Developmental disruption of recurrent inhibitory feedback results in compensatory adaptation in the Renshaw cell - motor neuron circuit

Enjin A, Perry S, Hilscher MM, Nagaraja C, Larhammar M, Gezelius H, Eriksson A, Leão KE, Kullander K (2017) Developmental disruption of recurrent inhibitory feedback results in compensatory adaptation in the Renshaw cell - motor neuron circuit. J Neurosci. May 8. pii: 0949-16. doi: 10.1523/JNEUROSCI.0949-16.2017.When activating muscles, motor neurons in the spinal cord also activate Renshaw cells, which provide recurrent inhibitory feedback to the motor neurons. The tight coupling with motor neurons suggests that Renshaw cells have an integral role in movement, a role that is yet to be elucidated. Here we used the selective expression of the nicotinic cholinergic receptor alpha 2 (Chrna2) in mice to genetically target the vesicular inhibitory amino acid transporter (VIAAT) in Renshaw cells. Loss of VIAAT from Chrna2Cre expressing Renshaw cells did not impact any aspect of drug-induced fictive locomotion in the neonatal mouse, nor did it change gait, motor coordination or grip strength in adult mice of both sexes. However, motor neurons from neonatal mice lacking VIAAT in Renshaw cells received spontaneous inhibitory synaptic input with a reduced frequency, showed lower input resistance and had an increased number of proprioceptive glutamatergic and calbindin labeled putative Renshaw cell synapses on their soma and proximal dendrites. Concomitantly, Renshaw cells developed with increased excitability and a normal number of cholinergic motor neuron synapses indicating a compensatory mechanism within the recurrent inhibitory feedback circuit. Our data suggest an integral role for Renshaw cell signaling in shaping the excitability and synaptic input to motor neurons

Deciphering the Locomotor Network : The Role of Spinal Cord Interneurons

In the spinal cord, an intricate neural network generates and coordinates the patterning of limb movements during locomotion. This network, known as the locomotor central pattern generator (CPG), comprises of various cell populations that together orchestrate the output of motor neurons. Identification of CPG neurons through their specific gene expression is a valuable tool that can provide considerable insight to the character, intrinsic properties and role of a population, which represents a step toward understanding locomotor circuit function and correlating neural activity to behaviour. We selectively targeted two inhibitory CPG populations to investigate their molecular characteristics, circuitry and functional role; Renshaw cells (RCs) marked by their specific expression of the cholinergic nicotinic receptor α2 (Chrna2) and a subset of the dI6 population derived by their selective expression of the Doublesex and mab-3 related transcription factor 3 (Dmrt3). We found that RCs have hyperpolarisation-activated cation (Ih) and small calcium-activated potassium (ISK) modulatory currents that differentially regulate their excitation and firing properties, which influence the instantaneous feedback to motor neurons through the recurrent inhibition circuit. Due to previous difficulties isolating RCs from the surrounding locomotor circuits, their functional role remains poorly defined. For the first time, we selectively silenced RC inhibition and found that all aspects of motor behaviour, including coordination and gait were normal. The deletion of RC signalling instead altered the electrical and synaptic properties of the recurrent inhibitory circuit, suggesting that developmental plasticity compensates for the loss of RC inhibition. We reveal Dmrt3 neurons comprise a population of glycinergic inhibitory, spike-frequency adapting commissural interneurons active during locomotion. Conditional silencing of the Dmrt3 population resulted in considerable gait abnormalities in the neonatal and adult mouse. This manifested as an uncoordinated CPG output in vitro, impaired limb coordination in pups and increased fore- and hindlimb synchrony in adults that was exacerbated at faster locomotor speeds. Dmrt3 mediated inhibition subsequently impacts locomotion and suggests the Dmrt3 population contribute to coordinating speed dependent left-right limb alternation. This thesis provides cellular, circuit and behavioural insights into the Renshaw cell and Dmrt3 populations and enhances our knowledge regarding their probable function within the locomotor CPG

The alpha2 nicotinic acetylcholine receptor, a subunit with unique and selective expression in inhibitory interneurons associated with principal cells

Nicotinic acetylcholine receptors (nAChRs) play crucial roles in various human disorders, with the alpha 7, alpha 4, alpha 6, and alpha 3-containing nAChR subtypes extensively studied in relation to conditions such as Alzheimer's disease, Parkinson's disease, nicotine dependence, mood disorders, and stress disorders. In contrast, the alpha 2-nAChR subunit has received less attention due to its more restricted expression and the scarcity of specific agonists and antagonists for studying its function. Nevertheless, recent research has shed light on the unique expression pattern of the Chrna2 gene, which encodes the alpha 2-nAChR subunit, and its involvement in distinct populations of inhibitory interneurons. This review highlights the structure, pharmacology, localization, function, and disease associations of alpha 2-containing nAChRs and points to the unique expression pattern of the Chrna2 gene and its role in different inhibitory interneuron populations. These populations, including the oriens lacunosum moleculare (OLM) cells in the hippocampus, Martinotti cells in the neocortex, and Renshaw cells in the spinal cord, share common features and contribute to recurrent inhibitory microcircuits. Thus, the alpha 2-nAChR subunit's unique expression pattern in specific interneuron populations and its role in recurrent inhibitory microcircuits highlight its importance in various physiological processes. Further research is necessary to uncover the comprehensive functionality of alpha 2-containing nAChRs, delineate their specific contributions to neuronal circuits, and investigate their potential as therapeutic targets for related disorders

Spinal Cord Interneurons Expressing the Gastrin-Releasing Peptide Receptor Convey Itch Through VGLUT2-Mediated Signaling

Itch is a sensation that promotes the desire to scratch, which can be evoked by mechanical and chemical stimuli. In the spinal cord, neurons expressing the gastrin-releasing peptide receptor (GRPR) have been identified as specific mediators of itch. However, our understanding of the GRPR population in the spinal cord, and thus how these neurons exercise their functions, is limited. For this purpose, we constructed a Cre line designed to target the GRPR population of neurons (Grpr-Cre). Our analysis revealed that Grpr-Cre cells in the spinal cord are predominantly excitatory interneurons that are found in the dorsal lamina, especially in laminae II-IV. Application of the specific agonist gastrin-releasing peptide induced spike responses in 43.3% of the patched Grpr-Cre neurons, where the majority of the cells displayed a tonic firing property. Additionally, our analysis showed that the Grpr-Cre population expresses Vglut2 mRNA, and mice ablated of Vglut2 in Grpr-Cre cells (Vglut2-lox; Grpr-Cre mice) displayed less spontaneous itch and attenuated responses to both histaminergic and nonhistaminergic agents. We could also show that application of the itch-inducing peptide, natriuretic polypeptide B, induces calcium influx in a subpopulation of Grpr-Cre neurons. To summarize, our data indicate that the Grpr-Cre spinal cord neural population is composed of interneurons that use VGLUT2-mediated signaling for transmitting chemical and spontaneous itch stimuli to the next, currently unknown, neurons in the labeled line of itch.Title in thesis list of papers: Spinal Cord Interneurons Expressing the Gastrin Releasing Peptide Receptor Convey Itch through VGLUT2-mediated Signaling</p

Neuron-specific inactivation of Wt1 alters locomotion in mice and changes interneuron composition in the spinal cord

Locomotion is coordinated by neuronal circuits of the spinal cord. Recently, dI6 neurons were shown to participate in the control of locomotion. A subpopulation of dI6 neurons expresses the Wilms tumor suppressor gene Wt1. However, the function of Wt1 in these cells is not understood. Here, we aimed to identify behavioral changes and cellular alterations in the spinal cord associated with Wt1 deletion. Locomotion analyses of mice with neuron-specific Wt1 deletion revealed a slower walk with a decreased stride frequency and an increased stride length. These mice showed changes in their fore-/hindlimb coordination, which were accompanied by a loss of contralateral projections in the spinal cord. Neonates with Wt1 deletion displayed an increase in uncoordinated hindlimb movements and their motor neuron output was arrhythmic with a decreased frequency. The population size of dI6, V0, and V2a neurons in the developing spinal cord of conditional Wt1 mutants was significantly altered. These results show that the development of particular dI6 neurons depends on Wt1 expression and that loss of Wt1 is associated with alterations in locomotion