J. Cell Set. 50, 245-258 (1981)
Printed in Great Britain © Company of Biologists Limited ig8i
245
FUNCTIONAL INTERDEPENDENCE OF
PSEUDOPODIA IN AMOEBA PROTEUS
STIMULATED BY LIGHT-SHADE DIFFERENCE
ANDRZEJ GREBECKI AND WANDA KLOPOCKA
Department of Cell Biology, Nencki Institute of Experimental Biology, ul. Pasteura
3, 02 093 Warszawa, Poland
SUMMARY
Polytactic cells of Amoeba proteus were exposed to localised photic stimulation. When a
pseudopodium is stimulated to advance, by shading ,it, other pseudopodia are retracted.
Activation of the shaded front is the primary response, and contraction of other fronts the
secondary one. When a pseudopodium is inhibited by illuminating its frontal segment, or
when it is allowed to enter the bright zone in the course of migration, it slows down and stops
but its eventual retraction depends on the existence of other possible directions for the endoplasmic flow. Therefore, if other active pseudopodia are lacking, the front suppressed by light
cannot retreat effectively until new fronts arise in other body regions kept in shade. In all
experimental situations the development of new fronts or the activation of forward flow in
lateral pseudopodia precedes the contraction of the former leading pseudopodium. Also the
reversal of direction of the endoplasmic streaming begins at the new front, and then it gradually
extends until it reaches the former front.
The results confirm the interdependence of different pseudopodia in the same individual
and they contradict the concept that pseudopodia behave as separate functional units. On the
other hand, they indicate that formation of new pseudopodia should not be explained as a
simple secondary effect of contraction of the older ones but, on the contrary, as a phenomenon
that initiates the changes in the pattern offlowin amoeba. The general interpretation is based
on this variant of the pressure-flow theory of amoeboid movement, which attributes the motive
power to the contractile activity of the whole cell cortex and the steering role to events taking
place in the front of the migrating cell.
INTRODUCTION
Analysis of time relationships between the phases of activity of different pseudopodia in freely moving polytactic amoebae demonstrated that they are correlated
(Ktapocka & Gre_becki, 1980). It is not in accord with the assertion that each pseudopodium ' ...behaves as an independent functional unit' (Allen & Allen, 1978), but
it confirms the classical view that 'amoeba acts as an organized u n i t . . . ' (Mast, 1932).
Our earlier experiments allowed us to conclude, in particular, that the moments of
initiation of new pseudopodia and the moments of inhibition or reversal of movement
of older fronts are distinctly correlated in time. However, the methods of analysing
the spontaneous motion of amoeba applied in the preceding study did not permit us
to answer the important question as to which of these 2 events takes place first. Is
the formation of a new pseudopodium the necessary condition preceding the retraction of the older one or, on the contrary, is the reversal of functions in the formerly
advancing pseudopodium the primary factor that initiates a new direction of flow?
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It is possible to promote the development of pseudopodia or to inhibit their
extension by using external factors applied locally to them. Such experiments may
provide an answer to the question raised above, when they are run with a simultaneous
recording of the effects produced in other, non-stimulated frontal areas. Stimulation
by light promises to be particularly convenient in such a study, because it is easy to
control its intensity, localization, and time of action. Localized photic stimuli have
already been applied to amoeba in the form of narrow vertical light beams by Mast
(1910, 1932) and Schaeffer (1917), and recently by one of us (Gre_becki, 1980) in the
form of bright and dark areas projected upon the plane of migration of amoebae,
within the field of view of the microscope. This method permits us to induce a new
pseudopodium or re-activate an old one by local application of shade, and to inhibit
an advancing pseudopodium by local exposure to light.
MATERIAL AND METHODS
Amoebae were grown in Pringsheim medium (Chapman-Andresen, 1958) and fed on Tetrahymenapyriformis. They were used for experiments 2-4 days after feeding, when they manifest the
highest locomotive activity. Polytactic individuals (terminology of Gre,becki & Gre.becka, 1978)
were transferred to a slide, together, with some drops of the original culture medium. Several
glass beads, 0-5 mm in diameter, were added to keep the coverslip at the necessary distance
from the slide. The slide was then placed, on the stage of the microscope ready for the experiment and left for 10-15 IT^n before the beginning of the observations; this allowed the amoebae
to adapt to the new conditions of luminosity and to recover the polytactic mode of locomotion
(they became heterotactic during the earlier manipulations). The microscope was equipped
with the differential interference contrast device of the Pluta system (PZO Warsaw). The
experiments were run in a semi-dark room at 18 ± 2 deg. C.
The light intensity at the plane of cell migration (measured with the photosensitive element
before mounting the slide with amoebae on it on the stage) ranged from 6000 to 8000 lux. It
was white light produced by a standard incandescent lamp with a heat filter. Small rectangles
cut out from neutral gelatine filters, sealed between 2 glass slides in order to facilitate manipulation, were placed on the top of the lighting apparatus of the microscope, and their image was
reduced by a factor of 10 and focused upon the plane of cell migration using the substage
condenser. The filters used were characterized by 75 % light absorption. The luminosity was
therefore reduced to 1500-2000 lux in the screened parts of the field of view. These 2 levels
of illumination (6000-8000 lux and 1500-2000 lux, respectively) will be referred to in the text
simply as light and shade. In one group of experiments amoebae were allowed to move freely
in light at the beginning, for 10-30 s, and then a single one of their pseudopodia was locally
shaded (by changing the position of the filter in respect to the cell). In other experiments the
same procedure served to expose locally to the light one advancing pseudopodium of a specimen
that was initially migrating entirely in the shade. This method is based on the original idea of
Mast (1910) to focus the image of the source of light with the substage condenser, and in the
present form it was developed by Gre.becki (1980) to study the behaviour of amoebae stimulated by a well-defined light-shade difference established across their bodies.
The reactions of the amoebae were filmed with a frequency of 4 frames/s with a 16 mm
camera and time-lapse equipment from Bolex. The films were analysed frame-by-frame with
the LW International Photo-optical Data Analyzer. The successive stages of motion were
redrawn at intervals corresponding to 2-5 or 5 s of real time, and used to plot the activity
curves of all the stimulated and unstimulated pseudopodia produced by an individual in the
course of the experiment.
Each experiment described in this article has been repeated at least 50 times, and recorded
cinematographically at least 30 times. In a few cases (2-4 in each series of experiments) the
amoebae failed to react promptly to the photic stimulus. Such experiments were eliminated.
The selected examples of behaviour of amoebae analysed in this study represent 80-90 % of
Interdependence of pseudopodia in Amoeba
247
the tests, depending on the type of experiment. It appears obvious, however, that examples
of behaviour different from the dominating pattern must happen sporadically, because an
amoeba can ' spontaneously' form a pseudopodium or withdraw it at any moment, which may
distort the picture of its response to the stimulus.
The cinematographic films were also used to produce photokimographically recorded
shadowgraphs of the endoplasmic streaming. The method used in producing them was the
same as that used in the study of Kamiya (1950) on protoplasmic flow in the slime mould
Physarum polycephalum, and those of Rinaldi & Jahn (1963) and Kanno (1965) on endoplasmic
flow in amoebae. It consists of projecting the image of the axial part of a pseudopodium (or of
a slime mould channel) through a narrow slit on a continuously running strip of photo-sensitive
material. The slit must be oriented in parallel to the direction of flow, and at right angles to
the shift of the film strip. The only innovation in the present application of this method consists of using cinematographic pictures, instead of recording in vivo the image of endoplaamic
streaming projected directly from the microscope. This makes it possible to produce several
synchronized shadowgraphs from different body regions of the same individual, by changing
the position of the slit during successive projections of the same film. The equipment used in
this laboratory for the photokimographic analysis of cinematographic records was described
earlier (Cieslawska & Gr?becki, 1978; Gre.becki & Moczon, 1978).
Photographs of the successive stages of motion of amoebae exposed to local stimulation
by light or by shade (Figs. 1-6) were taken under the same experimental and optical conditions
as described above for filming.
RESULTS
Activation of a single pseudopodium by shade
Reactions of different pseudopodia of amoebae observed after shading the tip of
one of them are presented in Fig. 1. The serial pictures show the successive phases of
response manifested by the shaded pseudopodium, as well as the behaviour of
pseudopodia kept out of the area of stimulation. The gradually changing pattern of
extension and retraction of different fronts of amoeba (as indicated by arrows)
strongly suggests that the withdrawal of pseudopodia that remain in light is delayed:
it begins when the extension of the shaded pseudopodium is already well advanced.
A more detailed study of this phenomenon was undertaken by means of frame-byframe analysis of cinematographic records. Series of contour drawings representing
the successive stages of motion at intervals of 5 s were used to construct graphs of the
activity curves of all the pseudopodia analysed. Two such graphs are given as examples in Fig. 7.
Three successive phases may be distinguished in the course of each experiment.
The first one represents the period of free locomotion of an unaffected amoeba before
stimulation. The second one corresponds to the lapse of time (5-15 s) between the
application of shade and the beginning of the visible reaction of the shaded pseudopodium. In the third phase the pseudopodium exposed to the lower level of luminosity reacts vigorously and the effects become manifest in others.
The graph in Fig. 7 A illustrates the sequence of events (similar to that recorded
photographically in Fig. 1) when one of the older pseudopodia, which has already
begun to contract, is exposed to shade. This results in the re-activation of its progressive movement, which returns with some delay after the onset of stimulation
(15 s in the case described). Then the rapid progression of the shaded front is followed
A. Grgbecki and W. Klopocka
Interdependence of pseudopodia in Amoeba
249
by inhibition and withdrawal of other pseudopodia, which were formerly extended.
The stimulated pseudopodium initially behaves as a leading one, but soon it becomes
the only advancing front of the moving cell.
The graph in Fig. 7B shows an example of a different experimental situation, where
the shade is applied to the tip of one of three well-progressing frontal pseudopodia.
The stimulated pseudopodium competes favourably, and it accelerates soon after
shading and takes over the leading role in cell locomotion. The increase in its activity
exerts a clear influence on the other fronts. Between 5 and 10 s after acceleration of
the stimulated front both the other pseudopodia begin to retreat.
In general, the results of frame-by-frame analysis confirm the suggestion based on
visual estimations that the re-activation of forward movement and the intensive
growth of a shaded contracting pseudopodium (Fig. IA, B) are the first responses to
stimulation of amoebae, whereas the retraction of advancing pseudopodia that remain
in the bright part of the field comes as a later consequence. The reaction of unstimulated pseudopodia is a secondary response to the original reaction of the stimulated
pseudopodium, and not the primary response to the stimulus.
Inhibition of a single pseudopodium by light
Parallel experiments were undertaken to reveal the time relations between the local
response of an advancing pseudopodium to the increased illumination (its inhibition
and retreat) and the distant effects manifested in other body regions kept outside the
field of stimulation, i.e. in shade. The main objective was to check whether the retraction of the illuminated pseudopodium is the first reaction of amoebae, and to
learn at which moment the new advancing fronts are formed (or older ones
activated) in unstimulated areas.
Two different experimental situations were studied. In the first the anterior segment of a leading pseudopodium of an amoeba migrating in shade was suddenly
exposed to light by an appropriate movement of the shading filter (Figs. 2, 3). In the
second the leading pseudopodium was allowed to reach and eventually to cross the
shade-light borderline in the course of its own free progressive movement (Figs. 4-6).
Fig. 1. Amoeba stimulated locally by shade after a period of free migration in full light.
The shade was applied to a lateral pseudopodium at an early stage of its retraction
(A). Note that the stimulated pseudopodium is activated before other fronts begin to
contract (B). At the later stages the stimulated front advances vigorously and others
retreat (C-E). Figs. 1-6, bars, 150 fim; arrows point in the direction of actual endoplasmic flow; (#) cases of momentary cessation of flow.
Fig. 2. Amoeba with a branched pattern of streaming in the frontal zone, at the initial
stage of migration in shade (A) ; the moment of exposure of its leading pseudopodium
to light (B) ; retraction of the stimulated front and further growth of the unstimulated
ones (C-D).
Fig. 3. Amoeba with a single advancing pseudopodium kept in shade (A) ; the beginning
of stimulation of its frontal segment by light (B) ; the temporary cessation of movement
(c); the formation and further development of 2 lateral pseudopodia in shade, which
enables the withdrawal of the former front (D-E).
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Interdependence of pseudopodia in Amoeba
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In the first as well as in the second variant of the experiment, the amoeba may react
to the increased illumination of its leading front in 2 slightly different ways, depending
on the actual configuration of its frontal body region. An amoeba that had, at the
moment of stimulation, more than one actively advancing pseudopodium (Figs. 2 A, B,
4A, B) may rather easily and promptly withdraw one of them when it is illuminated.
This may be explained by the fact that, in such a case, the flowing mass of endoplasm
can be directed immediately to other actively extending frontal pseudopodia, as is
demonstrated in Figs. 2C, D, 4c, D, E, The cell shown in Fig. 4 presents a particularly
good example of behaviour demonstrating the interdependence of the motion of
different pseudopodia. Each one of its 3 frontal pseudopodia approaches the shadelight borderline and stops; but they approach separately at different moments and
the cessation or the reversal of flow in one of them is always correlated with activation
of forward streaming in another.
The second mode of reacting is observed in cells that had not developed any lateral
active pseudopodia at the moment when their front was exposed to light (Fig. 3 A, B)
or when it met the shade-light borderline as it moved forward (Figs. 5 A, B, 6A). In
this case the stimulated pseudopodium continues for a while to progress across the
illuminated territory, but soon it slows down and eventually stops (Figs. 3 c, 5 c and
6 B). The decrease of frontal velocity after exposing the anterior regions of amoebae
to a higher light intensity was studied quantitatively by one of us and is
described elsewhere (Gre_becki, 1981). At the next stage, one or two new
pseudopodia are formed at the basal region or along the middle part of the old one
(Figs. 3D, 5c, 6c). This means that they always arise with some delay after the partial
or complete suppression of activity of the former front, but before its visible contraction and retreat. The new pseudopodia develop, as a rule, on the shaded part of
the old pseudopodium (otherwise they are subsequently retracted). Only after
initiating such new directions for the endoplasmic flow can the illuminated former
front be effectively withdrawn, as is seen in Figs. 3D, E, 5D, E and 6D.
Activity curves of pseudopodia produced by frame-by-frame analysis demonstrate
the same picture. Two examples of these (shown in Fig. 8) show this type of response
where amoeba reacts by the formation of new pseudopodia. Again, such an experiment
Fig. 4. Amoeba with 2 equipotential fronts migrating in shade (A); the approach of
1 front to the bright part of the field and the division of the other in two (B); the
retreat of the first front away from light and the approach of the second one to it (c);
the stopping of the second front after crossing the shade-light borderline and further
extension of the third one (D) ; the cessation of flow in the second and third fronts at the
limits of illuminated zone and re-activation of forward streaming in the first one (E).
Fig. 5. An amoeba approaching the bright field with its unique advancing pseudopodium (A); crossing the shade-light borderline (B); cessation of further progress
across the light zone and formation of a new front in shade (c); further development
and extension of the new front and retreat of the old one (D-E).
Fig. 6. Higher magnification of a leading pseudopodium penetrating onto the illuminated territory (A) ; cessation of its further progression (B) ; formation of a lateral front in
shade (c); its development and retraction of the former one from light (D).
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A. Gr§becki and W. Kiopocka
252
400 -
200
100
20
30
Time (s)
40
50
60
20
10
Time (s)
Fig. 7. Graphs composed of the extension curves of all pseudopodia active before
stimulation, and developed later, in amoebae exposed to the localized action of shade.
The beginning of stimulation is indicated by the long vertical arrow. The activity
curve of the shaded pseudopodium is marked by a heavy line for the stimulation
period. Moments of retraction of other pseudopodia are indicated by smaller arrows
falling down obliquely. Note that their retreat follows the reactivation of the front
stimulated by shade (A) or the acceleration of its progressive movement (B).
may be divided into 3 distinct phases, similar to those distinguished in the case of
stimulation by shade (cf. Fig. 7). The first phase represents the spontaneous movement of non-stimulated amoeba; during the second the illuminated pseudopodium
slows down and stops but never retracts, and in the meantime new pseudopodia
are formed; at the third stage, when new pseudopodia have already developed, the
old front, exposed to light, is retracted.
It should be added that in many cases the response of amoeba may represent a
mixture of both the types of behaviour described above, i.e. the withdrawal of an
illuminated leading pseudopodium may be partly facilitated by the presence of
another active front, and partly dependent on initiation of new pseudopodia.
All the experiments of this group, independently of the manner of application of
the light stimulus, demonstrate that local inhibition of an illuminated front cannot
Interdependence of pseudopodia in Amoeba
253
200 -
200 E
a.
E
a.
0
10
20
40
0
20
30
50
30
Time (s)
Time (s)
Fig. 8. Diagrams, similar to those shown in Fig. 7, constructed for amoebae locally
stimulated by light. The activity curve of the illuminated pseudopodium is marked
by a double line for the stimulation period. Moments of formation of new pseudopodia
or of acceleration of the pre-existing ones are indicated by arrows pointing up obliquely.
Other symbols are used as in Fig. 7. Note that, as well as after exposing the leading
pseudopodium to light (A), and, after allowing it to enter the bright zone during its
own migration (B), the formation of new pseudopodia (or acceleration of older ones)
precedes the definitive retraction of the illuminated front.
lead to the final retraction of the stimulated pseudopodium until new advancing
pseudopodia are formed in the unstimulated regions of the cell.
Observations of flow phenomena
Microscopic observations of moving amoeba in vivo, as well as cinematographic
records analysed either frame-by-frame or by producing shadowgraphs (Fig. 9),
allowed us to supplement the data concerning the extension or retraction of whole
pseudopodia by adding some information about the mode of changing the direction
of endoplasmic flow inside them. The changes of streaming were followed also in
freely migrating non-stimulated cells, as in amoebae locally stimulated by light or by
shade.
The reversal of endoplasmic streaming in a pseudopodium is most commonly
preceded by a brief cessation of flow. It may often happen as a consequence of illumination of the pseudopodial tip. It seems that in such a situation the streaming stops
almost simultaneously along the whole length of the pseudopodium (or in the whole
amoeba, in the case of orthotactic individuals). The wave of 'gelation' proceeding
from the stimulated tip backwards, which has been described by Mast (1932), was
never observed in the present experiments. After the brief period of rest the streaming
starts in the opposite direction. Usually the reversed streaming is observed first at
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A. Gr§becki and W. Klopocka
the base of the pseudopodium. Then, the zone of reversal is gradually extended forwards in a wave-like form, from the proximal segment of the pseudopodium to its tip.
In many cases it was evident that, in fact, the wave of reversal started from a new
front formed elsewhere during the period of the inhibition of motion of the older
leading pseudopodium (as described in the preceding section).
A slightly different situation, characterized by the absence of a perceivable phase
of complete cessation of flow, was also quite frequently observed. In such a case, in
the middle and frontal segments of the pseudopodium the endoplasm continues to
flow toward the tip (which may still advance), while the streaming has already
reversed in its proximal segment. The wave of reversal progresses to the distal end,
and it usually takes a few seconds before the uniform new streaming direction is
30
60
Time (s)
90
Fig. 9. Shadowgraph of the endoplasmic streaming in an advancing pseudopodium
that was initially kept in shade, illuminated between the 25th and 60th second of the
experiment, and then shaded again. The ascendant tendency of streaks produced by the
moving endoplasmic granules corresponds to the forward streaming, and their inclination downward corresponds to the reversed direction of flow. Note that the flow
reversal during illumination, and its later re-reversal after shading, begin from the
basal region of the pseudopodium (bottom of the shadowgraph), and then extend
to the pseudopodial tip (the upper edge of the record).
established along the whole length of the pseudopodium. The observed sequence of
events is in this case fully consistent with the description given by Allen (1973), based
on the unpublished study of Breuer.
One of the shadowgraphs confirming the gradual character of streaming reversal
is shown in Fig. 9. The recorded pseudopodium initially advanced in shade, then the
position of the filter was changed and it was exposed to light, which provoked the
reversal of flow and withdrawal of the tip, and finally it was shaded again, which
resulted in the recovery of forward movement. The pattern of the streaks produced
by moving endoplasmic granules indicates that in both critical periods (reversal and
re-reversal of streaming) the direction of flow changed earlier at the basal part of the
pseudopodium than at its more distal regions.
When the anterior segment of a contracting or inactive pseudopodium is exposed
to shade the activation of forward streaming begins in the shaded zone, i.e. it starts
from the pseudopodial tip.
Interdependence of pseudopodia in Amoeba
255
DISCUSSION
The experiments and observations presented in this study allow us to re-evaluate
the reliability of 2 statements made by Allen (1961a, b, 1968, 1973) in support of his
theory of frontal zone contraction of amoeboid movement. According to the first
statement, the motor phenomena in different pseudopodia of a polytactic amoeba
are unrelated. According to the second, the reversal of endoplasmic streaming always
begins at the tip of a new pseudopodium, and then it is propagated as a wave toward
the former leading front. Both these statements were considered by Rinaldi & Jahn
(1963) as contradicting the tail-contraction theory, but qualified as 'incompatible
with the evidence' and 'erroneously assumed, to support the explanation' given for
amoeboid movement by Allen.
The present experiments demonstrate that, in fact, the motor phenomena observed
in different pseudopodia in amoebae locally stimulated by light or shade are mutually
related, as was shown previously (Klopocka & Gre_becki, 1980) in the case of spontaneous locomotion of unaffected cells. The reinforcement of the activity of an
advancing pseudopodium or the re-activation of a contracting one, which are induced
by shading pseudopodial tips, invariably result in the inhibition and retraction of
other formerly active pseudopodia (Figs. 1, 7). The beginning of contraction of
pseudopodia exposed to local illumination is correlated in time with the formation of
new pseudopodia (Figs. 3, 5, 6, 8), or at least it appears to be dependent on the
activity of other advancing fronts that were present before (Figs. 2, 4).
Therefore, the assumption that pseudopodia are independent (because each one is
pulled forwards separately by its own frontal zone) seems to be untenable. The effects
of localized photic stimulation confirm, on the contrary, our earlier conclusion
(Klopocka & Gre_becki 1980) that the motor phenomena in different pseudopodia are
interdependent, as might be expected according to the classical concept that endoplasmic streaming in amoebae follows intracellular pressure gradients.
However, the time relations between the beginning of retraction of older pseudopodia and the initiation of new ones are hardly compatible with the most common
version of the pressure-flow theory, which limits the contraction site to the tail of
the amoeba and to the distal parts of its contracting pseudopodia (e.g. Goldacre &
Lorch, 1950; Wehland, Weber, Gawlitta & Stockem, 1979). Rather, it should be
expected, from this last point of view, that the contraction of an old pseudopodium
would seem to be the original cause inducing the formation of a new one. The present
experiments demonstrate that in reality the formation of a new pseudopodium begins
earlier, and it is followed (but not preceded) by the effective retraction of the older
front. When a pseudopodium is stimulated by shade (Figs. 1, 7) it begins to extend
forward vigorously before other pseudopodia react. The activation of such pseudopodia appears not as the consequence but as the necessary condition of withdrawal
of other cell parts. When an advancing front is inhibited by light it may slow down
or stop, but it effectively retracts only after the development of new pseudopodia at
its bases (Figs. 3, 5, 6, 8).
These results are in good agreement with the finding of Seravin (1966), who demon-
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A. Gr§becki and W. Klopocka
strated that in amoebae immobilized by different physical and chemical shocks the
resumption of endoplasmic streaming is initiated by the formation of new pseudopodial tips. They are also consistent with the observation reported by Kalisz-Nowak
(1978), that during the formation of a new pseudopodium the local gap in the ectoplasmic cylinder appears first, before and not after the change in the streaming
pattern in amoeba. This sequence of events is also confirmed by the studies in which
the withdrawal of older fronts was observed as a further consequence of the experimental initiation of a new direction of flow: by sucking the endoplasm with a micropipette (Gre.becka, 1980), by perforating the peripheral cell layers (Gr^becka, 1981),
by local rupture of the contractile cell cortex by an injected droplet of paraffin oil
(Goldacre, 1961; Gre_becka, 1977), by its local disorganization by an injection of
DNase I (Wehland et al. 1979), and by local application of anaesthetics (Korohoda,
1977)The fact that old pseudopodia begin to retract after the new ones are formed at
their bases makes credible the second assumption of Allen, that the wave of reversal
of streaming travels along the pseudopodium from its basal region to the distal end.
As a matter of fact, the present observations and records generally support this
particular statement of Allen (1961 a, 1973) based on the unpublished data of Breuer,
rather than the objection of Rinaldi & Jahn (1963). When a new pseudopodium arises,
or when an old one is re-activated by shade, the new direction of flow appears first
at its tip. When the former leading pseudopodium is subsequently retracted, the flow
usually (but not always) stops simultaneously along its axis, and then the direction of
streaming becomes gradually reversed from the base of the pseudopodium towards
its tip.
The wave-like propagation of streaming reversal from the new front towards the
old one is, in fact, incompatible with the restrictive tail-contraction theories, but it
seems to be admissible within the concept of generalized peripheral contraction
(Gre_becki, 1979). According to this theory the whole microfilamentous cortex of
amoeba is contractile and creates increased hydrostatic pressure inside the cell. But
the cortex is discontinuous and dissociated from the outer cell membrane at the tips
of advancing pseudopodia (Gre_becka, 19780,6; Gre,becka & Hrebenda, 1979;
Wehland et al. 1979), which lowers the pressure at the cell front and creates hydrostatic gradients that promote the endoplasmic streaming. When a new gap opens in
the cortical envelope, as it probably does in the case of stimulation by shade, the
outflow of endoplasm in the new direction will locally reduce the intracellular pressure, which may result in the inversion of pressure differences in the vicinity of the
new front, i.e. at the base of the old pseudopodium. It seems feasible that the zone
of the reversed pressure gradient may extend gradually from the new front towards
the old advancing tip and gradually reverse the direction of flow, as was recorded by
Breuer (according to Allen, 1973) and frequently observed by us. The weakening of
the endoplasmic inflow into the former front creates conditions for the rebuilding of
the contractile cortex around its tip (as shown by Grqbecka, 19786), which accomplishes the transformation of the advancing pseudopodium into a contracting one.
In the second situation, when a unique advancing pseudopodium is strongly in-
Interdependence of pseudopodia in Amoeba
257
hibited (as in the case of its local illumination) the rebuilding of cortex around its tip
probably results in raising the intracellular pressure at the front, and consequently
leads to the disappearance of the pressure gradient and to the temporary cessation of
streaming. Therefore, the formation of a new front becomes a necessary condition for
restoring pressure differences inside the cell and for the retraction of the inhibited
pseudopodium.
It may be stated in general that the relationships between the formation of new
pseudopodia and the retraction of old ones provide no arguments in favour of the
concept that the motive force of endoplasmic streaming is localized in the frontal
zone of amoeba. They may be explained adequately by the pressure-flow concept of
amoeboid movement, provided that the theory takes account of the motor functions
of the whole cell cortex (instead of localizing the motive force in the tail region alone)
and that it recognizes the steering role of the frontal zone. The mechanism for opening
new gaps in the contractile cortical cell envelope is not yet well understood, but it
seems evident that their formation and obturation play an essential role in the frontal
control of movement.
This study has been supported by Research Project PAN II.i of the Polish Academy of
Science.
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(Received 10 October 1980 - Revised 19 January 1981)