Protoplasmic Action and Nervous Action

* Cf. Cremer's [article, op. cit., p. 904, for historical account and discussion. view has more recently been emphasized by Cremer in an important series of studies on the ''Kernleiter" theory, pubhshed at intervals since 1899. He assumes in nerve, in addition to the physical polarization which this tissue exhibits in common with artificial systems of the coreconductor type, the presence of a special '^ physiological polarization,"^ by which termi he means some active process (in the nature of a response or reaction) exhibited at the regions of entrance and exit of the current; this process he conceives as based on a chemical change of some kind, which secondarily may alter polarization and hence serve as the source of a current. In this manner the polarization effect may be renewed at successive areas of a nerve, and transmission to an indefinite distance becomes possible.^

In the foregoing form the Kernleiter theory requires only slight modification in order to make it entirely consistent with the present form of the ''membrane" theory. Both theories agree that a change of polarization is the critical or primary event in the local stimulation process. Evidently if the polarization is confined to the surface of the protoplasmic element (as also of the metal in a core-conductor), this critical change is a surface change. According to the membrane theory, the variation of polarization in stimulation is the result of a sudden change in those features of structure, composition, or permeabiHty which determine the normal electromotor properties of the protoplasmic surface-film or plasma membrane. Such a change may result from

2 See Cremer's exposition in his article in Nagel's Handhuch, pp. 930 flf. a chemical change in the substance of the membrane; the theory further assumes that this change is of such a kind as to be readily induced by the passage of a current. The analogy between this hypothetical type of process and the phenomenon known as "local action" at metallic surfaces is a close one, to which I have recently called attention.^ An example is the spread of corrosion in metals like iron in contact with electrolyte solutions; this spread is the result of local chemical action under the influence of local electrical circuits between the altered and the unaltered areas of the metal. The possibility is thus suggested that in the irritable protoplasmic element the primary change in electrical excitation is also of the nature of an electrolysis. Through the chemical change thus induced the properties of the surface-film are altered in such a manner as to render it ''negative" to unaltered areas; a local circuit then arises at the boundary between altered and unaltered areas and causes electrolysis in the latter, in the same manner as the original current; and by a repetition of this effect the chemical and electromotor change spreads. Evidently a wavelike transmission without decrement is theoretically possible under these conditions. The chief requirement is the presence of a uniform and chemically unstable film forming the boundary layer of the irritable element.

Conditions of essentially this kind are in fact realized in the passive iron wire in nitric acid solution; and in this simple inorganic system the phenomena of activation and transmission exhibit a surprisingly detailed resem- ^ " Electrolytic Local Action as the Basis of Propagation of the Excitation-Wave," American Journal of Physiology, XLI (1916), 126. blance to those observed in nerve and other conducting protoplasmic systems. The possibihty of transmissions of the general type above is thus demonstrated, and the problem becomes chiefly one of determining the special nature of the conditions present in the living system. It is obvious that polarization effects are present at the surface of the iron wire when a current is passed through the system, just as in the case of the platinum wire in the core-conductor experiments of Hermann, Matteucci, and Boruttau. Changes of polarization, however, can give rise to unlimited transmission only in so far as they form the condition of chemical effects which alter the electromotor properties of the surface layer and themselves cause further changes of polarization. The general conditions of transmission in protoplasmic systems will now be discussed briefly, with more particular reference to the case of nerve, where the phenomena of protoplasmic transmission appear to exhibit themselves under the simplest conditions. The fundamental problem, however, is the same for all forms of protoplasm.

According to the law of polar stimulation, the direction of the current between active and resting areas is such, relatively to the surface of the irritable element, that its normal physiological effects— assuming them to be the same as those of an external current led into the tissue — would be to initiate excitation at the resting region adjoining the excited area and to repress activity in the excited area itself. An inspection of the diagram (Fig. 4) will show this. The direction of the local bioelectric current (relatively to the protoplasmic surface) at R is the same as that of the external current at the

cathode of a pair of stimulating electrodes. A current traversing the protoplasmic surface in this direction arises as soon as the local area of stimulation, S, is altered (e.g., by mechanical or chemical action) sufficiently to render it negative relatively to the areas adjoining. The appearance of this current, which acts at a distance from the directly altered area, is apparently the primary effect in stimulation. It initiates the propagated wavelike disturbance, because each secondarily stimulated

Fig. 4. — In A the arrows represent the direction of the current in the active-inactive circuit on one side of the stimulated region 5. In B the course of an external stimulating current from a battery is represented. Stimulation originates, on make, at the cathodal region, where the current has the same direction, relatively to the membrane, as at R. area, e.g., at region R, on itself becoming negative, produces automatically the same effect on regions beyond; and this effect is repeated at each boundary (or region of transition) between an active area and the resting area in immediate advance of it.

There is an analogy of a general kind between the spread of excitation over an irritable protoplasmic element and the spread of combustion along a fuse, with the difference that in the fuse the chemically active area initiates action in the adjoining area through the heat generated in the local reaction, while in the protoplasmic element transmission is effected by the electric current flowing between the two adjacent areas of different potential. In the fuse a temperature gradient exists between the burning area and the region adjoining, and the reaction begins wherever the temperature reaches the ignition point. The rate of transmission in such a case depends on the maximal distance (from the boundary of the burning area) at which this critical temperature is reached and on the rate at which heat is locally developed; i.e., V = Ksr where V is the speed of transmission, s the maximal distance, and r the rate at which temperature rises in the ignited area.^ Similarly in the nerve axone (or other protoplasmic element) the velocity of transmission will be a direct function (i) of the maximal distance, ^ (from the active area) at which the current of the local bioelectric circuit is effective as stimulus, and (2) of the rate, r, at which the current develops; again V = Ksr. Instead of the rate of development, r, we may consider its reciprocal, the time, /, required for the current at the secondarily stimulated point at distance, s, to attain a stimulating value; the shorter this time the more rapid the transmission, i.e., V = Ks/t.^

This equation also applies to the transmission in the passive iron model. Between the activated and the inactive areas of an iron wire immersed in dilute nitric ^ K represents constant limiting factors, such as the rate at which heat is conducted or radiated from the burning region. The distribution of temperature in the gradient on either side of a heated area in a wire (or similar heat-conductor) is in fact subject to the same quantitative law as the distribution of potential in a locally polarized core-conductor, as Cremer has pointed out (op. cit., pp. 906 ff.).

2 For a fuller discussion cf. my article in American Journal of Physiology, XXXIV (1914), 414; cf. pp. 436 ff.; also ihid., XXXVII (1915), 362 ff. Cremer has recently derived a more detailed formula for the acid there is a P.D. of about 0.7 volt; a local current flows between the two areas as indicated in Figure 3 (p. 253), the active area being anodal/ This current is most intense near the active-passive boundary, because the resistance of the portion of current traversing the surface at any point in the passive area increases with the distance of that point beyond the boundary; this resistance depends chiefly on the length and specific conductivity of the column of electrolyte intervening. Up to a certain critical distance, s, beyond the boundary the current wdll have sufficient intensity and local density to reduce the film; hence the time, t, required for the current to reach this intensity and the distance, s, from the boundary are the essential variables to be considered.^ The effect of increasing the resistance of the local circuit (and thus decreasing the distance, s) may be shown qualitatively by suspending the passive wire vertically, with a thin layer of acid adhering, and then touching it below with zinc.^ Under these condi-

velocity of transmission in nerve, introducing as factors the relative electrical resistances of axone and sheath and the specific electrical sensitivity of the tissue, as well as the rate of development of the electric variation. If certain reasonable assumptions are made, this formula agrees weU with observ^ation. Cremer's formula is consistent with the foregoing simpler expression, V = Ks/t, but attempts to define more closely the conditions determining the value of 5 {Ber. ges. Physiol., II [1920], 166; also Cremer's Beitrdge ziir Physiologic, II [1922], Heft i, 31).

^ Only the maximal distance from the boundary at which the current is effective need be considered, since both systems react in the "all or none" manner. This type of reaction is a necessar}^ condition for indefinite transmission without decrement. 3 It may also be shown quantitatively by inclosing the wires in glass tubes of different diameters; the rate varies directly with the sectional area of the tube. tions the wave of activation (marked by the darkening of the metallic surface) moves slowly upward at the rate of only a few centimeters per second; while in the case of a wire completely immersed in a large volume of acid the transmission is too rapid to follow with the eye, i.e., some hundred centimeters per second.

Rapidity in the local variation of potential in nerve or other conducting tissue is thus a necessary condition for rapidity of transmission. The table given on page 328 shows a general proportionaHty between the rate of development of the bioelectric currents in different tissues and the rate of transmission. The slowing of the bioelectric variation in a particular tissue, by cold, anaesthesia, or fatigue, involves a corresponding slowing in the transmission rate. An exact proportionality is hardly to be expected, because the other properties of the tissue, especially its irritabiHty and its electrical conductivity, are also affected by the change of conditions, and other variables enter; but that the rate of electromotor variation is the chief factor determining the speed of transmission seems clearly indicated by the observations cited in the table.

With regard to the other variable, s, the distance from the active-resting boundary through which the local bioelectric current is effective as a stimulus, definite information is difficult to obtain. I have attempted to estimate this distance in frogs' nerve by measuring the maximal distance between tw^o platinum electrodes, applied to the tissue and having a P.D. similar to that of the bioelectric variation (20 to 40 millivolts), at which stimulation occurs on the make and break of a

constant current.' With a P.D. of ten to twenty miUivolts, stimulation occurs at either make or break with the electrodes fifteen to twenty millimeters apart. If we regard the conditions of resistance as similar in a stimulating circuit of this kind and in the normal bioelectric circuit, we may infer that the normal action - current flowing between the active and the resting portions of an excited nerve is effective at a distance of three to four centimeters from the excited area. Other observations, by Hering and others, on secondary stimulation by the demarcation-current, support this conclusion.^

The local action-current in the frog's motor nerve reaches its maximum at about .001 of a second after its initiation (at 20°); this is the duration of the rising phase of the action-current curve, according to the observations of Garten and others.^ If we assume that this current, at the moment of reaching its maximum, has a stimulating effect on all resting regions of the nerve within a distance of three centimeters from the active 2 Cf. article just cited, p. 431. See also the recent observations of Spierling (Cremer's Beitrage zur Physiologie, I [1918], Heft 7) and Keil (Z. fiir Biologic y LXXV [1922], i), on the minimal P.D. required for the stimulation of frog's nerves. In Keil's experiments electrodes of varying form were used, and the stretch of nerve traversed by the current varied between 0.5 and 4 cm. in length. The values obtained were of a similar order to those found in my experiments just cited, but showed wide variation. The potentials required with stretches of nerve 2 to 4 cm. long varied between 20 and ca. 300 millivolts.

3 More recent determinations indicate a more rapid rise; cf. Gasser and Erlanger's observations with the cathode ray oscillograph {American Journal Physiology, LXII [1922], 496; also Plaut, Z. Jiir Biol., LXXVIII area, we have a transmission of stimulating effect through three centimeters in .001 of a second; this is equivalent to thirty meters per second, the usual transmission-velocity at this temperature. The results of this simple calculation are thus in agreement with observation and support the view that transmission is in reality a case of secondary stimulation by the current

Fig. 5. — Diagram of the momentary conditions in a frog's motor nerve axone at 20°. The shaded region marked A, between i?2 and R3, is occupied at the instant under consideration by the excitation-wave, which is regarded as advancing in the direction of the large arrow at the rate of 30 meters per second. Its length, assuming the total duration of the local process (as indicated by the duration of the local bioelectric variation) to be .002 second, is 6 cm. The excitation-process is just beginning at Rj, has reached its maximum at A 10, and has just subsided at R2. The curve represents the variation from the resting potential at different points in the active region; the maximum P.D., at Aio, is ca. 40 millivolts. The regions marked i? are in the resting state. The small arrows indicate the direction of the bioelectric current (positive stream) in a portion of the active-resting circuit. Between jRj and R4 its intensity is sufficient to excite the nerve; excitation is thus always being initiated at a distance 3 cm. in advance of the wave front (i.e., up to R4). For a somewhat similar distance RiRi in ihe wake of the excitation-wave the nerve is refractory to stimulation.

of the local bioelectric circuit. Figure 5 gives a diagrammatic representation of the conditions in a nerve during transmission. The course of the current in the bioelectric circuit should be noted; this course is partly extra-cellular, i.e., through the medium,^ and partly intra-cellular; ^ Part of this current passes through the galvanometer when the action-current is recorded by such an instrument, the remainder through the medium or other extracellular conducting path.

the current at its regions of entrance and exit also traverses the plasma membrane. The current passing lengthwise along the fibers of a tissue like a nerve is undoubtedly subject to electrostatic retardation, as in the analogous case of a cable; and Crehore and Williams^ have calculated that the speed with which its variations are transmitted would thus be reduced to a value similar to that of the nerve impulse. It is evident that an upper limit to the speed attainable by transmission of the kind above is set by the speed with which variations of currentintensity can be transmitted lengthwise along such a conductor, since the general physical conditions present in conductors with boundary surfaces having capacity are undoubtedly present in protoplasmic structures like nerves. Such conditions are common to all conducting paths having a certain structure (core-conductors). It would be erroneous, however, to infer that transmission in nerve is identical with transmission along a cable. The nerve impulse, or any other protoplasmic excitation-wave, is an active process whose energy is derived at each portion of its path from local chemical reactions. Its electrical component furnishes the conditions for transmission from each region to the next, but does not constitute the whole process in the physiological sense. Nevertheless the conclusion that certain physical factors are common to both systems must on general scientific grounds be regarded as correct.

Since a portion (''return path") of the bioelectric current traverses the external medium, we should expect that varying the electrical conductivity of the medium ^ Crehore and Williams, Proceedings of the Society oj Experimental Biology and Medicine, XI (1913), 59. would have a corresponding effect upon the speed of protoplasmic transmission. A lowering of the conductivity of the local bioelectric circuit should involve a corresponding decrease in this velocity, since the maximal distance, s, at which the local current still has stimulating intensity, would be proportionately decreased by any increase of electrical resistance. This critical distance should, other conditions being equal, be proportional to the conductivity of the circuit. Mayor's experiments on the rate of transmission in the nerve net of the medusa Cassiopea in dilute sea water show in fact that within a considerable range of dilutions (down to 50 volumes per cent sea water) a close proportionahty exists between the salt-content of the medium and the transmission rate.^ This result indicates a direct correlation of this rate with the electrical conductivity of the medium. The recent investigation of Pond'' on the speed of the contraction-wave in various forms of muscle (cardiac and voluntary of frog and heart of Limulus), using mixtures of balanced salt solution and isotonic sugar solution, has shown that in these tissues also the speed of transmission runs closely parallel with the electrical conductivity of the medium. The transfer of a muscle from a medium of low to one of high conductivity is followed by a corresponding increase in the speed of the contraction-wave, and vice versa. Mayor's and Pond's observations are difficult to explain except on the assumption that electric currents traversing the cell-media are a chief factor determining the rate at

which excitation is transmitted from one region of a protoplasmic element to another; this assumption, however, is a direct corollary of the local action theory of transmission. Further comparative observations in this field are desirable; the theoretical side also needs careful consideration, since both extracellular and intracellular conductivities are concerned and the precise relations between these two are insufficiently known/ Various other facts of comparative physiology also indicate the importance of the bioelectric variations in the different forms of protoplasmic transmission. The transmission of excitation between cells which are in contact or close proximity but not otherwise connected is a phenomenon difficult to explain except on the electrical theory; the ''rheoscopic frog" experiments have already been cited as examples of transmission demonstrably resulting from secondary electric stimulation by bioelectric currents. Several years ago, I called attention to a number of instances of apparently the same effect;^ for example, one active swimming plate in a ctenophore can influence another through several millimeters of sea water; spermatozoa collected in a clump are soon found beating synchronously;^ ciHated epithelial cells transmit waves of movement; the trans-

^ Brooks has recently studied the relation between the conductance of living cells and tissues ( Laminar ia, yeast, bacteria, Chlorclla) and the conductivity of the medium, and finds a close proportionality between the two (Jour. Gen. Physiol., V [1923], 365. 3 The detached cilia of Paramoecium show a similar behavior, according to recent obsrvations of Al verdes {Arch. ges. Physiol., CXCV missions between neurones in the central nervous system and between nerve-endings and muscle cells are evidently by contact. Such transmissions point almost certainly to electrical conditions. The sensitivity of certain irritable elements to electrical currents in the surroundings has in some cases been developed to a remarkable degree; for example, the catfish will respond to the dipping of a metallic rod into the aquarium at a distance of several centimeters from the fish.^ It is possible that such animals may detect Hving prey through the action-currents accompanying muscular movements.

Transmission of excitation or other physiological influence — implying transmission of chemical influence to a distance in protoplasm — may be called "physiological distance-action," after the analogy of ''chemical distance-action"; the latter is an electrical phenomenon depending on the mutual influence of the. electrode areas in circuits.^ A simple example will illustrate.^ When a copper or platinum wire, e.g., 25 centimeters long, is immersed in a vessel of dilute H2SO4 and touched at one point with a piece of zinc, instantly bubbles of hydrogen start out from the surface of the wire along its entire length. The zinc forms the anode in the local circuit produced by the contact, the copper or platinum is the cathode, hence hydrogen is formed from the surface of the latter at all points where the current-intensity is

^ Parker and von Heusen, American Journal of Physiology, XLIV 3 For other similar instances and a fuller discussion of the biological analogies cf. Biological Bulletin, XXXIII (1917), i35; cf. pp. 157 ff. sufficient. In other words, a reducing action comes instantly into play at a distance from the zinc as soon as the contact is made; this action depends on the passage of the current through the circuit, zinc, acid, and platinum. Similarly when a region of nerve is stimulated, that region becomes negative; and the presumption is that the current of the local circuit thus arising exerts chemical action at all points along the protoplasmic surface where its intensity is sufficient. According to the present theory, it is to this electrochemical action (which secondarily determines stimulation) that the transmission is due. Each area thus secondarily activated serves as a new point of departure for activation of the region beyond, and in this manner transmission to an indefinite distance becomes possible. As already pointed out, the conditions in the passive iron model are of the same general nature. The electrochemical modification of the surface layer (plasma membrane or passivating oxide film), through the electrolytic action of the local circuit, is in both systems the essential change determining transmission.

In the return to the resting or passive state after activation the current of the local circuit is also an essential factor; this current (as the diagram shows) passes in opposite directions (relatively to the surface) at active and resting regions; and correspondingly its physiological effect, which is excitatory at the resting region adjoining the region of acti\dty, is anti-excitatory or inhibitory at the active region itself. Any region, on becoming active, is thus automatically subjected to an electrical influence w^hich hmits or arrests its activity. Hence the local activity, in a muscle cell or nerve fiber,

is temporary, and the state of excitation appears to travel Kke a wave over the irritable element. In the wire model the automatic return of passivity in strong HNO3 is the direct result of the formation of a new surface-film by electrochemical (oxidative) action at the local anodal regions.^ The phenomena of the refractory period in irritable tissues, and the observations already described showing that the plasma membrane of egg cells changes from a temporarily unstable to a stable state after cell-division or insemination,^ indicate that in living irritable elements also the essential condition of recovery after stimulation is the formation of a new surface-film, or the return of the altered film to its original condition. Other physiological facts support this view; e.g., the delay in the return of irritability in a veratrinized muscle after stimulation is associated with a corresponding delay in the return phase of the bioelectric variation. The reversible change in the surface-film is the condition both of the normal bioelectric variation at any region and of the transmission of a similar change of state to adjoining regions.

The well-known interference with stimulation and transmission during the passage of a constant current lengthwise through a muscle or nerve is a further indication of the part played by electrical conditions in transmission. The region near the anode (region of anelectro tonus) acts as a block to an excitation-wave; this effect is undoubtedly complex, but it is probable ^ For a fuller account of the conditions in passive metals cf. the review of Bennett and Burnham, Journal oj Physical Chemistry, XXI

that it depends largely upon the direct physical compensation of the bioelectric current of the approaching excitation-wave, which in the region beyond the actual area of excitation traverses the surface in a direction opposed to that of the polarizing current (Fig. 6). An analogous effect is observed in a passive iron wire in nitric acid when a piece of platinum foil is pressed into close contact with it; an activation wave started Fig. 6. — Showing the opposed directions of action current and polarizing current through the resting portion of the nerve in the anelectrotonic region; the excitation wave is nearing the anodal region of a battery current led into the nerve by non-polarizable electrodes.

at another region of the wire is blocked in the vicinity of the platinum. This effect is dependent on the intersection of the two local circuits, active-passive and platinum-passive, which are opposed in direction. The mutual interference of excitation-waves in living tissues is probably to be explained in an essentially similar manner as an instance of mutual compensation of oppositely oriented bioelectric circuits. This phenomenon is best demonstrated in rings of medusa tissue^ or rings of heart muscle;^ two contraction-waves