ON THE NATURE OF THE SURFACE LAYER
The observations in the preceding chapters on the general movements of
the surface layer of amebas will afford a sufficient basis for an
inquiry into the nature of this layer. The mere demonstration of the
existence of this layer is, of course, interesting enough, for a number
of contradictory statements by various students of the amebas are
satisfactorily cleared up by these observations. But the problem of
ameboid movement affects other organisms besides amebas, and since the
movement of the surface layer is so intimately associated with ameboid
movement, it becomes of more than ordinary interest to learn something
of the nature and composition of this layer.
In the first place the property of carrying particles toward the
anterior end of amebas does not appear to be of any advantage. That is,
whatever the movements of the outer layer may be, the ameba does not
appear to be better off when particles are carried forward than when
none are carried, for such particles are very small and almost without
exception devoid of food value. The particles are masses of debris which
accidentally adhere to the ameba, and the ameba makes no visible effort
to make such particles adhere, nor to get rid of them. The ameba seems
to be quite indifferent to the presence of such particles.
On the other hand, as Schaeffer (’17) has pointed out, the capacity for
transporting particles cannot but be looked upon as a hindrance to
locomotion. As has been stated, the surface film moves in the same
direction as the ameba. Whenever the surface film comes against a solid
object, it pushes against the object, and nullifies to a certain, though
small, extent the energy expended in moving forward. And it will be seen
without further argument, of course, that the energy involved in
carrying particles forward is not only itself lost but consumes an
appreciable part of the energy available for forward movement. This
fact, together with the universal occurrence of this phenomenon among
amebas indicates beyond question that it is intimately associated with
ameboid movement as it is ordinarily understood in amebas, and that it
is almost certainly a “necessary” physical consequence of the more
fundamental physical processes involved in the movement of amebas.
That the third layer moves in the same general direction as the ameba
has already been mentioned. The direction of a moving particle is
however not necessarily parallel with the stream of endoplasm below. In
a retracting pseudopod that lies nearly parallel to and by the side of
the main advancing pseudopod, the particles on the far side and near the
base frequently move across the pseudopod at an angle (and therefore
also across the endoplasmic stream), and up the active pseudopod on the
near side. This shows conclusively that the direction of flowing
endoplasm by itself has no direct connection with the direction of flow
of the surface layer.
To say that the particles carried by the surface layer bring up at the
anterior ends of pseudopods or of the ameba when in clavate shape,
admits of further qualification. The advancing edge is not a straight
line but an arc, and the sides near the advancing edge are building at a
slower rate than the extreme tip. The most rapid formation of ectoplasm
is at that point of the ameba that is farthest ahead. At this point all
the ectoplasm to be made is still to be made, but as one passes back
along the side of the pseudopod more and more ectoplasm is encountered
and less and less remains to be made. There is therefore a gradient in
the rate and in the amount of ectoplasm formed as one passes back from
the forward end of the longitudinal axis of the pseudopod along the
side. This is especially the case with certain amebas like _Amoeba
discoides_, _A. laureata_ and others in which the pseudopods are more
nearly cylindrical. In such amebas as _A. proteus_ and _A. verrucosa_,
the factor of ridge formation complicates to some extent the
longitudinal gradient of ectoplasm formation. But in spite of these
specific differences, the general statement still holds that the rate of
ectoplasm formation at the extreme anterior end is higher than anywhere
else in the ameba, and that the rate gradually falls to zero as the
nearly straight and parallel sides of the pseudopod or ameba, as the
case may be, are approached.
Now we have seen that if a particle becomes attached to the outer layer
of such an ameba as discoides, which has nearly symmetrical pseudopods,
at some considerable distance from the tip of the pseudopod, it moves
forward until the tip of the pseudopod is reached. It does not tend to
come to rest near the tip of the pseudopod, where the rate of ectoplasm
formation is much higher than at the sides of the pseudopod, though not
as high as at the tip, but it moves on until the tip is reached. That
is, the movement of particles on the surface film is toward that small
area at the extreme anterior end where the rate of ectoplasm formation
is highest.
In such an ameba as _verrucosa_, however, the highest rate of ectoplasm
formation would be, not at a small circular area, but a very narrow
strip along the anterior edge; for the rate of ectoplasm formation over
a considerable portion of the width of the anterior end of the ameba is
practically the same, according to observation. Consequently we do not
find particles which are attached to the outer layer tending to move to
a point lying on the longitudinal axis, but their paths are found to be
straight and parallel with the longitudinal axis, if headed toward any
point over a considerable stretch of the anterior edge on either side of
the longitudinal axis.
All the evidence that is at hand therefore points to the conclusion that
the direction of movement of the surface film in a moving ameba is
toward that point where ectoplasm is formed most rapidly.
But where do the particles come from? At exactly what regions of the
ameba do they start to travel toward the anterior ends of the ameba? In
_sphaeronucleosus_ and its congeners, it is very difficult to determine
just when the particles begin to move toward the forward edge. Particles
near the posterior end on the upper surface of these amebas moved
forward slowly, much more slowly than particles near the middle.
Sometimes particles near the posterior end seem to be motionless for
some time, but the incessant though slow kneading process going on at
the posterior end makes accurate observation difficult. Only in a
general way it may be stated that particles begin their forward march at
or near the posterior end. In amebas that habitually form pseudopods
more accurate information can be obtained.
In _proteus_ or _discoides_, for example, projecting pseudopods are
often suddenly stopped and retracted, with a resultant change of an
anterior to a posterior end. Particles attached to the outer surface on
such pseudopods move toward the anterior end, of course, as long as the
pseudopod is building, in the manner described in the preceding pages.
But when the endoplasmic stream is arrested, the forward movement of the
particle likewise stops. When the endoplasm starts to flow back into the
main body of the ameba, the particle also starts moving back; but there
is a period of a few seconds after the endoplasmic stream is reversed
during which the particle remains quiet. And when it does start in to
move, it moves only slowly. Within a few seconds, however, the average
speed of movement is attained. This is true of particles located some
distance away from the tip of the pseudopod. If the particle has reached
the tip of the pseudopod before reversal of the endoplasmic stream takes
place, the particle often remains at the tip until the pseudopod is
almost completely withdrawn into the main body of the ameba (Figure 26,
p. 60). At other times such a particle becomes displaced, presumably by
irregular retraction of the tip of the pseudopod, and finds itself at
the side of the pseudopod. When this happens it moves slowly toward the
main body of the ameba, but faster than the tip of the pseudopod does.
It frequently happens, especially in _annulata_, but also in _proteus_
and other forms with many pseudopods, that when an advancing pseudopod
is about to be withdrawn, there intervenes a stage where the endoplasm
in the distal part moves away from the ameba, while that in the proximal
part moves toward the ameba, with a neutral or motionless zone between.
In such case a particle on the distal end moves slowly toward the tip
while a particle in the proximal region moves toward the base of the
pseudopod. Particles over the neutral zone are motionless. In these
cases, however, changes in the direction and speed of the ectoplasmic
stream are too frequent and the relative strengths of the distal and
proximal currents too variable, to enable one to secure very accurate
data by means of camera lucida drawings (a kinematograph is essential
for this purpose), so no figures of the speed of movement of such
particles are given. Nevertheless the general results of the
observations are as stated. It might be added that in some cases the
neutral zone for the particles attached to the surface did not coincide
exactly with the neutral zone of the endoplasm, but was located a little
further distally.
From these observations it appears that a rough index of the direction
of movement of the surface film is the direction of the streaming of the
endoplasm; and that the surface layer moves away from regions where
ectoplasm is in the process of being converted into endoplasm. Since a
particle attached to the surface may remain for some time at the tip of
a retracting pseudopod, while one that is attached to the sides of a
pseudopod moves toward its base, it appears that the speed of the moving
surface film is not directly correlated to the rate of transformation of
ectoplasm into endoplasm. The slower speed of particles near the
posterior end points also in this direction. The formation of ectoplasm
at the anterior end seems therefore to be much more intimately connected
with the movement of the surface film than the destruction of the
ectoplasm, though it is not yet clear that the liquefaction of the
ectoplasm is altogether without effect.
Now as to the speed with which the surface film moves. The foregoing
illustrations and figures show that the particles attached to a
_sphaeronucleosus_ on the upper surface move from 2.5 to 3.6 times as
fast as the ameba (Figure 19) while particles attached to a _discoides_
move only from 1.2 to 2 times as fast as the ameba moves. In _proteus_
the speed of the particles is still slower, because of the longitudinal
ridge-like waves of protoplasm which are continually being thrown out.
In this species it frequently happens that because of the numerous
ridges, the ameba moves faster than the particles attached to the outer
surface; but this is to be looked upon as a mechanical complication, not
as indicating a difference in the nature of the surface layer.
How is the difference in the speed of movement of the surface layer
between _sphaeronucleosus_ and _discoides_ to be explained? There are no
ridges to retard the movement of particles in _discoides_, while there
are ridges in _sphaeronucleosus_, where the particles move on the
average twice as fast as on _discoides_. In the first place the
advancing edge, the edge where ectoplasm is being made, is
proportionately much wider in _sphaeronucleosus_ than in _discoides_ as
compared with the amount of surface back of it. Figures 23 and 24 show
that the rate of movement of the surface film is directly proportional
to the amount of new ectoplasm forming. In the second place, the greater
part of the under surface in the forward half of _sphaeronucleosus_ is
attached to the substrate, so that the surface layer which flows toward
the anterior end is derived almost wholly from the upper surface; while
in _discoides_ the whole surface in free pseudopods, and nearly the
whole surface in attached amebas (cf. Dellinger’s observations described
on p. 56) possesses mobile surface protoplasm. Observation of moving
particles on these amebas proves this. Then again, the anterior edge of
a _sphaeronucleosus_ is not attached at the points farthest advanced,
but the point of attachment is some distance back, as indicated in
figure 20. The effect of this is to increase the amount of forming
ectoplasm in proportion to the surface of the ameba from which surface
protoplasm may be drawn. Still one other factor must be considered. As
is well known _sphaeronucleosus, verrucosa_ and their congeners possess
longitudinal ridges on the upper surface which consist of ectoplasm,
covered of course by the surface film. These ridges are formed near the
anterior edge, not by wrinkling, but by the construction of new
ectoplasm. Once formed, they remain until the ameba, so to speak, flows
out from under them. That is, the ridges undergo comparatively slight
changes until changed back into endoplasm at the posterior end of the
ameba. As the ameba flows ahead the ridges are of course continually
being added to or lengthened, by the conversion of some endoplasm into
ectoplasm. The ridges may thus retain their identity for a long time
although the substance composing them is changed every time the ameba
moves the length of its body. It is clear, therefore, that there is more
ectoplasm formed at the anterior end of a _sphaeronucleosus_ than would
be the case were the upper surface of the ameba plane; and the
conclusion therefore is obvious that the formation of ridges, occurring
as it does, chiefly at the anterior end, serves further to accelerate
the forward movement of the surface film.
If the form of _sphaeronucleosus_ were more regular than it is, the
amount of ectoplasm in the process of forming at any given moment could
be compared with a similar relation existing in _discoides_, to see
whether these respective ratios were proportional to the speed of the
moving surface films in the two amebas. As it is, the irregularity of
form of _sphaeronucleosus_ makes such computation subject to the
possibility of considerable error. In _discoides_ however the problem is
comparatively simple. I therefore did not go into this matter
extensively, but merely worked out the relations mentioned in one case,
and I mention it here to illustrate the method rather than to record the
result, which is not to be taken as very exact.
[Illustration:
Figure 28. A clavate _Amoeba discoides_, showing the amount of
ectoplasm that is constantly being made at the anterior end. Length
of the ameba, 310 microns.
]
Since the movement of the surface film is obviously a surface
phenomenon, only the surfaces of the amebas need to be taken into
account. In Figure 28 is illustrated a _discoides_ of such a shape as to
allow a fairly accurate computation of its surface. Three outlines of
the anterior end only are given; the rear portion of the ameba remained
approximately the same size and shape in the three outlines. The cross
lines at the anterior end divide the forming ectoplasm of the ameba from
the formed. As will be noticed the cross lines are drawn through the
intersections of two successive outlines. Computing the areas on both
sides of the cross lines for the two outlines and averaging them, there
is found a ratio of 1 to 10; one-eleventh of the total surface
represents forming ectoplasm, and ten-elevenths formed ectoplasm.
(One-twenty-second of the total surface was deducted for surface
attached to the substratum.) _Sphaeronucleosus_ stands in contrast with
_discoides_ for it is attached to the substratum over a much greater
area and in consequence only a slight amount of surface is drawn from
the under side. This ameba may therefore be regarded in this connection
as of only one surface, the upper. That part of outline 1 in Figure 14
cut by outline 2 indicates, as in _discoides_, the region of forming
ectoplasm, and the space between outlines 1 and 2 may be used as a basis
of computation. New ectoplasm is formed in this zone and far enough back
to include the tips of the longitudinal ridges, of which we have already
spoken (Figure 13). The zone of forming ectoplasm would therefore be
about twice as wide as the average width of the three zones between the
successive outlines in the figure, and of approximately the same shape.
On this basis, the surface occupied by forming ectoplasm is 1/5.8 of the
total surface, and the ratio of formed to forming ectoplasm is 4.8 to 1.
(For the sake of completeness, a few factors whose values cannot easily
be computed may be mentioned. 1. The anterior edge is not attached to
the substratum at its farthest point, but at some little distance back
of the edge, thus increasing the relative amount or forming ectoplasm;
but this is offset by the surface of a part of the under side at the
posterior end where the surface layer is active. 2. The ectoplasm
composing the ridges, which must be added to the formed ectoplasm, would
increase the ratio, though only slightly).
Approximately twice as much ectoplasm is therefore in the process of
formation in _sphaeronucleosus_ as in _discoides_ when compared with the
formed ectoplasm in the respective amebas, over which the surface film
is active. This ratio corresponds very well with the rate of movement of
the outer surface in these amebas, which as we have seen is about twice
as fast in _sphaeronucleosus_ as in _discoides_.
Where does the surface layer come from and what becomes of it after it
arrives at the anterior end? It moves continually forward as long as the
ameba moves forward. There would seem to be a tendency therefore for it
to accumulate at the end of a free pseudopod in such a form as
_discoides_, and even under ordinary conditions of locomotion where
there is occasional attachment to the substratum by very short
pseudopods, the surface layer is continually moving toward the anterior
end on practically all sides. Every time, therefore, that the ameba
moves a little less than its own length, there would accumulate at the
tip of the ameba, if it were not removed, an amount of surface layer
equivalent to that which covers the whole ameba. No such accumulation
can be detected however, from which we infer that it is removed as fast
as brought there. And the posterior region of the ameba, which is the
main source of the surface film, does not become poorer in this material
by reason of its continual flow forward, but new surface is made
continually to take the place of that moving forward. This process of
destruction and creation of surface is accordingly rapid during active
locomotion;--a _discoides_, moving approximately once its length at room
temperature in two minutes, destroying therefore the equivalent of its
entire coat of surface in that time; while a _sphaeronucleosus_, moving
once its length in two or three minutes, destroys all its surface every
minute.
From what has been said thus far, it must be apparent that there is
striking resemblance between the general movement of the surface layer
of the ameba, and of a surface tension layer in a drop of fluid in which
the tension is changed at some point. Let us now inquire briefly into
this resemblance.
As is well known the surface of a liquid in contact with another liquid,
solid or gas, with which it does not mix, behaves like a stretched
membrane, so that when the tension is reduced at any point the surface
layer moves away from that point. A good illustration of the effect of a
decrease of surface tension is found in a drop of clove or other oil
with which some substance that reduces the surface tension, such as
alcohol or soap, is brought into contact at one side. If previously some
dust particles have been placed on the surface of the oil drop, it will
be easy to see that the surface of the oil moves to the opposite side
from where the alcohol or soap solution touched the oil. In practice it
is a very simple matter to lower the surface tension of a drop of fluid
as described, so as to show the movement of particles on the surface.
Almost any liquid may be used for this purpose. But it is comparatively
very difficult to _increase_ the surface tension at some point of a
drop of fluid in such a way as to cause particles on the surface to move
toward that point. The principle underlying the movement of the surface
film in both cases is however exactly the same; so, although it would be
more desirable to compare the surface movements in a drop of fluid in
which the surface tension is increased at some point, because this is
what happens in an ameba during locomotion, we shall nevertheless find
it necessary to consider a drop of fluid in which the surface tension
has been lowered. The application of the illustration is readily made.
When the surface tension of a drop of fluid is lowered by bringing into
contact with it some other substance that possesses this power, the
surface rushes away at great speed in all directions from the point
where the tension is lowered, because usually the tension is reduced
very considerably. In this surface movement it is found that new surface
is made where the tension is lowered and old surface is destroyed, that
is, pulled into the interior over a large part of the surface opposite
to where the tension is lowered. The speed of the surface movement is
most rapid near the point where the tension is lowered and becomes
gradually slower as the opposite side of the drop is approached, where
there is no movement. This variation in speed of the moving surface
seems to be due largely to the small area in which the tension is
lowered as compared with the whole surface of the drop.
In the ameba the conditions are reversed. The surface layer moves
_toward_ a point with increasing speed, instead of away from a point. In
both the ameba and the drop the greatest speed is attained near the
small area where the change in surface tension occurs.
The behavior of large and heavy particles on the surface of a drop of
fluid and on an ameba are similar. A heavy particle as of sand, or a
small glass rod, laid on the ameba, is not moved by the surface layer.
It forms an island of surface matter around which the moving surface
layer flows. Precisely the same thing happens in surface layer movements
in inanimate fluids.
Again in point of thinness there is no disagreement so far as
microscopic observation goes. Neither the surface film on an ameba nor
the surface film on a fluid can be directly observed microscopically to
be different from the fluid below it. The surface layer is, as is
generally believed, of molecular dimensions, and its thickness is beyond
the limits of vision. Unless some special means is discovered therefore
for making visible the surface film, such as a process of staining, it
may be impossible to ascertain its ultimate structure directly, for it
overlies a mass of heterogeneous fluid whose composition is constantly
changing.
It seems to follow from what is observed of the surface tension layers
of the fluids of physics that such layers must be of the same
constitution as the body of the fluid over which the layer is formed,
although, as is well known, the proportion of the ingredients in the
surface layer is different from that in the body of the fluid. Now since
the resemblance between the surface layer of an ameba and a surface
layer on a drop of fluid has thus far been found to be complete, it is
pertinent at this point to discuss Gruber’s (’12) suggestion that the
movement of particles forward on an ameba is due to the forward movement
of an inert layer of mucus or gelatinous material secreted by the ameba.
To begin with, observation does not support Gruber’s suggestion. No such
layer can be seen. Such a layer, since it is shown to persist for
several minutes at least, should remain after an ameba bursts, under
experimental conditions, but no such remains can be seen. Its existence
should be demonstrable by the use of dyes, but the evidence is negative.
Indeed there is not any direct evidence that can be brought in support
of the suggestion that this surface layer is gelatinous in composition.
Moreover, as we have seen, the layer on the ameba that carries particles
forward seems to be destroyed at the anterior end, for in what other way
would particles remain at the anterior end after being brought there?
But the supposition that a gelatinous layer might be drawn into the
interior at the anterior end is also negatived by observation, for no
very small particles clinging to or imbedded in the surface substance
are ever drawn into the ameba, as would almost certainly be the case if
the substance composing the layer were gelatinous. And as to supposing
that this layer, if gelatinous, might behave essentially as a surface
tension layer and therefore be drawn in at the anterior end of the
ameba, this is contrary to the experience of physics; for the physical
nature of the ameba would make it impossible for the ameba to have a
surface layer of gelatinous matter. There do not seem to exist any
grounds therefore for supposing that the outermost layer of an ameba,
the layer that carries particles as described in the preceding pages,
can consist of an inert substance as Gruber suggests.[4]
From these considerations, then it appears that all the evidence
available, both direct and indirect, points to the conclusion that the
behavior of the surface layer on the ameba resembles in general and in
detail the behavior of a surface tension layer in an inert drop of
fluid, and that we must regard the surface layer on the ameba as a true
surface tension layer. This layer is therefore a dynamic layer,
containing free energy, and capable of performing work. It is
physiologically distinct from ectoplasm, as ectoplasm is distinct
physiologically from endoplasm. But the distinctive properties which the
surface layer possesses are functions of its position. These properties
clearly indicate that its constitution is protoplasmic, corresponding to
the fluid parts of the internal protoplasm.
The surface layer of the ameba is probably identical with what is
commonly called the _plasma membrane_ or semi-permeable membrane as
postulated by Overton (’07). The peculiar structure supposed to be
possessed by plasma membranes are held to be due chiefly to surface
forces. The fact that the surface layer of the ameba is continually
being destroyed and re-created during locomotion does not support the
view that the plasma membrane is of inert composition, as for example,
lipoidal, as has been suggested. The observations, on the contrary,
confirm Höber’s (’11) view that the plasma membranes generally are
living structures. But it may be regarded as certain that if lipoids are
present in the protoplasm of the ameba, these substances, according to
the principle of Willard Gibbs, will be found in higher concentration
in the surface film than in the body of the ameba.
Perhaps the most important question that arises in connection with the
surface layer of the ameba is: What causes it to move in the manner
described? But we can do little more than ask the question. It has been
seen that the surface film moves toward an area of increased tension
rather than from an area where the tension has been lowered. However,
since we are completely in the dark respecting the composition of the
surface layer or of the fluid parts of the ameba, it is exceedingly
hazardous to venture an explanation. If the surface layer should have
its tension lowered by a concentration of lipoids in it, we would be
faced by the necessity of explaining their removal at the anterior end.
If we turn to electrical causes we meet again with great difficulties.
An ameba moves with the electric current, when a current is passed
through the water. The surface layer under these conditions behaves
normally, as may be inferred from Jennings’ (’04) figure on page 198.
That is, the current controls the direction of the movement of the
ameba, with the current leaving the ameba at the point of highest
surface tension. This is contrary to the action of the mercuric
capillary electrometer, in which the mercury column also moves with the
current, but because of lowered surface tension where the current leaves
the mercury. The conditions surrounding these cases are so different
however, that very little can be gained by setting them in contrast to
each other.