form a complex and highly integrated system, or organic individual. Each cell in this organism is to be regarded as living and capable of independent existence under appropriate conditions.

The statement that single isolated cells are capable of independent life has been shown experimentally to be true, not merely of organisms which throughout their life are unicellular, but also of many of the cells of higher organisms when isolated under favorable conditions — leucocytes, ciliated cells, muscle cells, tissue-cells. Epithelial cells will grow in suitable culture-media;^ embryonic nerve cells, isolated in sterile plasma, send out axones in a characteristic manner; i.e., retain the normal power of growth, differentiation, and development;^ and many functional adult cells continue to live and grow when isolated under favorable conditions of food and oxygen supply.^

On the other hand, experiment shows that for normal and long-continued vital activity the cell must be complete, at least in the sense that both nucleus and cytoplasm (or portions of both) are present. This is shown by experiments on enucleated cells, such as egg cells; portions containing nuclei survive; the others die. But if enucleated portions are fertilized and thus furnished with nuclei, they continue to live.' An otherwise complete protozoon such as Stentor will die if deprived of its nucleus, while fragments of less than one-twentieth the normal size will survive and reform a complete organism if a portion of nucleus is present.^ Similar results have been obtained in experiments on other protozoa; e.g., Verworn's with Thalassicolla.^

There is a large body of similar experimental fact indicating that the continued interaction of nuclear and cytoplasmic components is an essential feature of normal cell-metabolism. It is usually supposed that the nucleus has special relations to synthetic metaboHsm; hence its special importance in growth and regeneration, but the whole problem of the relation of nucleus to cytoplasm is at present in an unsatisfactory state."* It is clear, nevertheless, that the nucleated cell of the higher organism is a complete and autonomous living unit. But in view of what has just been pointed out regarding the non-cellular or subcellular constitution of some organisms, we must avoid regarding the distinctively cellular features of protoplasmic organization

4 For a recent study cf. V. Lynch, American Journal of Physiology, as the all-essential ones. To do so would be to imply that in the early or precellular stages of organic evolution the assimilative or proliferative types of colloidal material, which presumably were then the only systems representing organisms, were not living. The formation of a particular kind of structure is not the essential criterion of vitality; the properties which underlie the formative or structure-building activities are the primary ones. In most animals and plants these activities give rise to a cellular type of structure, but this is not necessarily true of all.

There is a sense, therefore, in which we may regard the cellular type of structural organization as not so much the cause or necessary condition of the vital activities as their product or effect. Obviously all cellular organisms come into existence through the constructive processes of growth. This was pointed out by Huxley, in the early years of the cell theory, in a well-known passage in w^hich he speaks of the cells as being not the producers but simply the products or indicators of vital action. Like the shells on the sea beach the cells ''mark only w^here the vital tides have been and how they have acted. "^ This comparison is an apt one in that it emphasizes the primary importance of the structure-forming vital activity which expresses itself in the formation of cells; but it tends perhaps to subordinate the part played by the cellular structure, once it has been attained. There is no doubt that this

^ Cf. Huxley, "Review of the Cell Theory" in the British and Foreign Medico-chirurgical Review (1853). structure determines, in a quite special way, the character of the protoplasmic processes; i.e., has its own definite causative and controlling influence. The metabolic and other cell processes can be shown to be profoundly influenced by changes in the physical and other state of cell structures. For example, there is evidence that in most cells irritability depends primarily upon the special properties of the external protoplasmic layer or plasma membrane; the contractile, secretory, and similar mechanisms are cell structures; the special relation of the nucleus to constructive metabolism has already been mentioned. In general we may say that physiological activity in all higher organisms is intimately bound up with the special features of structure, chemical organization, and activity peculiar to cells.

A universal peculiarity of living matter, considered simply as a chemical reactionsystem, is that its principal chemical reactions, especially the specific constructive group, occur under the control of structural conditions. If protoplasmic structure is destroyed, mechanically or otherwise, these essential vital reactions at once cease. New structure as it arises in growth or development must therefore have a modifying influence on the metabolic processes and the other physiological processes dependent upon these. The structural characters peculiar to cells cannot fail to influence profoundly the chemical activity of all living systems having the cellular type of organization. One of the fundamental problems of general physiology has reference to the special nature of the relations existing between cellular structure and the chemical processes of the cell protoplasm.

The question^ Why is living matter so characteristically cellular in structure, seems to be equivalent to the question, Why is it partitioned, subdivided into minute, usually microscopical, portions (cells), or structurally discontinuous ? Each portion of protoplasm is separated from its surrounding medium or from adjoining cells by a thin, structurally distinct boundary layer usually called the ''plasma membrane"; and the presumption is that some definite physiological advantage attaches to this peculiarity. The most evident general answer is that this layer serves to separate or insulate the hving protoplasm from the surroundings, and thus to protect it from the disintegrative or otherwise adverse influence of the latter. This view regards the plasma membrane as primarily a protective structure. Through its presence each separate portion of living substance, or cell, is enabled to retain its special composition and individuality. But this answer, while undoubtedly correct in part, is too vague and general to be satisfactory. The recent experimental studies on protoplasmic permeabiUty have thrown a more definite light on the problem. They have shown that in typical living cells the external protoplasmic layer has the properties of a semi-permeable membrane; i.e., it is impermeable or difficultly permeable to the water-soluble substances of low molecular weight present in the protoplasm and surroundings (and to chemically similar substances), while freely permeable to w^ater. Free diffusion of soluble substances either into or out of the cell is thus prevented; the protoplasm can preserve a chemical composition different from that of the surrounding medium without the interference that would result

from unrestricted diffusive interchange/ It is evident that if a minute portion of protoplasm is to retain its special chemical organization, it must be protected against loss of its water-soluble constituents by diffusion, and also against the unregulated entrance of soluble substances from without. Chemical analysis shows in fact that the crystalloidal content of living cells is typically widely different from that of the surrounding medium.^ The presence of a diffusion-proof partition separating each small portion of living protoplasm from its surroundings is apparently an essential feature of the cellular organization.

Without such a diffusion-hindering type of structure, it is difficult to see how a high degree of chemical differentiation could be maintained in such a system as the living organism, consisting, as it does, in large part of an aqueous solution of diffusible substances. Differences in the distribution of soluble substances between protoplasm and surroundings would tend to equalize themselves by diffusion, and chemical differentiation would become difficult or impossible. Morphological differentiation has long been recognized as favored by the subdivision of the developing germ into cells; this condition permits morphogenetic processes in neighboring cells and cell groups to proceed in relative independence of one another.^ In a similar manner an essential

^ For a summary of work in this field, cf. Hober's Physikalische Chemie der Zelle und der Gewebe (1914), pp. 370, 491; cf. also Bottazzi's article in Winterstein's Handbuch der vergl. Physiol., I (191 1), 37. condition for the isolation of chemical and physiological processes in adjacent regions of the organism is the presence of the semi-permeable intercellular partitions. There is also evidence that the internal protoplasm of the single cell is frequently pervaded by a system of films or closed partitions giving a chambered type of structure ; and the possibility of intracellular chemical differentiation ('' chemical organization") has been referred to this condition.^ Such a chambered structure corresponds essentially to that of an emulsion-like or alveolar system. Apparently any physico-chemical system which is built up largely of water and substances in aqueous solution must be a partitioned system if it is to maintain within a small space a high degree of chemical differentiation together with a corresponding diversity of chemical activity.

In general, each living cell can be shown to possess a surface layer (plasma membrane) with properties different from those of the internal protoplasm. At the boundary between this surface layer and the adjoining medium the general phenomena characteristic of phase-boundaries are exhibited. A highly characteristic feature of the living cell is that its surface is sharply defined against the medium, like the surface of an oil drop, very much as if the surface layer consisted of water-insoluble material. This water-immiscible property of living protoplasm and the semi-permeability of its boundary layer are closely associated properties; together they constitute one of the most noteworthy physical peculiarities of living protoplasm. Especially significant is the

^ Hofmeister, Die chemische Organisation der Zelle, Braunschweig fact that they are preserved only while the cell remains living. All cells disintegrate on death; the vital semipermeability and water-immiscibility are then lost. Any living cell, such as a blood corpuscle, suspended in its normal medium, exhibits general physical properties similar to those of a suspended insoluble particle; e.g., an oil drop. These properties are largely an expression of general physical conditions present at all boundary surfaces between adjacent phases, and their consideration becomes of great importance to the physiologist.

In common with other boundary surfaces between mutually immiscible phases the cell surfaces have characteristic electrical properties (interfacial potential differences), exhibit surface tension, and possess the property of condensing or absorbing dissolved substances from the surrounding solution (adsorption). The general role of adsorption in protoplasmic activity is a highly important one, to be considered later in more detail; and undoubtedly this process is a chief factor in the catalytic or quasi-catalytic action of living matter. In general, the catalytic properties of finely divided substances, such as charcoal and colloidal metals, are referable — at least in large part — to adsorption, and the same is probably true of the catalytic properties of living cells. Adsorption appears also to be a factor in the collection of nutrient and other substances from very dilute solution, also a highly characteristic feature of protoplasmic activity.

These considerations show that in addition to limiting diffusion and thus providing for structural and chemical differentiation in the manner indicated, the cellular or partitioned structure of living matter is physiologically important because it furnishes the conditions for another highly characteristic group of properties, those dependent on surface conditions. The protoplasm is thus enabled to utilize (so to speak) the special physical properties exhibited by matter at boundary surfaces. With fine subdivision the proportion of surface protoplasm to the total mass of living substance is large, and the role of surface processes assumes corresponding importance. This general point of view recalls Herbert Spencer's explanation of cell-division as essentially a regulative process, the effect of which is to maintain a certain minimal surface-volume ratio in the protoplasmic mass. The living substance enters into relation with its surroundings through the intermediary of a surface layer, which has special physiological properties, in correspondence with the special nature of the physical conditions resident at boundary surfaces. Evidence will be presented later indicating that these electrical, adsorptive, and catalytic properties of the protoplasmic surface layers determine many of the most characteristic features of protoplasmic activity, especially the automatic and rhythmical processes, the susceptibility to electrical influence, and the various manifestations of irritability.

All organisms have the power of self -maintenance ; i.e., of maintaining their identity and a certain constancy of structure, chemical composition, and activity in spite of continual changes in their surroundings and in their own living substance. The degree of environmental change to which different organisms are exposed varies greatly, and many cells of higher animals pass their whole life in media which are automatically secured against all but slight variation. On the other hand, protoplasmic activity, implying chemical change, is uninterrupted during life; and, as already pointed out, is largely the expression of chemical reactions, chiefly oxidative in nature, by which energy is freed. In all organisms part of the energy thus freed takes such a form that the organism is enabled to maintain itself in equilibrium with its surroundings, grow, and eventually reproduce itself. A curious and highly characteristic cycle of activity is thus shown; thus the animal uses its muscular energy, derived from the oxidation of carbohydrate, to secure more carbohydrate and other materials which serve as sources of vital energy; and this cycle, regulated in accordance with the varying physiological requirements, is repeated continually throughout life. Such facts illustrate the general dependence of life upon the interchange of material and energy

with the environment and explain why so large a part of biological investigation has reference to the interrelations between organism and environment. We may here recall Spencer's characterization of life as essentially a continual adjustment of internal to external relations/ Such an abstract definition, however, applies to many other systems found in nature; e.g., to any system in *' dynamic equiHbrium," such as a candle flame, a whirlpool, or other physical system in which there is an automatically regulated balance between the material and energy supplied to the system and that lost to the environment. Nevertheless, it is pecuHarly true of organisms that their processes are of such a kind as to maintain constantly a certain special complex of structural and active characters in spite of internal and external changes. The requirements for such maintenance vary in the different cases, but certain conditions are universal. The primary condition is that material must be taken from the outside that will serve (i) as a source of energy (to replace substances consumed in. supplying this energy) and (2) as building material for the structural substratum (protoplasm) in which the energy-yielding transformations occur; in this second class are included substances which do not serve directly as sources of energy — e. g., inorganic salts. Considered from the most general point of view, therefore, the living organism exhibits (i) a continual transformation of material taken from its surroundings into its own specifically organized substance; and (2) a continual chemical decomposition of portions of this substance of such a kind as to furnish free energy which is utilized

by the organism in the characteristic activities (foodseeking, etc.) required for its individual maintenance and the perpetuation of its kind. From this general point of view the simplest cases are the most instructive; e.g., that of a single yeast cell or bacterium introduced into a nutrient medium. The organism grows and divides until eventually in place of the single cell there are thousands. Evidently the material of these additional cells comes from the surrounding medium, certain constituents of which are transformed into the living material or protoplasm. The total quantity of material in the whole system, organism plus culture-medium, is unaltered; but its condition has undergone a profound change. A typical nutrient solution for yeast (Pasteur's solution) contains sugar and various salts (NaK tartrate, chlorides, phosphates, and sulphates of Na and K) together with water and oxygen. From these relatively simple materials are built up proteins, lipoids, fats, and other complex bodies; not only are these characteristic substances synthesized but they are distributed or arranged (partly in solid form) in a definite and constant manner so as to give rise to numerous complex and uniformly constituted systems, the yeast cells. Each of these, once formed, becomes the seat of further transformations of the same kind; and by a repetition of this process the non-living material of the medium is progressively transformed into living protoplasm. The transformation is constant and specific, chemically, structurally, and physiologically ; ''heredity" receives here its simplest manifestation.^

^ Cf. my paper, "Heredity from a Physico-Chemical Point of View, Biological Bulletin^ XXXIV (19 18), 65. All organisms and all cells without exception possess this power, that of transforming certain materials selectively appropriated from the surroundings into their own specifically organized and chemically active living substance. The materials used by different organisms vary widely in chemical character and accessibility— contrast the case of a yeast cell growing in a culture-medium with man in his complex social environment— but in every case the essential process is the transformation of non-living environmental material into living substance of a constant and characteristic organization and activity.

The general as well as the special features ot the organization of any living being are an index of the nature and accessibility of the environmental materials required for its maintenance. This is well illustrated by the general morphological and physiological contrast between animals and plants. Since in plants constructive metabolism begins with simple mobile or diffusible materials (CO2, water, salts), present everywhere in the soil and atmosphere, there is no need for locomotion; and these organisms lead typically a stationary existence, remaining rooted to one spot where the necessary materials can reach them by diffusion. The typical radiating, branching, or dichotomous habit of growth, reaching out into all directions of space and thus providing a large area of surface for interchange, is an ''adaptation" to this general environmental condition. Sessile animals also tend to acquire a radiating plan of structure, as illustrated in coelenterates and echinoderms. In the great majority of animals, however, the food supplies have to be selected from an environment containing

relatively little utilizable and much non-utilizable material. In such a case self -maintenance demands, in addition to the ability to move from place to place, a selective power of reaction by which food materials may be picked out and incorporated. Accordingly the responsiveness to external changes (irritability, motor activity) reaches its highest development in this group of organisms. The development of locomotor powers is especially characteristic of animals; related to this is their great variety of reactions and instincts. From such general considerations we may see in a general way how the distinguishing or prevailing characters of each group have arisen in evolution in correspondence with the differences in their methods of nutrition.

In all organisms this selection of assimilable material from the environment and its transformation into living protoplasm proceed automatically and are regulated in correspondence with the physiological requirements, as these vary with the changes of activity and of external conditions. Both the automaticity and the regulated character of these activities are well illustrated by the changes in the reaction of animals to food materials during periods of ''hunger." Consumption of the energy-yielding reserves within the living protoplasm leads to an increased reactivity of the whole organism to these substances. Through this means the maintenance of the metabolic equilibrium is assured under the usual conditions. Regulation of this kind is shown to a greater or less degree by all organisms, and constitutes a fundamental condition of self-preservation; typically if the organism is deprived of any substance or condition necessary for maintenance, its reactivity and behavior

are altered in a manner tending to compensate or remove the deficiency. Thus hunger is, physiologically speaking, increased reactivity to food materials; thirst is increased reactivity to water; the respiratory center of vertebrates increases its rhythm as CO2 accumulates in the blood; when the oxygen in the water is decreased, the gill-cilia of the fresh-water clam beat more vigorously.^ These and many other instances illustrate the manner in which a physiological deficiency may itself furnish the means of setting in motion some physiological mechanism which remedies the deficiency.^ The end-effect of all such regulatory responses is to further the persistence of the organism in its environment. As already mentioned, the term adaptive is usually applied to those special peculiarities of structure and activity by which the organism is automatically conserved in spite of environmental change; hence, from the present generalized point of view any active adaptation may be regarded as a special kind of regulation. It is evident that all such regulations are based upon a highly developed irritability; this fundamental property of irritability, therefore, controls all of the active relations between organism and environment, including the interchange of material and energy which is the essential feature of such relations.

The constructive metabolic processes which build up the living system involve the synthesis of a multiplicity of new chemical compounds from the food materials and ^ In Pfliiger's aphorism, in living organisms "the cause of the need is the cause of the satisfaction of the need." other substances (oxygen, salts, water) furnished by the environment. Of these synthesized compounds the most individualized and specific are the proteins. These compounds, characteristically colloidal in their physical properties, constitute, together with certain other materials, chiefly lipoid, the relatively stable, solid, or^ permanent (structural) portion of the protoplasmic complex.

It is significant that the chief structure-forming compounds should be at the same time those which are chemically the most specific. Specific form and structure are the most obvious peculiarities of the living organism ; hence species are usually distinguished by their structural characters. It is to be remembered, however, that the chemical and physiological characters are equally constant and definite, and must be included in the complete characterization of any species. The essential fact, requiring physiological explanation, is that each individual animal or plant resembles, structurally, chemically, and physiologically other individuals of the same species, while differing from those of other species. As already indicated, the physiological basis of this specificity is to be sought in the specific nature of the chemical processes by which the organism is synthesized. We find in fact that a chemical specificity, corresponding to the specificity of the organism as a whole, is exhibited by its constituent proteins, and apparently by these compounds alone. The other chief biochemical compounds (carbohydrates, lipins) are chemically identical in widely differing species, while the proteins vary in their detailed chemical character from species to species. Apparently each native protein has a special composition

and stereo-chemical configuration, by which it is distinguished from the corresponding proteins of even nearly related species. This general fact of an association between specific chemical composition and specific organic structure indicates, together with other evidence, • that the specific chemical characters of the structural proteins of any organism determine, in a manner which cannot be defined in detail at present, its specific peculiarities as an organic species/ Apparently this chemical specificity determines the more intimate protoplasmic structure, and hence indirectly the protoplasmic activities, chemical and other, which are the correlative of that structure and determine ultimately the physiological and other peculiarities of the species.