REGENERATION

1. The action of the organism as a whole seems nowhere more pronounced than in the phenomena of regenera­tion, for it is the organism as a whole which represses the phenomena of regenera­tion in its parts, and it is the isola­tion of the part from the influence of the whole which sets in action the process of regenera­tion. The leaf of the Bermuda “life plant”—Bryophyllum calycinum—behaves like any other leaf as long as it is part of a healthy whole plant, while when isolated it gives rise to new plants. The power of so doing was possessed by the leaf while a part of the whole, and it was the “whole” which suppressed the formative forces in the leaf. When a piece is cut from the branch of a willow it forms roots near the lower end and shoots at the upper end, so that a tolerably presentable “whole” is restored. How does the “whole” prevent the basal end of the shoot from forming roots as long as it is part of the plant? A certain fresh-water flatworm has the mouth and pharynx in the middle of the body. When a piece is excised between the head and the pharynx a new head is formed at the oral end, a new tail at the opposite end, and in the middle of the remaining old tissue a new mouth and pharynx is formed. How does the “whole” suppress all this formative power in the part before the latter is isolated? It almost seems as if the isola­tion itself were the emancipa­tion of the part from the tyranny of the whole. The explana­tion of this tyranny or of the correla­tion of the parts in the whole is to be found, however, in a different influence. The earlier botanists, Bonnet, Dutrochet, and especially Sachs,[142] pointed out that the phenomena of correla­tion are determined by the flow of sap in the body of a plant. These authors formulated the idea that the forma­tion of new organs in the plant is determined by the existence of specific substances which are carried by the ascending or descending sap. Specific shoot-producing substances are carried to the apex, while specific root-producing substances are carried to the base of a plant. When a piece is cut from a branch of willow the root-forming substances must continue to flow to the basal end of the piece, and since their further progress is blocked there they induce the forma­tion of roots at the basal end. Goebel[143] and de Vries have accepted this view and the writer made use of it in his first experi­ments on regenera­tion and hetero­morphosis in animals.[144] At that time the idea of the existence of such specific organ-forming substances was received with some scepticism, but since then so many proofs for their existence have been obtained that the idea is no longer ques­tioned. Such substances are known now under the name of “internal secre­tions” or “hormones”; their connec­tion with the theory of Sachs was forgotten with the introduc­tion of the new nomenclature.

It may be well to enumerate some of the cases in which the influence of specific substances circulating in the blood upon phenomena of growth has been proven. One of the most striking observa­tions in this direc­tion is the one made by Gudernatsch on the growth of the legs of tadpoles of frogs and toads.[145] The young tadpoles have no legs, but the mesenchyme cells from which the legs are to grow out later are present at an early stage. From four months to a year or more may elapse before the legs begin to grow. Gudernatsch found that legs can be induced to grow in tadpoles at any time, even in very young specimens, by feeding them with the thyroid gland (no matter from what animal). No other material seems to have such an effect. The thyroid contains iodine, and Morse[146] states that if instead of the gland, iodized amino acids are fed to the tadpole the same result can be produced. We must, therefore, draw the conclusion that the normal outgrowth of legs in a tadpole is due to the presence in the body of substances similar to the thyroid in their action (it may possibly be thyroid substance) which are either formed in the body or taken up in the food.

Thus we see that the mesenchyme cells giving rise to legs may lie dormant for months or a year but will grow out when a certain type of substances, e. g., thyroid, circulates in the blood. There may exist an analogy between the activating effect of the thyroid substance and the activating effect of the spermato­zoön or butyric acid (or other partheno­genetic agencies) upon the egg, but we cannot state that the thyroid substance activates the mesenchyme cells by altering their cortical layer.

The fact that the substance of the thyroid may induce general growth in the human is too well known to require more than an allusion in this connec­tion. When growth stops in children as a consequence of a degenera­tion of the thyroid, feeding of the patient with thyroid again induces growth. It may also suffice merely to call atten­tion to the connec­tion between acromegaly and the hypophysis.

It was formerly believed that the nervous system acted as a regulator of the phenomena of metamorphosis in animals, but it was possible to show by simple experi­ments that the central nervous system does not play this rôle and that the regulator must be the blood or substances contained therein. In the metamorphosis of the Amblystoma larva the gills at the head and tail undergo changes simultaneously, the gills being absorbed completely. The writer showed that in larvæ in which the spinal cord was cut in two, no matter at which level,—the sympathetic nerves were in all probability also cut—the two organs continued to undergo metamorphosis simultaneously.[147] Uhlenhuth found that if the eye of a salamander larva is transplanted into another larva the transplanted eye undergoes its metamorphosis into the typical eye of the adult form, simultaneously with the normal eyes of the individual into which it was transplanted.[148] These and other observa­tions of a similar character leave no doubt that substances circulating in the blood and not the central nervous system are responsible for the phenomena of growth and metamorphosis.

An interesting observa­tion on the rôle of internal secre­tion in growth was made by Leo Loeb.[149] When the fertilized ovum comes in contact with the wall of the uterus it calls forth a growth there, namely the forma­tion of the maternal placenta (decidua). This author showed that the corpus luteum of the ovary gives off a substance to the blood which alters the tissues in the uterus in such a way that contact with any foreign body induces this deciduoma forma­tion. The case is of interest since it indicates that the substance given off by the corpus luteum does not induce growth directly, but that it allows mechanical contact with a foreign body to do so while without the interven­tion of the corpus luteum substance no such effect of the mechanical stimulus would be observable. The action of the substance of the corpus luteum is independent of the nervous system, since in a uterus which has been cut out and retransplanted the same phenomenon can be observed.

Bouin and Ancel[150] have shown that the corpus luteum, which in the case of pregnancy continues to exist for a long time, is responsible for the changes in the mammary gland in the first half of pregnancy, when an active cell prolifera­tion takes place in the gland. This process can be interrupted by destroying the corpus luteum artificially. During the second half of gravidity no further cell prolifera­tion takes place, but the cells begin to secrete milk while during the period of cell prolifera­tion such secre­tions do not occur.

Claude Bernard and Vitzou had shown that the period of growth and moulting of the higher crustacea is accompanied by a heaping up of glycogen in the liver and subdermal connective tissue. Smith[151] found that during the period between two moultings, when there is no growth, the storage cells are seen to be filled with large and numerous fat globules instead of with glycogen. He also found that in the Cladocera “the period of active growth is accompanied by glycogen—as opposed to fat—metabolism.” He observed, moreover, that if Cladocera are crowded at a low temperature the fat metabolism (with inhibi­tion to growth) is favoured, while at high temperatures and with no crowding of individuals the glycogen metabolism is favoured. In the latter case a purely partheno­genetic mode of propaga­tion is observed, while in the former sexual reproduc­tion takes place. The effect of crowding of individuals is possibly due to products of excre­tion, which then act on growth and reproduc­tion indirectly by changing the “glycogen metabolism” to “fat metabolism.”

All these cases agree in this, that apparently specific substances induce or favour growth, not in the whole body, but in special parts of the body. Sachs suggested that there must be in each organism as many specific organ-forming substances as there are organs in the body.

We will now show that the assump­tion of the existence of such “organ-forming” substances (which may or may not be specific) and of their flow in definite channels explains the inhibitory influence of the whole on the parts as well as the unbridled regenera­tion of the isolated parts.

2. We have seen that the resting egg can be aroused to development and growth by substances contained in a spermato­zoön or by certain other substances mentioned in the preceding chapter. We will assume that plants contain a large number of cells or buds which are comparable to the resting egg cell, but which can be aroused to action by certain substances circulating in the sap; and that the same is effected for animal cells by substances in the blood. In plants the cells which can be aroused to new growth have very often a rather definite loca­tion while in lower animals they are more ubiquitous. For experi­mental purposes organisms where these buds have a definite loca­tion are more favourable, since we are better able to study the mechanism underlying the process of activa­tion and inhibi­tion (correla­tion). When a leaf of the plant Bryophyllum calycinum is cut off and put on moist sand or into water or even into air saturated with water vapour, new plants will arise from notches of the leaf. This is the usual way of propagating the plant and in no other part of the leaf except the notches will new plants arise. These notches therefore contain cells comparable to seeds or to unfertilized eggs or to the mesenchyme cells which give rise to legs in the tadpole of the frog. The ques­tion arises: Why do notches in the leaf never begin to grow while the leaf is attached to an intact plant, and why do they grow when the leaf is isolated? To this we are inclined to give an answer in the sense of Bonnet, Sachs, de Vries, and Goebel, namely that the flow of (specific?) substances in the plant determines when and where dormant buds or anlagen shall begin to grow. Such substances may originate or may be present in the leaf; but as long as it is connected with a normal plant they will be carried by the circula­tion to the growing points of the stem and of the roots and they cannot reach the notches; while when we detach the leaf, either a new distribu­tion or a new flow of liquids will be established whereby the substances reach some of the notches; and in these notches new roots and a new shoot will be formed. When we cut off a leaf and put it into moist air, not all but only a few of the notches will, as a rule, grow out (Fig. 16); but when we isolate each notch leaving as much of the rest of the leaf as possible attached to it, each notch will give rise to a new plant.[152] (Fig. 17.) We see, therefore, that it does not even require a whole plant to cause inhibi­tion but that we may observe the tyranny of the whole over the parts in a single leaf. The explana­tion is as follows: When we isolate a leaf, some of the notches will commence to grow into new plants and this growth will arrest the development of the other notches of the leaf in the same way as their development was suppressed by the whole plant.

Fig. 16. Growth of roots and shoots in a few notches of an isolated leaf of Bryophyllum calycinum   Fig. 17. If all the notches of a leaf are isolated from each other each notch will give rise to roots and a shoot, but the growth will be less rapid than in Fig. 16. Figs. 16 and 17 were two leaves taken from the same node of a plant.

The explana­tion is the same; those notches which begin to grow first will attract the flow of substances to themselves, thus preventing the other notches from getting those substances. This idea is supported by the fact that if all the notches are isolated from the leaf each notch will give rise to a slowly growing plant, while if the leaf is not cut into pieces, and a few notches only grow out, their growth is much more rapid.

Fig. 18      Fig. 19     Fig. 20

In all these experi­ments the idea that the “isola­tion” in itself is responsible for the growth still presents itself. It can be disposed of by the following experi­ment which never fails. Three leaves of Bryophyllum calycinum are suspended in an atmosphere saturated with water vapour but their tips are submersed in water (Figs. 18, 19, 20). The first leaf, Fig. 20, is entirely separated from its stem, the second leaf, Fig. 19, remains connected with the adjacent piece of stem, and the third leaf, Fig. 18, remains also connected with this piece of stem but the latter still possesses both leaves. The first leaf, Fig. 20, produces new roots and shoots in the submerged part in a few days; the second leaf, Fig. 19, produces no roots or shoots for a long time. This might find its explana­tion by the assump­tion that the first leaf, being more isolated than the second, regenerates more quickly. But this explana­tion becomes untenable owing to the fact that the third leaf, Fig. 18, being less isolated than both (possessing a second leaf in addi­tion to the stem), forms new roots and shoots also more quickly than the second leaf. The phenomena become intelligible in the following way. The fact that in the second leaf shoots and roots are formed very late, if at all, finds its explana­tion not in the lessened isola­tion of this leaf, but in the fact that the forma­tion of a new shoot or of a callus in the piece of stem takes place more quickly than the forma­tion of roots and shoots in the notches of a completely isolated leaf. The stem acts therefore as a centre of suc­tion for the flow of substances from the leaf and this prevents or retards the forma­tion of roots and shoots in the notches. In the isolated leaf of Bryophyllum calycinum no callus forma­tion takes place and hence no flow of the sap away from the leaf will occur. This will allow one or more of the notch buds of this leaf to grow out and then a flow will be established towards these growing buds.

In the third specimen, Fig. 18, the presence of two leaves suppresses or, as a rule, retards the growth of a shoot on the stem and possibly also the flow from one leaf may block to some extent the flow from the opposite leaf if the piece of stem is very short. This puts the leaves in a condi­tion not as good as that in leaf Fig. 20, but better than in leaf Fig. 19.[153]

In the normal plant the buds in the notches of the leaf remain dormant since the flow of the “stimulating” substances takes place towards the tips of the stem and root, and because these substances are retained there in excess. This is probably the real basis of the mysterious dominance of the “whole” over its “parts” or of the anlagen of the tip of the stem over those farther below. When a piece of the stem of Bryophyllum is cut off and its leaves are removed, the two apical buds will grow out first. This “dominance” finds its explana­tion probably in the anatomical structure and the mechanism of sap flow which tend to bring the “stimulating” substances first to the anlagen in the tip. In Laminaria Setchell has been able to show directly that regenera­tion always starts from that tissue which conducts nutritive material.

When we cut out a piece of a stem of Bryophyllum, and remove all the leaves, new shoots will be formed from the two apical buds of the stem, and roots will arise from the most basal nodes; provided that the stem is suspended in air saturated with water vapour. The growth in such a stem deprived of all leaves is slow. If we remove all the leaves on such a piece of stem except the two at the apical end, the stem will form only roots, but these will develop much more rapidly than on a stem without leaves. If we remove all the leaves except the two at the basal end, the stem will only form shoots (at the apical end) but these will develop much more rapidly than in a leafless stem. Hence the leaves accelerate the growth of roots towards the basal end and inhibit it towards the apical end; and they favour the growth of shoots towards the apical end and inhibit it in the nodes located nearer the base.

We thus see that while the stem inhibits the growth of the leaves connected with it, the latter accelerate the growth in the stem. Both facts can probably be explained on the same basis; namely, on the assump­tion that it is the flow of substances from the leaf to the stem which inhibits the growth of the notches and accelerates the growth of the buds in the stem. On this assump­tion it would also follow that the leaves send root-forming substances towards the basal and shoot-forming substances towards the apex of the stem. It also seems to follow from recent as yet unpublished experi­ments by the writer that the root-forming substances are associated or identical with the substances which cause geotropic curvature in the stem.

These observa­tions show that the phenomena of correla­tion or of the influence of the whole over the parts is due to peculiarities of circula­tion or the flow of sap; and that the isola­tion prevents the sap from flowing away to other parts of the plant. There is no need for assuming the existence of a mysterious force which directs the piece to grow into a whole.

Fig. 21   Fig. 22

3. Phenomena of inhibi­tion or correla­tion such as we have described in Bryophyllum are not lacking in the regenera­tion of animals, as experi­ments on Tubularia show.[154] Tubularia mesembryanthemum (Fig. 21) is a hydroid consisting of a long stem terminating at one end in a stolon which attaches itself to solid bodies such as rocks, at the other end in a polyp. The writer found that if we cut a piece from a stolon and suspend it in an aquarium it forms as a rule a polyp at either end (Fig. 22), but the velocity with which the two polyps are formed is not the same, the polyp at the oral end of the piece being formed much more rapidly—a day or one or two weeks sooner—than the aboral polyp. The process of polyp regenera­tion at the aboral pole could, however, be accelerated and its velocity made equal to that of the regenera­tion of the oral polyp by suppressing the forma­tion of the latter. This was accomplished by depriving the oral pole of the oxygen necessary for regenera­tion, e. g., by merely putting the oral end of the piece of stem into the sand. It was, therefore, obvious that the forma­tion of the oral polyp retarded the forma­tion of the aboral polyp. This inhibi­tion might have been due to the fact that a specific organ-forming material needed for the forma­tion of a polyp existed in sufficient quantity in the stem for the forma­tion of one polyp only at a time. This idea, however, was found to be incorrect since when the stem was cut into two or more pieces each piece formed a polyp at once at its oral pole and regenerated the aboral polyps also, but again with the usual delay. It seemed more probable then that the cause of the difference in the rapidity of polyp forma­tion at both ends lay in the fact that certain material flowed first to the oral pole and induced polyp forma­tion here but that this flow was reversed as soon as the polyp at the oral pole was formed or as soon as the forma­tion of the oral polyp was inhibited by lack of oxygen. The partial or full comple­tion of the forma­tion of the oral polyp acted as an inhibi­tion to the further flow of material to this pole. This idea was supported by an observa­tion made independently by Godlewski and the writer that if a piece of stem be cut out of a Tubularia, and if the piece be ligatured somewhere between the two ends, the oral and the aboral polyps are formed simultaneously. This would be comprehensible on the assump­tion that the retarding effect which the forma­tion of the oral has on the aboral polyp was indeed of the nature of a flow of material towards the oral pole.

Fig. 23

Miss Bickford[155] found that the difference in time between the forma­tion of the two polyps disappears also when the piece cut from the stem becomes so small that it is of the order of magnitude of a single polyp. In that case two incomplete polyps are formed simultaneously at each end (Fig. 23). The new head in the regenera­tion of Tubularia arises, as Miss Bickford observed, from the tissue near the wound. At some distance from the wound in the old tissue two rows of tentacles arise, which are noticeable as rows of longitudinal lines inside the stem before the head is formed. Driesch noticed that the newly formed head is the smaller the smaller the whole piece. (This is true, however, only in rather small pieces.) There is, therefore, in small pieces a rough propor­tionality between size of head and size of regenerating piece. Driesch[156] uses this interesting fact to prove the existence of an entelechy, while we are inclined to see in it an analogue to the observa­tion of Leo Loeb, that the velocity of the process of healing in the case of a deficiency of the epithelium decreases when the size of the uncovered area diminishes. While we do not wish to offer any sugges­tion concerning the mechanism of these quantitative phenomena—they may be related in some way with the velocity of certain chemical reac­tions—we see no reason for assuming that they cannot be explained on a purely physico­chemical basis.

The writer noticed that certain pigmented cells from the entoderm of the organism always gather at that end where a new polyp is about to be formed. These red or yellowish cells always collect first at the oral end of a piece of stem. It may be that certain substances given off by the pigmented cells at the cut end are responsible for the polyp forma­tion, but this is only a surmise.

Another sugges­tion made by Child,[157] is that there exists an axial gradient in the stem whereby the cells regenerate the more quickly the nearer they are to the oral pole. If this were correct, and we cut a long piece from the stem of a Tubularia and bisect the piece, the oral pole of the anterior half should regenerate more quickly than the oral pole of the posterior half. According to the writer’s observa­tions on a Tubularian (T. crocea) growing in the estuaries near Oakland, California, both oral ends regenerate equally fast in such cases.

4. The phenomena of regenera­tion in Cerianthus membranaceus, a sea anemone, can be easily understood from the experi­ments on Tubularians, if we imagine the body wall of Cerianthus to consist of a series of longitudinal elements running parallel to the axis of symmetry of the animal from the tentacles to the foot. The number of these elements may be supposed to correspond to the number of tentacles in the outer row of the normal animal. Each such element behaves like a Tubularian, with this difference, however, that the elements in Cerianthus are more strongly polarized than in Tubularia, and that each one is able to form a tentacle at its oral pole only. This fact can be nicely illustrated in the following way: if a square or oblong piece (a b c d, Fig. 24) be cut from the body wall of a Cerianthus in such a way that one side, a c, of the oblong is parallel to the longitudinal axis of the animal, tentacles will grow on one of the four sides only; namely, on the side a b.[158] (Fig. 25.) The other three free edges are not able to produce tentacles. If an incision be made in the body wall of a Cerianthus, tentacles will grow on the lower edge of the incision (Fig. 26).

Fig. 24   Fig. 25   Fig. 26

The writer tried whether or not by tying a ligature around the middle of a piece of an Actinian this polarity could be suppressed; but the experi­ments did not succeed, inasmuch as the cells compressed by the ligature died, and were liquefied through bacterial action so that the pieces in front and behind the ligature fell apart. It is therefore impossible to decide whether or not a current or a flow of substances in a certain direc­tion through these elements is responsible for this polarity, though this may be possible. The writer found, however, that one condi­tion is necessary for the growth and regenera­tion of tentacles which also plays a rôle in the corresponding phenomena in plants, namely turgidity. The tentacles of Cerianthus are hollow cylinders closed at the tip, and by liquid being pressed into them they can be stretched and appear turgid. If, however, an incision is made in the body, the tentacles above the incision can no longer be stretched out. In one experi­ment the oral disk of a Cerianthus was cut off; very soon new tentacles began to grow at the top, and after having reached a certain size, an incision was made in the animal. The tentacles above the incision collapsed in consequence and ceased to grow, while growth of the others continued. On the lower edge of the incision new tentacles began to grow.

Fig. 27    Fig. 28   Fig. 29

It seems also possible that Morgan’s well-known experi­ment on regenera­tion in Planaria can be explained by a flow of substances. He[159] found that if a piece a c d b be cut out of a fresh-water Planarian at right angles to the longitudinal axis (Fig. 27), at the front end a new normal head, at the back end a new tail, will be regenerated (Fig. 28); but that if a piece a c d b be cut from a Planarian obliquely (Fig. 29) instead of at right angles to the longitudinal axis a tiny head is formed at the foremost corner of the piece a and a tiny tail at the hindmost corner b (Fig. [30]). Why is it that in the oblique piece the head is formed in the corner and not all along the cut surface as is the case when the cut is made at right angles to the longitudinal axis? The writer is inclined to believe that the right answer to this ques­tion has been given by Bardeen.[160] This author has pointed out the apparent rôle that the circulatory (or so-called digestive) canals in Planarians play in the localiza­tion of the phenomena of regenera­tion, inasmuch as the new head always forms symmetrically at the opening of the circulatory vessel or branch which is situated as much as possible at the foremost end of the regenerating piece of worm. He assumes that through muscular action the liquids of the body are forced to stream toward this end, and that this fact has some connec­tion with the forma­tion of a new head. There can be no doubt that the facts here mentioned agree with Bardeen’s sugges­tion. The oblique pieces in Morgan’s experi­ments which at first have the heads and tails outside the line of symmetry of the middle piece, gradually assume a normal posi­tion (Figs. 31, 32). The writer is inclined to believe that this is due to mechanical condi­tions. The head a e c of such an oblique piece is asymmetrical, the one side a e being less stretched than the other e c. The higher tension of the piece e c will have the effect of bringing e nearer c, since we know that acid forma­tion and hence energy produc­tion increases in propor­tion to surface, i. e., it must be the greater the more it is stretched. The reverse is true for the tail d f b, and the effect here will be that f will be pulled nearer d. In this way purely mechanical condi­tions are responsible for the fact that the soft tissues of the animal are gradually restored to their true orienta­tion.

Fig. 30   Fig. 31   Fig. 32

As a final possible example of the influence of internal secre­tion or substances contained in the blood may be mentioned the following curious observa­tion of Przibram.[161] In a crustacean, Alpheus, the two chelæ (pincers) are not equal in size and form, one being very much larger than the other. Przibram found that when he cut off the larger pincer in such crustaceans the remaining pincer assumes in the next moulting the size and shape of the removed large pincer; while in place of the removed pincer one of the small type is produced. Hence a reversal of the two pincers is thus brought about. If later on the large pincer is again cut off the process is repeated and the original dissymmetry is restored. Przibram was able to show that the nervous system has no connec­tion with this phenomenon.

The elements which have entered into the discussion thus far are, first, the flow of substances in preformed channels; second, the existence of general or specific substances required for the growing or regenerating organ. A third element is to be added; namely the “suc­tion” effect upon these substances of a developing organ. Thus we see that if one or a few of the notches in a leaf of Bryophyllum grow out the other notches of the leaf are inhibited from growing. There is enough material present in the leaf for all the notches to grow into shoots as is proved by the fact that all will grow out if they are isolated from each other. This was explained on the assump­tion that the notches of a whole which happen to develop first, create a flow of these substances from the rest of the leaf to themselves and thus prevent any getting to the other notches. We stated that this is supported by the fact that the few notches growing out in an undivided leaf grow more rapidly than the many shoots growing from each notch of a divided leaf. But why should a growing shoot or a growing point in general produce such a suction? I think this may be possible on the assump­tion that the consump­tion of these substances by the growing organs causes a low osmotic pressure of these substances in the growing region and this fall of osmotic potential will act as a cause for the further flow. This brings about the apparent “suc­tion” effect of the growing elements upon the flow of substances.

5. We mentioned that when a piece is cut from a Planaria between pharynx and head a new mouth is formed in the middle. It should also be mentioned that according to Child the piece after regenera­tion is smaller than it was before.[162] This indicates that material in the old cells has been digested or has undergone hydrolysis in order to furnish the nutritive material for the new head and tail, since the piece cannot take up any food from the outside before a mouth is formed. These phenomena of autodiges­tion—the process itself will be discussed in the last chapter—seem to occur in many (if not all) phenomena of regenera­tion. It may be that the collecting of red cells at the end in a Tubularian where regenera­tion is about to begin has to do with the furnishing of material by self-diges­tion, since these cells are partly at least destroyed in the process. It is of interest to look for more examples of autodiges­tion accompanying phenomena of regenera­tion.

Fig. 33   Fig. 34

Fig. 35

The writer has observed more closely the trans­forma­tion of an organ into more undifferentiated material in Campanularia (Fig. 33), a hydroid.[163] This organism shows a remarkable stereotropism. Its stolons attach themselves to solid bodies, and the stems appear on the side of the stolon exactly opposite the point or area of contact with the solid body. The stems grow, moreover, exactly at right angles to the solid surface element to which the stolon is attached. If such a stem be cut and put into a watch glass with sea water, it can be observed that those polyps which do not fall off go through a series of changes which make it appear as if the differentiated material of the polyp were trans­formed into undifferentiated material. The tentacles are first put together like the hairs of a camel’s-hair brush (Fig. 34), and gradually the whole fuses to a more or less shapeless mass which flows back into the periderm (Fig. 35). It follows from this that in this process certain solid constituents of the polyp, e. g., the cell walls, must be liquefied. This undifferentiated material formed from the polyp may afterward flow out again, giving rise to a stolon or a polyp; to the former where it comes in contact with a solid body, to the latter where it is surrounded by sea water. These observa­tions suggest the idea of reversibility of the process of differentia­tion of organs and tissues, in certain forms at least. We have to imagine that some of the cells or interstitial tissue is digested and that as a consequence the organ loses its characteristic shape.

Giard and Caullery have found that a regressive metamorphosis occurs in Synascidians, and that the animals hibernate in this condi­tion. The muscles of the gills of these animals are decomposed into their individual cells. The result is the forma­tion of a parenchyma which consists of single cells and of cell aggregates resembling a morula.[164]

Driesch,[165] experi­menting on the regenera­tion of an Ascidian, found that when he cut off the gills and siphons of the animal the portion removed was able to regenerate a whole animal. The gill-piece excised contained no heart, no intestine, and no stolon, and all these organs were regenerated from the gills. In a number of cases the regenera­tion took place by bud forma­tion at the edge of the wound, but in other cases the gills were trans­formed into an undifferentiated mass of tissue from which the missing parts of the animals arose by budding and new gills were formed.

It is probable that the two cases are only quantitatively different. In both, autodiges­tion of certain cell constituents and possibly of whole cells must take place in order to obtain material for the forma­tion of the lost part of the Ascidian. If an interstitial tissue is digested it becomes a ques­tion of how much of this tissue undergoes hydrolysis. If there is little destroyed the old shape of the gills remains, if too much is digested the old gills become a shapeless mass in which a certain number of the old cells are maintained and give rise to the new animal by cell division. The material for the new organs must of course be furnished from old cells which have been digested.

If regenera­tion takes place in pieces which take up no food the newly formed organs must originate from material absorbed from cells of the animal which are hydrolyzed and whose material serves as food for those cells which grow. Very often this process of diges­tion takes place without loss of the total form of the organ and is overlooked by the pure morphologists. In Campanularia also the process of collapse described above is only apparent in a fraction of the cases as in Driesch’s observa­tions on Clavellina.[166] It is also possible that the red and yellow entoderm cells which gather at the end where the new polyp forms furnish the material which is utilized for the process of growth of the cells from which the tentacles arise (with or without giving off specific “hormones” besides).

6. We have mentioned the ideas concerning a design, or “entelechy,” acting as a guide to the developing egg and have shown that this revival of Platonic and Aristotelian philosophy in biology was due to a misconcep­tion; namely, that the egg consisted of homogeneous material which was to be differentiated into an organism. For this supernatural task supernatural agencies seemed required. But we have seen that the unfertilized egg is already differentiated in a way which makes the further differentia­tion a natural affair. This idea of a quasi superhuman intelligence presiding over the forces of the living is met with in the field of regenera­tion, and here again it is based upon a misconcep­tion. The lens of the eye is formed in the embryo from the epithelium lying above the so-called optic cup (the primitive retina). Where this retina touches the epithelium the latter begins to grow into the cup, the ingrowing piece of epithelium is cut off and forms the lens, which probably under the influence of substances secreted by the optic cup becomes transparent. Certain animals like the salamander are able to form a new lens when the old one has been removed by opera­tion, but the new lens is formed in an entirely different way; namely, from the upper edge of the iris. G. Wolf, who observed this regenera­tion used it to endow the organism with a knowledge of its needs; the idea of a Platonic preconceived plan or an Aristotelian purpose suggested itself. But it can be shown that the organism does in this case what it is compelled to do by its physical and chemical structure.

Uhlenhuth[167] has shown by way of tissue culture that the cells of the iris cannot grow and divide as long as they are full of pigment granules as they normally are. When the fine superficial membrane of the iris is torn the pigment granules fall out and the cells can now grow and multiply. If the lens is taken out of the eye of the salamander the fine membrane of the iris is torn and the pigment cells at the edge (especially the upper edge) lose their pigment granules which fall down on account of their specific gravity. As soon as this happens the cells will proliferate. A spherical mass of cells is formed which become transparent and which will cease to grow as soon as they reach a certain size. The unanswered ques­tion is: Why does the mass of cells become transparent so that it can serve as a lens? The answer is that young cells when put into the optic cup always become transparent no matter what their origin; it looks as if this were due to a chemical influence exercised by the optic cup or by the liquid it contains. Lewis has shown that when the optic cup is transplanted into any other place under the epithelium of a larva of a frog the epithelium will always grow into the cup where the latter comes in contact with the epithelium; and that the ingrowing part will always become transparent. This leaves us then with one puzzle still: Why is the growth of the lens limited? The limita­tion in the growth of organs is one of the most important problems in growth and organ forma­tion, though unfortunately our knowledge of this topic is inadequate.

7. The botanist J. Sachs was the first to definitely state that in each species the ultimate size of a cell is a constant, and that two individuals of the same species but of different size differ in regard to the number, but not in regard to the size of their cells.[168] Amelung, a pupil of Sachs, determined the correctness of Sachs’s theory by actual counts. Sachs, in addi­tion, recognized that wherever there were large masses of protoplasm, e. g., in siphoneæ and other cœloblasts, many nuclei were scattered throughout the protoplasm. He inferred from this that “each nucleus is only able to gather around itself and control a limited mass of protoplasm.”[169] He points out that in the case of the animal egg the reserve material—fat granules, proteins, and carbohydrates—are partly trans­formed into the chromatin substances of the nuclei, and that the cell division of the egg results in the cells reaching a final size in which each nucleus has gathered around itself that mass of protoplasm which it is able to control. Morgan[170] and Driesch[171] tested and confirmed the idea of Sachs for the eggs of Echinoderms. We stated in the previous chapter that Driesch produced artificially larvæ of sea urchins of one-eighth, one-fourth, and one-half their normal size by isolating a single cleavage cell in one of the first stages of segmenta­tion of the fertilized sea-urchin egg. He counted in each of the dwarf gastrulæ resulting from these partial eggs the number of mesenchyme cells and found that the larvæ from a one-half blastomere possessed only one-half, those from a one-fourth blastomere only one-fourth, and those from a one-eighth blastomere only one-eighth of the number of cells which a normal larva developing from a whole egg possessed. Moreover, he could show that when two eggs were caused to fuse so as to produce a single larva of double size, the gastrulæ of such larvæ had twice the number of mesenchyme cells. Driesch drew the conclusion from his observa­tions that each morphogenetic process in an egg reaches its natural end when the cells formed in the process have reached their final size.

Since each daughter nucleus of a dividing blastomere has the same number of chromo­somes as the original nucleus of the egg, it is clear that in a normally fertilized egg each nucleus has twice the mass of chromo­somes that is contained in the nucleus of a merogonic egg, i. e., an enucleated fragment of protoplasm into which a spermato­zoön has entered and which is able to develop. Such a fragment has only the sperm nucleus. This phenomenon of merogony was discovered by Boveri and was elaborated by Delage.[172] Boveri, in comparing the final size of the cells in normal and merogonic eggs after the cell divisions had come to a standstill, found that this size is always in propor­tion to the original mass of the chromatin contained in the egg; the cells of the merogonic embryo, e. g., the mesenchyme cells, are only half the size of the same cells in the normally fertilized embryo. Driesch furnished a further proof of Boveri’s law, that the final ratio of the mass of the chromatin substance in a nucleus to the mass of protoplasm is a constant in a given species. Driesch compared the size of the mesenchyme cells in a sea-urchin embryo produced by artificial parthenogenesis with those of a normally fertilized egg and found them half of the size of the latter. When the fertilized eggs and the partheno­genetic eggs are equal in size from the start,—which is practically the case if eggs of the same female are used,—the process of the forma­tion of mesenchyme cells comes to a standstill when their number in the normally fertilized eggs is half as large as the final number in the partheno­genetic egg.[173] Boveri’s results as well as those of Driesch were obtained by counting the cells formed by eggs of equal size and not by simply measuring the size of the cells. It is most remarkable that certain apparent excep­tions to Boveri’s law which Driesch has actually found had been predicted by Boveri.

These facts show that the growth of an organ comes to a standstill when a certain size is reached or a certain number of cells are formed. We cannot yet state why this should be, but we are able to add that the forma­tion of a lens of normal size in the regenera­tion of the eye is in harmony with the phenomena in the embryo. There seems therefore no reason for stating that the regenera­tion of the lens cannot be explained on a purely physico­chemical basis. The only justifica­tion for such a statement on the part of Wolf is that he was not in possession of the more complete set of facts now available through the work of Fischel and Uhlenhuth.

The healing of a wound is a process essentially similar to the regenera­tion of the lens. Normally the cells which begin to proliferate after a wound is made in the skin lie dormant, inasmuch as they neither grow nor divide. When a wound is made certain layers of epidermal cells undergo rapid cell division. Leo Loeb[174] has studied this case extensively. He found that if the skin is removed anywhere, epidermis cells from the wound edge creep upon the denuded spot and form a covering. This may be a tropism (stereotropism) or it may be a mere surface tension phenomenon. Next a rapid process of cell division begins in the cells adjacent to the wound these cells having been heretofore dormant. He is inclined to attribute this increase in the rate of cell division to the stretching of the epithelial cells, and he is supported in this reasoning by the observa­tion that the larger the wound the more rapid the process of healing.[175] During wound healing the mitoses first increase markedly in the old epithelium. With the closure of the wound a sudden fall in the mitoses takes place. The closure of the wound causes an increase in the number of epithelial rows over the defect. This increase is therefore reached at an earlier period in the larger wound since the process of mitosis is more rapid here. Leo Loeb thinks that the pressure of the epithelial cells upon each other leads to a rapid diminu­tion in the mitotic prolifera­tion.[176]

Should it be possible that this is more generally the case, e. g., also in the lens after it has reached a certain size? The condi­tions limiting growth require further investiga­tion.

It is hardly necessary to point out that in these cases we are seemingly dealing with cases of the inhibi­tion of growth which cannot be explained by the tyranny of the whole over the parts, and that there must be condi­tions at work other than the mere flow of substances which can cause a cessa­tion of growth. This can be illustrated by certain observa­tions on the egg.

8. The history of the egg shows a reversible condi­tion of rest and of activity. The primordial egg cell multiplies actively until a large number of eggs are formed in the ovary which may reach into the millions in the case of sea urchins or certain annelids. These cell divisions then stop and the egg goes into the resting stage in which it deposits the reserve material for the development of the embryo. From this condi­tion it can only be called into activity again by the spermato­zoön or the agencies of artificial parthenogenesis.

It seemed of interest to find out whether or not the development of the egg may be reversed once more after it has been activated. From all that has been said in the chapter on artificial parthenogenesis, such a reversal should take place in the cortical layer. The result of these experi­ments seems to be that if a complete destruc­tion or change in the cortical layer has once taken place—such as that caused by the entrance of a spermato­zoön into the egg—no reversal is possible; although the development of the fertilized egg may be suppressed for a long time by either low temperature or lack of oxygen, or, in the case of seeds and spores, by lack of water. But as soon as the condi­tions for the chemical reac­tions in the egg are normal again, the development may go on unless the egg has suffered by the methods used to prevent development or by the long dura­tion of the suppression. With an incomplete destruc­tion of the cortical layer both development as well as reversal of development are possible. Thus the writer has shown that in the egg of Arbacia the effect of the cortical altera­tion of the egg induced by the butyric acid treatment or by the treatment with bases can be reversed. When unfertilized eggs of Arbacia are put for from two to five minutes into 50 c.c. sea water + 2.0 c.c. N/10 butyric acid they will all form a gelatinous, somewhat atypical fertiliza­tion membrane; when put back into normal sea water all will perish in a few hours unless they are submitted to the short treatment with a hypertonic solu­tion mentioned in the previous chapter, while if submitted to this treatment they will develop. If, however, these eggs are transferred from the butyric acid sea water not into normal sea water but into sea water containing some NaCN (10 drops of 1⁄10 per cent. NaCN or KCN in 50 c.c. sea water), and if they remain here for some time (e. g. overnight) they will not perish when subsequently transferred back to normal sea water. Such eggs will develop when fertilized with sperm. The activating effect of the membrane forma­tion has, therefore, been reversed and the eggs have gone back into the resting stage.[177] Wasteneys has found that the rate of oxida­tion which was raised considerably by the artificial membrane forma­tion goes back to the value characteristic for the resting eggs after the reversal of their developmental tendency.[178] Similar results were obtained in eggs activated with NH4OH. It appears from this as though the change in the cortical layer which leads to the development of the egg and the increase in the rate of oxida­tions were reversible in the egg of Arbacia.[179]

The writer had previously noticed that eggs of Strongylo­centrotus purpuratus, which had been treated for two hours with hypertonic sea water, not infrequently began to divide into two, four, or eight cells (and sometimes more) and then went back into the resting state (except that they possessed the second factor required for development as stated in Chapter V). It may be remarked incidentally that such eggs at the time of cell division contained the centrosomes and astrospheres, and yet went back into a resting state, thus showing that the centrosomes are only transitory organs or organs which are only active under certain condi­tions. It is quite possible that in these phenomena of reversal not the whole of the cortical layer has undergone altera­tion.

The writer must leave it undecided whether the changes from the resting to the active state in body cells can also be explained in analogy with these experi­ments.

9. In the formation of the lens we have already noticed an instance where the adjacent organ influences growth inasmuch as the optic cup controlled the forma­tion of the lens. Such influences are quite commonly observed. A piece of Tubularia when cut out from a stem and suspended in water will regenerate at the aboral pole not a stolon but a polyp, so that we have an animal terminating at both ends of its body in a head. The writer called such cases in which an organ is replaced by an organ of a different kind hetero­morphosis.

Fig. 36

Contact with a solid body favours the formation of stolons. Fig. 36 shows a piece of a stem of Pennaria another hydroid, which was lying on the bottom of an aquarium and which formed stolons at both ends a and b. In Margelis, another hydroid, the writer observed that without any opera­tion the apical ends of branches which were in contact with solid bodies continued to grow as stolons, while those surrounded by sea water continued to grow as stems.

Herbst discovered a very interesting form of hetero­morphosis in certain crustaceans; namely, that in the place of an eye which was cut off, an entirely different organ could be formed, namely, an antenna. He showed that the experi­menter has it in his power to determine whether the crustacean shall regenerate an eye or an antenna in place of the eye. The latter will take place when the optic ganglion is removed with the eye, the former when it is not removed. These experi­ments were carried out successfully on Palæmon, Palæmonetes, Sicyonia, Palinurus, and other crustaceans.

The influence of gravitation is very familiar in plants; in stems of Bryophyllum placed hori­zon­tally the roots usually come out from the lower end of the callus. Such phenomena are not often found in animals but they exist here too as the following observa­tion shows.

Fig. 37

Fig. 38

If we cut a piece a b (Fig. 37), from the stem s s of Anten­nu­laria an­ten­nina (Fig. 38), a hy­droid, and put it into the water in a hori­zon­tal pos­i­tion, new stems c d (Fig. 37) may arise on its upper side. The small branches on the under side of the old stem a b begin sud­den­ly to grow ver­ti­cal­ly down­ward.[180] In ap­pear­ance and func­tion these downward-growing elements are entirely different from the branches of the normal Antennularia; they are roots. In order to understand better the trans­forma­tion which thus occurs in these branches, it may be stated that under normal condi­tions they have a limited growth (see Fig. 38), are directed upward, and have polyps on their upper side. The parts which grow down (Fig. 37) have no polyps, but attach themselves like true roots to solid bodies. Thus the changed posi­tion of the stem alone, without any opera­tion, suffices to trans­form the lateral branches, whose growth is limited, into roots with unlimited growth. The lateral branches on the upper side of the stem do not undergo such a trans­forma­tion into roots except in the immediate surroundings of the place where a new stem arises. It seems that the forma­tion of a new stem also causes an excessive growth of roots, possibly because the forma­tion of new branches causes the removal of substances which naturally inhibit the forma­tion of roots. If a piece from the stem be put vertically into the water with top downward, the uppermost point may continue to grow as a stem, while the lowest point may give rise to roots. In this case, therefore, a change in the orienta­tion of organs has the effect of changing the character of organs.

There are only two ways by which we can account for these influences of gravita­tion. Either certain substances flow to the lowest level and collecting there induce growth and possibly changes in the character of growth (as in Anten­nularia) or if the cells have elements of different specific gravity the relative posi­tion of these elements may possibly change and influence in this way the condi­tions for growth. The influence of gravita­tion as well as of contact upon life phenomena are at present little understood.

In all these cases of hetero­morphosis the original form is not restored. It is needless to say that they are incompatible with the theory of natural selec­tion.

The reader will have noticed that in this chapter one term has not been mentioned which is commonly met with in the literature, namely the “wound stimulus.” As the writer had indicated in a former publica­tion,[181] the word “stimulus” is generally used to disguise our ignorance of (and also our lack of interest in) the causes which underlie the phenomena which we investigate. Regenera­tion very often does not take place near the wound but at some distance from it. But even when the regenera­tion takes place at the edge of the wound the latter only serves to create condi­tions for regenera­tion, and these condi­tions cannot be expressed by the word “stimulus.”

While our knowledge of the rôle of the whole in regenera­tion is incomplete in a great many details it seems that the known facts warrant the statement that the phenomena of regenera­tion belong as much to the domain of determinism as those of any of the partial phenomena of physi­ology.