Messages conveyed by descending tracts during scratching in the cat. II. Activity of rubrospinal neurons

Brain Research, 159 (1978) 111-123 © Elsevier/North-Holland Biomedical Press

111

MESSAGES CONVEYED BY DESCENDING TRACTS DURING S C R A T C H I N G IN T H E CAT. II. ACTIVITY OF R U B R O S P I N A L N E U R O N S

YU. I. ARSHAVSKY, G. N. ORLOVSKY, G. A. PAVLOVA and C. PERRET* Institute of Problems of Information Transmission, Academy o f Sciences, Moscow and Moscow State University, Moscow (U.S.S.R.)

(Accepted April 6th, 1978)

SUMMARY (1) The activity of rubrospinal (RS) neurons giving axons to the lumbosacral spinal cord was recorded during actual and fictitious a scratching in thalamic cats. (2) During both actual and fictitious scratching, the discharge frequency of many RS neurons was rhythmically modulated. Different neurons were active in different parts of the scratch cycle, but most neurons were active in the flexor phase. (3) The discharge frequency within the bursts during fictitious scratching was, on the average, equal to that during actual scratching. Immobilization usually resulted only in a small displacement of the burst position in the scratch cycle. Therefore, rhythmical modulation of RS neurons is determined mainly by central mechanisms and not by a rhythmical sensory input. (4) In decerebellate cats, the overwhelming majority of RS neurons had no rhythmical modulation. Very weak modulation was found only in a few neurons. (5) Transection of the ventral spinocerebellar tract (VSCT) resulted in considerable reduction or complete cessation of rhythmical modulation in RS neurons during fictitious scratching. On the contrary, transection of the spino-reticulocerebellar pathway (SRCP) resulted in just a small decrease of modulation. Therefore, of the two pathways (the VSCT and SRCP) transmitting messages about intraspinal processes to the cerebellum during scratching ~,~, the VSCT is of major importance for modulating RS neurons.

INTRODUCTION The activity of RS neurons giving axons to the lumbosacral spinal cord was * Present address: Laboratory of Comparative Neurophysiology, Paris University VI, Paris, France.

112 studied earlier during locomotion in thalamic 2z and decorticate "7 cats. The firing rate of most RS neurons was found to be rhythmically modulated in relation to the step cycle. In thalamic cats, modulation of RS neurons disappeared almost completely after ablation of the cerebellum. Therefore, in this preparation, the rhythmical drive to RS neurons is provided mainly by the cerebellum, which is known to be one of the main inputs to the red nucleus (for literature see refs. 10, 16, 21 and 28). Rhythmical modulation of RS neurons was observed not only during actual but also during fictitious locomotion in curarized cats 27. The efferent pattern of fictitious locomotion is similar to that of the actual one, but any rhythmical afferent signals are absent. Therefore, in this case, rhythmical modulation of RS neurons is determined by central factors. The study of actual locomotion also presented evidence indicating the central origin of the rhythmical activity of RS neurons 93. The present paper is concerned with the activity of RS neurons during actual and fictitious scratching in thalamic cats. This study has confirmed the bulk of the results obtained in the study of locomotion. We found that rhythmical modulation of RS neurons is determined mainly by signals coming from the central spinal mechanism of scratching, and that rhythmical influences of the central spinal mechanism upon RS neurons are mediated by the cerebellum, it was found earlier that signals from the central spinal mechanism of scratching reached the cerebellum through the ventral spinocerebellar tract (VSCT) 3 and the spino-reticulocerebellar pathway (SRCP) relayed through the lateral reticular nucleus e. In the present study we found that signals conveyed by the VSCT played a crucial role in modulating RS neurons. Corresponding results (i.e. the central origin of rhythmical modulation and the crucial role of the cerebellum and the VSCT in modulation) were obtained for the vestibulospinal tract and described in the first paper of this series 4. METHODS The methods were described in previous papersZ,4,s, 23. Briefly, a thalamic cat, fixed in a stereotaxic device, performed scratching movements with its hindlimb when the pinna was stimulated. To facilitate the scratch reflex, tubocurarine chloride was applied to the dorsal surface of the spinal cord at the CI levelg, x2. In experiments with fictitious scratching, animals were immobilized with Flaxedil. RS neurons were recorded extracellularly with platinum microelectrodes 13. Since RS axons are known to cross the midline (for literature see ref. 21), RS neurons were identified by their antidromic activation from the contralateral lateral funiculus of the spinal cord at the L1 level (Fig. 3G and F). Scratching was evoked on the side contralateral to a neuron (except for a few cases which are indicated in Results). Activity of RS neurons was recorded simultaneously with either the electromyogram (EMG) of m. gastrocnemius lateralis (during actual scratching) or the electroneurogram (ENG) of the corresponding nerve (during fictitious scratching).

113 RESULTS

(a) Activity of RS neurons during actual scratching The activity of many RS neurons was modulated during scratching: it increased in one part of the scratch cycle and decreased (usually to zero) in another part. We cannot estimate the relative number of modulated neurons, since usually we managed to find these neurons only during the first 2-4 h of an experiment, then modulation became weaker and, finally, disappeared. A similar phenomenon was observed in vestibulospinal neurons 4. It seemed likely that, at least in some cases, the absence of modulation was accounted for by a bad functional state of the red nucleus or the cerebellum, which was demonstrated (see Results, (c)) to play a crucial role in modulating RS neurons. We shall therefore concern ourselves with a description of modulated neurons exclusively. In 8 experiments with actual scratching, we recorded 35 modulated neurons. Most (82 ~ ) had resting discharge with frequencies of 1-30 pps; the mean value (with silent neurons included) was 10.1 pps (REST in Fig. 2B). After the beginning of pinna stimulation, the firing rate in most (67 ~o) of RS neurons increased, sometimes up to 70-80 pps. The mean value of the firing rate just before the beginning of rhythmical hindlimb movements was 29.8 pps. Examples of the activity of RS neurons during actual scratching are presented in Figs. 1 and 6A, C and E. The neurons fired in bursts separated by intervals of silence. Discharge frequency within the bursts was 30-100 pps. A few neurons generated two bursts per cycle. Fig. 2A shows the phase distribution of 35 RS neurons (for method see ref. 4); the flexor ('long') and extensor ('short') phases of the scratch cycle (L and S) s are also indicated. One can see that bursts of activity of RS neurons are distributed over the scratch cycle, but most neurons are active in the L-phase. These peculiarities of the phase distribution are also reflected in Fig. 2B, where the frequency curve of the 'average' RS neuron is presented (for methods see ref. 4). The curve characterizes, to some extent, the overall activity of the RS tract, i.e. the 'flow' of impulses coming through the RS tract to the lumbosacral spinal cord at various phases of the scratch cycle. The 'flow' is minimum at the beginning of the L-phase and maximum in the middle of the L-phase. However, the difference between the maximum and minimum

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Fig. 2. Relationship between the activity of RS neurons and the phase of the cycle during actual scratching. A: the phase distribution of neurons. The burst position of each RS neuron is plotted in relation to the normalized scratch cycle; the cycle is considered to start with the termination of the extensor burst. The flexor (L) and estensor (S) phases of the cycle are marked. The neurons are put in order according to the onset of their activity. This order is broken for only a few neurons which begin firing at the very end of the cycle but are shown in the upper part of the graph. B: the discharge frequency of the 'average' RS neuron (pulses/sec) as a function of the phase of the cycle (for method see ref. 4). REST: the average frequency of the resting discharge. The time calibration is presented for the 'average' scratch cycle. a c t i v i t i e s is n o t g r e a t . T h i s is a c c o u n t e d f o r b y t h e f a c t t h a t d i f f e r e n t R S n e u r o n s a r e a c t i v e i n d i f f e r e n t p h a s e s o f t h e cycle.

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115 value (with silent neurons included)was ll.7 pps (REST in Fig. 4B), i.e., it was practically equal to that in non-immobilized animals (10.1 pps). As in the case of actual scratching, activity of most ( 7 0 ~ ) RS neurons increased during the latent period of fictitious scratching (as in a neuron in Fig. 3A). The mean frequency just before the beginning of rhythmical generation was 41.9 pps. Examples of the activity of RS neurons during fictitious scratching are presented in Figs. 3A-E, 6B, D and F, 8A, C and E and 9A and C. The neurons fired in bursts, separated by intervals of silence. The discharge frequency in the bursts was usually 30-100 pps. Fig. 3C and D shows activity of the same neuron during weak (C) and intensive (D) scratching. During more intensive scratching, both the discharge frequency and the burst duration increased. Fig. 4A shows the phase distribution of 94 RS neurons. One can see that bursts of activity of RS neurons are distributed over the scratch cycle, but the number of neurons active in the L-phase is somewhat greater than that of neurons active in the Sphase. If 7 RS neurons generating only 1-2 spikes per cycle (the shortest lines in Fig. 4A) are excluded, the rest of the RS neurons may be divided into two groups, according to the positions of their bursts in the cycle. The 'early' group comprises the neurons presented in the upper part of the phase distribution (Fig. 4A). They begin firing rather synchronously in the S-phase and at the beginning of the L-phase. They

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stop firing in the second half of the L-phase, so that the activity of the 'early' group completely disappears by the beginning of the S-phase. Examples of 'early' neurons are presented in Figs. 3A-D, 6B and F, 8A and C and 9C. The 'late' group comprises the neurons presented in the lower part of the phase distribution (Fig. 4A). They are recruited rather evenly during the L-phase (except for the very beginning). They stop firing rather synchronously at the end of the S-phase and at the beginning of the subsequent L-phase. Examples of 'late' neurons are presented in Figs. 3E, 6D, 8E and 9A. The two groups of RS neurons are more distinctly seen in the two-dimension phase distribution (Fig. 5A). In this graph, each neuron is presented by a circle, the abscissa being the phase of the burst onset, the ordinate being the phase of the burst termination. Seven neurons generating 1-2 spikes per cycle are presented by black dots. Two neurons generating two bursts per cycle are presented by two circles corresponding to each burst. One can see that all circles in Fig. 5A fall into one of two groups. The circles situated in the left lower part of the graph correspond to 'early' neurons (51 units) - - those in the right upper part, to 'late' neurons (38 units). From the two-dimension distribution (Fig. 5A) we can more exactly indicate the border between groups in Fig. 4A; the 'early' group is above the arrow - - the 'late' one, under it (except for 3 neurons marked by dots which also belong to the 'late' group). Fig. 4B shows the discharge frequency of the 'average' RS neuron as a function of the phase of the cycle. In Fig. 4C, frequency curves are presented separately for the 'early' group (open circles) and for the 'late' one (filled circles). One can see that the 'early' and 'late' groups are active almost reciprocally. Had they been equal to each other, the overall activity would not have changed during the cycle. But, as the 'early' group is somewhat larger (51 neurons against 38), this results in some periodical modulation of the overall activity of the RS tract with a maximum in the L-phase (Fig. 4B). The comparison of the activity of RS neurons during fictitious scratching with

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Fig. 6. Activity of 3 RS neurons (A, B; C, D and E-G respectively) during actual (A, C and E) and fictitious (B, D, F and G) scratching. The neuron shown in E-G was recorded after immobilizing an animal not only during contralateral scratching (F) but also during ipsilateral one (G). The lower trace in A, C and E is the gastrocnemius EMG - - in B, D, F and G, the ENG.

that during actual scratching shows that rhythmical modulation of the discharge is distinctly expressed in both cases. Most neurons fire in bursts (30-100 pps) separated by periods of silence (cf. Figs. 1 and 3). In both cases, the overall activity of the RS tract is higher in the L-phase as compared to the S-phase (cf. Figs. 2A and 4A as well as 2B and 4B). However, immobilization of animals affects RS neurons: their bursts are somewhat shifted in the cycle, the shift being different for different neurons. This is seen in Fig. 6, which shows the activity of 3 RS neurons recorded firstly during actual scratching (A, C and E) and then during fictitious scratching after immobilizing the animals (B, D and F). In the first neuron, the burst began after the extensor burst when scratching was actual (Fig. 6A) and during the extensor burst when scratching was fictitious (Fig. 6B). In the second neuron, the burst occupied the second half of the Lphase during actual scratching (Fig. 6C) and the whole L-phase during fictitious scratching (Fig. 6D). In the third neuron, the burst was shifted towards the beginning of the cycle after immobilizing the animal (cf. Fig. 6E and F). The effects of immobilizing the animals can also be seen while comparing the two-dimension phase

118 distribution for actual scratching (Fig. 5B) with that for fictitious scratching (Fig. 5A). In non-immobilized animals, in contrast to immobilized ones, one cannot divide RS neurons into the 'early' and 'late' groups. This can be explained by the fact that sensory inflow from the moving limb does not affect burst positions of different RS neurons uniformly, leading to spreading of the 'initial' (centrally determined) phase distribution. Eight RS neurons were recorded not only during fictitious scratching evoked on the side contralateral to a neuron but also during ipsilateral scratching. In the latter case, 6 neurons also exhibited discharge modulation. However, the rhythmical activity was weaker than during contralateral scratching (cf. F and G in Fig. 6).

(c) Activity of RS neurons in decerebellate cats Thirty-two RS neurons were recorded in 7 decerebellate cats (6 neurons during actual scratching and 26 during fictitious scratching). About a half had resting discharge with frequencies of 1-10, sometimes up to 30 pps; the mean value (with silent neurons included) was 4.0 pps, i.e. about 2.5 times lower than in cats with the intact cerebellum. This reduction of the resting discharge may be attributed to the elimination of a tonic excitatory inflow to RS neurons from the nucleus interpositus of the cerebelluml0,Sl,3L During the latent period of scratching, the activity of most RS neurons increased. During scratching (both actual and fictitious) the firing of the overwhelming majority of RS neurons (29 out of 32) was not rhythmically modulated (as in neurons in Fig. 7A-D). In only 3 neurons weak modulation was observed as with the neuron in

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Fig. 7E, which was scarcely more active (on the average) in the L-phase than in the Sphase. The mean frequency for all RS neurons recorded in decerebellate cats during scratching was 13.0 pps.

(d) Effects of partial transections of the spinal cord The data described above show that the cerebellum is essential for rhythmical modulation of RS neurons, the signals coming from the central spinal mechanism of scratching being of crucial importance for modulation. These signals reach the cerebellum through the VSCT 3 and SRCPL To estimate the contribution from each of these pathways to modulation of RS neurons during fictitious scratching, we studied the effects of their transections. VSCT neurons are located in the L4-L6 segments; their axons cross the midline and then ascend on the contralateral side of the spinal cord16,24, zS. Therefore, for the VSCT interruption we performed hemisection of the spinal cord at the L1 level on the side contralateral to scratching (ipsilateral to RS neurons). Fig. 8 shows the activity of 3 RS neurons (two from the 'early' group (A, B and C, D) and one from the 'late' group (E, F)) before (A, C and E) and after (B, D and F) hemisection. In the first and second neurons, modulation disappeared after hemisection altogether, while in the third one it sharply decreased. The SRCP, in contrast to the VSCT, ascends in the ipsilateral lateral funiculus 16,

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during fictitious scratching. Transections were performed on the side ipsilateral to scratching (contralateral to a neuron) at the C5 level, the extent of lesions is shown as black areas on the spinal cord cross-sections. Activity of RS neurons was recorded before (A and C) and after (B and D) lesions. 20,z5. Previously it was shown that, after transection of this funiculus at the C4-C5 level, signals coming through the SRCP did not reach the cerebellum 2. Fig. 9 shows activity of 2 RS neurons (one from the 'late' group (A, B) and another from the 'early' group (C, D)) before (A and C) and after (B and D) this kind of transection. The transection resulted in only minor reduction of rhythmical modulation of RS neurons. This reduction could be accounted for not only by the SRCP interruption but also by some decrease of the intensity of scratching. DISCUSSION

Role of central and peripheral factors in rhythmical modulation of RS neurons The main finding of the present study is that the activity of RS neurons is rhythmically modulated during fictitious scratching, i.e. in the absence of any rhythmical sensory inflow. This modulation is similar to that seen during actual scratching: neurons fire in bursts separated by periods of silence, the discharge frequency within the bursts being approximately the same (on the average) in immobilized and non-immobilized animals. Immobilization usually results only in some displacement of the burst position in the scratch cycle. From these observations one can conclude that (a) rhythmical modulation of RS neurons during actual scratching is determined mainly by signals coming from the central spinal mechanism of scratching, and (b) rhythmical sensory inflow from the moving limb is of minor importance for modulation. The study of the activity of RS neurons during actual and fictitious locomotion had also led to the conclusion that rhythmical modulation of the neurons was determined by signals coming from the central spinal mechanism controlling stepping movementsZ3, 2v. While studying the activity of vestibulospinal and reticulospinal neurons during scratching, we found that their modulation also persisted after immobilizing an animal 4,26. We may thus conclude that rhythmical modulation of neurons of the 3

121 principal descending tracts originating from the brain stem is determined mainly by the signals coming from the central spinal mechanism of scratching, while sensory input is of minor importance for modulation.

Role of cerebellum and different ascending pathways in modulation of RS neurons The present study has demonstrated that rhythmical modulation of RS neurons during both actual and fictitious scratching is almost completely absent in decerebellate cats. This finding means that it is the cerebellum that mediates influences of the central spinal mechanism of scratching upon RS neurons. This means also that direct spinorubral fibers11, 22 are of little importance for modulating RS neurons. Corresponding results were obtained in walking thalamic decerebellate cats: modulation with the rhythm of stepping was almost absent 23. Thus, it can be concluded that, in both types of motor activity, the cerebellum is the main 'modulator' of RS neurons. However one should remember that, in our preparations, the rostral parts of the brain are removed. Rhythmical modulation found in a part of RS neurons during fictitious locomotion of decorticate decerebellate cats z7 could be accounted for by some extracerebellar inputs to the red nucleus which are absent or uneffective in thalamic preparations. The influences upon the red nucleus are exerted by the interposed nucleus which, in turn, is controlled by the intermediate area of the cerebellar cortex 1°,16. This area has the following inputs from the spinal cord: the VSCT, the SRCP, the dorsal spinocerebellar tract (DSCT) and the spino-olivocerebellar pathway (SOCP) 6,7,16,~s, 19,z4,zs. Let us consider the role of each of these inputs for modulating RS neurons. Olivocerebellar neurons were found to be either not active or non-modulated during fictitious scratching 3. Therefore, the SOCP does not participate in modulation of RS neurons during fictitious scratching. It is hardly possible that the behavior ot the SOCP in non-immobilized preparations would be fundamentally changed. Indeed, olivary units were found to have very weak periodical modulation during locomotion of thalamic cats 5. The DSCT is usually assumed to be the main input to the intermediate area of the cerebellar cortex16,18,24, 25. This tract conveys sensory information, mainly that concerning activity of the limb musclesl,24, zS. Therefore, the functional involvement of the DSCT discontinues after immobilizing an animal. Our finding indicates that rhythmical activity of RS neurons changes only slightly after immobilizing an animal. Consequently, the DSCT does not play any significant role in rhythmical modulation of RS neurons during scratching. The data obtained in the study of the activity of RS neurons during actual and fictitious locomotion2a, z7 also suggested that the DSCT was of minor importance for their modulation. The present study has shown that the SRCP, which conveys messages about the activity of the central spinal mechanism of scratching 2, does not play any important role in the control of RS neurons. On the contrary, another pathway carrying signals from the central spinal mechanism of scratching to the cerebellum, the VSCT a, was found to be responsible for rhythmical modulation of RS neurons. Corresponding results were obtained in the study of the vestibulospinal tract 4. The possible reasons

122 for different efficiency o f the V S C T a n d S R C P are discussed in the preceding p a p e r 4. Phases o f activity o f R S neurons The present study has shown t h a t phases o f m a x i m u m activity o f different RS n e u r o n s are distributed over the scratch cycle. The d i s t r i b u t i o n is uneven: m o s t RS

n e u r o n s are active in the flexor phase o f the cycle. This finding c o r r e s p o n d s to the fact t h a t the m a i n effect p r o d u c e d by the RS t r a c t in the spinal c o r d is an increase o f the flexor activity15,17,21, 29. W e have also f o u n d t h a t a c o n s i d e r a b l e n u m b e r o f R S n e u r o n s are active in the extensor phase. This can be c o r r e l a t e d with the fact that there is a fraction o f the RS tract which exerts excitatory influences u p o n extensor m o t o neurons14,15,29. The studies o f the activity o f RS neurons d u r i n g l o c o m o t i o n showed t h a t in t h a l a m i c p r e p a r a t i o n s m o s t RS n e u r o n s were active in the flexor phase o f the step cycle z3 - - in decorticate p r e p a r a t i o n s , s o m e w h a t later 27. These findings, t o g e t h e r with those o f the present study, suggest t h a t the RS t r a c t can exert influences u p o n the spinal c o r d in different phases o f the cycle. The possible functional m e a n i n g o f the m o d u l a t e d activity in descending p a t h w a y s during scratching was discussed in the preceding p a p e r 4.

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