RhoA regulates morphine associated NMDA receptor signaling in the prelimbic region of medial prefrontal cortex pyramidal neurons

Opioid dependence (OD) involves maladaptive neuroplasticity in brain reward circuits, particularly within the medial prefrontal cortex (mPFC). While RhoA and NMDA receptors (NMDARs) are implicated in addiction-related synaptic plasticity, their specific interaction within mPFC subregions remains unclear. Using male Sprague-Dawley rats (6 weeks old), we investigated the role of RhoA signaling in the prelimbic cortex (PLC) via behavioral, molecular biological, and electrophysiological assays. Intra-PLC infusion of the RhoA inhibitor Rhosin significantly attenuated morphine-induced conditioned place preference and locomotor sensitization. Furthermore, repeated morphine administration (RMA) upregulated RhoA expression in layer 5 pyramidal neurons. In vitro whole-cell patch-clamp recordings of layer 5 neurons, stimulated at layer 2/3, revealed that Rhosin reduced the amplitude of synaptic NMDAR-mediated excitatory postsynaptic currents. Additionally, using an activity-dependent MK-801 block to isolate extrasynaptic components, we demonstrated that RhoA inhibition significantly attenuated extrasynaptic NMDAR activation, likely by limiting glutamate spillover during high-frequency stimulation. These findings elucidate a critical mechanism by which RhoA mediates opioid-induced neuroadaptations through the regulation of both synaptic and extrasynaptic NMDAR activity, identifying RhoA in the PLC as a promising therapeutic target for opioid dependence.