Asa A. Schaeffer

Perhaps none of the factors influencing the streaming of the endoplasm mentioned above exercises as profound and constant an influence as its capacity to form ectoplasm. As has been intimated earlier (p. 3-9) streaming as observed during locomotion is not supposed to be possible at all unless accompanied by the formation of ectoplasm at the forward ends of pseudopods, and its transformation into endoplasm at the posterior end of the ameba. We may therefore next consider the rÔle ectoplasm plays in locomotion, and in some other fundamental activities of the ameba.

In the first place it is necessary to define the word ectoplasm, for two entirely different meanings are sometimes given to it. It is used often to designate the clear non-granular layer of protoplasm which thinly covers some of the commoner amebas, and is especially prominent in some of the small species, where the larger part of the anterior end often consists of protoplasm quite free from granules. The other use to which the word is put is to designate the layer of protoplasm on or near the outside of the ameba which is more or less rigid and motionless, resembling the gel state of a colloid. It is the latter meaning that is given the word as used in this discussion, while I shall follow Jennings (’04) and other, earlier, writers in using the word hyaloplasm in speaking of the outer clear layer. It may be necessary to add that neither of these two words is strictly definable, for in some cases, at least, hyaloplasm is not more rigid than the endoplasm, while in other cases it is. Strictness of definition can, of course, come only as investigation proceeds; and these words as well as the word endoplasm, should not be taken as defining the properties of the substances to which they refer, but only as labels.

The demonstration of the most conspicuous and important property of ectoplasm in Amoeba proteus is easily made. With the high power of the microscope one focusses on the upper surface of an active pseudopod, paying especial attention to the small crystals imbedded in the protoplasm. These crystals, although they dance about slightly (Brownian movement) and otherwise change position to a slight extent, nevertheless appear to be held in place by a very viscous medium. Such movement as is observed in these crystals appears more or less erratic; it is not coordinated and it is only by chance in the direction of locomotion of the ameba. While observing the practically stationary crystals of the ectoplasm one can at the same time, though indistinctly, see the forward sweep of the crystals and other granules in the endoplasm below. But observation fails to detect a definite line of separation between the stationary ectoplasm and the mobile endoplasm; the one grades off insensibly into the other.

The formation of ectoplasm in proteus is a much more complicated process than in almost any other ameba, excepting the large species Amoeba carolinensis[1] discovered by Wilson (’00). We shall have occasion however to refer at length to the method of ectoplasm formation in proteus later on, so we may consider proteus first from this point of view, and then take up a few other species in which the process is simpler.

It is a fact more or less familiar to observers of amebas that proteus, as distinguished from the other amebas, has a number of large irregular, roughly longitudinal folds or ridges on its pseudopods and on its main body (Figure 3). Under normal conditions these are never absent. They are not found at the free ends of advancing pseudopods, but they take their origin at some little distance from the ends. It is this characteristic of ridge formation that complicates the process of the transformation of endoplasm into ectoplasm; for instead of having to deal with ectoplasm formation at the anterior ends of pseudopods only, we find this process taking place irregularly all over the surface of the ameba.

These folds or ridges were first observed by Leidy (’79) and it is an eloquent tribute to the keen observation of this

Figure 3. Formation of longitudinal ridges and grooves in the ectoplasm of Amoeba proteus. A, B, C, D, showing stages in the development of a single pseudopod. a, b, c, d, d¹, cross sections of pseudopods at the levels indicated. The arrows show the direction of endoplasmic streaming with special reference to the formation of ridges. The numerals 1 to 7 indicate the order in which the ridges were formed. Note the tongues of ectoplasm which extend into the endoplasm, in the cross sections.

sympathetically-minded naturalist, that of the large number of subsequent writers on ameboid movement only one (Penard, ’02, p. 63) seems to have noticed these folds. Leidy says that “ ... the main trunk and larger pseudopods of the same ameba (proteus) assumed more or less the appearance of being longitudinally folded. The endosarc axially flowed as if in the interior of thick walled canals, of which the walls appeared to be composed of finer granular matter with scattered imbedded crystals. In the flow, all the contents did not move with the same rapidity, and usually the smaller particles were swept quickly by the larger ones. Other matter, including some of the largest elements appeared to stick to the inner surface of the extemporaneous tubes, but successively became detached to be carried along with the rest of the contents (p. 46).” “The endosarc appeared to flow within thick walls of ectosarc which often seemed to be longitudinally folded (p. 326).” Penard (’02) confirms Leidy’s observation as to the existence of these folds: “The current (of endoplasm) indeed is not unified, but there exist many currents at the same time because of the fact that the endosarc is divided into a certain number of longitudinal canals or grooves by dense walls, which are of a temporary nature, being broken down and built up from time to time. It is easy to distinguish one canal from the other in this species, the currents being at first more or less parallel, but terminating at the forward end, by their coalescence, as a single mass of liquid (p. 63).” But Penard questions Leidy’s conclusion that the walls are of ectoplasm: “Moreover Leidy deceives himself without any doubt in considering these partitions as folds of the ectosarc. The latter, in the rhizopods, is not a special substance, it is a plasma of surface, specialized for the functions which it has to perform, capable of modification as to its intimate structure, but only so temporarily (p. 63).”

Although it is a very simple matter to prove to one’s satisfaction the mere existence of these folds—a few minutes’ observation under the high power of the microscope will do that—it is a much more difficult matter to observe how these folds originate, because of the incessant changes going on, as recorded by Leidy.

Very young or small pseudopods in proteus have the same general appearance as the pseudopods of other large species (dubia, laureata, discoides, annulata, etc.); that is, there is a central axial stream of endoplasm surrounded by a layer of ectoplasm. But there is one difference even here, and that is the greater thickness of the ectoplasmic walls in proteus in proportion to the diameter of the pseudopod. The ectoplasmic tube however is not solid throughout, but is more or less honeycombed, somewhat like a network, with the spaces filled by endoplasm.

If the ectoplasm is actually endoplasm that has passed into the gel state, then the honeycomb condition just described resembles an intermediate stage where only a part of the endoplasm has been transformed. This network of endoplasm is strong enough however to impede the flow of the main stream of endoplasm along the sides of the pseudopod; but when large objects, such as the nucleus or food masses, too large to be readily carried in the endoplasmic stream, impinge against the imperfectly solidified sides of the tube of ectoplasm, the innermost strands of the spongy network of ectoplasm snap, usually with readiness, allowing the large object to pass by.

The surface of a young pseudopod is smooth, a cross section being oval in shape (Figure 3, a); but as the pseudopod increases in size, large folds or ridges begin to make their appearance. Usually the first ridges to appear are lateral. They begin as small waves of hyaloplasm which flow out along the sides of the pseudopod for a short distance and then continue to move forward. The endoplasm then flows in a number of small parallel streams amid numerous obstructions through the ectoplasmic tube of the pseudopod into the wave of ectoplasm. After the ridge is well begun, there is frequently observed a slow forward-moving stream of endoplasm within it, but the ridge is never closed from the main endoplasmic stream, as is readily proved by the numerous small streams of endoplasm which continually filter through the ectoplasm into the ridge.

In addition to the lateral ridges, which, as stated, are usually formed first, there appear ridges on the upper side of the pseudopod as well, and presumably also on the under side. So far as could be determined these ridges are all formed in much the same way; that is, by the projection of a small wave of protoplasm from some part of the surface of the pseudopod. The ridges do not always grow by extension at the anterior end as described above. Not infrequently a ridge ten to twenty times as long as wide is pushed out along its whole length at once. This is especially likely to happen in a slender pseudopod that suddenly becomes the main pseudopod. The width of a ridge, especially on the upper surface, does not change much after formation. One can frequently find two or three ridges of about the same width, which run the whole length of the ameba with the exception of a short distance at the anterior end, where, as before stated, there are no ridges.

As the figure indicates, new ridges may be formed from previous ones, either by lateral or endwise extension. In such case the walls of the ridge send out thin waves of hyaloplasm followed by streams of endoplasm, as described above in the formation of the first ridge on a pseudopod. When a pseudopod forms a branch, the ridges on the old pseudopod do not likewise branch, but new ridges are formed which have no connection with old ones, but they may later coalesce with old ridges. Such coalescence is however exceptional. Once a ridge is formed, it retains its identity as a rule; that is, as the ameba moves forward, the ridge in effect moves back over the ameba to lose itself in the wrinkles at the posterior end (See Figure 11, A). The number of ridges on any random selection of amebas is variable, and is moreover difficult to state. A large ameba may have as many as six or seven side by side on its upper surface. The number on the sides and on the lower surface are difficult to estimate. The space between ridges is about equal to the width of the ridges, but as one passes toward the posterior end, the ridges become more closely crowded together.

From these observations on the formation of ridges it is evident that they do not represent a wrinkling of the surface such as occurs in a semi-rigid curved surface when it is made to occupy a smaller space. The ridges are wrinkles only in appearance, not in origin. The surface of the ridges is younger than the space between them. It appears as if the pseudopod which has to widen as it increases in length, could not liquify the ectoplasm uniformly all around, but only in longitudinal strips here and there, and that through these openings the ectoplasm then flows. There is no question about the greater readiness with which ectoplasm is formed in this ameba as compared with many others, but after a careful comparison of proteus and carolinensis, where ridges are formed, with discoides (Figure 11, B), dubia (Figure 11, C), laureata (Figure 4) and annulata, where none are formed, the only conclusion presenting itself is that the visible physical properties of the protoplasm of proteus and carolinensis give no hint as to the cause of the presence of ridges in these species. The protoplasm of discoides and laureata is about as viscous as that of proteus, yet in these there is never any ridge formation.

The ridges in proteus recall, of course, the ridges always observed in verrucosa, sphaeronucleosus (Figure 13) and their congeners, especially while the latter are in locomotion. A sphaeronucleosus is especially favorable for study in this connection because of its greater activity. This ameba has four or more longitudinal ridges on its upper surface, while in locomotion, which strongly resemble those in proteus and carolinensis. The chief difference lies in the fact that in sphaeronucleosus the ridges are extended at their anterior ends continually, and unless the direction of locomotion is changed, the ridges may retain their identity while the ameba moves several scores of times the length of its body. Along the sides, however, new ridges are continually replacing older ones. When the direction of locomotion is changed, the old ridges usually all disappear into a jumble of ridges and crinkles running in every conceivable direction, and with the reestablishment of locomotion along a more or less straight path, a new set of ridges appears. In sphaeronucleosus and its congeners, the ridges are also not wrinkles, but ridges that are formed later than the surface contiguous to them.

It is interesting to recall also that the ectoplasm in sphaeronucleosus, verrucosa and the rest of this group, is much firmer than in most other amebas.