Download nerve impulse patterns and reflex control in the motor system

y. Exp. Biol. (1965), 43, 193-210
With 12 text-figures
^rinUd in Great Britain
NERVE IMPULSE PATTERNS AND REFLEX CONTROL
IN THE MOTOR SYSTEM OF THE CRAYFISH CLAW
BY D. M. WILSON AND W. J. DAVIS
Department of Zoology, University of California, Berkeley
(Received 4 December 1964)
INTRODUCTION
With the excellent description of anatomy and neuromuscular physiology which is
available for crayfish claws and legs they seem appropriate objects for detailed analysis
of the nervous control mechanisms which operate them. We wish to report on two
aspects of this control which seem especially significant, namely, the production of a
specially patterned motor output to which the muscles are sensitive, and a role for the
peripheral neuromuscular inhibition.
It has been shown (Wiersma & Adams, 1950) that the tension developed by certain
crustacean muscles is influenced not only by the average frequency of electrically
induced excitatory nerve impulses, but also by the spacing, or pattern, of the impulses.
For example, when the excitatory axon to the opener muscle of the crayfish claw was
stimulated 12 times in a second, more tension resulted when the impulses were
delivered in pairs than when all the intervals between impulses were equal. Within
limits the tension increased as the size of the smaller intervals (between paired
impulses) decreased. Such preparations have been cited as examples of patternsensitivity at a myoneural junction. It has not been shown previously that the animals
which possess this synaptic property can utilize it by generating suitably patterned
excitatory impulses, since recordings of the normally generated motor activity have
not been analysed until now.
Reflex and motor control of crab and crayfish legs have been studied especially by
Bush (1962a, b, 1963), Eckert (1959), and Atwood (personal communication). A
striking finding has been that often the excitatory and inhibitory motor neurons to the
same muscle are active concurrently rather than showing a temporal alternation as
expected for antagonists. We have regularly found this synergism between inhibitor
and excitor. By studying the effects of several input pathways and using additional
techniques of analysis it is possible to relate this phenomenon to adaptively organized
proprioceptive reflexes and exteroceptive reflexes.
MATERIAL AND METHODS
The observations were made upon commercially supplied Procambarus clarkii. The
animals tended toward lethargy, especially after a long period in the storage aquaria or
after some handling or dissection. Some of the results are possibly influenced by
this perhaps unnatural state (see later).
For the recording of muscle activity under the most nearly normal behavioural
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D. M. WILSON AND W. J. DAVIS
conditions attainable, fine insulated wires were inserted through tiny holes in the
skeleton of the propodite, into the claw muscles, and fixed by wax. The animals were
left free to move and were observed both during spontaneous activity and during
various kinds of mechanical stimulation. Usually they became quiescent rather soon
and only the most vigorous stimulation elicited strong activity. Records were made
from both the opener and closer muscle of the dactyl, but those from the latter were
mostly impossible to interpret. This appears to be because of temporal interaction of
the three innervating neurons. Sometimes the 'fast' muscle action potentials antifacilitate and the 'slow' facilitate to nearly the same size, so that they are indistinguishable, and both are affected by superimposed inhibition. Potentials of the claw opener
muscle, with only one exciting and one inhibiting axon (Van Harreveld & Wiersma,
1937), were not ambiguous, and it was usually possible to recognize the temporal
sequence of excitor motor neuron impulses from the muscle action potential record.
Since this is of crucial importance a later paragraph will be devoted to impulse
identification.
For nerve recordings and stimulation the animal was tied to a wax block and a few
square millimetres of skeleton was removed to expose the nerve. This was done either
in the abdominal, oesophageal, or claw region (inner surface of the propodite). Van
Harreveld's solution was used to keep the tissues moist. In some cases the gills were
perfused with water. The nerves were lifted into air or oil on bare silver wires.
Electrical records were either filmed or stored on magnetic tape to be filmed later.
For some of the statistical analysis the films were processed by punching time of
occurrence of impulses on IBM cards. The data was then processed automatically by
programs developed by Wyman (1964).
RESULTS
Identification of the nervous units
The detailed results in this paper will deal only with the neurons which innervate the
opener muscle of the dactyl of the cheliped of crayfish. It is well known that this
muscle is doubly innervated by an excitor axon (with properties intermediate between
fast and slow) and an inhibitor axon (van Harreveld & Wiersma, 1937). It has also
been observed that in extracellular nerve recording the inhibitor axon has the spike of
larger amplitude (Bush, 1962).
Since the conclusions of this paper rest heavily upon the unambiguous identification
of these nervous units in the electrical records we will review the criteria we have used,
even though on the basis of the work of the above investigators this would seem
superfluous.
The best identification is possible during simultaneous recording from nerve and
muscle such as is illustrated in Fig. 1. A fine nerve branch near the opener muscle has
little if any sensory activity. Two large efferent spikes may be recorded and these
always differ in amplitude. Their relative amplitudes remain nearly the same during
hours of recording even though the absolute size varies with electrode position,
amount of saline, etc. Over short periods the absolute amplitude is constant unless
there is movement of the preparation or unless the frequency of discharge is very high.
In the latter case a diminution of amplitude occurs. Spikes of the two amplitudes can
Motor system of the crayfish claw
195
bear any temporal relationship to each other, including simultaneity (the record sums);
hence, they represent two different axons. Spikes of the same amplitude never occur
separated by less than about 4 msec.; hence, they do not represent activity in independent units. Even electrical stimulation cannot synchronke spikes of a single
amplitude; therefore, only a single axon is implicated. The smaller of these nerve
spikes is always associated with a muscle action potential in the opener muscle. The
muscle action potentials are not of constant amplitude. In the absence of nerve spikes
of the larger kind the amplitude of the muscle potential shows a regular dependence on
previous activity (Marmont & Wiersma, 1938). During long trains of small nerve
impulses the muscle potentials grow or facilitate gradually; the extent of the facilitation
is dependent both on length of the train and on frequency. A closely spaced pair of
impulses results in a conspicuous augmentation. At low frequencies of the small
HI
0-1 sec
Fig. 1. Records of efferent nerve (below) and muscle action potential (above) during
electrical stimulation of the circumoesophageal connective. The smaller nerve potential is
associated with excitatory muscle action potentials which show facilitation upon repetition.
The larger nerve potentials cause a reduction in the muscle potentials when they occur a few
milliseconds before the excitatory impulse. The nerve potentials often occur in pairs following
a single stimulus. The inhibitor has the greater latency. Therefore it often depresses only the
second of a pair of muscle potentials, giving the illusion of antifacilitation.
impulse only, the muscle action potentials can be very small and there may be no
detectable motion of the dactyl. Most often, however, the dactyl opens due to contraction of the opener muscle and it can therefore be concluded that the small efferent
nerve impulses are those of an excitatory motor axon.
It is usually impossible to record muscle action potentials when only the large nerve
impulses occur. Occasionally, very small potentials are recorded by the extracellular
wire leads. Unlike the larger excitatory potentials these may be of either polarity and
when they occur in a train superimposed on a shifting baseline, their amplitude and
polarity vary with baseline shifts. Although extracellular a.c. (but long time-constant)
recording was used these potentials seemed to exhibit the reversal phenomenon
characteristic of inhibitory postsynaptic potentials.
When the large nerve spikes occur a few milliseconds before the small ones the
muscle action potentials are diminished relative to those due to small spike innervation
only (Marmont & Wiersma, 1938; Wiersma & Ellis, 1942). The amount of diminution
is dependent upon the temporal relationship between the two nerve spikes. Unless the
two are very closely spaced the muscle potentials are uninfluenced by the larger nerve
spike. The inhibitory impulse has greater effect upon the excitatory muscle potential
13-2
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D. M. WILSON AND W. J. DAVIS
if it precedes the excitatory nerve impulse by about 5 msec. This near concurrence is
not necessary in order to inhibit contraction. Since the large efferent nerve impulses
never cause contraction of the muscle, but do diminish the excitatory muscle action
potentials, it can be concluded that they are the recordable activity of the inhibitory axon.
No other nerve activity has been found to influence the muscle activity. Therefore,
even when the electrical record from the muscle is quite complex, it is possible to
interpret it as a sequence of activations by a single excitatory unit with the resulting
muscle action potentials having amplitudes which depend upon the state of facilitation
and presence or absence of nearly simultaneous inhibition. Since it is difficult to
record from the nerves of moving animals we have made such an interpretation of the
muscle records to obtain information on the normal patterns of use of a single
excitatory motor neuron. In the stationary preparations, we have recorded the nerve
potentials themselves, thus eliminating the need for one part of the argument.
Temporal pattern of motor discharge
Muscle potentials from freely moving animals
By the time the attachment of electrodes was complete the reactivity of the animals
was already low. It was nearly impossible to induce a hard claw pinch, but some
walking and defensive reactions occurred. The defence reaction, which involves
raising and opening the claws, was associated with strong electrical activity in the
opener muscle. In the absence of known stimuli usually no activity occurred, but
sometimes relatively low frequency 'spontaneous' muscle potentials were observed
and recorded. Medium and high levels of activity could be elicited by touching,
pinching or scratching various parts of the body. The film records were studied for
evidence of a patterned motor output by entering the values for the time intervals
between the muscle potentials into interval histograms. Separate histograms, corresponding to different overall levels of activity, were constructed. The histograms in
Fig. 2a-c represent low, medium and high levels of activity, respectively. Sample
records appear with the proper histograms. As noted above, variation in the amplitude
of the single-unit opener muscle action potentials can be attributed to neuromuscular
facilitation and to the occurrence of inhibitory activity during motor excitation.
At the two extreme levels of activity the histograms are typical of single-unit
behaviour, but the bimodal interval distribution for the medium level, with the
minimum value at 13 msec, between the modes, is unexpected. The bimodality could
arise simply because a small number of records contained long continuous runs of
intervals smaller than 13 msec., but examination of all the records tabulated in
histogram b shows that in only two cases did the short intervals occur in runs as long
as three. In 14 cases short intervals occurred in runs of two, and in 125 cases the short
intervals occurred singly, separated on each side from other short intervals by at least
one interval longer than 13 msec. The probability that the observed sequence of
intervals, or a less random one, could be drawn from a population of intervals occurring in random order, is 0-006 (two-tailed one-sample run test for large samples,
Siegel, 1956). It is therefore probable that the bimodality reflects the occurrence of the
kind of pattern to which the opener muscle has been shown to be sensitive, namely,
short-term variations in the size of the intervals. It is interesting that the modal value
Motor system of the crayfish claw
197
of the shorter interval class is near to the value most effective in increasing muscle
tension (Ripley & Wiersma, 1953). Many of the records which are tabulated in
histogram c, including the one illustrated, indicate that patterning of the impulses,
expecially into pairs, occurs at the highest average frequencies also, but since the
average interval size is small here, two modes did not separate in the histogram.
0-2 see
240
Interval (msec.)
Fig. a. Interval histograms for three average intensities of activity in the motor innervation
to the claw opener muscle, and short representative samples of the oscilloscope records.
Facilitation of muscle action potentials occurs at higher frequencies, (a) Low, (b) medium,
and (c) high levels of activity. The bimodal histogram in (b) is unusual for single-unit nervous
activity. In this record the micro-temporal pattern is most conspicuous.
198
D. M. WILSON AND W. J. DAVIS
It seems highly unlikely that some input to the animal had the same temporal
distribution as the output shown in Fig. zb and it is therefore probable that the
closely spaced impulses are generated by ganglionic mechanisms independent of
input pattern. However, this conclusion is not rigorous without the controlled
experiments described in the next section.
Motor neuron and muscle potentials due to preganglionic electrical stimulation
Electrical stimulation of the circumoesophageal connectives or abdominal nerve
cord elicits activity in the claw opener muscle. The muscle potentials may follow the
input shocks 1:1 with a latency that varies around a value of 25 msec^ Usually many
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Fig. 3. Muscle action potentials during electrical stimulation of the circumoesophageal
connective. Upper lines, 10 marks/sec.; middle lines, stimulus marker; lower lines, muscle
recording, (a) Single-to-triple responses; (6) several cases of apparent antifacilitation due to
inhibition during the second excitatory potential; (c) some activity which has escaped
dependence upon the stimulus timing. This activity continues as an after-discharge.
cycles of input are needed before the output begins. During the steady response which
finally develops the output may consist of multiple discharges due to a single input
shock. These doublets and triplets resemble in temporal characteristics those recorded
from the intact animal, the separating interval being about 4-12 msec. They are
clearly not due to a similar temporal micro-structure in the input. Fig. 3 gives an
illustration of this effect. The irregularities in amplitude of the muscle potential are
rationalized by comparison with the records of Fig. 1 in which the nerve and muscle
recordings are presented. In Fig. 1 it can be seen also that a single input shock often
gives rise to two small efferent nerve impulses which are correlated with a muscle
action-potential doublet, and that there is often another rather later impulse. In
Fig. 3 c, which is taken after a long period of stimulation, extra excitatory muscle
potentials occur which are not correlated closely with the stimulus, and these continue
as an after-discharge.
Multiple discharges of the large (inhibitory) spike can also be caused by single
preganglionic shocks.
The diversity of possible input pathways, large dependence on repetition of input,
variation of output with input voltage, and long after-discharge suggest that these
Motor system of the crayfish claw
199
motor neurons fire only as the result of a complex integrative activity which includes
both spatial and temporal summation as well as local pattern production.
Reflexes and central control of the opener muscle
General body stimulation
Mechanical stimulation nearly anywhere on the body surface can elicit activity in
both the excitor and the inhibitor axons of the claw opener. This is true also of electrical
stimulation of the nerve cord either ahead of or behind the thorax. The two units may
4—4
Fig. 4. Various excitor—inhibitor relationships, (a) A single unit (the excitor) is active; (6)
excitor dominates inhibitor; (c) inhibitor dominates excitor; (d) excitor and inhibitor exhibit
all phase relationships; (e) inhibitor shows some naturally induced two- or three-spike bursts
during low background activity. Both units increase in frequency from left to right. Time
marker, io/sec.
not respond equally and they may have different background (pre-stimulation) levels
of activity. Nevertheless, for most stimulation sites their responses are positively
correlated. Fig. 4 shows some various relationships between excitor and inhibitor
activity, and Fig. 5 shows the effect of mechanical stimulation of the body surface. In
Fig. 5 it is seen that both axons are activated together, but that the larger (inhibitor) is
later, and that either may produce a doublet following a single stimulus. During a long
series of mechanical stimuli stimulus-independent discharge begins, as it does during
electrical stimulation.
A more detailed knowledge of the behaviour of the nerve units comes from statistical
analysis of long trains of activity. From the several histograms (Figs. 2, 6 a, b) it can be
D. M. WILSON AND W. J. DAVIS
200
seen that, as in most nerve spike trains, low-frequency runs are associated with more
scatter. However, if the degree of scatter (the standard deviation) is compared to the
mean interval it is found that this relative measure of variation (the coefficient of
variation, S.D.-=-£) increases with frequency, not with interval (Fig. 7). This is not
necessarily to be expected. Werner & Mountcastle (1963) found the coefficient of
variation of some sensory projection neurons to be independent of frequency. The
increased variability at high frequency may not represent simply increased randomness but may reflect the addition of the new class of shorter intervals due to the
doublets and triplets of spikes. At any rate the increased variability should lead to an
enhancement of muscle tension in the absence of other influences such as neuromuscular inhibition.
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Fig. 5. Response to mechanical stimulation of body. Prodding of ventral abdominal surface
about 4-5 times/sec, produces bursts (1-3) of excitatory impulses often followed by one or two
inhibitory impulses. After a prolonged series of stimuli the excitor shows additional activity
which has escaped fixed latency dependence (c and d). Time marker, io/sec.
Spontaneous behaviour
In Fig. be are shown the first ten orders of serial correlation coefficient for the
intervals between impulses on the excitatory and inhibitory axons during a run of concurrent activity without known input. For both units all ten orders of correlation are
significantly positive, indicating frequency trends lasting more than 10 cycles.
(Alternatively, it is possible that the impulse sequence was not a stationary time-series
over the period examined, and that there was only a single long trend.) In addition to
the average high correlation values the first few orders are especially high for the
excitor axon. This indicates short-term frequency trends superimposed upon the
longer ones. A meaning to be attached to this is that the large standard deviation for
interval length is not to be regarded as due primarily to random cycle-to-cycle
fluctuations but to a somewhat longer-lasting process. Over the short term the
excitatory axon at this frequency showed a tendency to rather smooth rhythmic
behaviour. From the record (not shown) and the histogram (Fig. 6 b) it can be seen
that no paired, short-interval firings occurred.
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Fig. 6. Analysis of results of a long run of' spontaneous' output, (a), (4) Interval histograms
for the inhibitory and excitatory trains respectively, (c) Auto-correlograms consisting of ten
orders of serial correlation coefficient for each train, (d) Phase histogram comparing percentage time of occurrence of spikes of one train in intervals of the other, (e) Cross-correlograma consisting of eleven orders of interval correlation chosen as illustrated. Intervals of one
train (a single interval is shaded) are correlated to the preceding, but overlapping, interval of
the other train for the zeroth-order correlation. Comparison is made to the succeeding, but
overlapping, interval for the first-order correlation, and with successive intervals for higher
serial orders. Different correlation values are found, especially for the first two orders,
depending upon whether the cross-correlation is made from the excitor to the inhibitor
(•
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D. M. WILSON AND W. J. DAVIS
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Fig. 7. Plot of the coefficient of variation (standard deviation divided by mean) for several
runs of activity of the excitor motor neuron. The numbers near the points give sample size.
Although the standard deviation of interval length decreases with increase in frequency, this
measure, relative to the mean, increases with increase in frequency.
The interval distribution for the inhibitory impulses during the same period shows
a higher average frequency, but a lower tendency to smooth change in frequency as
judged from the autocorrelogram (Fig. 6 c). This is contrary to the behaviour of at least
some other related neuron pairs (Wilson & Wyman, 1964). Relatively more of the
variation in interval lengths may be attributed to uncorrelated cycle-to-cycle changes
than in the case of lower-frequency behaviour.
If the excitatory spike intervals are compared serially with the inhibitory ones a
cross-correlogram may be constructed (Fig. 6e). It can be seen directly that the two
are positively correlated in frequency, that is, trend together both over short and
longer periods. It should also be noted that if excitatory intervals are compared to
preceding inhibitory ones the correlation is not significant, but that only when excitatory ones are compared to succeeding inhibitory ones is the correlation high. The
opposite is true if inhibitory intervals are compared to excitatory ones. This means
that frequency changes generally begin to occur in the excitatory line before they do in
the inhibitory one, even though they usually occur in both at about the same time.
The phase histogram (Fig. 6d) shows that the detailed time of firing of one unit is
not related to that of the other even though they undergo related short-term frequency
changes. Similar behaviour has been seen in pairs of synergistic insect motor neurons
(Wyman, 1964; Wilson & Wyman, 1964). An indication of the phase independence
can be seen in Fig. ^d.
Motor system of the crayfish claw
203
Stimulation of the claw
Passive opening of the claw or mechanical stimulation of the hair beds along the
inner edge of the propodite (facing the dactyl) may also cause increased firing of both
excitor and inhibitor axons, but these stimuli cause much more pronounced increases
in the inhibitor. Passive opening of the dactyl presumably stimulates the joint
receptor organ. This proprioceptive reflex is a dependable way of eliciting inhibitor
activity. The background activity may be dominated by either the excitor or inhibitor,
but the result is the same; both increase in frequency (see Fig. 8), but the inhibitor
activity is probably sufficient to prevent tension development since inhibitor/excitor
ratios of less than one can be sufficient to block contraction (Ripley & Wiersma, 1953).
jOpen
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Fig. 8. Effect of passive claw opening. The movement lasted only a few tenths of a second.
(a) The inhibitor was already active and both units increased. (6) The excitor was already
active and both units increased, (c) There was no background activity. The inhibitor fires
first. The concurrence of activity in the two units during this reflex is not due to spontaneity
or stimulation from another cause. Time marker, io/sec. Amplitude changes are associated
with movement of the preparation.
Electrical stimulation of the claw nerve where it contains both sensory neurons and
the motor axons to the opener muscle produces the effects illustrated in Fig. 9. The
stimulus is followed by a compound nerve potential which varies more or less smoothly
with stimulus voltage. At least at higher stimulus strengths this wave of activity should
include the proprioceptive and hair-bed fibre spikes as well as antidromic motor axon
spikes. Many milliseconds later than this compound wave there may be an efferent
discharge in the motor axons. It is characteristic that this discharge will be in the
inhibitory axon only, or in both inhibitor and excitor. In the latter case the inhibitory
spikes are earlier and usually greater in number. In contrast to other sources of input,
for this afferent pathway the inhibitor seems to have the lower threshold (and shorter
latency).
Ratio of inhibitor to excitor activity
When only the excitatory motor neuron is active, muscle activity can be related to
the frequency of the discharge. For combined inhibitory and excitatory innervation
D. M. WILSON AND W. J. DAVIS
204
50 msec.
Fig. 9. Effect of electrical stimulation of the mixed nerve peripheral to the branching to the
opener muscle. The stimulus artifact is followed by a compound wave of activity which
presumably carries both sensory and motor axon spikes centrally. There is a later rebound of
individual spikes which can be identified as both efferent and motor. The first is the larger
inhibitor. With greater input or a longer period of facilitation, later, smaller excitor axon spikes
occur.
35
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15
20
25
30
35
40
45
Excitor (impulses/sec.)
Fig. 10. Inhibitor frequency plotted against excitor frequency for a large number of samples.
T h e points fall along two slopes depending upon general input source. T h e broken line
represents the maximum ratio needed for total inhibition according to Ripley & Wiersma (1953).
Motor system of the crayfish claw
205
iRipley & Wiersma (1953) have demonstrated that the frequency ratio is a useful index
for determining some aspects of muscle activity. For example, they find that I/E ratios
above about 0-4-0-8 result in no tension in their crayfish-claw preparations. If we plot
the naturally occurring inhibitory and excitatory frequencies for a number of data
samples we find that the points fall into two groups (Fig. 10). One group includes
results of passive claw opening and the other includes results of general body stimulation (without claw movement). The slope of the former group is steep, showing that
the effect of passive claw opening is to cause inhibitory dominance. The result will be
total inhibition during vigorous passive opening. The group of data points arising
from other kinds of stimulation have a slope of less than one. I/E ratios appear to
decrease with increased excitation and the latter should dominate. Because the
dominance changes depending upon whether the input is from the claw (primarily
proprioceptive) or from the rest of the body (primarily exteroceptive) the system can
work adaptively even though the apparently antagonistic units show a general
synergism.
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Fig. 11. Two examples of the sequence of motor neuron events
associated with active opening of the claw.
Natural claw opening
In all of the above reflex studies the dactyl was either moved passively, held still, or
did not move because inhibition dominated excitation. In these circumstances the
proprioceptive reflex does not operate as a feedback loop; the loop is open. If the
claw is left free to move, the loop does operate and the sequence of events due to a
transient input is the following (Fig. 11). If the animal is stimulated in any of a large
variety of ways there is first a noticeable rise in frequency of the excitor motor axon to
the opener muscle. The inhibitor may also increase in frequency due to the primary
input, but inhibitor activity does not become pronounced until later when the dactyl is
moving. The effect is that an exteroceptively or centrally driven movement is stopped
or limited by proprioceptive feedback. The effects are mostly phasic; the claw may
stay open, but the rate processes decline.
DISCUSSION
The lethargic preparations
The unresponsiveness of many crayfish in captivity may not be due to a deficiency
of nervous response to stimuli. In many of our preparations it was true that there was
a continual flow of efferent nervous activity and that this flow rate was sensitive to
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D. M. WILSON AND W. J. DAVIS
input. However, there was often no claw movement because of the effect of peripheral
motor inhibition. The extent of this inhibitory background and response varied from
animal to animal and over long periods in a single preparation. We have no systematic
results on this long-term variation, but it may be possible to correlate it to the animal's
behavioural state. One important function of the muscle inhibition may be to control
this behavioural state. For the crayfish it may be appropriate to speak not of a
' central excitatory state', but rather of a peripheral excitatory (or even inhibitory) state.
Not only can one say that' a crayfish thinks with his claws' but also it may be possible
that a significant part of this general tendency to behavioural response is regulated at
the neuromuscular level in addition to efferent sensory control or central facilitating
mechanisms.
The motor output pattern
Not only is the neuromuscular junction sensitive to changes in pattern of temporal
sequence of impinging impulses (shown by earlier workers), but it also receives,
under natural circumstances, trains of impulses containing similar pattern characteristics. Under the natural circumstances the pattern does not consist of neatly spaced
pairs of impulses but rather of an irregular train containing an unusual number of
closely spaced doublets and triplets. From the results of earlier workers we can conclude that any deviation from a smoothly rhythmic sequence will enhance tension
development, and that the naturally occurring sequence will produce a greater rate of
tension increase than a rhythmic one of the same average frequency. The fact that the
relative variance increases with impulse frequency may be adaptively related to the
need for rapid adjustment of tension. The use of two sequence parameters, average
frequency and variance, narrows the frequency spectrum needed for a given range and
sensitivity of control. It is possible, of course, that other parameters are significant as
well. For example, different patterns with the same frequency and variance have not
been investigated.
A possible weakness in the argument that the pattern-sensitive junction receives an
appropriate pattern, and that therefore the whole arrangement has behavioural significance, lies in the possibility of a negating action by simultaneous inhibition. Simultaneous action was seen to be common in our records of activity driven by electrical
input when there was more or less fixed latency-dependence between stimulus and
response. However, the finding that the excitor and inhibitor impulses bear no special
phase relationship one to the other shows that under some natural circumstances the
inhibitor only subtracts excitation with some average effect, and not an effect specially
related to the relative timing of the potentials, even if such a special effect were
characteristic of nerve-muscle preparation.
It is possible to make a reasonable speculation about the mechanism which generates
the pattern of pairs and triplets. This can be done in part by analogy with motor
neurons of another system, the locust flight system, in which paired impulses also
result in a bimodal (or trimodal) interval histogram, but one in which the two modes
are separated by a region of non-occurring interval values (Wilson, 1964). In that case
patterning of the excitatory impulses appears to be caused by an oscillatory input to
the motor axon at a frequency corresponding to the longer intervals, with production
of paired impulses occurring during the period of a single fundamental input cycle.
Motor system of the crayfish claw
207
'The shorter intervals between paired impulses are probably caused by a relaxation
oscillation due to some recovery process in the motor neuron. Either relative refractoriness or depression of excitatory membrane potential following impulse
initiation could give the observed effect. The same sort of mechanism would explain
the results obtained with the crayfish, but if this is the case then the fundamental
(input) rhythm is subject to more variation, since the bimodal histogram is continuous
rather than disjoint. The duration of the small intervals is consistent with expected
times for refractory recovery of arthropod motor neurons. By this multiple discharge
process, which occurs primarily during periods of high levels of excitation, it is possible
for the relative variance to increase with frequency not by a real increase in noisiness
but by the addition of a new aspect of temporal patterning. Thus these neurons do not
really violate the more usual finding that rhythmicity increases with frequency if one
considers that at higher levels of activity there is some frequency multiplication
following the fundamental oscillation and that the brief burst represents a single
excitatory cycle in the driving mechanism.
A model of the reflex organization
The excitor and inhibitor neurons seem to form (part of) a single motor neuron pool.
For all kinds of inputs their outputs are positively correlated in frequency (unless one
chooses very short or highly irregular samples). For most inputs, and during spontaneous activity, the apparently smaller excitor neuron has the lower threshold and
shorter stimulus-to-response latency. (By spontaneous we mean that there is no
known input to the animal which is correlated with the output. The motor neurons
probably received input from other parts of the C.N.s.) In this respect the neuron pool
is probably of the general type. For special inputs from the claw the larger inhibitor
neuron responds more readily. However, from the point of view of the central
connexions it is not necessary to postulate antagonistic interactions between these two
neurons. They may be synergistically driven neurons supplying the same muscle but
their inputs are differentially weighted, depending upon source. All of our observations lead to this conclusion. It should be remembered that we are talking about a
single pair of neurons to one muscle. The extent to which other muscles and other
animals use the same arrangement of excitor-inhibitor innervation remains to be seen.
For a contrary example see Kennedy & Takeda (1965), who find reciprocal behaviour between the inhibitor and excitors of the slow flexor muscles of the crayfish
abdomen.
Fig. 12 shows how this common but differentially weighted input can be put to use.
The proprioceptive loop has the characteristic of a negative feedback system not in the
well-known ways of inhibiting the excitor neurons of the muscles which would
supplement the on-going movement, or of exciting the excitor neurons of the antagonistic muscles, but instead the negativity is achieved by exciting the inhibitor of
the muscle producing synergistic movement. Here is a second adaptive role for
peripheral neuromuscular inhibition. It replaces the central inhibition which is missing in this case. In addition to this effect of the proprioceptive feedback there is also
excitation of the excitor neurons of the closer muscle and the two effects are
supplemental.
Whether the feedback loop as a whole serves a useful function is open to further
208
D. M. WILSON AND W. J. DAVIS
discussion. First it should be pointed out that it is not clear what sort of stability it
provides. Several factors will make an analysis tedious. First of all, the muscles
develop tension rather slowly in response to a change of input frequency. This means
that during short bursts or trends it is rate of change of tension and not steady-state
tension which is affected by the motor output. Secondly, it is true that in the absence
of antagonism or skeletal blocks or elasticity a steady tension proportional to some
innervation frequency should produce continual movement until muscle shortening is
impossible, but since the other things are not absent it is difficult to predict how
position or rate of movement will be related to motor frequency. Thirdly, the reflex
Opener
muscle
Input
Joint
receptor
Excitor
Inhibitor
Fig. 12. Model summarizing possible organization of the reflex system controlling the
opener muscle. Interneurona could intervene in the chain and provide the fundamental
oscillatory drive with multiplet production by the motor neurons themselves.
efficacy adapts fairly rapidly. The equilibrium value for the loop may therefore change
as a derivative function of its state. Our intuitive guess is that by reason of sensory and
central adaptation the reflex has phasic characteristics which tend simply to oppose
movement and to bring the claw to rest wherever it is, and that only to the extent that
there are non-adapting tonic elements will the feedback loop tend to maintain a given
position. It seems probable that tonic effects are more prominent in the extremes of
the range of claw positions.
A behavioural interpretation
A common reaction of crayfishes is the defensive reaction in which claws are raised
high and held open. This reaction may be elicited by visual or tactile stimulation or by
electrical stimulation of a special single fibre in the nerve cord (Wiersma, 1961). As
we have seen, many sources of input can result in activation of the claw opener muscle
which takes part in this whole reaction. Once in the defensive posture the crayfish
may respond to prodding of the anterior regions by sweeping the claw(s) medially.
Either by this action or otherwise objects may come between the dactyl and propodite.
Without further visual stimulation it is now possible for the claw to close because
stimulation of the sensory hairs on the propodite inhibits opening and excites the
closer muscle. The resulting pinch may cause positive feedback due to more stimulation of the hair beds (which are incidentally not arranged so that the dactyl itself can
touch them). Once closed the claw muscles need not exert maximum effort in order
to remain so but may relax until opening just begins. The phasic proprioceptive loop
will then operate to inhibit opening and excite closing. The object may be held for a
Motor system of the crayfish claw
209
tanaximum time with less than a maximum expenditure of energy and each effort by the
captive object to open the claw is countered by more holding force. This is only a
single example of how the reflex mechanisms may be integrated into a whole sequence,
but its overt aspects are probably sufficiently well known to all who handle crayfishes
that the example is well taken.
SUMMARY
1. The opener muscle of the crayfish claw receives, under nearly natural conditions,
a train of excitatory nerve impulses which may show a temporal patterning to which
the muscle is specially sensitive. Especially at high frequencies the impulse train
contains doublets which form a separate class in the interval distribution. Their
appearance at high frequencies gives rise to an increase in the coefficient of variation of
interval lengths.
2. Excitatory and inhibitory motor neurons to the same opener muscle seem to be
part of the same commonly excited motor neuron pool. Frequency changes in the two
axons generally show positive correlations. For most inputs and for 'spontaneous'
central drive the excitor has the lowest threshold and shortest latency, and it gives the
earliest indication of changes of excitatory state.
3. Proprioceptive input from the claw may excite both motor neurons, but generally
the inhibitory one gives the earlier, bigger response. The peripheral inhibition completes a negative feedback loop.
4. Inhibitory frequencies plotted against concurrent excitatory frequencies give
points falling into two groups with different slopes dependent upon input source. For
general body inputs the slope is less than one and for proprioceptive claw input it is
much more than one. This divergence leads to greater effectiveness of either excitation
or inhibition as overall level of output increases, with a good separation of antagonistic
functions in spite of the apparent lack of reciprocal interaction.
We wish to thank R. J. Wyman for help with the data processing, E. Reid for
preparing several illustrations, and D. Kennedy for criticizing a draft of the manuscript. Financial support came from NSF grant number GB2116 and NIH grant
number NB03927.
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