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Brain Research, 549 (1991) 268-2'74 © 1991 Elsevier Science Publishers B.V. 0006-8993/91/$03.50 ADONIS 000689939116616K
BRES 16616
Instability of motor unit firing rates during prolonged isometric contractions in human masseter M.A. Nordstrom and T.S. Miles Department of Physiology, University of Adelaide, Adelaide, S.A. (Australia) (Accepted 11 December 1990)
Key words: Masseter; Motor unit; Firing rate; Motoneuron; Isometric contraction; Recruitment; Fatigue
The firing patterns of up to 4 concurrently active masseter motor units were studied with intramuscular electrodes during a continuous isometric contraction of 15 min duration, in which the subject maintained the mean firing rate of one selected unit at 10 Hz. With this paradigm the net excitation (i.e. mean firing rate) of one unit in the muscle was controlled. This served as the reference for the functional state of other active units during the prolonged contraction. With the mean firing rate of one unit in the muscle fixed, 58% of other active units showed a slow, statistically-significant change in mean firing rate over the 15 min. The initial firing rate of the units did not influence the change in rate. The original firing rate hierarchy, which in short-term contractions reflects the recruitment order, was altered during the prolonged contraction. The explanation for these differential changes in motoneuron net excitation is not clear; they could be intrinsic to the motoneurons or perhaps mediated by reflex pathways. The selective facilitation or suppression of some motor units with continuous activation means that the original size-structured combination of motor units can be modified during a prolonged contraction.
INTRODUCTION The control of muscle force during voluntary contractions is achieved by variation of the n u m b e r of m o t o r units active (recruitment), and by modulation of the firing p a t t e r n of the active m o t o r units (rate-coding). M o t o r unit recruitment has been extensively studied. It is now known that during isometric contractions m o t o r units are recruited and de-recruited in an orderly sequence which correlates with their size (reviewed in ref. 10). In addition, the recruitment o r d e r is relatively fixed, although it may be altered under certain conditions 9'H'28'29. In contrast to the large body of information concerning m o t o r unit recruitment, comparatively little is known about the relative firing patterns of the m o t o r units once they are activated for the performance of a particular task. In an isometric contraction of short duration the recruitment hierarchy is reflected in the relative firing rates of the concurrently active units. That is, if one looks at a cross-section of m o t o r unit activity within a muscle at a given total force level, the last-recruited units are not only larger, but also have on average a lower discharge rate than the earlier-recruited units 2°'27. This finding has been verified in the masseter 7'17. This is a consequence of
the tendency for most units in a muscle to fire at similar mean rates just above recruitment threshold, and for all active units to exhibit a roughly p r o p o r t i o n a l increase in firing rate as force is increased 5'19'2°'24. H o w e v e r , it is not k n o w n w h e t h e r the hierarchy of m o t o r unit firing rates f o u n d in brief isometric contractions, which reflects the recruitment order, is fixed during contractions of longer duration. If it were modified, it would indicate a dynamic flexibility of m o t o r unit usage patterns analogous to changes in the recruitment order. In o r d e r to answer this question, it is necessary to follow the activity of identified pairs of m o t o r units for long periods. A l t h o u g h there have been several reports of single unit behavior during an a t t e m p t e d constant-force contraction 2'3'8'14'15'24'26, there a p p e a r to be no reports in the literature of comparison of the activity of pairs of m o t o r units during p r o l o n g e d contractions. This und o u b t e d l y reflects the technical difficulty of such an approach. In the present study the firing rate of one m o t o r unit in the masseter muscle was controlled voluntarily by the subject at a constant prescribed rate. W i t h this p a r a d i g m the net excitation (i.e. the m e a n firing rate) of one unit in the muscle was controlled. This b e c a m e the reference
Reprint requests: T.S. Miles, Department of Physiology, University of Adelaide, GPO Box 498, Adelaide SA 5001, Australia. Correspondence: M.A. Nordstrom. Present address: Department of Physiology, University of Arizona, Tucson, AZ 85724, U.S.A.
269
for the functional state of between 1 and 4 concurrently active units whose firing patterns were monitored during a c o n t i n u o u s i s o m e t r i c c o n t r a c t i o n o f l o n g d u r a t i o n (15 min). In this way the long-term stability of the original size-structured firing rate hierarchy was assessed for tonic motor unit activation.
MATERIALS AND METHODS
Apparatus and recording procedure Data is reported from 10 volunteer subjects (4 males, 6 females) aged 18-40 years with normal dentitions and no history of masticatory dysfunction. All subjects gave informed consent, and the experimental procedures were consistent with the recommendations of the Declaration of Helsinki for Human Experimentation. The analyses reported herein were part of a series of experiments concerned with the functional characteristics of human masseter motor units 21"22'23. The experimental protocol was identical to that previously reported in our study of the fatigue properties of masseter units 2~. Many of the units whose firing patterns are analyzed in the present work were part of the fatigue study. The subjects bit on stainless steel bite bars with their incisor teeth. The vertical separation of the incisor teeth was fixed at 6 mm for the duration of the contraction. Isometric biting force was measured by strain gauges mounted on the bars and recorded on FM tape (bandwidth 0-2500 Hz). The relationship of the jaws to the bars was kept constant by means of small, acrylic impressions of the subject's upper and lower incisal surfaces on the bars. This 3-dimensional fixation via the teeth ensured that the joint angle and direction of application of force did not change during the experiment. This was an important advantage of using a jaw muscle for the present study, as such a stable arrangement is unlikely to be accomplished in experiments utilizing the limbs or fingers. Motor unit activity was usually recorded with a single bipolar electrode inserted percutaneously into the right masseter muscle. In two experiments a second electrode was inserted at least 1 cm away from the first. Each electrode consisted of 2 Teflon-insulated, stainless steel wires (70/~m core diameter) threaded through the lumen of a 26-gauge disposable needle. Needles were inserted deep into the main body of the muscle. After insertion, the needle was removed leaving the fine wires in place. The surface electromyogram (EMG) of the right masseter muscle was recorded with bipolar Ag/AgCI electrodes. The muscle EMG signals were amplified (1000x; bandwidth 20-5000 Hz) and recorded on analog FM tape (bandwidth 0-2500 Hz).
Protocol The subject was seated comfortably with the incisor teeth on the bite bars so that he/she could observe an oscilloscope screen on which was displayed the mean firing rate of the masseter unit selected by the experimenters for the subject to control. The unit controlled by the subject will be referred to as the "feedback" unit. Motor unit action potentials were discriminated on-line using a computer-aided spike separation device (SPS 8701; Signal Processing Systems, 23 Airlie Ave., Prospect 5082, Australia) which used a template-matching algorithm, and was developed in this laboratory TM. Following a few short trial runs to optimize the spike recognition parameters, the subject was instructed to bite on the bars with the force necessary to keep the feedback unit firing continuously at a mean rate of 10 Hz for 15 min with the aid of audio and visual feedback. The total biting force during these contractions was usually less than 20% of maximal. While the subject controlled the feedback unit at 10 Hz it was usually possible to detect the activity of other units in the same, or separate intramuscular electrodes: these units are referred to as "background" units. Subjects received no feedback regarding the activity of background units.
Analysis The intramuscular E M G records were analyzed off-line to determine the firing pattern of all the single units recorded during the contraction using the SPS 8701. This device allowed recognition of motor unit action potentialwaveforms with very few falsepositive errors (i.e.few spikes were incorrectlyclassified),but in multi-unit recordings some motor unit potentialswere missed due to occasional superimposition of synchronous action potentialsof differentunits. Only individual spike trains that could be discriminated with high reliability were used for analysis. Data were rejected if the intramuscular record could not be discriminated with less than 5 % of total spikes unclassified due to recognition errors (almost exclusively superimpositions). In many intramuscular records only one or two unit potentialswere present, and this accuracy criterion was easily met. In other cases, this criterion usually restricted analysis to the 3 units with the largest amplitude in any particular intramuscular record, and a maximal perrnissable error of approximately 3 % in missed recognition of spikes from any one unit (of three). As an additional check on discrimination accuracy the interspike intervals (ISis) for each unit were checked for the absence of abnormally short ISis ( 0.01) using Student's t-test. (3) Following verification from tests 1 and 2 that the mean firing rate of the feedback unit was appropriate during the epochs of interest, the mean ISI ± S.D. was calculated for each background unit from the corresponding epoch in the first and 15th minute of the contraction. For each background unit, Student's t-test was used to assess the statistical significance of the change in mean ISI between the two epochs. A P value of less than 0.01 was taken to indicate a significant difference in mean ISI.
RESULTS Subjects were able to control the firing rate of the s e l e c t e d u n i t w i t h o u t difficulty t h r o u g h o u t t h e 1 5 - m i n
270 contraction, provided that they received reliable feedback. Off-line analysis showed that the mean firing rate of the feedback unit was usually less than 10 Hz, but rarely differed significantly from the target rate in any 30-s epoch. Each b a c k g r o u n d unit that could be discriminated with high accuracy was paired with its corresponding feedback unit for comparison of their firing patterns throughout the contraction. Thirty-three feedback/background unit pairs were followed for the full 15 min. In each case, the mean ISI of the feedback unit did not change significantly with time. In 16 b a c k g r o u n d units (48%) the mean ISI was not significantly different in the first and 15th minute; in 11 b a c k g r o u n d units (33%) the mean ISI decreased (firing rate increased) significantly; and in 6 pairs (19%) the mean ISI increased (firing rate decreased) significantly. A n example of a feedback and background unit pair in which the m e a n discharge rate of both r e m a i n e d relatively constant over the 15 min is shown in Fig. 1A. The feedback unit was controlled voluntarily by the subject at a m e a n rate just below 10 Hz for the 15 rain. The b a c k g r o u n d unit fired at a mean rate of 16 Hz for this period. T h e r e was some short-term firing rate irregularity
for each unit, but in both cases the mean ISI (calculated over 30-s epochs) was not significantly different between the first and 15th minutes. Segments of the intramuscular E M G record from which these two units were discriminated are shown in Fig. l B . The larger-amplitude unit is the one whose rate was voluntarily controlled by the subject. A n example of a b a c k g r o u n d unit exhibiting a significant change in mean rate over the 15 min is shown in Fig. 2. As was c o m m o n l y observed in such cases, the mean firing rate of the b a c k g r o u n d unit showed a slow drift with time while the firing rate of the feedback unit was controlled by the subject. The m e a n rate of the feedback unit (trace " a " ) did not change significantly over the 15 min, although the trace shows some shortterm fluctuations. The b a c k g r o u n d unit (trace " b " ) initially fired at a mean rate below that of the feedback unit, but its firing rate slowly increased until it reached 13
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Fig. 1. Example of a pair of masseter units which maintained relatively constant firing rates for 15 min. A: the mean firing rate is plotted against time for two units. The firing rate records have beeen low-pass filtered in order to show long-term trends more clearly. The lower trace belongs to the feedback unit. B: segments of the inframuscular EMG records from the first and 15th minute of the contraction. The feedback unit has the larger-amplitude action potentials. Both units were discriminated without difficulty throughout the 15-min contraction.
15th Minute
I t[
1 ll 1 I I00 ms
Fig. 2. Example of a pair of units in which the mean firing rate of the background unit increased significantly over 15 min. A: the smoothed firing rate is plotted against time for the feedback unit (trace "a") and background unit (trace "b"). B: segments of the intramuscular EMG records from the first and 15th minute of the contraction. The feedback unit (marked "a") has larger-amplitude action potentials than the background unit (marked "b"). Although in each case there was a reduction in action potential amplitude over the 15 min, both units were clearly discriminated throughout the 15-min contraction.
271 Hz after 7 min, and then remained relatively constant. The mean ISI from 30-s epochs in the first and 15th minutes were significantly different (P < 0.01). In a number of experiments additional background units were present which could not be reliably discriminated for the full 15 min, due to changes in action potential waveform or abrupt cessation of activity, presumably resulting from electrode movement. These units were excluded from the analysis. However, there was a total of 10 background units that ceased firing before the end of the test period, but could be reactivated later by a more forceful bite. In 8 of these units there was a progressive reduction in firing rate prior to the cessation of firing, and the mean firing rate in the last full 30-s epoch in which the unit was tonically active was significantly below its initial mean rate. In the other 2 units, the initial firing rate was too low for the change in rate to become statistically significant before the unit ceased firing. The evidence suggests that in these 10 units the drop-out was not due to electrode movement, but rather that the unit ceased firing because the net excitation fell below the unit's tonic firing threshold. These 10 units were therefore included in the analysis, and the distribution of the changes in mean firing rate over the 15 min becomes: in 18 background units (42%) the mean ISI did not change significantly; in 11 background units (26%) the mean ISI decreased significantly (firing rate increased); and in 14 pairs (32%) the mean ISI increased significantly (firing rate decreased). Thirtynine of 43 pairs were recorded from the same intramuscular electrode, and thus represent motor unit activity from a localized region of the muscle. In 4 pairs the background and feedback unit were detected in separate electrodes, and in each case the mean firing rate of the background unit changed significantly over 15 min. The
changes in firing rate for the 43 background units are summarized in the histogram in Fig. 3. The threshold for a significant change in rate was about + 1 Hz, regardless of the initial firing rate. The mean ( + S.D.) change in rate for the 43 background units was +0.1 + 2.2 Hz. The largest change of rate observed in a background unit was an increase of 6.6 Hz. When more than one background unit was recorded during the same contraction, the mean firing rate of each background unit was relatively independent of the others. This can be seen in Table I. Each numerical value in the table is the number of b a c k g r o u n d - b a c k g r o u n d unit pairs with a particular combination of firing rate changes over the 15 min. The even distribution of units falling in each category in Table I does not suggest any link between the firing rate behavior of both background units over the 15 min. In only 3 cases in 24 (12.5%) did both background units of a pair behave in the same manner as the feedback unit; i.e. no significant change in mean firing rate after 15 min. This is consistent with the proportion expected (11%) if it is hypothesized that the change in mean firing rates of the 3 motor units were independent variables. A n example of divergent changes in mean firing rates of two background units is illustrated in Fig. 4. Unit " a " was the feedback unit controlled by the subject, and its mean firing rate remained relatively constant at about 10 Hz for the duration of the test. There was no significant difference between the mean rates from the first and 15th minutes. Unit " b " progressively increased in rate from an initial 12 Hz to 14 Hz after 9 min. Unit " c " decreased in rate from 17 Hz to 15 Hz over the same interval. The changes in mean ISI of units b and c between the first and 10th minute were statistically significant (P < 0.01). The mean firing rate of the 43 background motor units in the first minute of the contraction ranged from 7.3 to
10TABLE I Direction of change in mean firing rate for pairs of background units, in experiments where 2 or more background units were recorded in addition to the feedback unit
"2
No change, firing rate did not change significantly after 15 min. Rate 1', firing rate increased significantly after 15 min. Rate ~,, firing rate decreased significantly after 15 min. * Values indicate the number of background unit pairs with a particular combination of firing rate changes over the 15 min.
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Fig. 4. Three concurrently-active motor units showing divergent changes in mean firing rate with time. The feedback unit (trace a) was controlled at 10 Hz by the subject. During this contraction the mean firing rate of one background unit increased (trace b), while the mean firing rate of another background unit decreased (trace c).
20.8 Hz. A histogram showing the distribution of initial mean firing rates of the background units included in this study is shown in Fig. 5A. Most background units were firing considerably faster than the feedback unit; only about 14% of background units fired at 10 Hz or less, while 61% had firing rates above 13 Hz. The relationship between the background unit's initial mean firing rate and its subsequent change in rate after 15 min is shown in Fig. 5B. The values for the units which "dropped out" before the end of the contraction, but which could be reactivated later by a harder bite, are indicated by the open circles in Fig. 5B. For the population as a whole, linear regression revealed no significant correlation be-
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Fig. 5. The initial m e a n firing rate of the background units, and its influence on the change in m e a n rate with time. A: histogram of the distribution of the initial m e a n firing rate of the background units. B: the relationship between the initial mean firing rate of the background unit, and its subsequent change in m e a n rate after 15 min of activity. The open circles represent the 'drop-out' units.
tween the initial firing rate and the change in rate over the 15 min (r = 0.09; P > 0.05). Note that the units which ceased firing had initial mean firing rates both above and below that of the feedback unit. That is, the tendency for units to drop-out was not related to their recruitment threshold. If the units which "dropped out" (open circles) were considered separately, there was a significant tendency for units with the higher initial mean firing rate to show a greater reduction in firing rate prior to drop-out (r = -0.76; P < 0.05); this is not surprising as most units ceased firing tonically at about the same mean rate (around 7 Hz). DISCUSSION
The main finding in the present study was that the mean firing rates of all active motor units in the human masseter were not stable with time while the mean firing rate of one unit in the muscle was kept constant voluntarily by the subject during an isometric contraction. Only 42% of background units maintained a constant mean firing rate over the 15 min. A change of rate of 1 Hz was in most cases sufficient for a statistically significant difference between the initial and final mean firing rates (Fig. 3). In some cases the differential changes were large, and sufficient to reverse the original size-structured rank order of motoneuron firing rates (e.g. Fig. 2). The initial mean firing rates of the background units were distributed over a large range (Fig. 5A), which emphasizes the importance of firing rate modulation for force production in the masseter. Most background units fired at a higher rate than the feedback unit, because the required rate of 10 Hz for the feedback unit is just above the threshold for tonic firing in the masseter. The actual distribution of firing rates for active masseter units would be skewed towards even higher rates, but many small, high-rate units could not be discriminated with sufficient accuracy for inclusion in the present study. The recruitment force threshold for the units controlled at 10 Hz by the subject in these experiments was predominantly in the range from 0.1 to 20% of maximal bite force zl. The present results are therefore representative of the activity of low- and moderate-force threshold motor units in the masseter. The relative firing rates of a pair of units during a short-term, tonic isometric contraction reflects their recruitment order, and hence their relative sizes. When concurrently active units are considered together, the unit recruited at the lower force level will tend to have the higher mean firing rate of the pair at any level of total f o r c e 8'17'27. This is a consequence of the fact that in human voluntary contractions motor units in most mus-
273 cles begin firing at about the same mean rate, and gross variation in muscle force is accompanied by a roughly proportional change in the firing rate of all the active motor units in the muscle, although units firing at high rates may reach a plateau 5'19'2°'24. In the masseter, background units with an initial firing rate above 10 Hz can be assumed to have been recruited earlier (on average) than the feedback unit, and background units with an initial firing rate less than 10 Hz can be assumed to have been recruited at a higher force than the feedback unit. Units with very close activation thresholds will have similar mean firing rates at a given force level under these conditions. The change in firing rate of the background units with time did not correlate with the unit's initial firing rate (Fig. 5B). This means that the factors underlying the relative change in net excitation of the feedback and background units were not influenced by, or organized systematically in accordance with, the size of the motor unit. In some cases the changes accentuated the sizerelated differences in initial firing rate of some unit pairs, yet in other pairs the changes reduced the size-related differences in initial mean firing rate, even to the extent of reversing the original size-determined rank order of firing rates. The present results show that, following isometric contractions of long duration in the masseter, the original recruitment order cannot be inferred from the relative firing rates of the motor units. For the series of experiments, no net trend was evident for either an increase or decrease in the overall level of excitation of other units in the muscle while the mean firing rate of one unit was held constant, since the mean change in rate of the 43 background units was near zero (Fig. 3). In a given experiment, however, the approach of keeping the mean firing rate of one unit constant did not achieve constant excitation of the whole muscle (i.e. the same motor units active at the same rates) over a 15-min isometric contraction. In some background units firing slowed, and even ceased, while in others the firing rate increased. Others have shown inhomogeneous motor unit recruitment in different regions of a muscle with different tasks 28'29. The present findings extend this by demonstrating that even within small regions of muscle the original size-structured combination of motor units for performance of a very rigid task is not stable over time. The change in mean firing rate of the other active units while the mean firing rate of one unit is kept constant indicates that there is a differential change in the net excitation of a proportion of motor units in the muscle with prolonged activity. Factors determining motor unit firing rates to a particular excitatory input drive include those determining its functional threshold for activation (e.g. input resistance, synaptic weightings),
but in addition factors related to repetitive firing (e.g. adaptation). The non-uniform long-term firing rate behavior of the masseter units is unlikely to be the result of a differential central excitatory drive to the motoneurons. Features such as the existence of a size-related recruitment order, and a roughly proportional increase in firing rate of active units in the muscle as the force is increased 19'2°'24 together with the simultaneous common modulation of motor unit firing rates during attempted constant-force contractions in limb muscles 6, and short-term synchronization 26, all support the notion that the motoneuron pool receives a widely-distributed, uniform, command signal with a large number of common inputs. One possible explanation for the present results is a differential adaptation of the motoneurons to excitatory currents (c.f. Kernell and Monster12'13). However, the changes in firing rate in the background units observed in the present study did not fit this pattern of adaptation. The changes in rate occurred with a slow time course, and frequently began after several minutes of repetitive firing (e.g. Figs. 2 and 4), whereas in the intraceUular current injection experiments of Kernell and Monster the adaptation was virtually over after one minute of activation. In addition, the adaptation reported by KerneU and Monster would tend to bring motor unit firing rates closer together, because units with a higher initial rate showed the greater reductions in rate. In the present study the change in firing rate in the background units was not influenced by their initial firing rate (Fig. 5B). Inputs which are unequally distributed amongst the motoneurons in the pool could mediate the observed differential changes in motor unit firing rates. One such system is Renshaw cell recurrent inhibition, which has been implicated as a possible explanation for the nonuniform changes in firing rates in tibialis anterior motor units associated with the recruitment of a new unit during slow, isometric force-ramps4. However, the rnotoneurons of the jaw muscles are considered to lack Renshaw-type recurrent inhibition 16,25, so these circuits are unlikely to be responsible for the present results. Another possibility is the muscle spindle afferent inputs which in the cat are not uniformly distributed to homonymous jaw-elevator motoneurons ~, in contrast to the uniform distribution found in the limb muscles 1°. If this arrangement were also found in human masseter, the muscle spindle afferent connections (or perhaps the tendon organs) would have the potential to produce differential effects on motoneurons, as the muscle force changed during the prolonged contraction. As reported previously, the 15-rain contraction induced fatigue in most units; the twitch force declined by at least 25% in 81% of units tested2L Another possibility is cutaneous afferents, which have
274 b e e n s h o w n to alter the r e c r u i t m e n t o r d e r of m o t o r units
contraction.
in the first dorsal interosseus muscle in h u m a n s 11. A
large d e v i a t i o n s f r o m the original r a n k o r d e r of firing
Although
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rates w e r e f o u n d for s o m e units with p r o l o n g e d activity.
w o u l d t h e r e f o r e be e x p e c t e d to h a v e similar n o n - u n i f o r m
T h e functional significance of t h e s e o b s e r v a t i o n s r e m a i n s
effects on the firing rates of tonically active m o t o r units.
to be elucidated. It also r e m a i n s to be s e e n w h e t h e r these
T h e s e r e c e p t o r s , o r o t h e r s in the oral o r m a s t i c a t o r y
firing p a t t e r n s are f o u n d in o t h e r muscles, an i m p o r t a n t
structures c o u l d possibly m e d i a t e the o b s e r v e d differen-
q u e s t i o n in v i e w of the d i f f e r e n c e s in n e u r a l o r g a n i z a t i o n of the t r i g e m i n a l m o t o r pool.
tial firing rate changes in m a s s e t e r m o t o r units if their synaptic c o n n e c t i o n s to m a s s e t e r m o t o n e u r o n s are not u n i f o r m , and also not size-structured. R e g a r d l e s s of the s o u r c e of the differential changes in m o t o r unit activity with p r o l o n g e d activation, the p r e s e n t results d e m o n s t r a t e that m o t o r unit firing p a t t e r n s are not
static,
but
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or
s u p p r e s s i o n of s o m e units during a c o n t i n u o u s isometric REFERENCES 1 Appenteng, K., O'Donovan, M.J., Somjen, G., Stephens, J.A. and Taylor, A., The projection of jaw elevator muscle spindle afferents to fifth nerve motoneurons in the cat, J. Physiol., 279 (1978) 409-423. 2 Bigland, B. and Lippold, O.C.J., Motor unit activity in the voluntary contraction of human muscle, J. Physiol., 125 (1954) 322-335. 3 Bigland-Ritchie, B., Cafarelli, E. and Johansson, R., Long-term discharge pattern of motor units, Soc. Neurosci. Abstr., 11 (1985) 1028. 4 Broman, H., De Luca, C.J. and Mambrito, B., Motor unit recruitment and firing rate interaction in the control of human muscles, Brain Research, 337 (1985) 311-319. 5 De Luca, C.J., Le Fever, R.S., McCue, M.E and Xenakis, A.E, Behaviour of human motor units in different muscles during linearly varying contractions, J. Physiol., 329 (1982) 113-128. 6 De Luca, C.J., Le Fever, R.S., McCue, M.P. and Xenakis, A.P., Control scheme governing concurrently active human motor units during voluntary contractions, J. Physiol., 329 (1982) 129-142. 7 Defiler, B. and Goldberg, L.J., Spike train characteristics of single motor units in the human masseter muscle, Exp. Neurol., 61 (1978) 592-608. 8 Freund, H.-J. B/idingen, H.J. and Dietz, V., Activity of single motor units from human forearm muscles during voluntary isometric contractions, J. Neurophysiol., 38 (1975) 933-946. 9 Garnett, R. and Stephens, J.A, Changes in the recruitment threshold of motor units produced by skin stimulation, J. Physiol., 311 (1981) 463-473. 10 Henneman, E. and Mendell, L.M., Functional organization of motoneuron pool and its inputs. In Handbook of Physiology, Sect. l, Vol. 2, American Physiological Society, Bethesda, MD, 1981, pp. 423-507. 11 Kanda, K., Burke, R.E. and Walmsley, B., Differential control of fast and slow twitch motor units in the decerebrate cat, Exp. Brain Res., 29 (1977) 57-74. 12 Kernell, D. and Monster, A.W., Time course and properties of late adaptation in spinal motoneurons in the cat, Exp. Brain Res., 46 (1982) 191-196. 13 KerneU, D. and Monster, A.W., Motoneurone properties and motor fatigue. An intracellular study of gastrocnemius motoneurones of the cat, Exp. Brain Res., 46 (1982) 197-204. 14 Kranz, H. and Baumgartner, G., Human alpha motoneurone discharge, a statistical analysis, Brain Research, 67 (1974) 324-329.
Acknowledgements. This research was supported by a grant from the National Health and Medical Research Council of Australia. M. Nordstrom was a Dental Postgraduate Scholar of the N.H. & M.R.C. The assistance of Dr. K.S. Tiirker is gratefully acknowledged. The authors wish to thank Dr. A.J. Fuglevand for helpful comments on the manuscript. 15 Lindsley, D.B., Electrical activity of human motor units during voluntary contraction, Am. J. Physiol., 114 (1935) 90-99. 16 Lorente De N6, R., Action potentials of the motoneurones of the hypoglossal nucleus, J. Cell Comp. Physiol., 29 (1947) 207-288. 17 Miles, T.S. and Ttirker, K.S., Decomposition of the human electromyogramme in an inhibitory reflex, Exp. Brain Res., 65 (1987) 337-342. 18 Miles, T.S., Smith, N.J., Tfirker, K.S. and Nordstrom, M.A., A fast, accurate, on-line system for discriminating action potentials, based on a template matching algorithm, Soc. Neurosci. Abstr., 13 (1987) 1215. 19 Milner-Brown, H.S., Stein, R.B. and Yemm, R., Changes in firing rate of human motor units during linearly changing voluntary contractions, J. Physiol., 230 (1973) 371-390. 20 Monster, A.W. and Chan, H., Isometric force production by motor units of extensor digitorum communis muscle in man, J. Neurophysiol., 40 (1977) 1432-1443. 21 Nordstrom, M.A. and Miles, T.S., Fatigue of single motor units in human masseter, J. Appl. Physiol., 68 (1990) 26-34. 22 Nordstrom, M.A. and Miles, T.S., Discharge variability and physiological properties of human masseter motor units, Brain Research, in press. 23 Nordstrom, M.A., Miles, T.S. and Tiirker, K.S., Synchronization of motor units in human masseter during a prolonged isometric contraction, J. Physiol., 426 (1990) 409-421. 24 Person, R.S. and Kudina, L.P., Discharge frequency and discharge pattern of human motor units during voluntary contraction of muscle, EEG Clin. Neurophysiol., 32 (1972) 471-483. 25 Shigenaga, Y., Yoshida, A., Tsuru, K., Mitsuhiro, Y., Otani, K. and Cao, C.Q., Physiological and morphological characteristics of cat masticatory motoneurons-intracellular injection of HRP, Brain Research, 461 (1988) 238-256. 26 Smith, O.C., Action potentials from single motor units in voluntary contraction, Am. J. Physiol., 108 (1934) 629-638. 27 Tanji, J. and Kato, M., Firing rate of individual motor units in voluntary contraction of abductor digiti minimi muscle in man, Exp. Neurol., 40 (1973) 771-783. 28 Ter Haar Romeny, B.M., Denier van der Gon, J.J. and Gielen, C.C.A.M., Relation between location of a motor unit in the human biceps brachii and its critical firing levels for different tasks, Exp. Neurol., 85 (1984) 631-650. 29 Zuylen, E.J. van, Gielen, C.C.A.M. and Denier van der Gon, J.J., Coordination and inhomogeneous activation of human arm muscles during isometric torques, J. Neurophysiol., 60 (1988) 1523-1548.
Brain Research, 549 (1991) 268-2'74 © 1991 Elsevier Science Publishers B.V. 0006-8993/91/$03.50 ADONIS 000689939116616K
BRES 16616
Instability of motor unit firing rates during prolonged isometric contractions in human masseter M.A. Nordstrom and T.S. Miles Department of Physiology, University of Adelaide, Adelaide, S.A. (Australia) (Accepted 11 December 1990)
Key words: Masseter; Motor unit; Firing rate; Motoneuron; Isometric contraction; Recruitment; Fatigue
The firing patterns of up to 4 concurrently active masseter motor units were studied with intramuscular electrodes during a continuous isometric contraction of 15 min duration, in which the subject maintained the mean firing rate of one selected unit at 10 Hz. With this paradigm the net excitation (i.e. mean firing rate) of one unit in the muscle was controlled. This served as the reference for the functional state of other active units during the prolonged contraction. With the mean firing rate of one unit in the muscle fixed, 58% of other active units showed a slow, statistically-significant change in mean firing rate over the 15 min. The initial firing rate of the units did not influence the change in rate. The original firing rate hierarchy, which in short-term contractions reflects the recruitment order, was altered during the prolonged contraction. The explanation for these differential changes in motoneuron net excitation is not clear; they could be intrinsic to the motoneurons or perhaps mediated by reflex pathways. The selective facilitation or suppression of some motor units with continuous activation means that the original size-structured combination of motor units can be modified during a prolonged contraction.
INTRODUCTION The control of muscle force during voluntary contractions is achieved by variation of the n u m b e r of m o t o r units active (recruitment), and by modulation of the firing p a t t e r n of the active m o t o r units (rate-coding). M o t o r unit recruitment has been extensively studied. It is now known that during isometric contractions m o t o r units are recruited and de-recruited in an orderly sequence which correlates with their size (reviewed in ref. 10). In addition, the recruitment o r d e r is relatively fixed, although it may be altered under certain conditions 9'H'28'29. In contrast to the large body of information concerning m o t o r unit recruitment, comparatively little is known about the relative firing patterns of the m o t o r units once they are activated for the performance of a particular task. In an isometric contraction of short duration the recruitment hierarchy is reflected in the relative firing rates of the concurrently active units. That is, if one looks at a cross-section of m o t o r unit activity within a muscle at a given total force level, the last-recruited units are not only larger, but also have on average a lower discharge rate than the earlier-recruited units 2°'27. This finding has been verified in the masseter 7'17. This is a consequence of
the tendency for most units in a muscle to fire at similar mean rates just above recruitment threshold, and for all active units to exhibit a roughly p r o p o r t i o n a l increase in firing rate as force is increased 5'19'2°'24. H o w e v e r , it is not k n o w n w h e t h e r the hierarchy of m o t o r unit firing rates f o u n d in brief isometric contractions, which reflects the recruitment order, is fixed during contractions of longer duration. If it were modified, it would indicate a dynamic flexibility of m o t o r unit usage patterns analogous to changes in the recruitment order. In o r d e r to answer this question, it is necessary to follow the activity of identified pairs of m o t o r units for long periods. A l t h o u g h there have been several reports of single unit behavior during an a t t e m p t e d constant-force contraction 2'3'8'14'15'24'26, there a p p e a r to be no reports in the literature of comparison of the activity of pairs of m o t o r units during p r o l o n g e d contractions. This und o u b t e d l y reflects the technical difficulty of such an approach. In the present study the firing rate of one m o t o r unit in the masseter muscle was controlled voluntarily by the subject at a constant prescribed rate. W i t h this p a r a d i g m the net excitation (i.e. the m e a n firing rate) of one unit in the muscle was controlled. This b e c a m e the reference
Reprint requests: T.S. Miles, Department of Physiology, University of Adelaide, GPO Box 498, Adelaide SA 5001, Australia. Correspondence: M.A. Nordstrom. Present address: Department of Physiology, University of Arizona, Tucson, AZ 85724, U.S.A.
269
for the functional state of between 1 and 4 concurrently active units whose firing patterns were monitored during a c o n t i n u o u s i s o m e t r i c c o n t r a c t i o n o f l o n g d u r a t i o n (15 min). In this way the long-term stability of the original size-structured firing rate hierarchy was assessed for tonic motor unit activation.
MATERIALS AND METHODS
Apparatus and recording procedure Data is reported from 10 volunteer subjects (4 males, 6 females) aged 18-40 years with normal dentitions and no history of masticatory dysfunction. All subjects gave informed consent, and the experimental procedures were consistent with the recommendations of the Declaration of Helsinki for Human Experimentation. The analyses reported herein were part of a series of experiments concerned with the functional characteristics of human masseter motor units 21"22'23. The experimental protocol was identical to that previously reported in our study of the fatigue properties of masseter units 2~. Many of the units whose firing patterns are analyzed in the present work were part of the fatigue study. The subjects bit on stainless steel bite bars with their incisor teeth. The vertical separation of the incisor teeth was fixed at 6 mm for the duration of the contraction. Isometric biting force was measured by strain gauges mounted on the bars and recorded on FM tape (bandwidth 0-2500 Hz). The relationship of the jaws to the bars was kept constant by means of small, acrylic impressions of the subject's upper and lower incisal surfaces on the bars. This 3-dimensional fixation via the teeth ensured that the joint angle and direction of application of force did not change during the experiment. This was an important advantage of using a jaw muscle for the present study, as such a stable arrangement is unlikely to be accomplished in experiments utilizing the limbs or fingers. Motor unit activity was usually recorded with a single bipolar electrode inserted percutaneously into the right masseter muscle. In two experiments a second electrode was inserted at least 1 cm away from the first. Each electrode consisted of 2 Teflon-insulated, stainless steel wires (70/~m core diameter) threaded through the lumen of a 26-gauge disposable needle. Needles were inserted deep into the main body of the muscle. After insertion, the needle was removed leaving the fine wires in place. The surface electromyogram (EMG) of the right masseter muscle was recorded with bipolar Ag/AgCI electrodes. The muscle EMG signals were amplified (1000x; bandwidth 20-5000 Hz) and recorded on analog FM tape (bandwidth 0-2500 Hz).
Protocol The subject was seated comfortably with the incisor teeth on the bite bars so that he/she could observe an oscilloscope screen on which was displayed the mean firing rate of the masseter unit selected by the experimenters for the subject to control. The unit controlled by the subject will be referred to as the "feedback" unit. Motor unit action potentials were discriminated on-line using a computer-aided spike separation device (SPS 8701; Signal Processing Systems, 23 Airlie Ave., Prospect 5082, Australia) which used a template-matching algorithm, and was developed in this laboratory TM. Following a few short trial runs to optimize the spike recognition parameters, the subject was instructed to bite on the bars with the force necessary to keep the feedback unit firing continuously at a mean rate of 10 Hz for 15 min with the aid of audio and visual feedback. The total biting force during these contractions was usually less than 20% of maximal. While the subject controlled the feedback unit at 10 Hz it was usually possible to detect the activity of other units in the same, or separate intramuscular electrodes: these units are referred to as "background" units. Subjects received no feedback regarding the activity of background units.
Analysis The intramuscular E M G records were analyzed off-line to determine the firing pattern of all the single units recorded during the contraction using the SPS 8701. This device allowed recognition of motor unit action potentialwaveforms with very few falsepositive errors (i.e.few spikes were incorrectlyclassified),but in multi-unit recordings some motor unit potentialswere missed due to occasional superimposition of synchronous action potentialsof differentunits. Only individual spike trains that could be discriminated with high reliability were used for analysis. Data were rejected if the intramuscular record could not be discriminated with less than 5 % of total spikes unclassified due to recognition errors (almost exclusively superimpositions). In many intramuscular records only one or two unit potentialswere present, and this accuracy criterion was easily met. In other cases, this criterion usually restricted analysis to the 3 units with the largest amplitude in any particular intramuscular record, and a maximal perrnissable error of approximately 3 % in missed recognition of spikes from any one unit (of three). As an additional check on discrimination accuracy the interspike intervals (ISis) for each unit were checked for the absence of abnormally short ISis ( 0.01) using Student's t-test. (3) Following verification from tests 1 and 2 that the mean firing rate of the feedback unit was appropriate during the epochs of interest, the mean ISI ± S.D. was calculated for each background unit from the corresponding epoch in the first and 15th minute of the contraction. For each background unit, Student's t-test was used to assess the statistical significance of the change in mean ISI between the two epochs. A P value of less than 0.01 was taken to indicate a significant difference in mean ISI.
RESULTS Subjects were able to control the firing rate of the s e l e c t e d u n i t w i t h o u t difficulty t h r o u g h o u t t h e 1 5 - m i n
270 contraction, provided that they received reliable feedback. Off-line analysis showed that the mean firing rate of the feedback unit was usually less than 10 Hz, but rarely differed significantly from the target rate in any 30-s epoch. Each b a c k g r o u n d unit that could be discriminated with high accuracy was paired with its corresponding feedback unit for comparison of their firing patterns throughout the contraction. Thirty-three feedback/background unit pairs were followed for the full 15 min. In each case, the mean ISI of the feedback unit did not change significantly with time. In 16 b a c k g r o u n d units (48%) the mean ISI was not significantly different in the first and 15th minute; in 11 b a c k g r o u n d units (33%) the mean ISI decreased (firing rate increased) significantly; and in 6 pairs (19%) the mean ISI increased (firing rate decreased) significantly. A n example of a feedback and background unit pair in which the m e a n discharge rate of both r e m a i n e d relatively constant over the 15 min is shown in Fig. 1A. The feedback unit was controlled voluntarily by the subject at a m e a n rate just below 10 Hz for the 15 rain. The b a c k g r o u n d unit fired at a mean rate of 16 Hz for this period. T h e r e was some short-term firing rate irregularity
for each unit, but in both cases the mean ISI (calculated over 30-s epochs) was not significantly different between the first and 15th minutes. Segments of the intramuscular E M G record from which these two units were discriminated are shown in Fig. l B . The larger-amplitude unit is the one whose rate was voluntarily controlled by the subject. A n example of a b a c k g r o u n d unit exhibiting a significant change in mean rate over the 15 min is shown in Fig. 2. As was c o m m o n l y observed in such cases, the mean firing rate of the b a c k g r o u n d unit showed a slow drift with time while the firing rate of the feedback unit was controlled by the subject. The m e a n rate of the feedback unit (trace " a " ) did not change significantly over the 15 min, although the trace shows some shortterm fluctuations. The b a c k g r o u n d unit (trace " b " ) initially fired at a mean rate below that of the feedback unit, but its firing rate slowly increased until it reached 13
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Fig. 1. Example of a pair of masseter units which maintained relatively constant firing rates for 15 min. A: the mean firing rate is plotted against time for two units. The firing rate records have beeen low-pass filtered in order to show long-term trends more clearly. The lower trace belongs to the feedback unit. B: segments of the inframuscular EMG records from the first and 15th minute of the contraction. The feedback unit has the larger-amplitude action potentials. Both units were discriminated without difficulty throughout the 15-min contraction.
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Fig. 2. Example of a pair of units in which the mean firing rate of the background unit increased significantly over 15 min. A: the smoothed firing rate is plotted against time for the feedback unit (trace "a") and background unit (trace "b"). B: segments of the intramuscular EMG records from the first and 15th minute of the contraction. The feedback unit (marked "a") has larger-amplitude action potentials than the background unit (marked "b"). Although in each case there was a reduction in action potential amplitude over the 15 min, both units were clearly discriminated throughout the 15-min contraction.
271 Hz after 7 min, and then remained relatively constant. The mean ISI from 30-s epochs in the first and 15th minutes were significantly different (P < 0.01). In a number of experiments additional background units were present which could not be reliably discriminated for the full 15 min, due to changes in action potential waveform or abrupt cessation of activity, presumably resulting from electrode movement. These units were excluded from the analysis. However, there was a total of 10 background units that ceased firing before the end of the test period, but could be reactivated later by a more forceful bite. In 8 of these units there was a progressive reduction in firing rate prior to the cessation of firing, and the mean firing rate in the last full 30-s epoch in which the unit was tonically active was significantly below its initial mean rate. In the other 2 units, the initial firing rate was too low for the change in rate to become statistically significant before the unit ceased firing. The evidence suggests that in these 10 units the drop-out was not due to electrode movement, but rather that the unit ceased firing because the net excitation fell below the unit's tonic firing threshold. These 10 units were therefore included in the analysis, and the distribution of the changes in mean firing rate over the 15 min becomes: in 18 background units (42%) the mean ISI did not change significantly; in 11 background units (26%) the mean ISI decreased significantly (firing rate increased); and in 14 pairs (32%) the mean ISI increased significantly (firing rate decreased). Thirtynine of 43 pairs were recorded from the same intramuscular electrode, and thus represent motor unit activity from a localized region of the muscle. In 4 pairs the background and feedback unit were detected in separate electrodes, and in each case the mean firing rate of the background unit changed significantly over 15 min. The
changes in firing rate for the 43 background units are summarized in the histogram in Fig. 3. The threshold for a significant change in rate was about + 1 Hz, regardless of the initial firing rate. The mean ( + S.D.) change in rate for the 43 background units was +0.1 + 2.2 Hz. The largest change of rate observed in a background unit was an increase of 6.6 Hz. When more than one background unit was recorded during the same contraction, the mean firing rate of each background unit was relatively independent of the others. This can be seen in Table I. Each numerical value in the table is the number of b a c k g r o u n d - b a c k g r o u n d unit pairs with a particular combination of firing rate changes over the 15 min. The even distribution of units falling in each category in Table I does not suggest any link between the firing rate behavior of both background units over the 15 min. In only 3 cases in 24 (12.5%) did both background units of a pair behave in the same manner as the feedback unit; i.e. no significant change in mean firing rate after 15 min. This is consistent with the proportion expected (11%) if it is hypothesized that the change in mean firing rates of the 3 motor units were independent variables. A n example of divergent changes in mean firing rates of two background units is illustrated in Fig. 4. Unit " a " was the feedback unit controlled by the subject, and its mean firing rate remained relatively constant at about 10 Hz for the duration of the test. There was no significant difference between the mean rates from the first and 15th minutes. Unit " b " progressively increased in rate from an initial 12 Hz to 14 Hz after 9 min. Unit " c " decreased in rate from 17 Hz to 15 Hz over the same interval. The changes in mean ISI of units b and c between the first and 10th minute were statistically significant (P < 0.01). The mean firing rate of the 43 background motor units in the first minute of the contraction ranged from 7.3 to
10TABLE I Direction of change in mean firing rate for pairs of background units, in experiments where 2 or more background units were recorded in addition to the feedback unit
"2
No change, firing rate did not change significantly after 15 min. Rate 1', firing rate increased significantly after 15 min. Rate ~,, firing rate decreased significantly after 15 min. * Values indicate the number of background unit pairs with a particular combination of firing rate changes over the 15 min.
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Fig. 4. Three concurrently-active motor units showing divergent changes in mean firing rate with time. The feedback unit (trace a) was controlled at 10 Hz by the subject. During this contraction the mean firing rate of one background unit increased (trace b), while the mean firing rate of another background unit decreased (trace c).
20.8 Hz. A histogram showing the distribution of initial mean firing rates of the background units included in this study is shown in Fig. 5A. Most background units were firing considerably faster than the feedback unit; only about 14% of background units fired at 10 Hz or less, while 61% had firing rates above 13 Hz. The relationship between the background unit's initial mean firing rate and its subsequent change in rate after 15 min is shown in Fig. 5B. The values for the units which "dropped out" before the end of the contraction, but which could be reactivated later by a harder bite, are indicated by the open circles in Fig. 5B. For the population as a whole, linear regression revealed no significant correlation be-
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Fig. 5. The initial m e a n firing rate of the background units, and its influence on the change in m e a n rate with time. A: histogram of the distribution of the initial m e a n firing rate of the background units. B: the relationship between the initial mean firing rate of the background unit, and its subsequent change in m e a n rate after 15 min of activity. The open circles represent the 'drop-out' units.
tween the initial firing rate and the change in rate over the 15 min (r = 0.09; P > 0.05). Note that the units which ceased firing had initial mean firing rates both above and below that of the feedback unit. That is, the tendency for units to drop-out was not related to their recruitment threshold. If the units which "dropped out" (open circles) were considered separately, there was a significant tendency for units with the higher initial mean firing rate to show a greater reduction in firing rate prior to drop-out (r = -0.76; P < 0.05); this is not surprising as most units ceased firing tonically at about the same mean rate (around 7 Hz). DISCUSSION
The main finding in the present study was that the mean firing rates of all active motor units in the human masseter were not stable with time while the mean firing rate of one unit in the muscle was kept constant voluntarily by the subject during an isometric contraction. Only 42% of background units maintained a constant mean firing rate over the 15 min. A change of rate of 1 Hz was in most cases sufficient for a statistically significant difference between the initial and final mean firing rates (Fig. 3). In some cases the differential changes were large, and sufficient to reverse the original size-structured rank order of motoneuron firing rates (e.g. Fig. 2). The initial mean firing rates of the background units were distributed over a large range (Fig. 5A), which emphasizes the importance of firing rate modulation for force production in the masseter. Most background units fired at a higher rate than the feedback unit, because the required rate of 10 Hz for the feedback unit is just above the threshold for tonic firing in the masseter. The actual distribution of firing rates for active masseter units would be skewed towards even higher rates, but many small, high-rate units could not be discriminated with sufficient accuracy for inclusion in the present study. The recruitment force threshold for the units controlled at 10 Hz by the subject in these experiments was predominantly in the range from 0.1 to 20% of maximal bite force zl. The present results are therefore representative of the activity of low- and moderate-force threshold motor units in the masseter. The relative firing rates of a pair of units during a short-term, tonic isometric contraction reflects their recruitment order, and hence their relative sizes. When concurrently active units are considered together, the unit recruited at the lower force level will tend to have the higher mean firing rate of the pair at any level of total f o r c e 8'17'27. This is a consequence of the fact that in human voluntary contractions motor units in most mus-
273 cles begin firing at about the same mean rate, and gross variation in muscle force is accompanied by a roughly proportional change in the firing rate of all the active motor units in the muscle, although units firing at high rates may reach a plateau 5'19'2°'24. In the masseter, background units with an initial firing rate above 10 Hz can be assumed to have been recruited earlier (on average) than the feedback unit, and background units with an initial firing rate less than 10 Hz can be assumed to have been recruited at a higher force than the feedback unit. Units with very close activation thresholds will have similar mean firing rates at a given force level under these conditions. The change in firing rate of the background units with time did not correlate with the unit's initial firing rate (Fig. 5B). This means that the factors underlying the relative change in net excitation of the feedback and background units were not influenced by, or organized systematically in accordance with, the size of the motor unit. In some cases the changes accentuated the sizerelated differences in initial firing rate of some unit pairs, yet in other pairs the changes reduced the size-related differences in initial mean firing rate, even to the extent of reversing the original size-determined rank order of firing rates. The present results show that, following isometric contractions of long duration in the masseter, the original recruitment order cannot be inferred from the relative firing rates of the motor units. For the series of experiments, no net trend was evident for either an increase or decrease in the overall level of excitation of other units in the muscle while the mean firing rate of one unit was held constant, since the mean change in rate of the 43 background units was near zero (Fig. 3). In a given experiment, however, the approach of keeping the mean firing rate of one unit constant did not achieve constant excitation of the whole muscle (i.e. the same motor units active at the same rates) over a 15-min isometric contraction. In some background units firing slowed, and even ceased, while in others the firing rate increased. Others have shown inhomogeneous motor unit recruitment in different regions of a muscle with different tasks 28'29. The present findings extend this by demonstrating that even within small regions of muscle the original size-structured combination of motor units for performance of a very rigid task is not stable over time. The change in mean firing rate of the other active units while the mean firing rate of one unit is kept constant indicates that there is a differential change in the net excitation of a proportion of motor units in the muscle with prolonged activity. Factors determining motor unit firing rates to a particular excitatory input drive include those determining its functional threshold for activation (e.g. input resistance, synaptic weightings),
but in addition factors related to repetitive firing (e.g. adaptation). The non-uniform long-term firing rate behavior of the masseter units is unlikely to be the result of a differential central excitatory drive to the motoneurons. Features such as the existence of a size-related recruitment order, and a roughly proportional increase in firing rate of active units in the muscle as the force is increased 19'2°'24 together with the simultaneous common modulation of motor unit firing rates during attempted constant-force contractions in limb muscles 6, and short-term synchronization 26, all support the notion that the motoneuron pool receives a widely-distributed, uniform, command signal with a large number of common inputs. One possible explanation for the present results is a differential adaptation of the motoneurons to excitatory currents (c.f. Kernell and Monster12'13). However, the changes in firing rate in the background units observed in the present study did not fit this pattern of adaptation. The changes in rate occurred with a slow time course, and frequently began after several minutes of repetitive firing (e.g. Figs. 2 and 4), whereas in the intraceUular current injection experiments of Kernell and Monster the adaptation was virtually over after one minute of activation. In addition, the adaptation reported by KerneU and Monster would tend to bring motor unit firing rates closer together, because units with a higher initial rate showed the greater reductions in rate. In the present study the change in firing rate in the background units was not influenced by their initial firing rate (Fig. 5B). Inputs which are unequally distributed amongst the motoneurons in the pool could mediate the observed differential changes in motor unit firing rates. One such system is Renshaw cell recurrent inhibition, which has been implicated as a possible explanation for the nonuniform changes in firing rates in tibialis anterior motor units associated with the recruitment of a new unit during slow, isometric force-ramps4. However, the rnotoneurons of the jaw muscles are considered to lack Renshaw-type recurrent inhibition 16,25, so these circuits are unlikely to be responsible for the present results. Another possibility is the muscle spindle afferent inputs which in the cat are not uniformly distributed to homonymous jaw-elevator motoneurons ~, in contrast to the uniform distribution found in the limb muscles 1°. If this arrangement were also found in human masseter, the muscle spindle afferent connections (or perhaps the tendon organs) would have the potential to produce differential effects on motoneurons, as the muscle force changed during the prolonged contraction. As reported previously, the 15-rain contraction induced fatigue in most units; the twitch force declined by at least 25% in 81% of units tested2L Another possibility is cutaneous afferents, which have
274 b e e n s h o w n to alter the r e c r u i t m e n t o r d e r of m o t o r units
contraction.
in the first dorsal interosseus muscle in h u m a n s 11. A
large d e v i a t i o n s f r o m the original r a n k o r d e r of firing
Although
they w e r e c o m p a r a t i v e l y
rare,
t i m e - d e p e n d e n t c h a n g e in the activity of these r e c e p t o r s
rates w e r e f o u n d for s o m e units with p r o l o n g e d activity.
w o u l d t h e r e f o r e be e x p e c t e d to h a v e similar n o n - u n i f o r m
T h e functional significance of t h e s e o b s e r v a t i o n s r e m a i n s
effects on the firing rates of tonically active m o t o r units.
to be elucidated. It also r e m a i n s to be s e e n w h e t h e r these
T h e s e r e c e p t o r s , o r o t h e r s in the oral o r m a s t i c a t o r y
firing p a t t e r n s are f o u n d in o t h e r muscles, an i m p o r t a n t
structures c o u l d possibly m e d i a t e the o b s e r v e d differen-
q u e s t i o n in v i e w of the d i f f e r e n c e s in n e u r a l o r g a n i z a t i o n of the t r i g e m i n a l m o t o r pool.
tial firing rate changes in m a s s e t e r m o t o r units if their synaptic c o n n e c t i o n s to m a s s e t e r m o t o n e u r o n s are not u n i f o r m , and also not size-structured. R e g a r d l e s s of the s o u r c e of the differential changes in m o t o r unit activity with p r o l o n g e d activation, the p r e s e n t results d e m o n s t r a t e that m o t o r unit firing p a t t e r n s are not
static,
but
may
be
modified
by
facilitation
or
s u p p r e s s i o n of s o m e units during a c o n t i n u o u s isometric REFERENCES 1 Appenteng, K., O'Donovan, M.J., Somjen, G., Stephens, J.A. and Taylor, A., The projection of jaw elevator muscle spindle afferents to fifth nerve motoneurons in the cat, J. Physiol., 279 (1978) 409-423. 2 Bigland, B. and Lippold, O.C.J., Motor unit activity in the voluntary contraction of human muscle, J. Physiol., 125 (1954) 322-335. 3 Bigland-Ritchie, B., Cafarelli, E. and Johansson, R., Long-term discharge pattern of motor units, Soc. Neurosci. Abstr., 11 (1985) 1028. 4 Broman, H., De Luca, C.J. and Mambrito, B., Motor unit recruitment and firing rate interaction in the control of human muscles, Brain Research, 337 (1985) 311-319. 5 De Luca, C.J., Le Fever, R.S., McCue, M.E and Xenakis, A.E, Behaviour of human motor units in different muscles during linearly varying contractions, J. Physiol., 329 (1982) 113-128. 6 De Luca, C.J., Le Fever, R.S., McCue, M.P. and Xenakis, A.P., Control scheme governing concurrently active human motor units during voluntary contractions, J. Physiol., 329 (1982) 129-142. 7 Defiler, B. and Goldberg, L.J., Spike train characteristics of single motor units in the human masseter muscle, Exp. Neurol., 61 (1978) 592-608. 8 Freund, H.-J. B/idingen, H.J. and Dietz, V., Activity of single motor units from human forearm muscles during voluntary isometric contractions, J. Neurophysiol., 38 (1975) 933-946. 9 Garnett, R. and Stephens, J.A, Changes in the recruitment threshold of motor units produced by skin stimulation, J. Physiol., 311 (1981) 463-473. 10 Henneman, E. and Mendell, L.M., Functional organization of motoneuron pool and its inputs. In Handbook of Physiology, Sect. l, Vol. 2, American Physiological Society, Bethesda, MD, 1981, pp. 423-507. 11 Kanda, K., Burke, R.E. and Walmsley, B., Differential control of fast and slow twitch motor units in the decerebrate cat, Exp. Brain Res., 29 (1977) 57-74. 12 Kernell, D. and Monster, A.W., Time course and properties of late adaptation in spinal motoneurons in the cat, Exp. Brain Res., 46 (1982) 191-196. 13 KerneU, D. and Monster, A.W., Motoneurone properties and motor fatigue. An intracellular study of gastrocnemius motoneurones of the cat, Exp. Brain Res., 46 (1982) 197-204. 14 Kranz, H. and Baumgartner, G., Human alpha motoneurone discharge, a statistical analysis, Brain Research, 67 (1974) 324-329.
Acknowledgements. This research was supported by a grant from the National Health and Medical Research Council of Australia. M. Nordstrom was a Dental Postgraduate Scholar of the N.H. & M.R.C. The assistance of Dr. K.S. Tiirker is gratefully acknowledged. The authors wish to thank Dr. A.J. Fuglevand for helpful comments on the manuscript. 15 Lindsley, D.B., Electrical activity of human motor units during voluntary contraction, Am. J. Physiol., 114 (1935) 90-99. 16 Lorente De N6, R., Action potentials of the motoneurones of the hypoglossal nucleus, J. Cell Comp. Physiol., 29 (1947) 207-288. 17 Miles, T.S. and Ttirker, K.S., Decomposition of the human electromyogramme in an inhibitory reflex, Exp. Brain Res., 65 (1987) 337-342. 18 Miles, T.S., Smith, N.J., Tfirker, K.S. and Nordstrom, M.A., A fast, accurate, on-line system for discriminating action potentials, based on a template matching algorithm, Soc. Neurosci. Abstr., 13 (1987) 1215. 19 Milner-Brown, H.S., Stein, R.B. and Yemm, R., Changes in firing rate of human motor units during linearly changing voluntary contractions, J. Physiol., 230 (1973) 371-390. 20 Monster, A.W. and Chan, H., Isometric force production by motor units of extensor digitorum communis muscle in man, J. Neurophysiol., 40 (1977) 1432-1443. 21 Nordstrom, M.A. and Miles, T.S., Fatigue of single motor units in human masseter, J. Appl. Physiol., 68 (1990) 26-34. 22 Nordstrom, M.A. and Miles, T.S., Discharge variability and physiological properties of human masseter motor units, Brain Research, in press. 23 Nordstrom, M.A., Miles, T.S. and Tiirker, K.S., Synchronization of motor units in human masseter during a prolonged isometric contraction, J. Physiol., 426 (1990) 409-421. 24 Person, R.S. and Kudina, L.P., Discharge frequency and discharge pattern of human motor units during voluntary contraction of muscle, EEG Clin. Neurophysiol., 32 (1972) 471-483. 25 Shigenaga, Y., Yoshida, A., Tsuru, K., Mitsuhiro, Y., Otani, K. and Cao, C.Q., Physiological and morphological characteristics of cat masticatory motoneurons-intracellular injection of HRP, Brain Research, 461 (1988) 238-256. 26 Smith, O.C., Action potentials from single motor units in voluntary contraction, Am. J. Physiol., 108 (1934) 629-638. 27 Tanji, J. and Kato, M., Firing rate of individual motor units in voluntary contraction of abductor digiti minimi muscle in man, Exp. Neurol., 40 (1973) 771-783. 28 Ter Haar Romeny, B.M., Denier van der Gon, J.J. and Gielen, C.C.A.M., Relation between location of a motor unit in the human biceps brachii and its critical firing levels for different tasks, Exp. Neurol., 85 (1984) 631-650. 29 Zuylen, E.J. van, Gielen, C.C.A.M. and Denier van der Gon, J.J., Coordination and inhomogeneous activation of human arm muscles during isometric torques, J. Neurophysiol., 60 (1988) 1523-1548.
