Long-lasting recovery of locomotor function in chronic spinal rat following chronic combined pharmacological stimulation of serotonergic receptors with 8-OHDPAT and quipazine.

In chronic spinal rats, long-term stimulation of 5-HT receptors with quipazine or 8-OHDPAT by means of daily injection, promotes robust locomotor recovery. The question of a possible potentiation between treatments when applied together was addressed. Daily injections of both 8-OHDPAT and quipazine, were performed for a month in spinal animals. Animals were placed on a treadmill and the bipedal hindlimb locomotion was tested. Motor performances (behavioural test) and locomotor parameters (EMG and kinematic) were analysed weekly during the treatment. Furthermore, the locomotor performances were evaluated during two supplemental months following the end of the treatment. Our results suggest that association of both agonists induced long-lasting positive effects on locomotor function. Motor performances were significantly better after combined injection of both drugs than when the agonists were used separately. But, the most significant and new result is that the locomotor scores did not decrease during the weeks that followed the end of the treatment. These results suggests a long-lasting and 5-HT-dependent reorganisation of spinal networks.

5-HT1A receptors are involved in short- and long-term processes responsible for 5-HT-induced locomotor function recovery in chronic spinal rat.

After thoracic spinal cord transection, a paraplegic syndrome occurs. Previous data showed that an acute administration of a 5-HT2 agonist (quipazine) could promote motor function recovery in spinal rats. However, continuous subdural perfusion of quipazine via an osmotic pump over 1 month proved to be more effective. The present study was designed to investigate the possible involvement of 5-HT1A receptors in such recovery. Motor performances and locomotor parameters were analysed in spinal animals receiving daily, for 1 month, a dose of the 5-HT1A agonist 8-OHDPAT. The results were compared to those obtained in spinal rats receiving either a placebo or quipazine in the same conditions. Using daily injections instead of continuous perfusion of either receptor agonist to spinal animals allowed characterization of short- and long-term consequences of pharmacological stimulation of 5-HT1A and 5-HT2 receptors on motor function recovery. Our data demonstrate that daily injections of a 5-HT1A agonist induce long-term, cumulative, positive effects on motor function recovery, as assessed by the improvement in the walking parameters observed before the 'day-test' injection. This might involve use-dependent processes depending on a chronic and/or repetitive stimulation of the spinal network for locomotion in relation to 5-HT receptor activation. A further improvement in the motor parameters, transiently observed following the injection, suggests a more direct action of 5-HT1A and 5-HT2 receptor activation on spinal neurons involved in motor pattern generation.

Locomotor recovery in the chronic spinal rat: effects of long-term treatment with a 5-HT2 agonist.

A complete transection of the spinal cord at a low thoracic level induces a paraplegic syndrome that is accompanied by a loss of spinal cord serotonin content. Former experimental data suggest that the central pattern generator for locomotion, located in the lumbar segments of the spinal cord, might be able to generate rhythmic motor outputs (similar to automatic walking under certain circumstances) involving exteroceptive stimulations and activation of serotonergic receptors. In the present study, we investigated the effects of a chronic treatment using a serotonergic agonist, delivered continuously to the sublesionned spinal cord, and its effect on motor function recovery. The data obtained from behavioural, kinematic and electromyographic measurements suggest that the chronic stimulation of 5-HT2 type receptors allows motor function recovery. Behavioural measurements show a clear improvement in motor performances when compared to spinal animals (confirmed by kinematic observations): alternating steps and foot placement is recovered in these animals. However, electromyographic data demonstrate that the pattern of activation of the muscles is only restored partially.

Serotonin-induced activation of the network for locomotion in adult spinal rats.

The biogenic amine serotonin has been described in the literature as a powerful modulator of the spinal central pattern generator for locomotion. In the present study, we tested whether administration of serotonin or its agonist quipazine could restore motor activity in a model of paraplegia. One to three weeks after a complete transection of the spinal cord at a low thoracic level, rats were given either intrathecal injections of serotonin (5 mM, 15 microL) or intraperitoneal injections of quipazine (400-600 microg/kg). Both treatments allowed recovery of locomotor activity on a treadmill in response to tail pinching. As compared with the activity elicited before treatment, the locomotor activity produced by spinal animals was characterised by longer locomotor sequences with a larger number of successive steps, better body support, better interlimb coordination, and a higher amplitude of electromyographic bursts. These results suggest that serotonergic drugs could be used for the recovery of motor functions after lesions of the spinal cord.

Modulation of the spinal network for locomotion by substance P in the neonatal rat.

The tachykinin substance P (SP) is present in the ventral and medial area of the lumbar spinal cord. Its localisation suggests that it could modulate the spinal network for locomotion. We have investigated its effects on motor outputs by applying SP, in vitro, to the lumbosacral segments of an isolated spinal cord of new-born rats. SP was applied to the lumbosacral spinal cord either on a quiescent preparation or during episodes of fictive locomotion induced by N-methyl-D,L-aspartate. When applied on quiescent preparations, SP induced a slow rhythmic activity (period >30 s). During fictive locomotion, SP increased both the locomotor frequency and the duration of the bursts of cyclic activity. Furthermore, SP stabilised the locomotor rhythm. These results demonstrate that SP is able to modulate both the "clock" and the pattern generator for locomotion.

[Can transplantation of neurons facilitate motor recovery in paraplegics? Study of an animal model]. / Les transplantations de neurones peuvent-elles faciliter la récupération motrice chez les paraplégiques? Etude d'un modèle animal.

This review strives forward at least two goals. First, to take from the literature the arguments demonstrating that hindlimbs locomotion is controlled by a spinal network of neurons (the so-called Central Pattern Generator for locomotion--CPG) known to be able to generate locomotor activity independently of the control of supraspinal nervous structures, as it is after thoracic lesions of the spinal cord. The principles of work of the CPG and its intrinsic possibilities to adapt its working are reviewed. Special reference is made to the various ways used during experiments to activate the CPG in spinal animals or clinical practice in paraplegic men: training to walk, electrical stimulations, pharmacological stimulations. Second, to show, from our own results, obtained from the study of an animal model of paraplegia, the adult spinal rat, how it could be possible to take advantage of the autonomy of the CPG, with special reference to its sensibility to monoamines, to obtain locomotor recovery in hindlimbs after section of the thoracic spinal cord, by means of transplantation of noradrenergic and/or serotonergic embryonic neurons in the lumbo-sacral spinal cord. Section of the spinal cord at a thoracic level results in an important locomotor deficit in hindlimbs, likely linked to degeneration of monoaminergic terminals in the lumbar enlargement. In the adult spinal rat, sub-lesional injection of a suspension of embryonic nervous cells, taken from either locus coeruleus or raphe sites, leads to reinnervation of the lumbar enlargement with monoaminergic terminals. Despite the fact that connections with supraspinal structures are not reestablished, transplanted animals recover progressively a posture convenient for locomotion. The hindlimbs, which are in an extended position a few days after the lesion, become progressively flexed and able to support the body weight. This evolution does not appear in spinal but non transplanted animals. But, the main point is that transplanted animals develop, within the few weeks that follow transplantation, a good-quality locomotor activity in hindlimbs which had no equivalent in spinal but non transplanted animals. The reality of a lumbar CPG for locomotion and the efficacy of pharmacological treatments and training to walk, to elicit recovery of stepping, are discussed in man, in connection with the relevance to use transplantation of monoaminergic nervous cells in the spinal cord of paraplegics.

Central modulation of stretch receptor neurons during fictive locomotion in lamprey.

1. In lamprey, stretch receptor neurons (SRNs), also referred to as edge cells, are located along the lateral margin of the spinal cord. They sense the lateral movements occurring in each swim cycle during locomotion. The isolated lamprey spinal cord in vitro was used to investigate the activity of SRNs during fictive locomotion induced by bath-applied N-methyl-D-aspartate (NMDA). Intracellular recordings with potassium acetate filled electrodes showed that 63% of SRNs had a clear locomotor-related modulation of their membrane potential. 2. Of the modulated SRNs, two-thirds had periods of alternating excitation and inhibition occurring during the ipsilateral and the contralateral ventral root bursts, respectively. The phasic hyperpolarization could be reversed into a depolarizing phase after the injection of chloride ions into the cells; this revealed a chloride-dependent synaptic drive. The remaining modulated SRNs were inhibited phasically during ipsilateral motor activity. 3. Experiments with barriers partitioning the recording chamber with the spinal cord into three pools, allowed an inactivation of the locomotor networks within one pool by washing out NMDA from the pool in which the SRN was recorded. This resulted in a marked reduction, but not an abolishment, of the amplitude of the membrane potential oscillations. Both the excitatory and the inhibitory phases were reduced, resulting from removal of input from inhibitory and excitatory interneurons projecting from the adjacent pools. If the glycine receptor antagonist strychnine (1 microM) was applied in one pool, the phasic hyperpolarizing phase disappeared without affecting the excitatory phase. 4. Bath application of the gamma-aminobutyric acid (GABA)A receptor antagonist, bicuculline (50-100 microM) blocked the spontaneous large unitary inhibitory postsynaptic potentials, which occurred without a clear phasic pattern. Bicuculline had no significant effect on the peak to peak amplitude of the locomotor-related membrane potential oscillations. The inhibition in SRNs therefore has a dual origin: glycinergic interneurons provide phasic inhibition, while the GABA system can exert a tonic inhibition via GABAA receptors. 5. These data show that, in addition to the stretch-evoked excitation, which SRNs receive during each locomotor cycle, most of them also receive excitation from the central pattern generator network during the ipsilateral contraction, which may ensure a maintained high level of sensitivity to stretch during the shortening phase of the locomotor cycle. This arrangement is analogous to the efferent control of muscle spindles exerted by gamma-motoneurons in mammals, which as a rule are coactivated with alpha-motoneurons to the same muscle (alpha-gamma linkage).

Voltage-dependent block of NMDA responses by 5-HT agonists in ventral spinal cord neurones.

1. Modulation by 5-hydroxytryptamine receptor agonists of the NMDA responses of ventral spinal cord neurones was studied by use of the whole-cell patch-clamp technique. 2. In a Mg-free solution containing tetrodotoxin and glycine, 5-hydroxytryptamine (5-HT, 10-100 microM) reduced the NMDA response, the block increasing with hyperpolarization. Kainate responses were little affected. 3. Some classical agonists of 5-HT receptors induced similar blocking effects. At 10 microM, both a selective agonist of 5-HT2 receptors, (+/-)-2,5-dimethoxy-4 iodo amphetamine (DOI), and a selective agonist of some 5-HT1 receptors, (+/-)-8-hydroxy-2(n-dipropyl amino) tetralin (8-OH-DPAT), induced pronounced blocking effects, of 48% and 33% respectively at -100 mV, whereas another 5-HT1 agonist, 5-carboxamidotryptamine (5-CT) was ineffective. At 100 microM, 5-methoxytryptamine (5-MeOT) induced a complete block of the NMDA responses recorded at -100 mV. The order of potency was: 5-MeOT congruent to DOI > 8-OH-DPAT > 5-HT > 5-CT. 4. Neither spiperone nor ketanserin (1 microM) prevented the blocking effect of 5-HT or DOI. 5. Prolonged preincubations with 5-HT did not block the response if NMDA was applied without 5-HT. When 5-HT agonists were applied both by preincubation and with NMDA, the degree of block increased during the NMDA application. 6. Lowering the NMDA concentration (from 100 to 20 microM) slightly decreased the blocking effect of 5-MeOT. 7. External Mg2+ ions (1 mM) also reduced the blocking effects of 5-HT and 5-MeOT. 8. The blocking effects described appear to be independent of classical 5-HT receptors. Their voltage-dependence suggests a mechanism of open channel block consistent with all the results obtained.

Neurotensin-induced modulation of spinal neurons and fictive locomotion in the lamprey.

1. Neurotensin containing interneurons are present in the spinal cord of both mammalian and nonmammalian vertebrates, but as yet little is known about their functional role. In this study we examine the effect of neurotensin on spinal cells and on the central pattern generator for locomotion in the lamprey spinal cord. 2. Bath application of neurotensin (10(-8) to 10(-6) M) slowed down the fictive locomotor activity induced by the glutamate agonist N-methyl-D-aspartate in the isolated spinal cord. The duration of the bursts of activity in the ventral roots increased in proportion to the increase of the locomotor cycle duration. 3. Intracellular recordings from grey matter neurons and intraspinal stretch receptors neurons showed that neurotensin induced a depolarization [4.4 +/- 0.5 (SE) mV, n = 19]. This depolarization could still be obtained after a blockade of voltage-sensitive sodium channels with tetrodotoxin (1.5 +/- 0.5 mV; n = 6), and after removal of calcium (2.8 +/- 0.4 mV; n = 5). Moreover no consistent change occurred in the fast and slow phase of the afterhyperpolarization (AHP) both of which are carried by potassium currents.

In vitro, proctolin and serotonin induced modulations of the abdominal motor system activities in crayfish.

An in vitro thoraco-abdominal preparation of the crayfish (Procambarus clarkii) ventral nerve cord was used to study the sites of action and the effects of proctolin and serotonin on the nervous activities of the two abdominal motor systems, namely the swimmeret and the abdominal positioning systems. In this preparation spontaneous motor activity was recorded corresponding to continuous rhythmic bursts in the swimmeret motor nerves and tonic discharge of motoneurons in both abdominal extensor and flexor motor nerves. Proctolin applied on the abdominal ganglia elicited bursts of spikes in the flexor motor nerve which were able to disturb and even stop the swimmeret activity. Increasing concentrations of serotonin applied on the thoracic ganglia were able, first, to increase the period durations of the swimmeret bursting activity and, second, to stop it. In this last condition, continuous swimmeret activity resumed by application of proctolin on the abdominal ganglia although period durations stayed slightly longer than in control. The actions of serotonin and proctolin on the two abdominal motor systems were discussed in terms of modulations and interactions between central neuronal networks and behaviors.

Dopamine and motor activity in the lobster Homarus gammarus.

Motor activity similar to agonistic behaviour is obtained after dopamine (DA) injection in lobster. Specially vigorous swimmeret beatings are observed and can be compared to the 'in vitro' motor activity elicited by DA superfusion of the isolated abdominal nervous system. DA-immunoreactive neurons stained by monoclonal antibodies in abdominal ganglia may be involved in swimmeret activation during the agonistic behavior.