Altered dynamics of urination-related ON and OFF neurons in the primary motor cortex during detrusor overactivity

The coordination of bladder function with behavior depends on distributed neural circuits, but cortical activity associated with detrusor overactivity (DO) remains incompletely understood. We used miniature two-photon calcium imaging to record neuronal activity in the primary motor cortex (M1) of freely moving mice during urination. Individual neurons were classified as Direct-ON, Direct-OFF, or Direct-Other according to a permutation-based comparison between their calcium activity and synchronized urination behavior. In DO mice, Direct-OFF neurons exhibited increased activity around urination onset, whereas Direct-ON neurons exhibited increased activity around urination termination. Differences in the spatial distribution of these functionally classified neuronal populations were also observed. Trospium chloride altered urination behavior, while selected abnormalities in M1 neuronal activity and spatial organization remained detectable. Pyridostigmine bromide did not produce clear changes in the measured behavioral or neuronal abnormalities. In trospium-treated DO mice, region-wide chemogenetic inhibition of M1 was associated with additional changes in urination frequency and duration. Because functionally defined ON/OFF neuronal subtypes were not selectively manipulated and a DO plus chemogenetic-inhibition-only group was not included, the present findings identify altered M1 neuronal dynamics as a cortical correlate of bladder dysfunction but do not establish M1 activity as its primary cause or demonstrate the therapeutic efficacy of combined pharmacological and chemogenetic intervention.