Interaction between metabotropic and ionotropic glutamate receptors regulates neuronal network activity.

Experimental and computational techniques have been used to investigate the group I metabotropic glutamate receptor (mGluR)-mediated increase in the frequency of spinal cord network activity underlying locomotion in the lamprey. Group I mGluR activation potentiated the amplitude of NMDA-induced currents in identified motoneurons and crossed caudally projecting network interneurons. Group I mGluRs also potentiated NMDA-induced calcium responses. This effect was blocked by a group I mGluR-specific antagonist, but not by blockers of protein kinase A, C, or G. The effect of group I mGluRs activation was also tested on NMDA-induced oscillations known to occur during fictive locomotion. Activation of these receptors increased the duration of the plateau phase and decreased the duration of the hyperpolarizing phase. These effects were blocked by a group I mGluR antagonist. To determine its role in the modulation of NMDA-induced oscillations and the locomotor burst frequency, the potentiation of NMDA receptors by mGluRs was simulated using computational techniques. Simulating the interaction between these receptors reproduced the modulation of the plateau and hyperpolarized phases of NMDA-induced oscillations, and the increase in the frequency of the locomotor rhythm. Our results thus show a postsynaptic interaction between group I mGluRs and NMDA receptors in lamprey spinal cord neurons, which can account for the regulation of the locomotor network output by mGluRs.     

A role for slow NMDA receptor-mediated,intrinsic neuronal oscillations in the control of fast fictive swimming in Xenopus laevis larvae

In larvae of the amphibian, Xenopus laevis, spinal neurons which are active during fictive swimming also display tetrodotoxin-resistant membrane potential oscillations following the coactivation of N-methyl-dl -aspartate (NMDA) and 5-hydroxytryptamine (serotonin or 5-HT) receptors ( 1 ;Eur. J. Neurosci., 9, 1473–1482). The oscillations are slow (≈0.5 Hz) compared with swimming (≈7–35 Hz) raising doubt over their contribution to the cycle by cycle depolarizations occurring during swimming. We investigated an alternative: that the intrinsic oscillations modulate swimming activity over many consecutive cycles. Bath application of NMDA induced continuous fictive swimming that differed between embryonic and larval preparations. In 81% of larval preparations (n = 36), there was a slow (approximately every 2 s) rhythmic modulation of ventral root activity in which burst durations and intensities increased as cycle periods decreased. This pattern of activity was enhanced rather than abolished following blockade of glycine and γ-aminobutyric acid (GABA) A receptors and presumably therefore resulted from a periodic increase in the excitation of motor neurons. To determine whether this slow rhythm resulted from intrinsic, 5-HT-dependent membrane potential oscillations, larvae were spinalized to prevent the release of 5-HT from brainstem raphe neurons. The resulting pattern of NMDA-induced activity lacked any slow modulation. The slow modulation could also be enhanced by the bath application of a 5-HT receptor agonist (5-carboxamidotryptamine) and abolished either by the addition of an antagonist (pindobind-5-HT_1A) or by removal of magnesium ions, providing more direct evidence for a contribution of intrinsic oscillations. Thus, the 5-HT-dependent intrinsic oscillations modulate NMDA-induced swimming activity over several consecutive cycles.     

Voltage Oscillations in Xenopus Spinal Cord Neurons: Developmental Onset and Dependence on Co-activation of NMDA and 5HT Receptors

The development of intrinsic, N-methyl-D-aspartate (NMDA) receptor-mediated voltage oscillations and their dependence on co-activation of 5-hydroxytryptamine (5HT) receptors was explored in motor neurons of late embryonic and early larval Xenopus laevis. Under tetrodotoxin, 100 μM NMDA elicited a membrane depolarization of around 20 mV, but did not lead to voltage oscillations. However, following the addition of 2–5 μM 5HT, oscillations were observed in 12% of embryonic and 70% of larval motor neurons. The voltage oscillations depended upon co-activation of NMDA and 5HT receptors since they were curtailed by selectively blocking NMDA receptors with D-2-amino-5-phosphonovaleric acid (APV) or by excluding Mg²⁺ from the experimental saline. 5HT applied in the absence of NMDA also failed to elicit oscillations. Oscillations could be induced by the non-selective 5HT_1a receptor agonist, 5-carboxamidotryptamine (5CT) and both 5HT- and 5CT-induced oscillations were abolished by pindobind-5HT₁, a selective 5HT_1a receptor antagonist. To test whether 5HT enables voltage oscillations by modulating the voltage-dependent block of NMDA channels by Mg²⁺, membrane conductance was monitored under tetrodotoxin. Although 5HT caused membrane hyperpolarization of 4–8 mV, there was little detectable change in conductance. NMDA application caused an approximate 20 mV depolarization and an ‘apparent’ decrease in conductance, presumably due to the conductance pulse bringing the membrane into a voltage region where Mg²⁺ blocks the NMDA ionophore. 5HT further decreased conductance, which we propose is due to its enhancement of the voltage-dependent Mg²⁺ block. When the membrane potential was depolarized by ～20 mV via depolarizing current injection (to mimic the NMDA-induced depolarization), 5HT increased rather than decreased membrane conductance. Furthermore, 5HT did not affect the increase in membrane conductance following NMDA applications in zero Mg²⁺ saline. The results suggest that intrinsic, NMDA receptor-mediated voltage oscillations develop in a brief period after hatching, and that they depend upon the co-activation of 5HT and NMDA receptors. The enabling function of 5HT may involve the facilitation of the voltage-dependent block of the NMDA ionophore by Mg²⁺ through activation of receptors with 5HT_1a-like pharmacology.     

The spinal 5-HT system contributes to the generation of fictive locomotion in lamprey

Activation of NMDA receptors evokes sustained fictive locomotion in the isolated spinal cord of the sea lamprey Petromyzon marinus (P. marinus), but in the river lamprey Lampetra fluviatilis (L. fluviatilis) the ventral root activity is often irregular. A previous study showed that the number of 5-HT immunoreactive fibres, neurones and varicosities are much lower in the spinal cord of L. fluviatilis than in P. marinus. To further analyse the underlying mechanisms, the present study investigated the role of the 5-HT system in stabilising fictive locomotion. In P. marinus a blockade of 5-HT1A receptors by spiperone reversibly increased the frequency and the coefficient of variation. This implies that there is an endogenous release of 5-HT during fictive locomotion that is important for the generation of locomotor activity. In L. fluviatilis bath applied NMDA or -glutamate evoked in most cases irregular activity. An addition of 5-HT (0.5–2 μM) rapidly stabilised the burst generation and led to a sustained fictive locomotion. In a split-bath configuration, NMDA administered to the rostral part of the spinal cord in P. marinus evoked fictive locomotion in both the rostral part and the first few segments of the caudal part. When spiperone was added to the caudal part, the burst activity changed into tonic activity within 10 min. Taken together, these results indicate that activity in the intrinsic 5-HT system in the lamprey spinal locomotor network contributes significantly to the rhythm generation. The quantitative differences with regard to the 5-HT plexus between P. marinus and L. fluviatilis may account for the observed discrepancy between the two species.     

N-methyl-D-aspartate receptor-induced, inherent oscillatory activity in neurons active during fictive locomotion in the lamprey

Bath application of N-methyl-aspartate induces fictive locomotor activity in the isolated spinal cord preparation of the lamprey, as well as TTX-resistant membrane potential oscillations in many individual neurons. This inherent oscillatory activity is shown to depend on a specific activation of N-methyl-D-aspartate (NMDA) receptors. This activation initiates voltage-dependent, magnesium-requiring membrane potential bistability, presumably due to a development of a region of negative slope conductance in the current-voltage relation of the neuron. When sodium ions were removed from the bathing solution, oscillations disappeared, and the membrane potential was maintained at a hyperpolarized level, suggesting that the depolarizing current during the oscillatory cycle is mainly carried by sodium ions. Replacing Ca2+ with Ba2+ also leads to a cessation of oscillatory activity, with the membrane potential remaining at the more depolarized level. This indicates an involvement of a Ca2+-dependent K+ current during the repolarization phase. These findings, together with the voltage dependence, can account for the main characteristics of the NMDA receptor-induced, TTX-resistant membrane potential oscillations. This oscillatory behavior has been demonstrated in motoneurons and in several interneurons including CC interneurons but has not been found in edge cells, dorsal cells, or lateral interneurons. The possibility that inherent oscillatory membrane properties may contribute to the activity pattern during fictive locomotion was investigated in experiments with intracellular current injection in the absence of TTX. The stimulation effects obtained required the presence of magnesium ions and were analogous to the stimulation effects seen during oscillations after TTX blockade. Together with similarities in, for instance, frequency and amplitude between the locomotor oscillatory activity and the TTX-resistant oscillations, the results are compatible with an involvement of inherent, oscillatory membrane properties during fictive locomotion in the lamprey spinal cord.     

Nicotine‐mediated neuroprotection of rat spinal networks against excitotoxicity

Activation of neuronal nicotinic acetylcholine receptors (nAChRs) by nicotine is reported to protect brain neurons from glutamate excitotoxicity. We inquired whether a similar phenomenon can occur in the rat isolated spinal cord (or spinal slice culture) challenged by a transient (1 hr) application of kainate (a powerful glutamate receptor agonist) to induce excitotoxicity mimicking spinal injury in vitro. We recorded spinal reflexes and fictive locomotion generated by the locomotor central pattern generator before and 24 hr after applying kainate. We also monitored network activity with Ca²⁺ imaging and counted neurons and glia with immunohistochemical methods. In control conditions, nicotine (1 μM; 4 hr) depressed reflexes and fictive locomotion with slow recovery and no apparent neurotoxicity at 24 hr although synchronous Ca²⁺ transients appeared in slice cultures. Kainate nearly halved neuron numbers (while sparing glia), decreased reflexes and Ca²⁺ transients, and suppressed fictive locomotion. When nicotine was applied (4 hr) after washout of kainate, fictive locomotor cycles appeared 24 hr later though with low periodicity, and significant protection of neurons, including motoneurons, was observed. Nicotine applied together with kainate and maintained for further 4 hr yielded better neuroprotection, improved fictive locomotion expression and reversed the depression of Ca²⁺ transients. nAChR antagonists did not intensify kainate neurotoxicity and inhibited the neuroprotective effects of nicotine. These data suggest that nicotine was efficacious to limit histological and functional excitotoxic damage probably because it activated and then desensitized nAChRs on excitatory and inhibitory network neurons to prevent triggering intracellular cell death pathways.     

Voltage clamp analysis of lamprey neurons — role of N-methyl-d-aspartate receptors in fictive locomotion

Spinal neurons in the lamprey have been subjected to a voltage clamp analysis of the excitatory currents generated during fictive locomotion with particular reference to the phasic activation of voltage dependent N-methyl-D-aspartate (NMDA) receptors. Voltage-clamped neurons observed during NMDA-induced fictive swimming show excitatory and inhibitory synaptic currents in phase with the ipsilateral and contralateral ventral root discharges, respectively. The excitatory synaptic currents showed a marked voltage dependence suggesting that potential sensitive conductances such as the NMDA ionophore are involved in the synaptic events underlying rhythmic locomotor activity. The effect of NMDA receptor activation during application of tetrodotoxin has also been analyzed during NMDA-induced pacemaker-like oscillations. Such NMDA-induced oscillations are essentially abolished during the voltage clamp. In the presence of NMDA current voltage plots reveal a negative slope conductance in the potential range of the inherent oscillations. The addition of tetraethyl ammonium (TEA) to NMDA solution enhanced a net steady state inward current by more than 10-fold due to a partial block of the outward currents. A kinetic analysis was done with a frequency domain technique using a white noise stimulus to linearly perturb the membrane potential over a wide range of frequencies. The analysis revealed that the induced negative conductance leads to a response which is nearly 180 degrees out of phase with the stimulus at low frequencies. This is an unstable condition which leads to the depolarizing phase of the induced oscillations.     

Spino-bulbar neurons convey information to the brainstem about different phases of the locomotor cycle in the lamprey

Lamprey retriculospinal neurons show phasic oscillations of their membrane potential during fictive locomotion. This modulation originates from the spinal cord locomotor networks. The aim of the present study was to elucidate the pattern of discharge of the spino-bulbar axons responsible for this modulation. Experiments were performed on in vitro brainstem/spinal cord preparations. Two baths were formed in the recording chamber. The caudal one was perfused with 150 microM N-methyl-D-aspartate (NMDA) solution to induce fictive locomotion. The rostral bath containing the brain and the first 3-5 segments of the spinal cord was exposed to a 0 Ca2+ + 2.6 mM Mn2+ solution to block synaptic transmission and therefore to abolish any rhythmic descending activity. Spinobulbar axons were recorded intracellularly at the level of the brain/spinal cord junction. They exhibited phasic discharges correlated with the ongoing motor activity in the caudal pool. Some discharged in phase with either the ipsilateral or the contralateral ventral root bursts, others with either of the transition phases between these two bursts. These spinal cells with ascending axons, running in the ventrolateral spinal cord, may be important for modulating the activity of supraspinal neurons to match the ongoing locomotor activity.     

Endogenous activation of metabotropic glutamate receptors contributes to burst frequency regulation in the lamprey locomotor network

The effect of metabotropic glutamate receptor (mGluR) agonists and antagonists on the spinal cord network underlying locomotion in the lamprey has been analysed. The specific group I mGluR agonist (R,S)-3,5-dihydroxyphenylglycine (DHPG) and the broad-spectrum mGluR agonist (1S,3R)-1-aminocyclopentane-1,3-dicarboxylic acid (1S,3R-ACPD) both increased the burst frequency of N-methyl-d -aspartic acid (NMDA)-induced fictive locomotion and depolarized grey matter neurons. The burst frequency increase induced by the mGluR agonists was counteracted by the mGluR antagonists (+)-alpha-methyl-4-carboxyphenylglycine ((+)-MCPG), cyclopropan[b]chromen-1a-carboxylic acid ethylester (CPCCOEt) and (RS)-1-aminoindan-1,5-dicarboxylic acid (AIDA). Application of CPCCOEt alone reduced the locomotor burst frequency, indicating that mGluRs are endogenously activated during fictive locomotion. The mGluR antagonist CPCCOEt had no effect on NMDA-, or (S)-α-amino-3-hydroxy-5-methyl-4-isoazolepropionic acid (AMPA)-induced depolarizations. The mGluR agonists 1S,3R-ACPD and DHPG increased the amplitude of NMDA-induced depolarizations, a mechanism which could account for the increase in burst frequency. The group III mGluR agonist L-2-amino-4-phosphonobutyric acid reduced intraspinal synaptic transmission and burst frequency.     

Cellular bases of a vertebrate locomotor system – steering, intersegmental and segmental co-ordination and sensory control

The isolated brainstem–spinal cord of the lamprey is used as an experimental model in the analysis of the cellular bases of vertebrate locomotor behaviour. In this article we review the neural mechanisms involved in the control of steering, intersegmental co-ordination, as well as the segmental burst generation and the sensory contribution to motor pattern generation. Within these four components of the control system for locomotion, we now have good knowledge of not only the neurones that take part and their synaptic interactions, but also the membrane properties of these neurones, including ion channel subtypes, and their contribution to motor pattern generation.     

The respective contribution of lumbar segments to the generation of locomotion in the isolated spinal cord of newborn rat

Various studies on isolated neonatal rat spinal cord have pointed to the predominant role played by the rostral lumbar area in the generation of locomotor activity. In the present study, the role of the various regions of the lumbar spinal cord in locomotor genesis was further examined using compartmentalization and transections of the cord. We report that the synaptic drive received by caudal motoneurons following N-methyl-d-l-aspartate (NMA)/5-HT superfusion on the entire lumbar cord is different from that triggered by the same compounds specifically applied on the rostral segments. These differences appear to be due to the direct action of NMA/5-HT on motoneuron membrane potential, rather than on premotoneuronal input activation. In order to assess the possible participation of the caudal lumbar segments in locomotor rhythm generation, the segments were over-stimulated with high concentrations of NMA or K+. We find that significant variations in motor cycle period occurred during the over-activation of the rostral segments. Over-activation of caudal segments only si+gnificantly increased the caudal ventral roots burst amplitude. We find that low 5-HT concentrations were unable to induce fictive locomotion under our experimental conditions. When a hemi-transection of the cord was performed between the L2-L3 segments, rhythmic bursting in the ipsilateral L5 disappeared while rhythmicity persisted on the contralateral side. Sectioning of the remaining L2-L3 side totally suppressed rhythmic activity in both L5 ventral roots. These results show that the thoracolumbar part of the cord constitutes the key area for locomotor pattern generation.     

The effect of 4-aminopyridine on the spinal locomotor rhythm induced byl-DOPA

In high spinal curarized cats rhythmic motor output similar to locomotion (‘fictive locomotion’) of all 4 limbs was obtained after intravenous application of the noradrenergic precursor,l-DOPA, and nialamide combined with 4-aminopyridine (4-AP). The activity was recorded from muscle nerves.In the presence of 4-AP, which enhances transmission at various excitatory and inhibitory synapses, reduced amounts of DOPA were sufficient to evole fictive locomotion. 4-AP alone did not elicit locomotion.The burst rate increased up to 6 Hz with the amount of 4-AP given (0.5–50 mg/kg).The cycle frequency of high spinal cats exhibiting either fictive locomotion or walking on a treadmill was accelerated by 4-AP.After a supplementary transection of the spinal cord at the upper lumbar level both fore- and hindlimbs generally continued to show fictive locomotion with similar frequencies.In the presence of high doses of clonidine (alpha-receptor-activator, > 4mg/kg), the locomotor pattern was replaced by regular (2 Hz) synchronous discharges in all flexors and extensors.     

Mechanism of synchronized Ca oscillations in cortical neurons

Dissociated rat cortical neurons reassociate in vitro to form synaptically connected networks. Removal of Mg²⁺ from the extracellular medium then induces neurons in the network to undergo synchronized oscillations of cytoplasmic calcium. Previous studies have shown that these calcium oscillations involve the activation of NMDA receptors and that the rising phase of each calcium spike is coincident with a brief burst of action potentials (Robinson et al., Jpn. J. Physiol. 43 (Suppl. 1) (1993) S125–130; Robinson et al., J. Neurophysiol. 70 (1993) 1606–1616; Murphy et al., J. Neurosci. 12 (1992) 4834–4845). We have found that these calcium oscillations are dependent on an influx of extracellular calcium but are independent of mobilization of calcium from intracellular stores. The influx of extracellular Ca²⁺ occurs primarily through L-type voltage-gated calcium channels (VGCCs), since diltiazem inhibits calcium oscillations under all conditions. On the other hand, N-, P/Q-, and T-type VGCCs are not required for calcium oscillations, although inhibitors of these channels may act as partial antagonists. In addition to removal of Mg²⁺, oscillations can also be induced by the inhibition of voltage-gated K⁺ channels with 4-aminopyridine (4-AP), a treatment known to increase neurotransmitter release. In the presence of 4-AP, synchronized calcium oscillations become independent of NMDA receptor activation, although they continue to require activation of AMPA/KA receptors. A model for the mechanism of neuronal calcium oscillations and the reason for their synchrony is presented.     

The contribution of the NMDA receptor glycine site to rhythm generation during fictive swimming in Xenopus laevis tadpoles

The presence of the N-methyl-D-aspartate (NMDA) receptor glycine-binding site and its role in locomotor activity have been examined using fictive swimming in stage 42 Xenopus laevis frog tadpoles as a simple model system. The specific NMDA/glycine site blocker L-689560 (0.1-20 microm) impaired swimming rhythm generation and abolished NMDA-induced locomotor-like ventral root activity. D-serine (50 microm), an agonist at the NMDA/glycine site, increased the duration of skin stimulus-induced fictive swimming episodes, and produced slow modulations of burst frequency and amplitude. These effects of D-serine were reversed by L-689560. In some animals, D-serine also induced an alternative intense, non-locomotory form of rhythmic motor output termed struggling. Glycine (100 microm), another endogenous agonist at this site, triggered similar effects to D-serine, but only when applied in the presence of strychnine. Manipulations of endogenous glycine levels using sarcosine or ALX 5407 (inhibitors of the glycine re-uptake protein, GlyT1b), produced similar effects to glycine site agonists, including increased episode durations, and modulations in cycle period and burst amplitude. Sarcosine and ALX 5407 also induced struggling. In summary, these experiments support the hypothesis that NMDA receptors in the swimming network of Xenopus laevis tadpoles possess glycine-binding sites, not all of which are fully occupied under normal circumstances. Altering the strength of the NMDA receptor-mediated component of the synaptic drive for swimming by increasing or decreasing occupancy of this site potently influences the locomotor pattern.     

Rostrocaudal distribution of 5-HT innervation in the lamprey spinal cord and differential effects of 5-HT on fictive locomotion

5-hydroxytryptamine (5-HT) is known to modulate the locomotion generator network in the lamprey spinal cord, but little is known about the pattern of 5-HT innervation along the spinal cord. The distribution of 5-HT-immunoreactive (5-HT-ir) cells and fibers, as well as the effects of 5-HT on the locomotor network in the rostral and caudal parts of the spinal cord were compared in two lamprey species, Lampetra fluviatilis and Petromyzon marinus. Intraspinal 5-HT cells form a very dense ventromedial plexus in which the dendrites of neurons forming the locomotor network are distributed. The number of 5-HT cells and varicosities in this plexus decreases in the fin area (segments 70–90), and then increases somewhat in the most caudal segments. The descending 5-HT fibers from the rhombencephalon are located in the lateral and ventral columns, and their numbers gradually decrease to around 50% in the tail part of the spinal cord. In contrast, the number of 5-HT-ir axons in the dorsal column remains the same along the spinal cord. Bath application of both N-methyl-D-aspartic acid (NMDA; 20–250 μM) and D-glutamate (250–1000 μM) was used to induce fictive locomotion in the isolated spinal cord. Bath application of 5-HT (1 μM) reduced the burst frequency in the presence of NMDA. The 5-HT effect was, however, significantly greater in the rostral as compared to the caudal part. With D-glutamate, the 5-HT effect was instead more pronounced in the caudal spinal cord. To account for this difference in 5-HT effects on NMDA- and D-glutamate-induced fictive locomotion, the cellular effect of D-glutamate was further investigated. It activates not only NMDA, but also alpha amino-3-hydroxy-5-methyl-4-isoxyl propionate (AMPA)/kainate and metabotropic glutamate receptors. In contrast to NMDA, D-glutamate did not elicit tetrodotoxin (TTX)-resistant membrane potential oscillations. This difference in action between NMDA (selective NMDA receptor agonist) and D-glutamate (mixed agonist) may partially account for the differences in effect of 5-HT on the locomotor pattern. © Wiley-Liss, Inc.     

Phasic modulation of transmission from vestibular inputs to reticulospinal neurons during fictive locomotion in lampreys

The aim of this study was to determine whether the transmission from sensory inputs to reticulospinal neurons is modulated during fictive locomotion in lampreys. Reticulospinal neurons play a key role in the control of locomotion; modulation of sensory transmission to these neurons might be of importance for the adaptation of the control they exert during locomotion. In this series of experiments, intracellular synaptic responses of reticulospinal neurons of the posterior rhombencephalic reticular nucleus elicited by electrical stimulation of vestibular nerves on each side were studied during fictive locomotion induced by 50 microM N-methyl-D-aspartate (NMDA). Interestingly, shortly after NMDA had reached the bath and much before locomotor discharges were apparent in the recorded ventral roots, there was a significant depression of the synaptic transmission from vestibular nerves. The effect was reversed by washing out the NMDA and persisted in the isolated brainstem after spinal transection at the first segmental level. As locomotor discharges appeared in the ventral roots, synaptic responses elicited by vestibular nerve stimulation showed a clear phasic modulation of their amplitude during the locomotor cycle. Responses to stimulation of the ipsilateral vestibular nerve were smaller during the ipsilateral burst discharge than during the contralateral activity, whilst responses to stimulation of the contralateral vestibular nerve were minimal during contralateral activity and maximal during ipsilateral activity. This opposite pattern of modulation observed in the same reticulospinal neuron suggests that the phasic modulation of vestibular transmission is not due to changes in the membrane properties of the reticulospinal cell but is produced at a pre-reticular level.(ABSTRACT TRUNCATED AT 250 WORDS)     

Anesthetic effects on fictive locomotion in the rat isolated spinal cord

General anesthetic mechanisms are poorly understood. Anesthetic immobilizing effects occur in the spinal ventral horn. However, a detailed analysis of anesthetic effects on ventral motor networks is lacking. We delivered isoflurane, desflurane, or propofol during NMDA/5-HT-induced, or noxious tail stimulus-evoked, fictive locomotion in neonatal rat isolated spinal cords. Anesthetics changed the frequency, amplitude, and regularity of fictive locomotion with little effect on phase-lag. Isoflurane abolished pharmacologically-induced versus noxious stimulus-induced motor output at similar concentrations. Propofol abolished pharmacologically-induced fictive locomotion through a γ-aminobutyric acid type A-receptor mechanism. Anesthetic effects on pharmacologically-elicted fictive locomotion appear clinically-relevant, and support a ventral horn immobilizing effect on locomotor rhythm generation.     

The role of spinal cord inputs in modulating the activity of reticulospinal neurons during fictive locomotion in the lamprey

Lamprey reticulospinal neurons are rhythmically modulated during fictive swimming. The present study examines the possibility that this modulation may originate from the spinal cord locomotor networks rather than from the brainstem. To test this, the in vitro preparation of the lamprey brainstem-spinal cord was separated into two compartments which could be exposed to different chemical environments. Locomotor activity was induced pharmacologically in the caudal spinal cord compartment and reticulospinal (RS) neurons from the posterior rhombencephalic reticular nucleus (PRRN) were recorded intracellularly in the rostral compartment containing normal lamprey Ringer. Under these conditions, the membrane potential of RS neurons showed clear rhythmic oscillations which are correlated with the ongoing locomotor activity in the caudal spinal cord bath, although no locomotor discharges were present in the ventral roots of the rostral bath. Such oscillations were not present in the absence of locomotion. These results indicate that the spinal cord locomotor networks can contribute to the rhythmic oscillations which occur in RS neurons during fictive locomotion. Moreover, the latter oscillations of membrane potential are due to both phasic excitation and Cl- -dependent inhibition in the opposite phase.     

The action of 5-HT on calcium-dependent potassium channels and on the spinal locomotor network in lamprey is mediated by 5-HT1A-like receptors

5-HT has a powerful modulatory action on the firing properties of single neurons as well as on locomotor activity. In lamprey, 5-HT increases the neuronal firing frequency in spinal neurons by reducing the conductance in Ca²⁺-dependent K⁺ channels (K_Ca) underlying the slow afterhyperpolarization (sAHP), and it also lowers burst frequency of the spinal locomotor network. To elucidate which type of 5-HT receptor mediates these effects, different specific receptor agonists and antagonists were applied during intracellular current lamp recordings and during NMDA-induced fictive locomotion in the lamprey spinal cord in vitro preparation. The 5-HT_1A receptor agonist 8-OH-DPAT ((±)-8-hydroxy-dipropylaminotetralin hydrobromide), the 5-HT₁ receptor agonist 5-CT (5-car☐yamidotryptamine maleate) and the 5-HT₂ receptor agonist α-CH₃-5-HT (α-methylserotonin maleate) all reproduced the actions of 5-HT at both the cellular and the network levels. The effects of all agonists were completely or partially blocked by the 5-HT_1A and 5-HT₂ receptor antagonist spiperone (spiroperidol hydrochloride) while selective 5-HT₂ receptor antagonists were ineffective. The selective 5-HT_1A receptor antagonist S(−)-UH301 (S(−)-5-fluoro-8-hydroxy-dipropylaminotetralin hydrochloride) also counteracted the effect of 5-HT on the sAHP. 5-HT₃ and 5-HT₄ receptor agonists and antagonists were without effects. The intracellular coupling mechanism was not sensitive to pertussis toxin nor to the cAMP dependent protein kinase blocker (Rp)-cAMPS. These results indicate that the intracellular coupling mechanism is not likely to be due to a down regulation of adenylate cyclase activity or through a direct modulation of K⁺ channels, as is common for 5-HT₁ receptors. The present results taken together with previous data indicates that the receptor responsible for the effects of 5-HT on the sAHP, and on the locomotor pattern generator in lamprey shares certain features, but is not identical to the mammalian 5-HT_1A receptor.