RhoA activation promotes ordered membrane domain coalescence and suppresses neuronal excitability

Abstract

The formation of ordered proteolipid membrane domains (OMDs) within the plasma membrane has emerged as a fundamental process that modifies membrane function, particularly in response to cell stresses that promote pathological states. Here, we identify a previously unrecognized role for the small GTPase RhoA to promote the coalescence of OMDs, thereby linking cytoskeletal remodeling and membrane mechanics to OMD formation. Pharmacological and optogenetic manipulation of RhoA rapidly altered OMD dimensions in both human cell lines and dorsal root ganglion (DRG) nociceptors. The RhoA-dependent OMD expansion required actin remodeling, changes in membrane mechanical state, and protein palmitoylation. Functionally, RhoA inhibition increased action potential firing and potentiated HCN channel activity in DRG neurons. Conversely, in a spared nerve injury model characterized by altered membrane mechanics, reduced OMD size, and hyperexcitability, RhoA activation enlarged OMDs, suppressed HCN channel activity, and reduced firing. These findings highlight alterations in plasma membrane physical properties, including changes in OMD organization and membrane tension, as key features of neuropathic stress. RhoA/ROCK-driven OMD remodeling may serve as a compensatory membrane adaptation that counteracts neuropathic hyperexcitability.