Download Neural Control of Interappendage Phase During Locomotion

Neural Control of Interappendage Phase During Locomotion
PAUL S. G. STEIN
Department of Biology, Washington University, St. Louis, Missouri 63130
SYNOPSIS. Interappendage phasing of crayfish swimmeret mo\ements depends upon a
central nervous system network of command, oscillator, and coordinating neurons. The
command neurons serve to set the general excitation level in each of the segmental
oscillators. The oseil'ator neurons in each hcnii-ganglion generate the rhythmic alternations of powerstroke and leturnstroke motor neuron activity. The coordinating neurons
transmit the precise timing information about the state of one oscillator to other
oscillators. This information can serve to advance or to delay the motor bursts driven
by the other oscillators. Which effect is observed depends upon the arrival time of the
coordinating neuron discharge within the cycle period of the modulated oscillator. This
type of modulation leads to the prediction that a stable interappendage phase can result
from situations where there is not a fixed excitability gradient among the segmental
oscillators. This prediction has been verified using a cut command neuron preparation.
Coordinated locomotion in many multiappendage animals (e.g., crayfish, cockroach,
chicken, cat) is characterized by the movement of a given limb having (i) the same
frequency as another limb and (ii) a regulated phase relationship with another limb
(Gray, 1968). Motor neuron recordings
from deafferented nervous systems have
shown that both frequency equalization and
phase regulation are properties of central
networks and need not depend on sensory
feedback (Evarts et al., 1971). The network
controlling swimmerets of the crayfish Procambarus clarkii (Girard) is one such system (Ikeda and Wiersma, 1964). Much
information has been gathered about the
functional properties of individual neurons
in this network. It is known that there exists
a set of command neurons, any one of which
when stimulated above a critical frequency
will drive coordinated swimmeret beating
(Hughes and Wiersma, 1960; Wiersma and
Ikeda, 1964; Davis and Kennedy, 1972). In
addition, the existence of oscillator neurons
within each segmental ganglion has been
inferred from the demonstration that an
This work has been supported from 1972 to the
present date by N.S.F. grant GB-35534 to the author
at Washington University. Dr. Theodore H. Bullock
of the Department of Neurosciences and Scripps
Xeurobiology Unit at the University of California at
San Diego provided support from 1969-1971. Dr.
Donald Kennedy of the Department of Biology at
Stanford University provided support from 19661969.
isolated segmental ganglion can spontaneously produce rhythmic bursts of motor
neuron discharge (Ikeda and Wiersma,
1964). Stimulation of one command neuron
can excite all the swimmeret segmental oscillators; however, this excitation does not
carry precise interlimb timing information.
This latter class of information is necessary
for interlimb phase control and is carried
by another class of intersegmental interneurons, the coordinating neurons (Stein, 1971).
These central cells have a discharge which
coincides with the motor activity of one set
of synergists in a limb. This discharge persists in the absence of sensory input. These
cells have input properties similar to corollary discharge or efference copy neurons
(Evarts et al., 1971). When the axons of
these cells are cut, interlimb phase control
is destroyed (Stein, 1971).
These results demonstrate that specific
neuronal elements are necessary for interlimb phase control. They neither explain
the mechanisms underlying this control, nor
do they make predictions about the magnitude of interlimb phase given knowledge of
other parameters of the network. More recent data gathered in the swimmeret system
(Stein, 1972, 1973, and unpublished) have
given insight into the possible mechanisms
underlying phase control and have made
possible qualitative predictions. These new
data are measurements of the phase shift
observed in the discharge of a swimmeret
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PAUL S. G. STEIN
FIG. 1. Normal intersegmental sequence of powerstroke (PS) motor neuron discharge to swimmeret
appendages of the crayfish abdomen. Simultaneous
recordings from homologous peripheral roots of the
right fifth (ps5, upper trace), right fourth (ps4,
middle trace), and right third ganglia (ps3, lower
trace) are displayed. Stimuli at 30 Hz were applied
via an electrode placed on the lateral region of the
right connective between the first and second ganglion. Suction electrodes were utilized for recording
and stimulation. Time mark, 500 ms. (From Stein,
1971.)
oscillator in response to single bursts of coordinating neuron activity (Stein, 1972, and
unpublished). These data can be utilized to
predict that when limb control centers are
producing rhythmic output, then stable interlimb phase relationships can exist in situations where an anteriad oscillator has
either a slightly higher, or a slightly lower,
or the same intrinsic frequency as a posteriad oscillator. These predictions have been
verified (Stein, 1973). They constitute a
direct proof of the statement that the command neurons need to set the intrinsic frequency of each oscillator only within a
rough range of each other and do not need
to establish a precise gradient of oscillator
intrinsic excitabilities along the neuraxis.
The present paper will review the experimental work with the crayfish, Procambarus
clarkii (Girard) and will discuss the similarity of this work to results obtained in other
locomotory systems. There are strong logical equivalences observed among the nerve
networks controlling locomotion. The existence of these relationships makes it reasonable to utilize experimental results from one
system as hypotheses in other systems.
swimmerets, I have selected the powerstroke
activity as an indicator of the state of the
limb activity (Fig. 1). The repetition rate of
powerstroke motor neuron bursts is equal
to the frequency of limb movement (Davis,
1968a). Recordings from homologous
powerstroke motor neurons in adjacent
segments show that the burst frequency
of motor neuron discharge is equal in all
segments. In addition, there is a phase lag
in the activation of powerstroke motor neurons in anteriad segments when compared
with powerstroke activation in a posteriad
segment (Hughes and Wiersma, 1960; Stein,
1971; Davis and Kennedy, 1972). The lags
in the motor neuron discharges correlate
well with the lags in limb movements observed using high-speed motion pictures
(Davis, 19686).
Limbs of greater complexity also utilize
the principle of alternate contraction of
agonists and antagonists but with some
modifications (Engberg and Lundberg,
1969; Szekely et al., 1969). Some muscles
may be active twice during a movement
cycle. Cocontractions of agonists and antagonists are sometimes observed. Muscles that
act around one joint within a limb may be
active with phase lag when compared to
similar muscles acting around a second
joint in the same limb.
Experiments which record normal limb
motor neuron activity during locomotion
are critical. These provide the results which
further investigations must explain. I will
discuss one class of investigations which relate to the problem of interlimb phase control. These experiments establish that each
limb is controlled by an anatomically discrete center.
NORMAL MOTOR DISCHARGE DURING
LOCOMOTION
Movements of the swimmeret of the crayfish or lobster during rhythmic beating
consist of contractions of the main powerstroke muscles which alternate with contractions of the main returnstroke muscles
(Hughes and Wiersma, 1960; Davis, 1968a).
Other movements of the limb occur and are
coactive with either powerstroke or returnstroke activity (Davis, 1968a). In my examination of interlimb coordination of crayfish
NEURAL CONTROL OF INTERAPPENDAGE PHASE DURING LOCOMOTION
IDENTIFICATION OF LIMB CONTROL CENTERS
It is desirable to determine the minimum
piece of nervous tissue which is capable of
producing the normal rhythmic output to a
limb. The technique which has been utilized in the crayfish swimmeret system is the
isolation of the presumptive control center
from the remainder of the CNS. The abdominal nerve cord of a crayfish consists of
a series of segmental ganglia which are anatomically discrete. Each ganglion is connected to the remainder of the nervous
system via two pairs of interganglionic connectives. One pair of connectives exits the
ganglion at the anteriad edge of the ganglion; the other pair exits at the posteriad
edge. In addition, the most anteriad set of
roots leaving the ganglion, termed the first
roots, innervate the swimmerets of that segment (Hughes and Wiersma, 1960). The
output of a ganglion, as observed by recording the powerstroke motor discharge in the
first root, can display a normal rhythm even
in a preparation which is isolated from the
rest of the CNS as the result of cutting of
the connectives at both ends of the ganglion
(Ikeda and Wiersma, 1964). This demonstrates that the rhythmic control center for
the pair of limbs innervated by that ganglion resides within the segment. There are
situations where the left swimmeret can
beat at a different frequency than the right
swimmeret in the same segment (Hughes
and Wiersma, 1960). This suggests that each
limb is controlled by its own center.
The isolation experiments (Ikeda and
Wiersma, 1964) were performed not only for
the fifth segmental ganglion, which innervates the most posteriad set of swimmerets,
but also for the fourth ganglion and for the
third ganglion. These data establish that
the neuronal network driving the swimmerets is distributed among a set of local
control centers. Wiersma and Ikeda (1964)
presented the hypothesis that the interlimb
coupling observed in the intact animal was
due to phase information which travelled
among the segmental control centers.
In the isolation experiments of Ikeda and
Wiersma (1964) all the peripheral roots
leaving the ganglion were cut, thus estab-
1005
lishing that the CNS control center resident
in the ganglion can produce its normal
motor output in the absence of sensory input, i.e., the rhythm is central (Evarts et al.,
1971). The concept of a distributed network
is independent of the concept of a central
network. Any given network can display
one property without showing the other
property. The fact that the swimmem system is both central and distributed is extremely useful and will be be discussed later
in this paper.
It is not possible in some preparations to
isolate totally a portion of the CNS from the
rest of the nervous system and still observe
the normal motor output. In certain cases
it has been possible to transect the nerve
cord and observe that there is normal
rhythmic movement both above and below
the transection (Wilson, 1961; Kristan, personal communication). There is no phase
relationship observed between movements
above the cut and below the cut, however.
These experiments establish that there are
at least two separate control centers which,
in the intact animal, must be coordinated.
There are still other situations where it is
impossible to intervene surgically between
pieces of nervous tissue which are suspected
to contain different control centers for each
limb. In these preparations, the animal may
spontaneously move appendages at different
frequencies (von Hoist, 1936; Hughes and
Wiersma, 1960). The simplest hypothesis
which can explain the observation that the
appendages can move at different frequencies is that each appendage has its own
rhythmic control center.
Another approach was elegantly utilized
by the Russian workers Kulagin and Shik
(1970) in their study of treadmill walking
in the cat. Their study relies on earlier observations that electrical stimulation of the
midbrain of a decerebrate cat will induce
walking, trotting, or gallopping on a treadmill (Shik et al., 1966). The speed of
locomotion is controlled in part by the
speed of the treadmill. Kulagin and Shik
(1970) utilize one treadmill for the left legs
and another treadmill for the right legs.
Their experiment consists of a trio of
stimulating conditions: (A) both treadmills
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PAUL S. G. STEIN
moving at the same low speed, (B) both
treadmills moving at the same high speed,
and (C) one treadmill moving at the low
speed of condition A and the other treadmill moving at the high speed of condition
B. In conditions A and B the right and left
legs alternate as in normal walking. The results in condition C depend on the relative
speed of the two treadmills. If one treadmill is moving only two or three times as
fast as the other, then in condition C there
is alternation between right and left legs. If,
on the other hand, one treadmill is moving
four to six times as fast as the other in condition C, then the leg walking on the faster
treadmill will walk at double the frequency
as the leg walking on the slower treadmill.
Every second "lift" phase of the fast moving
leg is coupled with each "support" phase of
the slowly moving leg.
These data suggest that each leg has its
own control center and that sensory feedback can influence its rate. These experiments also suggest that interlimb coupling
is the result of information which is transferred from one limb control center to the
other, rather than common phase information received by both centers. Selective
excitation of separate limbs has also been
achieved in the crayfish swimmeret system
by cutting command neuron tracts and
stimulating the cut ends of the command
neurons both above and below the cut with
different electrodes (Stein, 1971, 1973).
Transplantation of nervous tissue is still
another method which has been used to
demonstrate that there is a local pattern
generator resident in a piece of nerve cord
capable of producing the rhythmic movements in a limb during locomotion. Szekely
and Czeh (1971) have utilized the dorsal fin
of urodele larvae as a culture chamber
(Weiss, 1950) for an isolated piece of nerve
cord in association with a deplanted limb.
They have shown that if brachial nerve cord
is utilized to innervate the deplanted limb,
then when the limb becomes innervated, it
will display spontaneous movements characteristic of stepping in a normal limb.
They have also recorded electromyographic
activity from the muscles of the deplanted
limb. These myograms are similar to record-
ings obtained from normal limbs. In contrast, if thoracic nerve cord is deplanted
into the dorsal fin region, then the resultant
movements of the deplanted limb bear no
resemblance to normal limb movements.
A variation of this experiment has been
carried out in chick embryos (Straznicky,
1963; Narayanan and Hamburger, 1971).
These experiments rely on the observation
that chick wings "flap" in phase with each
other, whereas chick legs "walk" with alternate movements of each limb. When embryonic tissue from the brachial cord of a
donor is transplanted into a gap in the lumbosacral cord of a host and the donor CNS
is allowed to innervate the legs of the host,
then the legs of the host will move in phase
with each other. Conversely, when donor
lumbosacral cord is trasplanted into a gap
in the brachial region of a host and the
donor CNS is allowed to innervate the
wings of the host, then the wings move in
alternate fashion. The simplest explanation
of these experiments is that there is a specific set of pattern generators in the brachial
cord which generate in-phase movements of
the wings and another set of pattern generators in the lumbosacral cord which generate
alternate movements of the legs.
In summary, there are many independent
lines of evidence which establish that a set
of neuronal pattern generators distributed
along the CNS are involved in the control
of multiappendage locomotory movements.
The simplest hypothesis which can explain
the results observed in locomotory networks
is that each limb pattern generator is the
basic unit of the multiappendage control
system and that interlimb phase relationships emerge as the result of coupling information which is transferred among the
pattern generators. This latter hypothesis
will be amplified in the remaining sections
of this paper.
MECHANISMS OF INTERLIMB CONTROL
Consequences of the distributed property
If a locomotory network is distributed,
then there must exist a mechanism by which
the activity of one control center is coupled
with the activity of another control center.
NEURAL CONTROL OF INTERAPPENDAGE PHASE DURING LOCOMOTION
This follows directly from the observation
that if the movements of the limbs display
frequency equalization and phase regulation, then the control centers driving the
limbs must be coupled. The possible mechanisms of coupling are either an "internal
CNS" mechanism or a "sensory" mechanism. "Internal CNS" coupling can involve
the output of one control center comprising
part of the input to another control center.
Alternately, it might involve a common
phasic input to each of the control centers.
"Sensory" mechanisms can couple the discharge of different control centers in several
ways. Firstly, the central axons of sensory
neurons from one limb may project to the
motor center controlling another limb.
Secondly, sensory interneurons driven by
primary sensory cells from one limb may in
turn project to the motor center for another limb. Thirdly, the sensory cells of a
second limb can be mechanically linked to
the movement of the first limb. This
mechanical linkage can occur within
the body of the animal or via the substrate on which the animal is moving.
The final step required for this class of
coupling is that sensory cells of the second
limb must affect the motor center of the
second limb. A combination of "internal
CNS" and "sensory" mechanisms may also
be utilized. Sensory neurons from one limb
may alter the rhythm of the "central" control center of that limb. The concept of a
"central" control center relies on the observation that the center can produce rhythmic
output in the absence of sensory input. It is
still possible that the center can be modulated by sensory discharge. If so, then central interneurons driven by that control
center can also be rhythmic in the absence
of sensory input and can be modulated by
sensory discharge. If these central interneurons project to other motor centers, then
interlimb phase coupling is produced by a
hybrid "internal CNS-sensory" mechanism.
Given the possibility that all these mechanisms may in fact be employed in any one
organism, the task of sorting out all the possible mechanisms is a difficult one.
One important simplification of these
analyses can be made, however, if an addi-
1007
tional piece of information about the locomotory nerve network is established. If the
entire locomotory network is central and if
each local control center is also central,
then "internal CNS" mechanisms must be
sufficient to produce coordinated interlimb
movements. This does not rule out the possibility that sensory mechanisms also serve
to couple the limb centers. In fact, the cockroach walking system is both central and
distributed (Pearson and lies, 1970; Pearson,
1972); sensory input, however, is known to
produce major changes in the interleg phase
relationships observed during talking
(Hughes, 1957).
Two types of "internal CNS" coupling
are possible. I am not aware of any evidence
that limb control centers are coupled via
common phasic input from a "master"
center which itself is not related to any one
limb. All the available evidence suggests
that "internal CNS" coupling is accomplished by neurons which I have termed
coordinating neurons (Stein, 1971).
Basic properties of coordinating
neurons
The coordinating neuron is defined as a
cell whose discharge is (i) coactive with the
motor discharge of a set of synergistic muscles in one limb, (ii) persistent in the absence of sensory input from the limbs, and
(iii) able to modulate the activity of a motor
center(s) driving another limb(s) so that
normal interlimb phase is established
(Stein, 1971). Properties (i) and (iii) are a
consequence of the distributed nature of
the crayfish swimmeret system. Property
(ii) is a consequence of the central characteristics of the swimmeret system. Properties
(i) and (ii) are input properties of the coordinating neurons. Property (iii) is an output property of the coordinating neurons.
Thus, coordinating neurons have been specifically defined as central interneurons
which project from one motor center to a
homologous motor center. According to this
definition, they are distinct from corollary
discharge neurons or efference copy neurons
(Evarts et al., 1971). These latter neurons
have been defined as central neurons which
project from a motor center to a sensory
center.
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PAUL S. G. STEIN
Evidence for the existence of coordinating
neurons
It is first necessary to locate in the interganglionic connectives neurons whose activity is phase locked with limb motor discharge. Motor neuron discharge to the
swimmerets can be recorded from the first
root. Command neurons can be stimulated
to produce rhythmic motor discharge
(Wiersma and Ikeda, 1964). Fine bundles of
axons can be dissected free from the connectives and the activity of the bundle recorded. Neurons were found in a given
connective which' were correlated with
powerstroke motor neuron activity in a
more posteriad ganglion. No" cells were
located in any given connective which were
coactive with powerstroke motor activity
in a ganglion anteriad to the connective.
These ascending, powerstroke-related cells
were found in distinct regions of the connective (Stein, 1971). The largest number
of these cells were found in a tract running
near the midline. When this paradigm was
repeated in a deafferented preparation, the
discharge of these powerstroke-related cells
was still present (Fig. 2). These neurons,
therefore, satisfy the first two properties of
coordinating neurons. If these cells are coordinating neurons, their discharge must
modulate the activity of another control
center. It would be most satisfying to establish this latter property directly with an
intracellular recording of a control center
cell in an anteriad ganglion. This has not
been accomplished at this present date.
It is possible, however, to utilize indirect
techniques to establish the modulating ability of these central, powerstroke-related
cells. The tracts containing these neurons
can be cut between two segments. In such a
preparation, phase locking is destroyed between the limbs nearest the cut (Stein,
1971). It is important to note that the majority of command neurons to the swimmerets run in the lateral regions of the
connectives and are not destroyed by a
medial cut. This experiment further substantiates the claim that the command neurons do not carry the precise timing
information necessary to couple the limbs.
FIG. 2. Discharge of central nonsensory fibers related to the activity of swimmeret motor neurons.
Drawing shows the placement of recording suction
electrodes on the posterior portion of the left first
root from the third ganglion (ps3) and on a small
central bundle of neurons isolated from the medial
region of the left connective between the third and
second ganglia (ma). The anterior connection of
the fiber with the second ganglion was severed. All
first roots were cut, eliminating not only sensory
feedback from the swimmerets but also all swimmcret movement. The connectives posterior to the
third ganglion and anterior to the second ganglion
were also cut. Recordings show spontaneous activity
in powerstroke motor neurons (ps3, upper trace)
occurring during overlapping time intervals with
activity in the central fibers which ascend in the
medial region of the connective (ma, lower trace).
Time mark, 1 sec. (From Stein, 1971.)
The converse experiment can also be
utilized to establish the modulating capacity
of these medial powerstroke-related neurons. In this experiment the ipsilateral medial tract is spared while other cells in the
connectives between two ganglia are cut. In
particular all the command neurons in the
lateral regions of the connective are severed. This preparation is termed the cut
command neuron experiment (Fig. 3). In
this preparation if limb control centers both
above and below the cut are active, then
the motor discharge to the limb above the
cut will be phase regulated with motor discharge to the limb below the cut if neurons
in the medial portion of the connective act
as coordinating neurons. In fact, if the excitation level of the control center above the
cut is at roughly the same level as the center
below the cut, then normal interlimb phase
relationships are observed (Stein, 1971,
1973). In addition, if the control centers
have extremely different levels of excitation, then major phase shifts in the anteriad
NEURAL CONTROL OF INTERAPPENDAGE PHASE DURING LOCOMOTION
STIM 3
FIG. 3. Drawing of the "cut command neuron"
preparation of the crayfish nerve cord. The entire
contralateral connective and the lateral portion of
the ipsilateral connective between two segmental
ganglia, the third abdominal ganglion (G3) and
the fourth abdominal ganglion (G4), have been cut.
Recording suction electrodes have been placed to
record the activity of homologous motor neuron
discharge to the two limbs adjacent to the cut. With
the arrangement shown, the activities of motor
neurons controlling the powerstroke muscles of the
ipsilateral swimmerets in the third segment (PS 3)
and in the fourth segment (PS 4) may be recorded.
In addition, a recording electrode (MA 4-3) is
placed on the medial portion of the 4-3 connective
in order to monitor the discharge of the medial
ascending tract of coordinating neurons. Electrodes
utilized to stimulate cut command neurons were
placed on lateral regions of the nerve cord in the
3-2 connective (STIM 3) and in the 5-4 connective
(STIM 4).
limb cycle occur (Stein, 1971, 1972) and are
associated with powerstroke motor neuron
discharge to a posteriad limb. It is likely
that the powerstroke-related cells in the medial portion of the connective are mediating
these phase shifts (Stein, 1971). These indirect lines of evidence suggest strongly that
the ascending powerstroke-related interneurons in the medial portions of the connective are in fact coordinating neurons. Direct
verification awaits the proper intracellular
recordings.
Further information about the coupling
within the swimmeret system can be obtained with these indirect techniques. In
particular, the cut command neuron preparation (Fig. 3) has been most helpful since
it allows the experimenter to have separate
frequency control over individual limb
motor centers. For example, if low frequency swimmeret beating is required anterior to the cut, then a command neuron
anterior to the cut can be stimulated at 20
Hz. If high frequency swimmeret move-
1009
ments are required anterior to the cut, then
the command neuron anterior to the cut
may be stimulated at 60 Hz. On the other
hand, if a single burst of powerstroke motor
activity is required posterior to the cut,
then a short burst of high frequency command stimulation can be applied posterior
to the rnr. The cut command neuron preparation can be utilized to determine the
modulatory effect of a single burst of coordinating neuron discharge on an anteriad
control center.
Synaptic input to a neuron with bursting
discharge
Before I describe the results in the swimmeret system, I would like to discuss the
effect of a single burst of synaptic input on
the discharge of a bursting neuron in other
preparations. Kandel (1967) has examined
the modulatory effect of a burst of monosynaptic IPSP's on the activity of an Aplysia
cell with endogenous bursting discharge. If
the IPSP's arrive during the silent period
of the burster, then onset of the next burst
is delayed, i.e., the interburst interval is
lengthened. If the IPSP's are delivered during the burst, however, then a shorter burst
containing fewer impulses is observed and
the onset of the next burst is advanced. Kandel termed such responses contingent, since
the nature of the response varied according
to the arrival time of the modulating input.
Contingent effects can also be demonstrated utilizing depolarizing or hyperpolarizing currents aplied intracellularly
(Strumwasser, 1967; Kater and Kaneko,
1972). In addition, antidromic stimulation
of the large cells of the lobster cardiac ganglion will cause a contingent effect on the
cardiac rhythm (Mayeri, 1973). It is, therefore, likely that many neurons which generate bursting discharge will react in a
contingent manner when stimulated with a
burst of input at various times of its cycle.
Locomotory motor centers in the crayfish
swimmeret system will respond in a contingent manner to bursts of coordinating
neuron discharge (Stein, 1972, and unpublished) (Fig. 4). The cut command neuron
preparation was utilized to demonstrate this
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PAUL S. G. STEIN
MA 4-3
center associated with each burst of coordinating neuron discharge from the posteriad control center.
If the coordinating neuron burst related
to powerstroke (PS) activity in a posteriad
limb began during returnstroke activity
(PS-off) in an anteriad limb, then the onset
of the subsequent PS burst in the anteriad
limb was advanced (Fig. 4/4). If the coordinating burst began and ended during the
FIG. 4. Contingent response of a swimmeret control
center to a single burst of coordinating neuron
discharge. The cut command neuron design illustrated in Figure 3 is utilized. Simultaneous record15
ings of powcrstroke motor neurons from the fourth
ganglion (PS 4, upper trace), coordinating neurons
from the medial ascending tract of the 4-3 connective (MA 4-3, middle trace), and powerstroke motor
neurons from the third ganglion (PS 3, lower trace)
are displayed. Constant frequency stimulation (12.5
K
Hz) was applied via the STIM 3 electrode in order
to produce repetitive bursts of PS 3. A short burst
Q
(170 msec) of stimulation (45 Hz) was delivered via
the STIM 4 electrode in order to produce a single
burst of PS 4 discharge and a single burst of MA
4-3 discharge. The STIM 4 electrode was activated
E
at different times of the PS 3 cycle.
Q.
A, Activation of the STIM 4 electrode during the
PS 3-off (returnstroke) portion of the cycle was
associated with an advance in the onset of the next
powerstroke burst (PS 3). D, Activation of the
0.5 "—
STIM 4 electrode during the PS 3-on (powerstroke)
portion of the cycle results in a more vigorous
10
05
PS 3 burst, and the onset of the subsequent PS 3
STIMULUS PHASE (L/To)
burst was delayed. A systematic plot of these phase
shifts is presented in Figure 5. Time mark, 1 sec. FIG. 5. Plot of the phase shift of powerstroke motor
neuron discharge in the third abdominal ganglion
contingent effect. Constant frequency stim- (PS 3) correlated with the discharge of medial
coordinating neurons in the 4-3 conneculation was applied via the anteriad stimu- ascending
tive (MA 4-3). The triangles represent points in
lating electrode. This resulted in rhythmic which the MA 4-3 discharge began and ended in
motor neuron discharge to the anteriad the PS 3 burst (PS 3-on). The filled circles represent
swimmeret. A brief burst of stimulation points in which the MA 4-3 discharge began during
epoch of PS 3 (PS 3-off) and ended in
was applied via the posteriad stimulating athequiescent
subsequent PS 3-on. The open circles represent
electrode. This stimulation elicited one points in which MA 4-3 burst began during the
burst of powerstroke motor discharge to PS 3-on and ended during the following PS 3-off.
the posteriad swimmeret. In addition, a The phase shift in PS 3 is measured as the period
burst of ascending coordinating neuron ratio of the perturbed period (T,) to the arithmetic
average of the three prior unperturbed periods
discharge was coactive with the burst of (To). The stimulus phase is measured as the ratio
powerstroke motor discharge to the pos- of latency (L) to To. Latency (L) is defined as the
teriad swimmeret. There was a phase interval to the onset of the MA 4-3 burst as meashift in the timing of the anteriad control sured from the onset of the prior PS 3 burst.
NEURAL CONTROL OF INTERAPPENDAGE PHASE DURING LOCOMOTION
1011
PS burst in an anteriad limb, then the subsequent PS burst in the anteriad limb was
delayed (Fig. AB). The magnitude of the
phase shift in the anteriad control center
was expressed as a ratio in which the duration of the perturbed period (Tj) was divided by the duration of the unperturbed
period (To) (Fig. 5). Coordinating neuron
bursts arriving iate in the cycle of the anteriad control center were associated with
period ratios of less than one. Coordinating
neuron bursts arriving early in the cycle
and terminating during the PS of the
anteriad control center were associated with
ratios of greater than one. Coordinating
neuron bursts arriving early in the cycle
and terminating during the PS-off of the
anteriad control center were associated with
period ratios of less than one.
The simplest hypothesis to explain these
data is that the ascending powerstrokerelated coordinating neuron discharge synaptically excites the PS portion of the
anteriad swimmeret control center. Coordinating neuron input during PS-off causes an
earlier PS. Coordinating neuron input during PS causes a stronger PS which in turn
delays the onset of the subsequent PS.
FIG. 6. Example of effects of regularly applied coordinating neuron bursts to a swimmeret control
center. Black bars represent the powerstroke motor
neuron burst driven by a swimmeret control center.
Arrows represent coordinating neuron discharge
which modulates the timing of the control center.
A, There are no coordinating impulses and a
constant burst frequency is observed. B, There is a
short burst of coordinating input during the second
powerstroke burst. This causes a delay in the onset
of the third and all subsequent powerstroke bursts.
C, There is a second short burst of coordinating
neuron discharge. This occurs during the second
PS-off and causes an advance in the expected arrival
time of the third PS burst. In this example, the
advance caused by the second coordinating burst in
C precisely compensates for the delay caused by
the first coordinating burst in C, i.e. there is no net
phase shift. D, The second burst of coordinating
neuron discharge is long. The first half of this
burst is utilized to advance the onset of the third
PS burst as in C; the second half of this long burst
occurs during the third PS burst and causes a delay
in the onset of the fourth PS burst. E, The first
half of each long burst is utilized to compensate
for the effect caused by the second half of the prior
burst.
Interappendage phase control
These phase shift data form the basic
logic of the interlimb phase control system.
They may be utilized to predict phase behavior of the normal system. These predictions are based on an approach developed
for the study of a coupled constant frequency oscillator system (Moore et al., 1963;
Perkel et al., 1964). When applied to the
swimmeret system, this approach predicts
that repetitive bursts of coordinating neuron discharge can achieve frequency equalization and phase stability with an anteriad
swimmeret control center in situations
where the burst rate of the coordinating
neuron discharge is either greater than,
1012
PAUL S. G. STEIN
equal to, or less than the intrinsic frequency
of the modulated anteriad center. The intrinsic frequency of a control center, is the
frequency of motor output bursts observed
PS 3
when the center is not receiving coordinating neuron discharge from other centers.
I would like to illustrate (Fig. 6) how the
basic logic derived from the single burst experiments (Figs. 4, 5) can be utilized to predict the interlimb phase stability observed
during normal behavior of the crayfish
swimmerets. This illustration utilizes black
bars to represent powerstroke activity in an
anteriad oscillator. The arrows represent
coordinating activity which can modulate
the anteriad oscillator in the contingent
manner described in the previous para- FIG. 7. Stability of swimmeret system when anteriad
graphs. Each successive line of the figure control center has a higher intrinsic frequency than
posteriad control center. The cut command
illustrates the effect of additional coordi- the
neuron preparation of Figure 3 is used. A, Stimulanating neuron input. The last line illus- tion of posteiiad control center with 73 Hz stimulatrates the prediction that when a posteriad tion applied via the STIM 4 electrode. On somecontrol center and its associated coordinat- occasions, weak motor output in the anteriad coning neuron discharge have an intrinsic fre- trol center was observed with STIM 4 stimulation.
IS, Stimulation of the anteriad control center with
quency equal to that of an anteriad control 41 Hz stimulation applied via the STIM 3 electrode.
center (zero excitability gradient; Stein, C, Stimulation of both control centers with 73 Hz
1973) then the control centers will achieve applied \ia STIM 4 and 41 Hz applied via STIM 3
electrode.
phase stability with a non-zero phase lag.
The upper trace in each section is a stimulus
This logic also predicts that if the coordi- monitor.
Upward deflections mark pulses applied
nating input lias a higher intrinsic burst via the STIM 4 electrode. Downward deflections
frequency than the modulated center, then mark pulses applied via the STIM 3 electrode.
a net advance of the modulated center is Placement of recording and stimulating electrodes
as in Figure 3 except that coordinating neuron disnecessary to achieve frequency equalization. charge
was not monitored.
In this situation the interlimb lag will be
large. If, on the other hand, the coordinating input has a lower intrinsic burst fre- charge to the anteriad limb is taken as the
quency than the modulated center, then a intrinsic frequency of the anteriad control
net delay of the modulated center is neces- center. Stimulation is again applied for 10
sary to achieve frequency equalization. In- sec and then 10 sec of rest is allowed. Then
stimulation is applied both anteriad to and
terlimb lag will be small in this case.
These predictions have been confirmed posteriad to the cut. In this situation freutilizing the cut command neuron prepara- quency of the posteriad control center may
tion. In these experiments (Stein, 1973) the be equal to the frequency of the anteriad
cut command neuron posteriad to the cut control center (Figs. 1C, 8C) even though
is stimulated at constant frequency (Figs. their intrinsic frequencies were different. In
1A, 8/4). The frequency of motor discharge fact, frequency equalization may occur in
to the posteriad limb is taken as the intrin- situations where the anteriad control center
sic frequency of the posteriad control center. had a higher intrinsic frequency than the
Stimulation is applied for 10 sec and then posteriad control center (Fig. 7) and in sitthe preparation is allowed to rest for 10 sec. uations where the anteriad control center
The cut command neuron anteriad to the had a lower intrinsic frequency than the
cut is stimulated at constant frequency posteriad control center (Fig. 8). In these
(Figs. IB, 8B). The frequency of motor dis- experiments, the intrinsic frequency of each
NEURAL CONTROL OF INTERAPPENDAGE PHASE DURING LOCOMOTION
1013
lag observed with zero excitability gradient
in the cut command neuron preparation is
often seen during natural movements of the
intact animal. These results suggest that
in the intact animal, the command neuron
PS 3
system may strive for a zero excitability
B
gradient among the oscillators driving the
swimmerets. Deviations away from zero
gradient will not destroy phase locking as
long as the deviations are not too great.
C
The existence of non-zero phase lag with
a zero gradient of oscillator excitability has
been independently demonstrated by Davis
and Kennedy (1972). They have isolated
different classes of command neurons in the
FIG. 8. Stability of swimmeret system when anterior swimmeret system of the lobster. One class
of command neurons preferentially excited
control center has a lower intrinsic frequency than
posterior control center. Same experimental condiposterior segments (Fig. 9A,B); another
tions as in Figure 7 except STIM 3 is applied at
class of command neurons produced bal15 Hz.
anced excitation among all the limbs (Fig.
9C); a third class of command neurons preferentially
excited the more anterior limbs
control center could be altered by altering
the stimulation frequency applied via one (Fig. 9D,E). The direction of the metaof the stimulating electrodes. Note that interlimb phase is small in Figure 1C and 2
large in Figure 8C as predicted. There
,i A l l '
is an additional effect observed in these ex- 5
periments which cannot be explained by a
consideration of the ascending coordinating
neuron data alone. This effect is that the
frequency of the posteriad oscillator increases when the anteriad oscillator is also
excited. A possible explanation of this effect
is that descending coordinating neurons
(Stein, 1971, see Table 1) may also influence
this system.
When the intrinsic frequencies of the two
control centers were extremely different,
then different forms of coupling were observed. On some occasions, every second
burst of the control center with high intrinsic frequency was phase locked with each
burst of the control center with low intrin,/
sic frequency. On other occasions, each third
burst of one control center was phase locked
with each second burst of the other control FIG. 9. Different segmcntal effects of five command
center. These properties are predicted by interncurons (d-E) from the same preparation in
coupled oscillator theory (Moore et al., the lobster. Recordings were made from the left
powerstrokc nerves of all four segments (2-5).
1963; Perkel et al., 1964).
Records were obtained in random order and orThese results demonstrate that phase ganized to show progressively stronger anterior
stability can exist in the absence of distinct effects from A to E. (From Davis and Kennedy,
oscillator excitability gradients. The phase 1972.)
1014
PAUL S. G. STEIN
chronal wave was from posterior to anterior
in all three cases. The magnitude of the lag
varied according to the balance of excitation. Since no command neurons were cut,
it was not possible in these experiments to
measure the intrinsic frequency of each oscillator. It is known, however, that the number of motor impulses per burst increases
as the burst frequency of the swimmeret
movements increases (Davis, 1971). It is
reasonable to infer that the number of impulses per burst is a rough measure of intrinsic oscillator frequency. The situation
illustrated in Figure 9C may be taken as an
example of non-zero phase lag observed
with a zero gradient of excitability. It is
striking that the Davis and Kennedy (1972)
result is similar to the Stein (1973) result,
even though different experimental designs
were utilized.
POSSIBLE GENERALITIES OF THE
SWIMMERET RESULTS
Many locomotory systems are controlled
by distributed networks. Some of these
systems are also central. In most of these
systems, each limb control center drives repetitive bursts of motor discharge. In some
systems, cells whose discharge is a central
projection of control center activity have
been located (Arshavsky et al., 1972; Pearson and lies, 1973). It remains to be shown
whether other systems will display the contingent response of a motor center to coordinating discharge. Stability of these systems in trje absence of a fixed gradient of
intrinsic oscillator excitability also needs to
be demonstrated.
If the approach utilized for the swimmerets of crayfish is useful for other systems,
it is important to note that it is likely that
the details of the coupling may vary from
animal to animal or even among systems
controlling different gaits in the same animal. For example, there is evidence that
neighboring ipsilateral legs in the cockroach are coupled via inhibitory mechanisms (Pearson and lies, 1973). On the other
hand, it is possible that the trot in a fourlegged mammal may be controlled by coordinating neurons which utilize inhibition
Command Fiber
FIG. 10. Diagram of hypothesized connections
among representative neuronal elements of the crayfish swimmeret system. Two adjacent ipsilateral
hemiganglia are shown. (From Stein, 1971.)
while the gallop in the same individual may
be controlled by coordinating neurons
which utilize excitation. In spite of the differences in detail, the logical similarities of
networks controlling locomotion are many.
The full extent of these similarities will
only be revealed by future work.
SUMMARY OF SWIMMERET SYSTEM
PROPERTIES
1) The CNS network (Fig. 10) driving
locomotory rhythms is distributed. This
means that there is a rhythmic control
center driving the movements of each limb.
Each center is anatomically separated from
the control center for each other limb.
2) The CNS network driving locomotory
rhythms is central. This means that a deafferented nervous system can produce the
coordinated interlimb and intralimb motor
patterns observed in the intact animal.
3) Each control center is central. This
means that each control center when isolated from the remainder of the CNS and
from limb sensory input can produce a
normal intralimb locomotory output.
4) The CNS network can be activated
by command neurons. These neurons set the
general level of excitability of the control
centers, but they cannot control the precise interlimb phase relationships.
5) The CNS network utilizes coordinating neurons to establish a regulated interlimb phase and to achieve frequency equalization of limb movements.
6) The discharge of a given coordinating
NEURAL CONTROL OF INTERAPPEXDAGE PHASE DURING LOCOMOTION
neuron is driven by the control center of
one link.
7) The discharge of coordinating neurons can modulate the activity of other limb
control centers. The nature of the modulation is contingent upon the arrival time of
the coordinating discharge in the cycle of
the modulated oscillator.
8) The CNS network producing locomotory output will be stable in the absence
of a fixed gradient of intrinsic oscillator excitabilities.
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