Renshaw Cells Research Articles

Cholinergic terminals in the ventral horn of adult rat and cat: Evidence that glutamate is a cotransmitter at putative interneuron synapses but not at central synapses of motoneurons

Until recently it was generally accepted that the only neurotransmitter to be released at central synapses of somatic motoneurons was acetylcholine. However, studies on young mice (P0-10) have provided pharmacological evidence indicating that glutamate may act as a cotransmitter with acetylcholine at synapses between motoneurons and Renshaw cells. We performed a series of anatomical experiments on axon collaterals obtained from intracellularly labeled motoneurons from an adult cat and labeled by retrograde transport in adult rats to determine if glutamate is co-localized with acetylcholine by these terminals. We could find no evidence for the presence of vesicular glutamate transporters in motoneuron axon terminals of either species. In addition, we were unable to establish any obvious relationship between motoneuron terminals and the R2 subunit of the AMPA receptor (GluR2). However we did observe a population of cholinergic terminals in lamina VII which did not originate from motoneurons but were immunoreactive for the vesicular glutamate transporter 2 and formed appositions to GluR2 subunits. These were smaller than motoneuron terminals and, unlike them, formed no relationship with Renshaw cells. The evidence suggests that glutamate does not act as a cotransmitter with acetylcholine at central synapses of motoneurons in the adult cat and rat. However, glutamate is present in a population of cholinergic terminals which probably originate from interneurons where its action is via an AMPA receptor.

Glycinergic Innervation of Motoneurons Is Deficient in Amyotrophic Lateral Sclerosis Mice: A Quantitative Confocal Analysis

Altered motoneuron excitability is involved in amyotrophic lateral sclerosis pathobiology. To test the hypothesis that inhibitory interneuron innervation of spinal motoneurons is abnormal in an amyotrophic lateral sclerosis mouse model, we measured GABAergic, glycinergic, and cholinergic immunoreactive terminals on spinal motoneurons in mice expressing a mutant form of human superoxide dismutase-1 with a Gly93-->Ala substitution (G93A-SOD1) and in controls at different ages. Glutamic acid decarboxylase, glycine transporter-2, and choline acetyltransferase were used as markers for GABAergic, glycinergic, and cholinergic terminals, respectively. Triple immunofluorescent labeling of boutons contacting motoneurons was visualized by confocal microscopy and analyzed quantitatively. Glycine transporter-2-bouton density on lateral motoneurons was decreased significantly in G93A-SOD1 mice compared with controls. This reduction was absent at 6 weeks of age but present in asymptomatic 8-week-old mice and worsened with disease progression from 12 to 14 weeks of age. Motoneurons lost most glycinergic innervation by 16 weeks of age (end-stage) when there was a significant decrease in the numbers of motoneurons and choline acetyltransferase-positive boutons. No significant differences in glutamic acid decarboxylase-bouton densities were found in G93A-SOD1 mice. Reduction of glycinergic innervation preceded mitochondrial swelling and vacuolization. Calbindin-positive Renshaw cell number was decreased significantly at 12 weeks of age in G93A-SOD1 mice. Thus, either the selective loss of inhibitory glycinergic regulation of motoneuron function or glycinergic interneuron degeneration contributes to motoneuron degeneration in amyotrophic lateral sclerosis.

Properties of Renshaw cells excited by recurrent collaterals of pudendal motoneurons in the cat

Properties of Renshaw cells excited by recurrent collaterals of pudendal motoneurons in the cat

Four Excitatory Postsynaptic Ionotropic Receptors Coactivated at the Motoneuron–Renshaw Cell Synapse

Renshaw cells (RCs) are spinal interneurons excited by collaterals of the axons of motoneurons (MNs). They respond to a single motoneuronal volley by a surprisingly long (tens of milliseconds) train of action potentials. We have analyzed this synaptic response in spinal cord slices of neonatal mice in light of recent observations suggesting that the MN axons release both acetylcholine and glutamate. We found that the RC synaptic current involves four components of similar amplitudes mediated by two nicotinic receptors (nAChRs, tentatively identified as alpha(7) homomers and alpha(4)beta(2) heteromers) and two glutamate receptors (AMPARs and NMDARs). The decay time constants of the four components cover a wide range: from 3.6 +/- 2.2 ms (alpha(7) nAChRs) to 54.6 +/- 19.5 ms (NMDARs, at -45 mV). The RC discharge can be separated into an initial doublet of high-frequency action potentials followed by later spikes with a variable latency and longer interspike intervals. The initial doublet involves the four ionotropic receptors as well as endogenous voltage-dependent conductances. The late discharge depends on NMDARs, but these receptors must be primed by the initial depolarization. The activation of the NMDARs is prolonged by the fact that their slow deactivation is further slowed by depolarization. The formation of the initial doublet is favored by hyperpolarization, whereas the late discharge is favored by depolarization. This suggests that in physiological conditions the pattern of discharge of the RC in response to a MN input may alternate between a phasic and a tonic response.

Birdsey Renshaw (1911–1948) and his eponym

Birdsey Renshaw was an American neuroscientist and made innovative contributions to neuroscience. He is credited with investigating the inhibitory interneurons in the ventral horn of grey matter of the spinal...

A differential developmental pattern of spinal interneuron apoptosis during synaptogenesis: insights from genetic analyses of the protocadherin-γ gene cluster

Although the role of developmental apoptosis in shaping the complement and connectivity of sensory and motoneurons is well documented, the extent to which cell death affects the 13 cardinal classes of spinal interneurons is unclear. Using a series of genetic manipulations in vivo, we demonstrate for the first time a differential pattern of developmental apoptosis in molecularly identified spinal interneuron populations, and implicate the adhesion molecule family encoded by the 22-member protocadherin-gamma (Pcdh-gamma) gene cluster in its control. In constitutive Pcdh-gamma null mouse embryos, many interneuron populations undergo increased apoptosis, but to differing extents: for example, over 80% of En1-positive V1 neurons are lost, whereas only 30% of Chx10-positive V2a neurons are lost and there is no reduction in the number of V1-derived Renshaw cells. We show that this represents an exacerbation of a normal, underlying developmental pattern: the extent of each population's decrease in Pcdh-gamma mutants is precisely commensurate both with the extent of its loss during normal embryogenesis and with the extent of its increase in Bax(-/-) mice, in which apoptosis is genetically blocked. Interneuron apoptosis begins during the first wave of synaptogenesisis in the spinal cord, occurring first among ventral populations (primarily between E14 and E17), and only later among dorsal populations (primarily after P0). Utilizing a new, conditional Pcdh-gamma mutant allele, we show that the gamma-Pcdhs can promote survival non-cell-autonomously: mutant neurons can survive if they are surrounded by normal neurons, and normal neurons can undergo apoptosis if they are surrounded by mutant neurons.

Seeking functions for spinal recurrent inhibition

In this issue of The Journal of Physiology, Lamy et al. (2008) describe a rhythmic modulation of recurrent inhibition during walking in man. This provides some insight into the function of recurrent inhibition, first described by Renshaw (1941) as an inhibition of cat motoneurones produced by antidromic stimulation of the axons of neighbouring motoneurones (homonymous action). Subsequent work, also in cats, showed that motor axon collaterals activate interneurones (the eponymous Renshaw cells) that in turn inhibit motoneurones. Actions extend to more distant muscle groups (heteronymous) and parallel the pattern of heteronymous monosynaptic excitation from muscle spindle Ia afferents. Renshaw cells receive peripheral sensory and descending inputs and have output connections to other spinal interneurones and to other Renshaw cells. So recurrent inhibition is more than just negative feedback from and to motoneurones and may have several functions depending upon motor context. Lamy et al. have focused on heteronymous actions (which are more widespread in man than in cats or monkeys) in the context of our unique bipedal form of walking. The knee extensor quadriceps is a source of recurrent inhibition to the ankle extensor and the flexor muscles soleus and tibialis anterior. But given that these ankle muscles are antagonists, how is differential activation achieved? The authors found that around heel strike and early in stance, when quadriceps is active and soleus activity is increasing, recurrent inhibition of soleus is weak. Conversely, later in stance, when quadriceps is active and soleus activity is declining, recurrent inhibition of soleus is strong. Recurrent inhibition of tibialis anterior motoneurones was neither task nor phase dependent. The reduction in soleus recurrent inhibition would assist the transition from swing to stance and enhanced inhibition later in stance would favour the transition from stance to swing by reducing reciprocal inhibition of tibialis anterior. The origin of these changes in recurrent inhibition remains uncertain but activity of peripheral sensory or descending motor pathways could be involved. Two interesting possibilities are the corticospinal pathway, known to depress homonymous recurrent inhibition of soleus in sitting subjects (Mazzocchio et al. 1994) and the vestibulospinal pathway that depresses recurrent inhibition of soleus in standing stance (Iles & Pisini, 1992). It might be possible to extend such studies to heteronymous actions in walking stance. More meticulous experiments built upon the framework of human research protocols pioneered by the Paris group will likely increase our understanding of recurrent inhibition even further.

Modulation of recurrent inhibition from knee extensors to ankle motoneurones during human walking

The neural control for muscle coordination during human locomotion involves spinal and supraspinal networks, but little is known about the exact mechanisms implicated. The present study focused on modulation of heteronymous recurrent inhibition from knee extensors to ankle motoneurones at different times in the gait cycle, when quadriceps (Quad) muscle activity overlaps that in tibialis anterior (TA) and soleus (Sol). The effects of femoral nerve stimulation on ankle motoneurones were investigated during treadmill walking and during tonic co-contraction of Quad and TA/Sol while standing. Recurrent inhibition of TA motoneurones depended on the level of background EMG, and was similar during walking and standing for matched background EMG levels. On the other hand, recurrent inhibition in Sol was reduced in early stance, with respect to standing, and enhanced in late stance. Reduced inhibition in Sol was also observed when Quad was coactivated with TA around the time of heel contact, compared to standing at matched background EMG levels in the two muscles. The modulation of recurrent inhibition of Sol during walking might reflect central and/or peripheral control of the Renshaw cells. These modulations could be implicated in the transition phases, from swing to stance to assist Sol activation during the stance phase, and from stance to swing, for its deactivation.

V3 Spinal Neurons Establish a Robust and Balanced Locomotor Rhythm during Walking

A robust and well-organized rhythm is a key feature of many neuronal networks, including those that regulate essential behaviors such as circadian rhythmogenesis, breathing, and locomotion. Here we show that excitatory V3-derived neurons are necessary for a robust and organized locomotor rhythm during walking. When V3-mediated neurotransmission is selectively blocked by the expression of the tetanus toxin light chain subunit (TeNT), the regularity and robustness of the locomotor rhythm is severely perturbed. A similar degeneration in the locomotor rhythm occurs when the excitability of V3-derived neurons is reduced acutely by ligand-induced activation of the allatostatin receptor. The V3-derived neurons additionally function to balance the locomotor output between both halves of the spinal cord, thereby ensuring a symmetrical pattern of locomotor activity during walking. We propose that the V3 neurons establish a regular and balanced motor rhythm by distributing excitatory drive between both halves of the spinal cord.

Simulation system of spinal cord motor nuclei and associated nerves and muscles, in a Web-based architecture

A Web-based simulation system of the spinal cord circuitry responsible for muscle control is described. The simulator employs two-compartment motoneuron models for S, FR and FF types, with synaptic inputs acting through conductance variations. Four motoneuron pools with their associated interneurons are represented in the simulator, with the possibility of inclusion of more than 2,000 neurons and 2,000,000 synapses. Each motoneuron action potential is followed, after a conduction delay, by a motor unit potential and a motor unit twitch. The sums of all motor unit potentials and twitches result in the electromyogram (EMG), and the muscle force, respectively. Inputs to the motoneuron pool come from populations of interneurons (Ia reciprocal inhibitory interneurons, Ib interneurons, and Renshaw cells) and from stochastic point processes associated with descending tracts. To simulate human electrophysiological experiments, the simulator incorporates external nerve stimulation with orthodromic and antidromic propagation. This provides the mechanisms for reflex generation and activation of spinal neuronal circuits that modulate the activity of another motoneuron pool (e.g., by reciprocal inhibition). The generation of the H-reflex by the Ia-motoneuron pool system and its modulation by spinal cord interneurons is included in the simulation system. Studies with the simulator may include the statistics of individual motoneuron or interneuron spike trains or the collective effect of a motor nucleus on the dynamics of muscle force control. Properties associated with motor-unit recruitment, motor-unit synchronization, recurrent inhibition and reciprocal inhibition may be investigated.

Early Postnatal Development of Reciprocal Ia Inhibition in the Murine Spinal Cord

The pathway mediating reciprocal inhibition from muscle spindle afferents (Ia axons) to motoneurons (MNs) supplying antagonist muscles has been well studied in adult cats, but little is known about how this disynaptic pathway develops. As a basis for studying its development, we characterized this pathway in mice during the first postnatal week, focusing on the projection of quadriceps (Q) Ia axons to posterior biceps-semitendinosis (PBSt) MNs via Ia inhibitory interneurons. Synaptic potentials in PBSt MNs evoked by Q nerve stimulation are mediated disynaptically and are blocked by strychnine, implying that glycine is the major inhibitory transmitter as in adult cats. The specificity of neuronal connections in this reflex pathway is already high at birth; Q afferents evoke inhibitory synaptic potentials in PBSt MNs, but afferents supplying the adductor muscle do not. Similar to this disynaptic pathway in cats, Renshaw cells inhibit the interposed Ia interneurons, as they reduce the disynaptic input from Q axons but do not inhibit PBSt MNs directly. Reciprocal inhibition functionally inhibits the monosynaptic excitatory reflex in PBSt MNs by P3, but this functional inhibition is weak at P1. Finally, deletion of the transcription factor Pax6, which is required for the development of V1-derived Renshaw cells, does not block development of this pathway. This suggests either that Pax6 is not required for the phenotypic development of all V1-derived spinal interneurons or that these inhibitory interneurons are not derived from V1 precursors.

Synaptic inputs to Renshaw cells from sensory pathways in the neonatal mouse spinal cord

Synaptic inputs to Renshaw cells from sensory pathways in the neonatal mouse spinal cord

Imaging the spatiotemporal organization of neural activity in the developing spinal cord

In this review, we discuss the use of imaging to visualize the spatiotemporal organization of network activity in the developing spinal cord of the chick embryo and the neonatal mouse. We describe several different methods for loading ion- and voltage-sensitive dyes into spinal neurons and consider the advantages and limitations of each one. We review work in the chick embryo, suggesting that motoneurons play a critical role in the initiation of each cycle of spontaneous network activity and describe how imaging has been used to identify a class of spinal interneuron that appears to be the avian homolog of mammalian Renshaw cells or 1a-inhibitory interneurons. Imaging of locomotor-like activity in the neonatal mouse revealed a wave-like activation of motoneurons during each cycle of discharge. We discuss the significance of this finding and its implications for understanding how locomotor-like activity is coordinated across different segments of the cord. In the last part of the review, we discuss some of the exciting new prospects for the future.

Hb9 Interneurons: Reply to Ziskind-Conhaim and Hinckley

reply: We thank Drs. Ziskind-Conhaim and Hinckley for their comments and are appreciative that they find our paper of interest. We, too, find it interesting that the conclusions of our two groups differ. We initially provided indirect evidence of electrotonic coupling involving Hb9 interneurons in

Relative Location of Inhibitory Synapses and Persistent Inward Currents Determines the Magnitude and Mode of Synaptic Amplification in Motoneurons

In some motoneurons, L-type Ca2+ channels that partly mediate persistent inward currents (PICs) have been estimated to be arranged in 50- to 200-microm-long discrete regions in the dendrites, centered 100 to 400 microm from the soma. As a consequence of this nonuniform distribution, the interaction between synaptic inputs to motoneurons and these channels may vary according to the distribution of the synapses. For instance, >93% of synapses from Renshaw cells have been observed to be located 65 to 470 microm away from the cell body of motoneurons. Our goal was to assess whether Renshaw cell synapses are distributed in a position to more effectively control the activation of the L-type Ca2+ channels. Using compartmental models of motoneurons with L-type Ca2+ channels distributed in 100-microm-long hot spots centered 100 to 400 microm away from the soma, we compared the inhibition generated by four distributions of inhibitory synapses: proximal, distal, uniform, and one based on the location of Renshaw cell synapses on motoneurons. Regardless of whether the synapses were activated tonically or transiently, in the presence of L-type Ca2+ channels, inhibitory synapses distributed according to the Renshaw cell synapse distribution generate the largest inhibitory currents. The effectiveness of a particular distribution of inhibitory synapses in the presence of PICs depends on their ability to deactivate the channels underlying PICs, which is influenced not only by the superposition between synapses and channels, but also by the distance away from the somatic voltage clamp.

Multiple Modes of Amplification of Synaptic Inhibition to Motoneurons by Persistent Inward Currents

The ability of inhibitory synaptic inputs to dampen the excitability of motoneurons is augmented when persistent inward currents (PICs) are activated. This amplification could be due to an increase in the driving potential of inhibitory synapses or the deactivation of the channels underlying PICs. Our goal was to determine which mechanism leads to the amplification of inhibitory inputs by PICs. To reach this goal, we measured inhibitory postsynaptic currents (IPSCs) in decerebrate cats during somatic voltage-clamp steps. These IPSCs were generated by tonic activation of Renshaw cells. The IPSCs exhibited a rapid rise and a slower decay to a plateau level. Activation of PICs always led to an increase in the peak of the IPSC, but the amount of decay after the peak of the IPSC was inversely related to the size of the IPSC. These results were replicated in simulations based on compartmental models of motoneurons incorporating distributions of Renshaw cell synapses based on anatomical observations, and L-type calcium channels distributed as 100-microm-long hot spots centered 100 to 400 microm away from the soma. For smaller IPSCs, amplification by PICs was due to an increase in the driving force of the inhibitory synaptic current. For larger IPSCs, amplification was caused by deactivation of the channels underlying PICs leading to a lesser decay of the IPSCs. As a result of this change in the time course of the IPSC, deactivation of the channels underlying PICs leads to a greater amplification of the total inhibitory synaptic current.

The continuing case for the Renshaw cell

Renshaw cell properties have been studied extensively for over 50 years, making them a uniquely well-defined class of spinal interneuron. Recent work has revealed novel ways to identify Renshaw cells in situ and this in turn has promoted a range of studies that have determined their ontogeny and organization of synaptic inputs in unprecedented detail. In this review we illustrate how mature Renshaw cell properties and connectivity arise through a combination of activity-dependent and genetically specified mechanisms. These new insights should aid the development of experimental strategies to manipulate Renshaw cells in spinal circuits and clarify their role in modulating motor output.

Effects of spinal recurrent inhibition on motoneuron short-term synchronization

Spinal recurrent inhibition linking skeleto- motoneurons (alpha-MNs) via Renshaw cells (RCs) has been variously proposed to increase or decrease tendencies toward synchronous discharges between alpha-MNs. This controversy is not easy to settle experimentally in animal or human paradigms because RCs receive, in addition to excitatory input from alpha-MNs, many other modulating influences which may change their mode of operation. Computer simulations help to artificially isolate the recurrent inhibitory circuit and thus to study its effects on alpha-MN synchronization under conditions not achievable in natural experiments. We present here such a study which was designed to specifically test the following hypothesis. Since many alpha-MNs excite any particular Renshaw cell, which in turn inhibits many alpha-MNs, this convergence-divergence pattern establishes a random network whose random discharge patterns inject uncorrelated noise into alpha-MNs, and this noise counteracts any synchronization potentially arising from other sources, e.g., common inputs (Adam et al. in Biol Cybern 29:229-235, 1978). We investigated the short-term synchronization of alpha-MNs with two types of excitatory input signals to alpha-MNs (random and sinusoidally modulated random patterns). The main results showed that, while recurrent inhibitory inputs to different alpha-MNs were indeed different, recurrent inhibition (1) exerted rather small effects on the modulation of alpha-MN discharge, (2) tended to increase the short-term synchronization of alpha-MN discharge, and (3) did not generate secondary peaks in alpha-MN-alpha-MN cross-correlograms associated with alpha-MN rhythmicity.

Functional connection from Mauthner cell to homologous reticulospinal neurons in the goldfish hindbrain

Crossed vestibular inhibition first described in mammals, also occurs in many species ranging from reptiles to mammals, but was nct observed in frog and toadfish. During a study of the inhibitory pathways of the Mauthner cell (M-cell) of a teleost, two classes of interneurons were individualized. Their identification was made possible by intracellular injection of horseradish peroxidase (HRP) after they had been physiologicaIly identified by the presence of an intracellularly recorded passive hyperpolarizing potential induced by the firing of their adjacent M-cell. Interneurons of the first group are part of the recurrent collateral network of the M-cell; they have some similarities with the Renshaw cells. The second group is composed of commissural vestibule-vestibular neurons. In a recent study of the neurons inhibiting the M-cell of teleosts, it was demonstrated that each bouton established on the M-cell by the commissural and recurrent collateral interneurons functions as an independent all-or-none unit. This result has prompted the investigation of the ultrastructure of these endings to determine if they were morphologically different from those found at other central chemically transmitting synapses.This chapter deals with deals with the relation that was found between the morphology and the function of a set of neurons that are known to inhibit the M-cell, in this case second order vestibulo-vestibular interneurons.

Inhibition of Renshaw cells during spinal reflexes in the neonatal mouse spinal cord

Inhibition of Renshaw cells during spinal reflexes in the neonatal mouse spinal cord