Neuroscience Letters, 111 (1990) 287-291 Elsevier Scientific Publishers Ireland Ltd.
287
NSL 06779
Cervical interneurones oligosynaptically excited from primary afferents and rhythmically active during forelimb fictive locomotion in the cat Mitsuyo Hishinuma and Takashi Yamaguchi Institute of Basic Medical Sciences, Tsukuba University, Tsukuba, Ibaraki (Japan) (Received 1 November 1989; Revised version received 14 December 1989; Accepted 15 December 1989)
Key words: Interneuron; Cervical enlargement; Fictive locomotion; Forelimb; Decerebrate cat; Oligosynaptic reflex pathway In C6-C7, activity of neurones oligosynaptically excited from primary afferents was examined during fictive locomotion. Two kinds of neurones were found: neurones with activity modulated and with activity not modulated. The minimum linkage of the pathway to modulated neurones was mono- and disynaptic from muscle and cutaneous afferents, respectively, while non-modulated neurones were monosynaptically excited from both afferents. Modulated neurones were mainly located in the ventral part of the dorsal horn and adjoining intermediate region, non-modulated ones located in the dorsal horn. Modulated neurones were found near motor nuclei as well. Active periods of modulated neurones were widely distributed in the step cycle.
A previous report [4] has demonstrated that short-latency excitation of elbow flexor motoneurones from primary afferents (radial nerves) is rhythmically facilitated in the flexor phase of fictive locomotion (fictive locomotion is efferent discharge of stepping evoked in immobilized animals [1]). The facilitation may occur at interneuronal level of the reflex pathways, because during fictive locomotion, long-latency excitation of the same motoneurone is differentially affected and motoneuronal inputresistances almost remain constant [3]. To examine this possibility, activity of interneurones receiving mono- or disynaptic excitation from the radial nerves was examined during fictive locomotion. We sought neurones in the C6 and C7 segments, where elbow flexor motoneurone pools are located [12]. Seven adult cats were precollicularly decerebrated under halothane anaesthesia, spinalized at the lowest thoracic level (Tl3), after having recovered from anaesthesia, immobilized with pancuronium bromide and artificially respired. Various nerves were dissected for stimulation and recording. Forelimb fictive locomotion, identified by alternating discharges of elbow extensor and flexor nerves with a frequency of Correspondence: T. Yamaguchi, Institute of Basic Medical Sciences, Tsukuba University, Tsukuba, Ibaraki 305, Japan. 0304-3940/90/$ 03.50 © 1990 Elsevier Scientific Publishers Ireland Ltd.
288
about 1 Hz, was induced by stimulation (a tungsten microelectrode was used: 0.5 ms duration, 33 Hz, lOq00/~A strength) of the midbrain locomotor region (P2.0-2.5, L4.0-4.5 and H0.0 to - 0 . 5 in Horsley-Clarke coordinates) [1]. C6-C7 neurones were sought with a glass capillary microelectrode (2 M potassium citrate), while dissected nerves were stimulated with strength of 10 times threshold (10 x 73 of nerve volleys. The recording electrode was left in situ, the tracks recovered in frozen transverse sections, and then the site of recording along the tracks interpolated from accurate depth readings during the penetration. Ninety-five interneurones were examined during fictive locomotion, 50 neurones showing rhythmic modulation of activity (modulated neurones) and 45 not (nonmodulated neurones). Active periods of modulated neurones distributed variously in the step cycle (see below). Twenty-eight out of the 50 modulated neurones and 31 out of the 45 non-modulated neurones responded oligosynaptically to nerve stimulation. Fig. 1 illustrates a modulated neurone receiving oligosynaptic input. The raw record (A) with raster display and histogram (B) shows that the neurone discharges bursts mainly at transition from the flexor to extensor phase. In this neurone, mono-
A
B
4~-
TMe
TMe
Br ii i,~ ....
i
Li,. ]L~.,L .L__,~
LI I1~1tllJ
JI
[ ii iiiiii i i ~i L i i IHLIIH iI iIII1~1 ~l
'
UNIT 100 msec
400 m s e c
C a
e
Bi
10T
DR
b
Br
c
f
SR
g
TLo
M
d
D
An
h
U
_•4mV
O~c6
10 rnsec a - h 2 msec
i - k
Fig. l. C6 interneurone with oligosynaptic input from forelimb afferents. A: activity of the neurone during fictive locomotion. Upper two traces, rectified nerve discharges of elbow extensor (TMe) and flexor (Br). The third, unit activity through an active filter (band-pass with - 3 dB points at 80 Hz and 5 kHz). B: raster diagram and histogram obtained from a part shown by dashed line in A. C: convergence of peripheral input. In each panel, the upper trace, unit recording without the active filter; the lower, cord dorsum potential. (i k), early response shown with fast sweep of (e g). D: the recording site. TMe, medial head of triceps brachii; TLo, long head of triceps brachii: An anconeus; Br, brachialis: Bi, biceps brachii; DR, deep radial: SR, superficial radial: M, medianus; U, ulnaris.
289
synaptic excitation is evoked from the deep radial (DR) and median (M) nerves; the latencies are 0.8 and 1.1 ms, respectively. Cutaneous afferent volleys (the superficial radial nerves, SR) produces disynaptic excitation, latency 1.9 ms. Excitation from the brachialis (Br) and ulnaris (U) nerves is rather late, latencies 5.3 and 4.0 ms, respectively. Further late excitation is evoked from all the nerves tested, latencies more than 9 ms. The neurone was located in the dorsolateral part of the intermediate region
(D). Fig. 2 summarizes neurones oligosynaptically excited from deep radial (DRexcited neurones) and superficial radial nerves (SR-excited neurones). Out of the 28 modulated neurones with oligosynaptic input, there were 9 DR-excited and 7 SRexcited neurones. Two neurones were excited from both nerves, unit 3,6 of DRexcited corresponding to unit 1,3 of SR-excited. Synaptic convergence was observed with the other nerves as well (input in A a and Ba). Degree of convergence was more extensive in SR-excited, modulated neurones than in DR-excited, modulated ones. The minimum linkage of the pathways from D R and SR to the modulated neurones was monosynaptic and disynaptic, respectively. Note that neurones excited monosynaptically from SR do not show modulation of activity. The low threshold afferents in both nerves seemed responsible for the effects, because in most cases threshold of the effects was low, less than 2 x T in 5 out of 7 DR-excited, modulated neurones A
a
Ext.
B
Input
OR-excited units
a SR-excited units
-
.
4
Input
Ext
. . . . .
.
o
~
.
.
-
•
_
.
.
.
.
o o
.
~ o
7 ~ ' ~ ~ - rK~ " ~ qL- ~
~
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o
b MonosynaDtically-excited C Dlsynaptlcally-exclted
o
elo
o
b Monosynaptlcally-exclted
.o0
modulate~7
C
Dtsynapticarly-excited
.o0u %
,.0 a
Fig. 2. Summary of C 6 - - C 7 n e u r o n e s responding oligosynaptically to stimulation o f DR (A) and SR (B). a: active period during fictive locomotion and input convergence of modulated neurones (filled circle, monosynaptic excitation; open circle, disynaptic), b,c: location of neurones with monosynaptic input and those with disynaptic one (in each cross-section, modulated neurones shown on the right, non-modulated on the left). Modulated neurones numbered in b and c correspond to those with the same number in a. The neurone of Fig. 1 is numbered 6 in A and 3 in B.
290 tested and 2 out of 3 SR-excited, modulated neurones tested, in the remaining cases threshold was between 2 × T and 3 x T. Active periods of modulated neurones were variously distributed in the step cycle (A a and Ba). Between DR-excited and SR-excited neurones no significant differences could be found in their distribution. The DR-excited, modulated neurones were located in the dorsal part of the intermediate region and near motoneurone pools. The SR-excited, modulated neurones were located in the ventrolateral part of the dorsal horn and in the adjoining intermediate region, the location being just dorsal to and overlapping with that of the DR-excited, modulated neurones. In the overlapping zone neurones with synaptic convergence from D R and SR were found. The DR-excited, non-modulated neurones and SR-excited, non-modulated ones were located mainly in the dorsal part of the dorsal horn (Ab.c and Bb,c). In the hindlimb, some reflex excitations of motoneurones are rhythmically modulated during fictive locomotion [2, 5, 6--9] and interneuronal mechanisms for the modulation have been supposed [2, 5, 6, 9]. Further, neurones interposed in an excitatory pathway to hindlimb motoneurones seem to be rhythmically active [10]. This present study suggests similar possibility in the forelimb segments. In the ventral region of the dorsal horn, where modulated neurones with oligosynaptic inputs from peripheral nerves were mainly located, interneurones antidromically excited from the flexor motor nuclei are found, and the last-order neurones are activated in the flexor phase of fictive locomotion (see the subsequent paper [1 1]). Thus, some of DR- and/ or SR-excited neurones with rhythmic burst in the flexor phase in this region may be interposed in the reflex pathways previously reported [4]. The synaptic linkage of the reflex pathways to flexor motoneurones is disynaptic from DR and trisynaptic from SR. Consistently, modulated neurones were monosynaptically and disynaptically excited from the respective nerves. It is noteworthy that neurones monosynaptically excited from SR were not rhythmically activated, suggesting that the facilitation of the SR-evoked trisynaptic excitation of flexor motoneurones can not occur at the first-order neurone in the pathway. The authors thank Mrs. Akiko Ohgami for her excellent technical assistance.
1 Amemiya,M. and Yamaguchi, T., Fictive locomotion of the forelimb evoked by stimulation of the mesencephalic locomotorregion in the decerebrate cat, Neurosci. Lett., 50 (1984) 91-96. 2 Andersson, O., Forssberg, H., Grillner, S. and Lindquist, M., Phasic gain control of the transmission in cutaneous reflex pathways to motoneurones during 'fictive' locomotion, Brain Res., 149 (1978) 503-507. 3 Hishinuma, M. and Yamaguchi, T., Facilitation of reflex pathways onto forelimb flexor motoneurons by the central locomotorpattern generator in cats, J. Physiol. Soc. Jpn., 50 (1988) 444. 4 Hishinuma, M. and Yamaguchi, T., Modulation of reflex responses during fictive locomotionin the forelimb of the cat, Brain Res., 482 (1989) 184-188. 5 Schmidt, B.J., Meyers, D.E.R., Fleshman, J.W., Tokuriki, M. and Burke, R.E., Phasic modulation of short latency cutaneous excitation in flexor digitorum longus motoneurons during fictive locomotion, Exp. Brain Res., 71 (1988) 568-578.
291 6 Schomburg, E.D. and Behrends, H.B., The possibility of phase-dependent monosynaptic and polysynaptic la excitation to homonymous motoneurones during fictive locomotion, Brain Res., 143 (1978) 533 537. 7 Schomburg, E.D. and Behrends, H.B., Phasic control of the transmission in the excitatory and inhibitory relfex pathways from cutaneous afferents to alpha-motoneurons during flctive locomotion in cats, Neurosci. Lett., 8 (1978) 277-282. 8 Schomburg, E.D., Behrends, H.B., and Steffens, H., Alteration of transmission in segmental pathways from flexor reflex afferents (FRA) to alpha-motoneurones during spinal locomotor activity, Neurosci. Lett., Suppl. 1 (1978) S103. 9 Shefchyk, S.J. and Jordan, L.M., Excitatory and inhibitory postsynaptic potentials in ~-motoneurons produced during fictive locomotion by stimulation of the mesencephalic locomotor region, J. Neurophysiol., 53 (1985) 1345-1355. 10 Shefchyk, S., McCrea, D., Kriellaars, D., Jordan, L. and Fortier, P., Activity of midlumbar group II interneurons during brainstem evoked flctive locomotion in the mesencephalic cat, Soc. Neurosci. Abstr., 14 (1988) 180. 11 Terakado, Y. and Yamaguchi, T., Last-order interneurones controlling activity of elbow flexor motoneurones during forelimb fictive locomotion in the cat, Neurosci. Lett., 111 (1990) 292-296. 12 Thomas, R.C. and Wilson, V.J., Recurrent interactions between motoneurones of known location in the cervical cord of the cat, J. Neurophysiol., 30 (1967) 661~74.
287
NSL 06779
Cervical interneurones oligosynaptically excited from primary afferents and rhythmically active during forelimb fictive locomotion in the cat Mitsuyo Hishinuma and Takashi Yamaguchi Institute of Basic Medical Sciences, Tsukuba University, Tsukuba, Ibaraki (Japan) (Received 1 November 1989; Revised version received 14 December 1989; Accepted 15 December 1989)
Key words: Interneuron; Cervical enlargement; Fictive locomotion; Forelimb; Decerebrate cat; Oligosynaptic reflex pathway In C6-C7, activity of neurones oligosynaptically excited from primary afferents was examined during fictive locomotion. Two kinds of neurones were found: neurones with activity modulated and with activity not modulated. The minimum linkage of the pathway to modulated neurones was mono- and disynaptic from muscle and cutaneous afferents, respectively, while non-modulated neurones were monosynaptically excited from both afferents. Modulated neurones were mainly located in the ventral part of the dorsal horn and adjoining intermediate region, non-modulated ones located in the dorsal horn. Modulated neurones were found near motor nuclei as well. Active periods of modulated neurones were widely distributed in the step cycle.
A previous report [4] has demonstrated that short-latency excitation of elbow flexor motoneurones from primary afferents (radial nerves) is rhythmically facilitated in the flexor phase of fictive locomotion (fictive locomotion is efferent discharge of stepping evoked in immobilized animals [1]). The facilitation may occur at interneuronal level of the reflex pathways, because during fictive locomotion, long-latency excitation of the same motoneurone is differentially affected and motoneuronal inputresistances almost remain constant [3]. To examine this possibility, activity of interneurones receiving mono- or disynaptic excitation from the radial nerves was examined during fictive locomotion. We sought neurones in the C6 and C7 segments, where elbow flexor motoneurone pools are located [12]. Seven adult cats were precollicularly decerebrated under halothane anaesthesia, spinalized at the lowest thoracic level (Tl3), after having recovered from anaesthesia, immobilized with pancuronium bromide and artificially respired. Various nerves were dissected for stimulation and recording. Forelimb fictive locomotion, identified by alternating discharges of elbow extensor and flexor nerves with a frequency of Correspondence: T. Yamaguchi, Institute of Basic Medical Sciences, Tsukuba University, Tsukuba, Ibaraki 305, Japan. 0304-3940/90/$ 03.50 © 1990 Elsevier Scientific Publishers Ireland Ltd.
288
about 1 Hz, was induced by stimulation (a tungsten microelectrode was used: 0.5 ms duration, 33 Hz, lOq00/~A strength) of the midbrain locomotor region (P2.0-2.5, L4.0-4.5 and H0.0 to - 0 . 5 in Horsley-Clarke coordinates) [1]. C6-C7 neurones were sought with a glass capillary microelectrode (2 M potassium citrate), while dissected nerves were stimulated with strength of 10 times threshold (10 x 73 of nerve volleys. The recording electrode was left in situ, the tracks recovered in frozen transverse sections, and then the site of recording along the tracks interpolated from accurate depth readings during the penetration. Ninety-five interneurones were examined during fictive locomotion, 50 neurones showing rhythmic modulation of activity (modulated neurones) and 45 not (nonmodulated neurones). Active periods of modulated neurones distributed variously in the step cycle (see below). Twenty-eight out of the 50 modulated neurones and 31 out of the 45 non-modulated neurones responded oligosynaptically to nerve stimulation. Fig. 1 illustrates a modulated neurone receiving oligosynaptic input. The raw record (A) with raster display and histogram (B) shows that the neurone discharges bursts mainly at transition from the flexor to extensor phase. In this neurone, mono-
A
B
4~-
TMe
TMe
Br ii i,~ ....
i
Li,. ]L~.,L .L__,~
LI I1~1tllJ
JI
[ ii iiiiii i i ~i L i i IHLIIH iI iIII1~1 ~l
'
UNIT 100 msec
400 m s e c
C a
e
Bi
10T
DR
b
Br
c
f
SR
g
TLo
M
d
D
An
h
U
_•4mV
O~c6
10 rnsec a - h 2 msec
i - k
Fig. l. C6 interneurone with oligosynaptic input from forelimb afferents. A: activity of the neurone during fictive locomotion. Upper two traces, rectified nerve discharges of elbow extensor (TMe) and flexor (Br). The third, unit activity through an active filter (band-pass with - 3 dB points at 80 Hz and 5 kHz). B: raster diagram and histogram obtained from a part shown by dashed line in A. C: convergence of peripheral input. In each panel, the upper trace, unit recording without the active filter; the lower, cord dorsum potential. (i k), early response shown with fast sweep of (e g). D: the recording site. TMe, medial head of triceps brachii; TLo, long head of triceps brachii: An anconeus; Br, brachialis: Bi, biceps brachii; DR, deep radial: SR, superficial radial: M, medianus; U, ulnaris.
289
synaptic excitation is evoked from the deep radial (DR) and median (M) nerves; the latencies are 0.8 and 1.1 ms, respectively. Cutaneous afferent volleys (the superficial radial nerves, SR) produces disynaptic excitation, latency 1.9 ms. Excitation from the brachialis (Br) and ulnaris (U) nerves is rather late, latencies 5.3 and 4.0 ms, respectively. Further late excitation is evoked from all the nerves tested, latencies more than 9 ms. The neurone was located in the dorsolateral part of the intermediate region
(D). Fig. 2 summarizes neurones oligosynaptically excited from deep radial (DRexcited neurones) and superficial radial nerves (SR-excited neurones). Out of the 28 modulated neurones with oligosynaptic input, there were 9 DR-excited and 7 SRexcited neurones. Two neurones were excited from both nerves, unit 3,6 of DRexcited corresponding to unit 1,3 of SR-excited. Synaptic convergence was observed with the other nerves as well (input in A a and Ba). Degree of convergence was more extensive in SR-excited, modulated neurones than in DR-excited, modulated ones. The minimum linkage of the pathways from D R and SR to the modulated neurones was monosynaptic and disynaptic, respectively. Note that neurones excited monosynaptically from SR do not show modulation of activity. The low threshold afferents in both nerves seemed responsible for the effects, because in most cases threshold of the effects was low, less than 2 x T in 5 out of 7 DR-excited, modulated neurones A
a
Ext.
B
Input
OR-excited units
a SR-excited units
-
.
4
Input
Ext
. . . . .
.
o
~
.
.
-
•
_
.
.
.
.
o o
.
~ o
7 ~ ' ~ ~ - rK~ " ~ qL- ~
~
~
~
o
b MonosynaDtically-excited C Dlsynaptlcally-exclted
o
elo
o
b Monosynaptlcally-exclted
.o0
modulate~7
C
Dtsynapticarly-excited
.o0u %
,.0 a
Fig. 2. Summary of C 6 - - C 7 n e u r o n e s responding oligosynaptically to stimulation o f DR (A) and SR (B). a: active period during fictive locomotion and input convergence of modulated neurones (filled circle, monosynaptic excitation; open circle, disynaptic), b,c: location of neurones with monosynaptic input and those with disynaptic one (in each cross-section, modulated neurones shown on the right, non-modulated on the left). Modulated neurones numbered in b and c correspond to those with the same number in a. The neurone of Fig. 1 is numbered 6 in A and 3 in B.
290 tested and 2 out of 3 SR-excited, modulated neurones tested, in the remaining cases threshold was between 2 × T and 3 x T. Active periods of modulated neurones were variously distributed in the step cycle (A a and Ba). Between DR-excited and SR-excited neurones no significant differences could be found in their distribution. The DR-excited, modulated neurones were located in the dorsal part of the intermediate region and near motoneurone pools. The SR-excited, modulated neurones were located in the ventrolateral part of the dorsal horn and in the adjoining intermediate region, the location being just dorsal to and overlapping with that of the DR-excited, modulated neurones. In the overlapping zone neurones with synaptic convergence from D R and SR were found. The DR-excited, non-modulated neurones and SR-excited, non-modulated ones were located mainly in the dorsal part of the dorsal horn (Ab.c and Bb,c). In the hindlimb, some reflex excitations of motoneurones are rhythmically modulated during fictive locomotion [2, 5, 6--9] and interneuronal mechanisms for the modulation have been supposed [2, 5, 6, 9]. Further, neurones interposed in an excitatory pathway to hindlimb motoneurones seem to be rhythmically active [10]. This present study suggests similar possibility in the forelimb segments. In the ventral region of the dorsal horn, where modulated neurones with oligosynaptic inputs from peripheral nerves were mainly located, interneurones antidromically excited from the flexor motor nuclei are found, and the last-order neurones are activated in the flexor phase of fictive locomotion (see the subsequent paper [1 1]). Thus, some of DR- and/ or SR-excited neurones with rhythmic burst in the flexor phase in this region may be interposed in the reflex pathways previously reported [4]. The synaptic linkage of the reflex pathways to flexor motoneurones is disynaptic from DR and trisynaptic from SR. Consistently, modulated neurones were monosynaptically and disynaptically excited from the respective nerves. It is noteworthy that neurones monosynaptically excited from SR were not rhythmically activated, suggesting that the facilitation of the SR-evoked trisynaptic excitation of flexor motoneurones can not occur at the first-order neurone in the pathway. The authors thank Mrs. Akiko Ohgami for her excellent technical assistance.
1 Amemiya,M. and Yamaguchi, T., Fictive locomotion of the forelimb evoked by stimulation of the mesencephalic locomotorregion in the decerebrate cat, Neurosci. Lett., 50 (1984) 91-96. 2 Andersson, O., Forssberg, H., Grillner, S. and Lindquist, M., Phasic gain control of the transmission in cutaneous reflex pathways to motoneurones during 'fictive' locomotion, Brain Res., 149 (1978) 503-507. 3 Hishinuma, M. and Yamaguchi, T., Facilitation of reflex pathways onto forelimb flexor motoneurons by the central locomotorpattern generator in cats, J. Physiol. Soc. Jpn., 50 (1988) 444. 4 Hishinuma, M. and Yamaguchi, T., Modulation of reflex responses during fictive locomotionin the forelimb of the cat, Brain Res., 482 (1989) 184-188. 5 Schmidt, B.J., Meyers, D.E.R., Fleshman, J.W., Tokuriki, M. and Burke, R.E., Phasic modulation of short latency cutaneous excitation in flexor digitorum longus motoneurons during fictive locomotion, Exp. Brain Res., 71 (1988) 568-578.
291 6 Schomburg, E.D. and Behrends, H.B., The possibility of phase-dependent monosynaptic and polysynaptic la excitation to homonymous motoneurones during fictive locomotion, Brain Res., 143 (1978) 533 537. 7 Schomburg, E.D. and Behrends, H.B., Phasic control of the transmission in the excitatory and inhibitory relfex pathways from cutaneous afferents to alpha-motoneurons during flctive locomotion in cats, Neurosci. Lett., 8 (1978) 277-282. 8 Schomburg, E.D., Behrends, H.B., and Steffens, H., Alteration of transmission in segmental pathways from flexor reflex afferents (FRA) to alpha-motoneurones during spinal locomotor activity, Neurosci. Lett., Suppl. 1 (1978) S103. 9 Shefchyk, S.J. and Jordan, L.M., Excitatory and inhibitory postsynaptic potentials in ~-motoneurons produced during fictive locomotion by stimulation of the mesencephalic locomotor region, J. Neurophysiol., 53 (1985) 1345-1355. 10 Shefchyk, S., McCrea, D., Kriellaars, D., Jordan, L. and Fortier, P., Activity of midlumbar group II interneurons during brainstem evoked flctive locomotion in the mesencephalic cat, Soc. Neurosci. Abstr., 14 (1988) 180. 11 Terakado, Y. and Yamaguchi, T., Last-order interneurones controlling activity of elbow flexor motoneurones during forelimb fictive locomotion in the cat, Neurosci. Lett., 111 (1990) 292-296. 12 Thomas, R.C. and Wilson, V.J., Recurrent interactions between motoneurones of known location in the cervical cord of the cat, J. Neurophysiol., 30 (1967) 661~74.
