RESUMEN
Spinal motoneurones (Mns) constitute the final output for the execution of motor tasks. In addition to innervating muscles, Mns project excitatory collateral connections to Renshaw cells (RCs) and other Mns, but the latter have received little attention. We show that Mns receive strong synaptic input from other Mns throughout development and into maturity, with fast-type Mns systematically receiving greater recurrent excitation than slow-type Mns. Optical recordings show that activation of Mns in one spinal segment can propagate to adjacent segments even in the presence of intact recurrent inhibition. While it is known that transmission at the neuromuscular junction is purely cholinergic and RCs are excited through both acetylcholine and glutamate receptors, here we show that neurotransmission between Mns is purely glutamatergic, indicating that synaptic transmission systems are differentiated at different postsynaptic targets of Mns.
Asunto(s)
Neuronas Motoras/fisiología , Transmisión Sináptica , Animales , Animales Modificados Genéticamente , Fenómenos Electrofisiológicos , Femenino , Masculino , Ratones Endogámicos C57BL , Músculo Esquelético/inervación , Vías Nerviosas/fisiologíaRESUMEN
In neonatal mice motoneurons excite Renshaw cells by releasing both acetylcholine (ACh) and glutamate. These two neurotransmitters activate two types of nicotinic receptors (nAChRs) (the homomeric α7 receptors and the heteromeric α*ß* receptors) as well as the two types of glutamate receptors (GluRs) (AMPARs and NMDARs). Using paired recordings, we confirm that a single motoneuron can release both transmitters on a single post-synaptic Renshaw cell. We then show that co-transmission is preserved in adult animals. Kinetic analysis of miniature EPSCs revealed quantal release of mixed events associating AMPARs and NMDARs, as well as α7 and α*ß* nAChRs, but no evidence was found for mEPSCs associating nAChRs with GluRs. Bayesian Quantal Analysis (BQA) of evoked EPSCs showed that the number of functional contacts on a single Renshaw cell is more than halved when the nicotinic receptors are blocked, confirming that the two neurotransmitters systems are segregated. Our observations can be explained if ACh and glutamate are released from common vesicles onto spatially segregated post-synaptic receptors clusters, but a pre-synaptic segregation of cholinergic and glutamatergic release sites is also possible.
Asunto(s)
Acetilcolina/metabolismo , Ácido Glutámico/metabolismo , Neuronas Motoras/fisiología , Células de Renshaw/fisiología , Sinapsis/metabolismo , Transmisión Sináptica , Animales , Ratones , Receptores de Glutamato/metabolismo , Receptores Nicotínicos/metabolismoRESUMEN
Renshaw cells represent a fundamental component of one of the first discovered neuronal circuits, but their function in motor control has not been established. They are the only central neurons that receive collateral projections from motor outputs, yet the efficacy of the excitatory synapses from single and converging motoneurons remains unknown. Here we present the results of dual whole-cell recordings from identified, synaptically connected Renshaw cell-motoneuron pairs in the mouse lumbar spinal cord. The responses from single Renshaw cells demonstrate that motoneuron synapses elicit large excitatory conductances with few or no failures. We show that the strong excitatory input from motoneurons results from a high probability of neurotransmitter release onto multiple postsynaptic contacts. Dual current-clamp recordings confirm that single motoneuron inputs were sufficient to depolarize the Renshaw cell beyond threshold for firing. Reciprocal connectivity was observed in approximately one-third of the paired recordings tested. Ventral root stimulation was used to evoke currents from Renshaw cells or motoneurons to characterize responses of single neurons to the activation of their corresponding presynaptic cell populations. Excitatory or inhibitory synaptic inputs in the recurrent inhibitory loop induced substantial effects on the excitability of respective postsynaptic cells. Quantal analysis estimates showed a large number of converging inputs from presynaptic motoneuron and Renshaw cell populations. The combination of considerable synaptic efficacy and extensive connectivity within the recurrent circuitry indicates a role of Renshaw cells in modulating motor outputs that may be considerably more important than has been previously supposed. SIGNIFICANCE STATEMENT: We have recently shown that Renshaw cells mediate powerful shunt inhibition on motoneuron excitability. Here we complete a quantitative description of the recurrent circuit using recordings of excitatory synapses between identified motoneuron and Renshaw cell pairs. We show that the excitation is highly effective as a result of a high probability of neurotransmitter release onto multiple release sites and that efficient neurotransmission is maintained at physiologically relevant firing rates in motoneurons. Our results also show that both excitatory and inhibitory connections exhibit considerable convergence of inputs. Because evaluation of the determinants of synaptic strength and the extent of connectivity constitute fundamental parameters affecting the operation of the recurrent circuit, our findings are critical for informing any future models of motor control.
Asunto(s)
Neuronas Motoras/fisiología , Inhibición Neural/fisiología , Células de Renshaw/fisiología , Médula Espinal/citología , Sinapsis/fisiología , Potenciales Sinápticos/fisiología , Animales , Animales Recién Nacidos , Biofisica , Calcio/metabolismo , Estimulación Eléctrica , Femenino , Proteínas de Transporte de Glicina en la Membrana Plasmática/genética , Proteínas de Transporte de Glicina en la Membrana Plasmática/metabolismo , Técnicas In Vitro , Masculino , Potenciales de la Membrana/genética , Ratones , Ratones Transgénicos , Vías Nerviosas/fisiología , Técnicas de Placa-ClampRESUMEN
Although Renshaw cells (RCs) were discovered over half a century ago, their precise role in recurrent inhibition and ability to modulate motoneuron excitability have yet to be established. Indirect measurements of recurrent inhibition have suggested only a weak modulatory effect but are limited by the lack of observed motoneuron responses to inputs from single RCs. Here we present dual recordings between connected RC-motoneuron pairs, performed on mouse spinal cord. Motoneuron responses demonstrated that Renshaw synapses elicit large inhibitory conductances and show short-term potentiation. Anatomical reconstruction, combined with a novel method of quantal analysis, showed that the strong inhibitory input from RCs results from the large number of synaptic contacts that they make onto individual motoneurons. We used the NEURON simulation environment to construct realistic electrotonic models, which showed that inhibitory conductances from Renshaw inputs exert considerable shunting effects in motoneurons and reduce the frequency of spikes generated by excitatory inputs. This was confirmed experimentally by showing that excitation of a single RC or selective activation of the recurrent inhibitory pathway to generate equivalent inhibitory conductances both suppress motoneuron firing. We conclude that recurrent inhibition is remarkably effective, in that a single action potential from one RC is sufficient to silence a motoneuron. Although our results may differ from previous indirect observations, they underline a need for a reevaluation of the role that RCs perform in one of the first neuronal circuits to be discovered.