Anna Velica: The role of Chrna2+ Martinotti cells in the primary motor cortex: A study using retrograde rabies virus-based tracing and chemogenetics.

Abstract

This thesis investigates the connectivity of Chrna2+ Martinotti cells (MCα2s) and their influence on cortical pyramidal cell (PC) assemblies during motor learning and execution. In paper I, we developed a systematic flowchart for control experiments in monosynaptic rabies virus–based tracing. Following the flowchart, we identified Cre-independent leakiness of the TVA receptor in RΦGT responder mice, resulting in nonspecific labeling. In paper II, we observed different experimental outcomes depending on the agonist dose during chemogenetic activation of MCα2s, illustrating the complexity of the tool. In paper III, we mapped monosynaptic presynaptic inputs to MCα2s in the forelimb area of the primary motor cortex using rabies virus-based tracing. Traced cells were predominantly PCs in the ipsilateral primary motor cortex (72.1%) and secondary motor cortex (12.9%), with additional input from the primary somatosensory cortex (9.8%), thalamus (3.3%), contralateral cortex (1.2%), basal ganglia and basal forebrain. Local inhibitory input was mainly originating from parvalbumin interneurons near the injection site. Chemogenetic activation of MCα2s increased falls in the hanging wire test, indicating a role in sensorimotor integration. In paper IV, we examined how MCα2 activity modulates PC assemblies during motor learning using chemogenetic activation combined with calcium imaging. During a session in the skilled forelimb single-pellet reaching task, in which the pellet presentation was more challenging, Chrna2-Cre+ mice exhibited a restricted cortical assembly area and reduced session-to-session changes in assembly composition compared to controls. These results suggest that elevated MCα2 excitability constrains PC assembly plasticity during relearning. Moreover, chemogenetic activation of MCα2s increased success rates in the skilled forelimb single-pellet reaching task in mice that had already learnt the task and enhanced theta-band local field potential power, indicating a role in stabilization and refinement of learned motor behaviors. Collectively, these studies provide an integrative view of MCα2 connectivity and function, indicating that MCα2s influence both sensorimotor integration and PC assembly plasticity, with distinct roles during learning versus execution. They further emphasize the necessity of careful controls and the combined use of viral tracing, chemogenetics and population calcium imaging to dissect the circuit mechanisms underlying skilled motor behavior.