1Department of Medical Physiology, Section of Neurophysiology, University of Copenhagen, 2200 Copenhagen N, Denmark; and 2Institute of Neurophysiology, University of Oslo, N-0317 Oslo, Norway
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ABSTRACT |
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Gorassini, Monica,
Torsten Eken,
David J. Bennett,
Ole Kiehn, and
Hans Hultborn.
Activity of Hindlimb Motor Units During Locomotion in the
Conscious Rat.
J. Neurophysiol. 83: 2002-2011, 2000.
This paper compares the activity of hindlimb motor units
from muscles mainly composed of fast-twitch muscle fibers (medial and
lateral gastrocnemius: MG/LG, tibialis anterior: TA) to motor units
from a muscle mainly composed of slow-twitch muscle fibers (soleus:
SOL) during unrestrained walking in the conscious rat. Several
differences in the activation profiles of motor units from these two
groups of muscles were observed. For example, motor units from fast
muscles (e.g., MG/LG and TA) fired at very high mean frequencies of
discharge, ranging from 60 to 100 Hz, and almost always were recruited
with initial doublets or triplets, i.e., initial frequencies 100 Hz.
In contrast, the majority of SOL units fired at much lower mean rates
of discharge,
30 Hz, and had initial frequencies of only 30-60 Hz
(i.e., there were no initial doublets/triplets
100 Hz). Thus the
presence of initial doublet or triplets was dependent on the intrinsic
properties of the motor unit, i.e., faster units were recruited with a
doublet/triplet more often than slower units. Moreover, in contrast to
units from the slow SOL muscle, the activity of single motor units from
the fast MG/LG muscle, especially units recruited midway or near the end of a locomotor burst, was unrelated to the activity of the remainder of the motoneuron pool, as measured by the corresponding gross-electromyographic (EMG) signal. This dissociation of activity was
suggested to arise from a compartmentalized recruitment of the MG/LG
motoneuron pool by the rhythm-generating networks of the spinal cord.
In contrast, when comparing the rate modulation of simultaneously
recorded motor units within a single LG muscle compartment, the frequency profiles of unit pairs were modulated in a
parallel fashion. This suggested that the parent motoneurons were
responsive to changes in synaptic inputs during unrestrained walking,
unlike the poor rate modulation that occurs during locomotion induced
from brain stem stimulation. In summary, data from this study provide
evidence that the firing behavior of motor units during unrestrained
walking is influenced by both the intrinsic properties of the parent
motoneuron and by synaptic inputs from the locomotor networks of the
spinal cord. In addition, it also provides the first extensive
description of motor-unit activity from different muscles during
unrestrained walking in the conscious rat.
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INTRODUCTION |
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One way to study the central processes involved in
the production of locomotion is to record the activity of single
motoneurons during walking. Traditionally this has been done by
examining locomotor-like behavior induced by stimulation of the
mescencephalic locomotor region (MLR) in the decerebrate cat
(Brownstone et al. 1992; Orsal et al.
1986
; Severin et al. 1967
; Tansey and
Botterman 1996
; Zajac and Young 1980
).
Single-motor-unit activity recorded in unrestrained, conscious
preparations during self-initiated walking is rare because of the
technical difficulties in obtaining such data (cat, Hoffer et
al. 1987
; rat, Eken 1998
; human, DeSerres et al. 1995
; Grimby 1984
; Jakobsson et
al. 1988
; Ogawa et al. 1991
). Because of this,
detailed information from only a limited sample of motor-unit types is
available, including the anterior thigh muscles in the intact cat and
the tibialis anterior and the short toe extensor muscles in the human.
One aim of the present study therefore was to record from various types
of motor units in the conscious animal that differ in both action
(extensors vs. flexors) and motor-unit type (slow vs. fast).
Single-motor -unit activity was recorded in the medial and lateral
gastrocnemius (MG and LG), soleus (SOL), and tibialis anterior (TA)
muscles of the rat hindlimb. We used the rat because of the increasing
number of laboratories using the neonatal rat preparation to study
locomotion (Kiehn and Kjaerulff 1998) and the need to
establish what the normal adult motoneuron patterns are during
unrestrained, self-initiated walking.
When comparing motoneuron behavior during MLR-induced activity and
self-initiated walking, several interesting discrepancies arise. For
example, during MLR-induced locomotion, there is little rate modulation
of motoneuron activity during a locomotor burst, i.e., the firing
frequency profiles are flat. In addition, firing rates are affected
only slightly by changes in either synaptic input (Severin et
al. 1967; Zajac and Young 1980
) or injected current (Brownstone et al. 1992
). In contrast during
self-initiated walking, motor-unit frequency profiles are more rounded,
and they follow the envelope of their corresponding gross
electromyograms (EMGs) (DeSerres et al. 1995
;
Eken 1998
; Hoffer et al. 1987
). Likewise,
mean firing rates tend to increase as the speed of locomotion increases
(Grimby 1984
; Hoffer et al. 1987
;
Ogawa et al. 1991
). In addition, during MLR-induced
locomotion, there tends to be a greater percentage of motoneurons that
fire with initial doublets (Zajac and Young 1980
,
although cf. Brownstone et al. 1992
) in comparison to
self-initiated walking (Hoffer et al. 1987
; Ogawa et al. 1991
) [an initial doublet is defined as the occurrence of a short interspike interval (i.e.,
10 ms) between the first two
motor-unit action potentials (MUAPs) in a burst (Zajac and Young
1980
)].
The poor rate modulation and high occurrence of initial doublets may be
due to stronger and more abrupt synaptic activation of motoneurons by
MLR input than during self-initiated walking. However, it is also
possible that motoneurons with different intrinsic properties (i.e.,
fast vs. slow) differ in their recruitment and firing behavior. This
latter possibility has not been excluded because the motoneurons
studied in the MLR preparations often were not identified with respect
to motoneuron type or muscle they supplied and the data from
self-initiated walking come from only a few muscle types. We therefore
compared the firing patterns of motor units in muscles with
predominantly fast twitch muscle fibers (90%; LG, MG, and TA) with
those of motor units in a muscle with 90% or more slow twitch muscle
fibers (SOL) (Gillespie et al. 1987
;
Tötösy de Zepetnek 1992
) to examine if there
were systematic differences in the recruitment and rate modulation patterns of the different motor-unit populations. Parts of this paper
have been presented in abstract form (Gorassini et al.
1995
).
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METHODS |
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Single-motor-unit recordings were obtained from 18 (of 24) adult
male Wistar rats weighing between 270 and 390 g. Intramuscular micro-EMG electrodes were implanted in the MG, LG, and TA muscles to
record single MUAPs. The firing behavior of five TA, three MG, and nine
LG motor units during unrestrained walking are described (the activity
of 3 of these units during imposed muscle stretch is described in
Gorassini et al. 1999). In all rats, gross (i.e., whole
muscle) EMG electrodes were implanted in both flexor (TA) and extensor
(MG or LG) muscles. In a separate series of experiments performed at
the University of Oslo by Torsten Eken, data from the SOL muscle were
obtained from four male adult Møll-Wistar rats (n = 5 units). Experimental details and a qualitative description of these SOL
motor units during tonic activity and walking can be found in
Eken (1998)
. Experiments were approved by the local ethics committees in Copenhagen and Oslo, respectively.
Implant procedures and data recording
Details of the construction and implantation of the micro- and
gross-EMG electrodes can be found in Eken (1998) and
Gorassini et al. (1999)
. Note that the gross-EMG
electrodes were inserted on either side of the corresponding micro-EMG
electrodes, separated by
1 cm, to record from the same population of
motor units.
All walking data were obtained from over-ground locomotion. Animals
walked freely in a 1.0 × 0.8 m glass aquarium with the walking surface covered by a rubber mat. In this space, the rat could
take 10 consecutive steps along the length of the aquarium. Gross-EMG activity from TA and LG or MG was used to assess the quality
of walking. The swing and stance phases of the step cycle were defined
as occurring during the periods of TA and MG/LG gross-EMG activity,
respectively. Unit activity was analyzed only when good alternation
between flexor and extensor muscles was present and during sequences of
at least four steps. For the SOL units, flexor activity was not
recorded but quality of walking was assessed from video recordings
taken during the walking sequences. Details of the recording set-up and
single-unit discrimination techniques can be found in Eken
(1998)
and Gorassini et al. (1999)
.
Discrimination of all MUAP waveforms in an entire locomotor burst was
possible in only 10% of the steps recorded due to movement of the
electrodes or from superimposition of other units. These steps were
distributed randomly in the recordings, i.e., there were no particular
speeds or gross-EMG profiles in which single MUAP waveforms could be
discriminated more easily. In steps where only a portion of the MUAPs
could be discriminated, the firing frequency profiles appeared very
similar to the fully analyzed steps.
Data analysis
MEAN FREQUENCIES.
For each unit, the mean interspike interval in a single locomotor burst
was calculated (interspike intervals 10 ms and
200 ms were omitted)
using Linux-based custom software and SigmaPlot 3.0. The individual
means for each step then were summed and averaged together [each unit
had an average of 16 ± 10.5 steps (mean ± SD)]. The
resulting mean interspike interval value for a given unit then was used
to calculate the mean firing-frequency value. Table
1 displays the group averages for units
in the various muscle and motor-unit groups. The mean firing-frequency
values for the initial doublet/triplets also were calculated from the mean interspike interval values and expressed as means ± SD.
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CORRELATION BETWEEN ACTIVITY OF SINGLE MOTOR UNITS AND
CORRESPONDING GROSS-EMG.
In 12 units (5 LG, 5 SOL, 2 TA), the amplitude of the rectified and
smoothed gross-EMG signal (30 Hz low-pass digitally filtered, 0°-phase shift) was plotted against the firing frequency of the single unit (Hoffer et al. 1987). This was done to test
if the firing frequency of a single motor unit was modulated in a
similar manner to the activity of the entire motoneuron pool, as
represented by the amplitude of the gross-EMG signal. The equation
Rate(t) =
*EMG(t) + b (where
t = time,
= the slope of the regression line,
and b = the horizontal offset) was fitted to the data,
using both Matlab custom-written software and Sigma Plot 3.0 (note that we did not force the regression line through the origin) (cf. Hoffer et al. 1987
). The coefficient of determination,
r2 (r = correlation
coefficient) was calculated to obtain a measure of the total variance
in motor-unit discharge that could be accounted for by fluctuations in
the gross-EMG profile (Hoffer et al. 1987
). The mean
absolute error (MAE) with respect to the regression line also was
calculated to determine the error in predicting frequency from the
preceding linear EMG relation. Initial doublets/triplets and other
frequencies >200 Hz and <5 Hz were omitted to remove outlying points
for the regression analysis.
CORRELATION BETWEEN ACTIVITY OF SIMULTANEOUSLY RECORDED MOTOR-UNIT PAIRS. The firing rate profiles of motor units that were discriminated from a single micro-EMG record also were compared to examine if the activity of motor units located close to one another were modulated in a similar manner. Three motor-unit pairs located in the superficial layer of the LG muscle were analyzed. The firing rate profiles of each unit in a pair were compared at similar points in time by fitting a fifth-order polynomial to the individual frequency profiles (Sigma Plot 3.0). Frequency values from the two polynomials were sampled every 25 ms and plotted against one another (see Fig. 8). Finally, a straight line was fitted to these data points and r2 values calculated, as described in the preceding text.
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RESULTS |
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In total, the activation profiles of 21 motor units were analyzed during locomotion (see Table 1 for summary). Two main groups of motor units were found in the extensor muscles MG/LG and SOL. One group was recruited at the beginning of a locomotor burst and was classified as "early-stance phase-activated units" and the second group was recruited later in the locomotor burst and was defined as "mid-late-stance phase-activated units." Two groups of TA motor units also were recorded, one type that fired only two or three MUAPs in a single locomotor burst (doublet/triplet only) and a second type that fired longer trains of MUAPs.
"Fast" mid-late-stance phase-activated MG/LG units
The majority of MG and LG units (10/12) that were discriminable
from the raw micro-EMG records were recruited either mid-way through or
at the end of the stance phase as the hindlimb pushed on the ground to
propel the rat forward. These units will be referred to as
mid-late-stance phase units. An example is shown in Fig. 1 where a large amplitude unit in the
single-unit recording (micro-EMG; 3rd trace) was recruited
halfway through the locomotor burst. The firing frequency profile of
the unit is shown in the bottom trace. A striking feature of
the mid-late-stance phase MG/LG units was the high mean firing rates
reached during walking. In the two steps in Fig. 1, the mean rate was
100 Hz (means for all steps, excluding doublets, ranged from 54 to
86 Hz, Table 1). For comparison, the firing rates of neonatal rat
motoneurons during transmitter-induced locomotion only reach 5-10 Hz
(Bertrand et al. 1998
; Hochman and Schmidt
1998
; MacLean et al. 1997
). The mid-late-stance
phase units fired briefly during a locomotor burst, with an average of
16 spikes per step (see 1st 5 MG/LG units in Fig. 5A for
individual values). The number of MUAPs fired related poorly to the
duration of the corresponding gross-EMG burst (Fig. 5B),
with r2 values ranging from 0.12 to
0.31.
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An unexpected finding, as demonstrated in Fig. 1, was the high
occurrence of very short interspike intervals (10 ms) of the first
two or three MUAPs. These initial doublets or triplets occurred in all
mid-late-stance phase recruited MG/LG units and in 82% of all steps
analyzed (87/106 steps, see Table 1 for MG and LG group values).
Initial doublets/triplets were present at all speeds of walking, and
thus data from the various walking speeds were grouped together. Even
when a unit fired two discrete bursts in a single step (3/12 units),
each burst often would start with an initial doublet (e.g., step 1, Fig. 1). Discrete bursts were defined as discharges separated by
200
ms (Hennig and Lømo 1985
).
The mid-late-stance phase units were most likely fast twitch motor
units as judged by their high mean firing rates and phasic discharge
pattern (Hennig and Lømo 1985). In addition, the sample of MG and LG units probably was biased toward large, fast units considering >90% of MG/LG units are fast-twitch (Gillespie et al. 1987
) and tend to have large-amplitude signals that are
easier to discriminate throughout the locomotor burst.
"Fast" early-stance phase-activated LG units
The unit in Fig. 2 was one of two LG
units that were recruited at the beginning of the stance phase
(referred to as early-stance phase units). These units displayed
activation properties that were intermediate to the mid-late-stance
phase MG/LG units and the SOL units described in the following text. As
seen in Fig. 2, the early stance phase units also commenced firing with
initial doublets (in 46 and 70% of all steps, Table 1) and fired more MUAPs per step cycle than other LG units (see * for these 2 units, Fig.
5A) despite their lower frequency of discharge (see Table 1). Note that in some of the step cycles in Fig. 2, changes in rate by
as much as 50% occurred within a single locomotor burst (e.g., last 2 step cycles). The second early-stance phase unit that was recorded (not
shown) was distinct from all other MG/LG units recorded in that the
number of MUAPs fired increased linearly as a function of the duration
of the corresponding gross-EMG activity (see unit , Fig.
5B).
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"Slow" early-stance phase-activated SOL units
Motor-unit activity recorded from the "slow" soleus muscle
showed very different activation patterns during walking as compared with the MG/LG motor units. In general, the frequency profiles of the
SOL units were more rounded in shape, and their activity was more
related to the corresponding gross-EMG signal (Fig.
3) (see also Fig. 9 in Eken
1998). Even though the firing rates were the lowest of all
extensor motor units recorded (25-31 Hz, Table 1), the SOL units fired
more MUAPs per locomotor burst than the mid-late-stance phase MG/LG
units (26 on average, see Fig. 5A for individual values),
for step cycles of similar duration. The SOL units also differed from
the MG/LG units in that the number of MUAPs increased linearly with the
duration of the corresponding gross-EMG activity (Fig. 5C),
with r2 values ranging from 0.72 to
0.91.
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None of the slow soleus units had initial interspike intervals that were <10 ms, but in two units the initial frequencies were, on occasion, twice that of the mean frequency in the locomotor burst (e.g., step 1, Fig. 3A). We also have classified this overshoot in frequency as an initial doublet, and it occurred in only 18% of the steps cycles for these two units (8/44 steps). In a large number of steps (50%), however, SOL units were recruited with frequencies above or close to the peak rate in the locomotor burst (e.g., steps 2 and 3 in Fig. 3A). A more gradual recruitment pattern was seen in the remainder of the steps (32% e.g., steps 1 and 3 for a different unit in Fig. 3B); however, initial frequencies rarely fell below 20 Hz.
These SOL units were considered to be slow-twitch, fatigue-resistant
(S) due to the high percentage of slow twitch muscle fibers in the SOL
muscle (90%) (Gillespie et al. 1987
) and because of
their low mean firing rates and ability to fire tonically for several
minutes (see Eken 1998
for recordings of these units
during static posture). The remaining SOL units have been shown to be fast, fatigue-resistant (FR) (Chamberlain and Lewis
1989
), and the SOL unit described in the next section may fall
into this category.
"Fast" SOL unit
There was one SOL unit that had activation properties similar to
the early-stance phase LG units described in the preceding text. This
unit was recruited at high levels of gross-EMG activity and had a
higher mean firing rate (45 Hz, Table 1) and a greater interval to
interval variability in firing rate than the other "slow" SOL
units. Unlike the slow SOL units, this unit fired initial doublets with
interspike intervals that were 10 ms. The proportion of initial
doublets (46% of all steps) and relationship between the number of
MUAPs to the gross-EMG duration (see unit
, Fig. 5C) were
similar to the early stance phase LG unit described above (unit
in
Fig. 5B). This characteristic firing profile suggested that
the recorded SOL unit was of the fast, FR type (Eken
1998
; Hennig and Lømo 1985
).
"Fast" TA flexor units
In contrast to the extensor units, the flexor motor units
(n = 5) fired very briefly when activated with an
average of 4.3 MUAPs per step cycle (see Fig. 5A for
individual unit means). In almost every step (99%, Table 1), the TA
units were recruited with a doublet or triplet (Fig.
4). In one unit, two to three very
rapid MUAPs were fired per step with an average initial interspike interval of only 2.4 ± 1.1 ms (417 Hz, n = 47 steps). The mean firing rates of the TA motor units were the highest of
all units investigated (see Table 1), ranging from 82 to
109 Hz (excluding initial doublets). The firing rates within a
locomotor burst were quite variable (Fig. 4), but a relative undershoot
in firing was often seen after the doublet/triplet (Fig.
6Bvii). The TA units recorded during walking were most
likely fast due to the firing properties described in the preceding
text and because 94% of unit types in the TA muscle are fast-twitch
(Tötösy de Zepetnek et al. 1992
). In
addition, the micro-EMG electrodes were implanted into the superficial
layers of the TA muscle where there is a preponderance of FF unit types
(Tötösy de Zepetnek et al. 1992
).
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Initial doublets and triplets
When comparing the initial activation patterns of motor units from the various muscles, it turned out that there was a positive relationship between the mean firing rate of the first two MUAPs and the mean firing rate during the sustained phase of a locomotor burst. For example, the group mean firing rates (i.e., the average of the individual unit means for a particular muscle or motor-unit group) for the first two MUAPs were TA 286 Hz > MG/LG 238 Hz > fast SOL 161 Hz > slow SOL 68 Hz (see Fig. 6A for individual values). Correspondingly, the group mean firing rates were TA 97 Hz > MG/LG 69 Hz > fast SOL 45 Hz > slow SOL 28 Hz (Table 1). An r2 value of 0.6 was obtained from the linear regression when the individual mean firing rates of 13 units were plotted against the corresponding mean initial firing rates (not shown).
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A variety of initial doublet and triplet firing patterns were recorded in the various units. Figure 6B gives examples of the four general patterns observed. A common occurrence, however, was that the interspike interval after the initial doublet/triplet was the longest in the spike train, resulting in a frequency that "undershot" all the others in the train (Fig. 6B, iii, iv, vi, and viii). A "rebound" in firing often would occur one or two intervals after the frequency undershoot (arrows, Fig. 6B, ii, iii, iv, vi, vii, and viii). Two patterns of triplet firing were observed, one in which the second interspike interval was longer than the first, but still under 10 ms (see triplet decrement: 6B, v and vi) and the second in which the second interspike interval was even shorter than the first, resulting in an initial increase in firing frequency ("triplet increment", 6B, vii and viii). Motor units with the highest mean firing rates (see preceding text) exhibited the highest percentage of initial triplets, i.e., TA units 30% (33/111 steps) > MG/LG units 18% (25/136 steps) > fast SOL unit 0.03% (1/33 steps) > slow SOL units 0% (0/65 steps), with an r2 value of 0.93 for this linear relation.
Relationship between single motor-unit firing frequency and rectified gross-EMG amplitude
Similar to that shown for cat motor units (Hoffer et al.
1987), the firing profiles of the slow rat SOL units roughly
followed the envelope of their corresponding rectified and smoothed
gross-EMG signals. This suggested that the firing frequency of a single unit was modulated in a similar manner to the activity of the remaining
motoneuron pool as represented by the amplitude of the gross-EMG
signal. Figure 7A, left,
demonstrates this for one of the slow SOL units where the firing
frequency profile of the unit was superimposed over the corresponding
rectified and smoothed gross-EMG profile. When these two parameters
were plotted against each other and a straight line was fit through the
data (Fig. 7A, right), a relatively positive correlation
emerged in comparison with the MG/LG units described below. The
r2 values obtained from the fitted lines for the
slow SOL units ranged from 0.14 to 0.35 (n = 4 units).
The MAE with respect to the fitted line also was calculated for the
slow SOL units to determine the average error in predicting frequency
from the linear EMG relation (see METHODS). The MAE values
ranged from 6.3 to 7.1 Hz,
16-18% of the modulation depth of the
units during stepping, which was 40 Hz on average (modulation
depth = maximum rate
minimum rate, excluding initial
doublet/triplets).
|
In comparison, the firing profiles of the mid-late-stance phase MG/LG
units related poorly to the corresponding gross-EMG signal, with
r2 values ranging from 0.0015 to 0.010 and MAEs from 20.2 to 31.0 Hz (see representative example of an LG unit
in Fig. 7B). The interval-to-interval variability in the
discharge rates of these units was greater than the slow SOL units, and
this may account for the poorer correlation to the gross-EMG signals.
Another contributing factor may be that because the MG/LG muscles are
compartmentalized (DeRuiter et al. 1996; English
and Letbetter 1982
), the gross-EMG signals probably were not
good indicators of overall motoneuron pool activity, i.e., different
compartments of the MG/LG motoneuron pools may have received different
drives from the locomotor networks. Because the gross-EMG electrodes
most likely were recording activity from these different compartments,
the correlation to a single motor unit in one compartment may have been
poor, unlike that for the more homogeneous SOL motoneuron pool
(Chamberlain and Lewis 1989
; Gillespie et al.
1987
).
To compare the rate modulation of motor units within a
single LG muscle compartment, the activation profiles of
pairs of motor units discriminated from a single micro-EMG record were
examined. We wanted to investigate if the firing rates of
simultaneously activated units were modulated in a parallel fashion and
thus responding in a similar manner to a common locomotor drive, unlike that seen for motoneurons during MLR-induced walking, which display flat frequency profiles (Brownstone et al. 1992).
Activity from a pair of LG motor units discriminated from a single
micro-EMG record (and thus probably from the same muscle compartment)
is shown in Fig. 8. A representative step
cycle is shown in Fig. 8A, which clearly demonstrates that
the firing profiles of both units were similarly modulated. The two
rate profiles were fitted with a fifth-order polynomial to obtain a
smoothed rate profile (see Fig. 8A,
, unit A and
, unit
B). When the smoothed rate of one unit was plotted against the smoothed
rate of the other at similar points in time, a positive linear
relationship emerged (see Fig. 8B,
r2 value of regression line = 0.62, n = 6 steps). A similar parallel rate modulation
was seen in two other motor-unit pairs that were discriminated from the
same micro-EMG electrodes, with r2
values of 0.53 and 0.71 from the linear regression.
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![]() |
DISCUSSION |
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Differences in firing properties between fast and slow motor units during locomotion
A consistent finding from this study was that motor units often
would jump to a relatively high-frequency of discharge when recruited.
Only the fast motor units (i.e., MG/LG, fast SOL and TA units),
however, initiated firing with a doublet or triplet. These fast motor
units therefore displayed similar recruitment patterns to the hindlimb
motor units recorded during MLR-induced locomotion (Brownstone
et al. 1992; Tansey and Botterman 1996
; Zajac and Young 1980
). The slow SOL units, on the other
hand, did not initiate firing with a doublet or triplet but had
frequencies that were above or close to the mean rate in a locomotor
burst, similar to the anterior thigh motor units recorded during
locomotion in the intact cat (Hoffer et al. 1987
). From
our data, therefore it appears that the firing of initial
doublets/triplets was dependent on the type of motor unit recorded
(i.e., fast vs. slow) rather than from the type of locomotion studied
(i.e., self-initiated vs. MLR-induced).
Likewise, the presence or lack of doublets/triplets was probably not a
result of different locomotor drives to the different muscles (i.e., an
abrupt synaptic drive to MG/LG and TA motoneuron pools compared with a
more gradually increasing drive to the SOL motoneuron pool). This was
supported by the fact that during gradual muscle stretch
(Gorassini et al. 1999), only units with very high mean
rates of discharge (>40 Hz and thus probably fast motor units) (Hennig and Lømo 1985
) would initiate firing with a
doublet, whereas units with mean firing rates of <40 Hz (and thus
probably slow motor units) did not. It is not known why motor units
jump to such high frequencies of discharge at recruitment. The
activation of intrinsic conductances at recruitment may be one
possibility and this is discussed in Initial
doublets/triplets.
Another difference between the slow and fast motor units observed in this study was that the slow motor units (i.e., early stance phase SOL) were activated as a more homogeneous group during locomotion in contrast to the MG/LG units. This was evident in the linear relationship that existed between the number of MUAPs fired and the duration of the corresponding gross-EMG for the SOL but not the mid-late-stance phase MG/LG motor units. In addition, only the firing rate profiles of the slow SOL units followed the envelope of their gross-EMGs, indicating that these units were being modulated in a similar manner by a common locomotor drive.
The dissociation of activity of the mid-late-stance phase MG/LG units
from the gross-EMG profiles may have resulted from a compartmentalized
recruitment of the MG/LG motoneuron pool. For example, anatomic and
physiological compartmentalization of the MG/LG muscle has been shown
in both cats and rats with FR unit types occupying deeper parts of the
muscle and FF unit types in the more superficial layers
(DeRuiter et al. 1996; English and Letbetter
1982
; Vanden Noven et al. 1994
). The majority of
MG/LG units recorded in this study were most likely from the
superficial compartment because the micro-EMG electrodes were inserted
very close to the surface of the muscle belly. These potentially large and fast units were activated preferentially midway through or toward
the end of the stance phase and may be activated especially to provide
the force needed to propel the animal forward at the end of stance. The
delayed recruitment of these motor units may have resulted from a
separate drive from the rhythm-generating networks in the spinal cord
(rather than from an orderly recruitment in response to a gradually
increasing central drive) because the recruitment of the
mid-late-stance phase MG/LG units could occur as the corresponding
gross-EMG activity (recorded in deeper muscle compartments) was
declining (e.g., Figs. 2 and 7B).
Rate modulation during locomotion
The parallel modulation between the firing rate profiles of the
SOL units and their corresponding gross-EMG activity also provides
further evidence that the firing rates of motor units can be modulated
during a locomotor burst in contrast to the poor rate modulation of
hindlimb motor units that occurs during MLR-induced locomotion in the
cat (Brownstone et al. 1992; Severin et al. 1967
; Zajac and Young 1980
). Further evidence
was provided by the parallel rate modulation of pairs of simultaneously
recorded motor units within a single LG muscle compartment. Perhaps the lack of rate modulation of motoneuron activity that occurs during MLR
stimulation in the cat results from a rate limiting of motoneuron discharge due to maximal synaptic excitation (Binder et al.
1996
). Alternatively, changes in intrinsic properties that
increase motoneuron excitability and repetitive firing (Krawitz
et al. 1997
) may also contribute given that firing frequencies
tend to be higher during MLR locomotion (40-50 Hz) (Brownstone
et al. 1992
) compared with intact walking (25-35 Hz)
(Hoffer et al. 1987
). In conclusion, it appears from
this and other studies (see also DeSerres et al. 1995
;
Eken 1998
; Hoffer et al. 1987
) that rate
modulation of motor-unit activity occurs during natural, self-initiated walking.
Initial doublets/triplets
The recruitment of the mid-late-stance phase MG/LG motor units
with doublets/triplets most likely was a result of intrinsic conductances activated at recruitment, rather than from an abrupt increase in excitatory synaptic drive. Evidence to support this comes
from observations in which initial doublets/triplets have been shown to
occur without abrupt increases in the corresponding gross-EMG activity
or in the firing rates of other concurrently active motor units
(Gorassini et al. 1999). One possible intrinsic mechanism may be the activation of voltage-dependent, noninactivating plateau potentials considering it recently has been shown that when
plateau potentials are activated near the threshold for action potential generation, the plateau can produce an abrupt increase in the
rate of rise of the membrane potential at the time of recruitment, resulting in high initial firing rates (Bennett et al.
1998
).
The added depolarization produced by the plateau at recruitment may
have acted like an abrupt current injection through a microelectrode,
which is known to cause doublets and triplets (Granit et al.
1963a). Furthermore in cat motoneurons it was shown that when
sodium spikes were inactivated with QX314, an overshoot in membrane
potential that lasted for
80 ms emerged when a plateau initially was
activated (Bennett et al. 1998
; Brownstone et al. 1994
). Thus two or three MUAPs could have been fired on the
crest of this overshoot, resulting in the initial doublets/triplets, provided that the motor units could fire fast enough. Taking this latter point into consideration, it is interesting that initial doublets (and especially initial triplets) were more prevalent in units
that were able to fire two to three MUAPs within this short time period
(e.g., TA and late-stance phase MG/LG units), i.e., their firing rates
were high enough to "sample" the underlying membrane potential. The
SOL units on the other hand only reached maximal initial rates of ~60
Hz, and perhaps they were unable to follow the fast transients in
membrane potential. Alternatively, the doublets/triplets may have been
produced from a postinhibitory rebound (Bertrand and Cazalets
1998
), from the activation of a low-threshold
Ca2+ spike (Llinás and Yarom
1981
; Russo and Hounsgaard 1996
), or from extra
spikes arising out of an afterdepolarization (Fulton and Walton
1986
: Granit et al. 1963b
; see also discussion
in Eken 1998
), which has been shown to be enhanced
during plateau potential activation (Bennett et al.
1998
).
Tension-frequency relation of single motor units during locomotion
Regardless of the mechanism(s) involved, a high initial firing
rate of a motoneuron has been shown to produce a more rapid onset in
motor unit force (i.e., catch property) (Burke et al. 1970). A rapid onset in force is essential during walking,
especially for flexor muscles in the rat. For example, during steady
walking (1 s/step), a rat must dorsiflex its ankle within
50 ms to
clear the foot at the beginning of the swing phase due to the very
flexed posture of both the hip and knee joints (Gruner et al.
1980
). Thus it is not surprising that a large number of motor
units are recruited with initial frequencies that are two to five times greater than the mean rate of discharge in a locomotor burst. In
addition, these doublets/triplets often are followed by an undershoot
in firing frequency, and this pattern of discharge has been shown to
optimize the speed at which maximum force is reached in a motor unit
(Stein and Parmiggiani 1979
). The gross-EMG patterns of
a variety of hindlimb muscles that show an initial peak followed by a
rapid decline may result from this pattern of motor-unit recruitment
(Gorassini et al. 1994
; Gruner et al. 1980
).
After recruitment, the mean firing rates of the various units during a
locomotor burst fell close to the top of their corresponding tension-frequency curves [the tension-frequency relation was taken from Fig. 3 in Henning and Lømo (1985) for the slow SOL
and fast extensor digitorum longus (EDL) muscles]. For example, in
this study, when the nerve to the SOL muscle was stimulated at a rate of 30 Hz (i.e., the mean rate at which SOL units fired during walking,
see Table 1), the muscle reached nearly 90% of its maximum tetanic
force at its optimal length. Similarly, in the EDL muscle, which has a
similar fast motor-unit type composition as the TA muscle, stimulation
of the nerve at 100 Hz (i.e., mean rate of TA units during walking, see
Table 1) also produced 90% of the maximum tetantic force. Stimulation
rates of 60-80 Hz (i.e., firing rates of MG/LG units during walking)
produced 60-80% maximum tetanic force in the EDL muscle and probably
would produce even higher forces in the LG/MG muscle because it
contains more FR unit types than EDL (DeRuiter et al.
1996
).
From these indirect estimates it appears that during walking, motor units (on average) fire at discharge rates that produce forces that are close to their maximum levels. These estimates were taken from experiments in which the muscle lengths were set to produce maximum force. Although motor units may not always be at their optimal lengths throughout a locomotor burst, this may be compensated for by the high rates at recruitment, as discussed in the preceding text.
Summary
The data from this study provide the first extensive description
of single motor-unit activity from different muscles during unrestrained walking in the conscious adult rat. It is important to
establish this considering the increasing number of studies looking at
the control of motoneuron activity during transmitter-induced locomotion in the neonatal rat. For example, neonatal motoneurons do
not initiate firing with initial doublet/triplets, and they fire at
frequencies that are 5-10 times slower than adult motoneurons during
unrestrained walking (Bertrand et al. 1998;
Hochman and Schmidt 1998
; MacLean et al.
1997
).
In summary, evidence was provided to suggest that the activation patterns of single motor units during unrestrained locomotion in the rat are shaped both by the intrinsic properties of the parent motoneuron and from synaptic inputs from the rhythm-generating networks of the spinal cord. The integration of these two effects on motor-unit recruitment and firing aids in the shaping of the final force output of the respective muscle fibers to produce the appropriate activation patterns required for locomotion.
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ACKNOWLEDGMENTS |
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The authors acknowledge the excellent technical assistance of I. Kjaer for construction of the micro-EMG wires, L. Grøndahl for help in animal care, and E. Gudbrandsen for building electrical equipment.
This research was supported by the Danish Medical Research Council and the Novo Nordisk Foundation. D. J. Bennett was supported by the Alberta Heritage Foundation for Medical Research and M. Gorassini by the Danish Research Academy. T. Eken was supported by the Norwegian Research Council for Science and the Humanities. O. Kiehn is a Hallas Møller Associate Professor supported by the Novo Foundation.
Present address of D. J. Bennett: Division of Neuroscience, Faculty of Rehabilitation Medicine, University of Alberta, Edmonton, Alberta T6G 2G4, Canada
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FOOTNOTES |
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Present address and address for reprint requests: M. Gorassini, 3-48 Corbett Hall, Faculty of Rehabilitation Medicine, University of Alberta, Edmonton, Alberta T6G 2G4, Canada.
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
Received 21 July 1999; accepted in final form 8 December 1999.
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REFERENCES |
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