AP5

Inhibitory control of external urethral sphincter motor neurons (EUS‐MNs) in the spinal dorsolateral nucleus (DLN), which corresponds to a portion of Onuf's nucleus in humans, is essential for normal micturition by inducing EUS relaxation during voiding; yet synaptic mechanisms remain poorly characterized. Using neonatal mice P8‐P12, we developed a slicing technique—cutting spinal cords at 150° from the coronal plane (30° from the horizontal plane in the agarose block), for maximizing EUS‐MNs captured per slice. Using transgenic mice co‐expressing channelrhodopsin‐2 in inhibitory interneurons (VGAT‐ChR2) and GFP in cholinergic neurons (ChAT‐GFP), we investigated inhibitory synaptic transmission onto EUS‐MNs. Optogenetic activation evoked robust inhibitory postsynaptic potentials (IPSPs), classified as sustained or transient based on temporal profiles. Pharmacology revealed that sustained IPSPs contained both glycinergic and GABAergic components, while GABAA receptors predominantly mediated transient IPSPs. Strychnine (1 μM) selectively blocked glycinergic transmission, while bicuculline (10 μM) eliminated GABAergic components. Insensitivity to glutamatergic antagonists (CNQX and AP5) confirmed purely inhibitory responses. Our findings demonstrate segregation of inhibitory inputs onto EUS‐MNs, with glycinergic and GABAergic transmission contributing to sustained and transient inhibition, respectively, establishing the methodological foundation for investigating inhibitory circuit dynamics in pathological conditions such as spinal cord injury with deficient inhibitory control.

Chronic pain is well-known for inducing anxiety and depression in humans. Enriched environment (EE) has shown promising effects in alleviating mood disorders; however, the precise mechanisms remain poorly understood. The medial prefrontal cortex (mPFC), a pivotal brain region associated with emotion regulation, particularly through excitatory synaptic transmission, is implicated in the processing of pain-related mood disorders.

First, adult male C57BL/6 mice 6 weeks after spared nerve injury surgery, were subjected to the behavioral tests and dendritic spine morphology staining. Then, 2 weeks after spared nerve injury (SNI), mice were housed in enriched environment, to establish model EE. Behavioral outcomes, dendritic spine maturation, and AMPA and NMDA receptors expression were assessed up to 4 weeks after EE treatment using behavioral tests, Golgi staining, Western blot, and pharmacological experiments.

Our results showed that SNI mice exhibit anxiety and depression-like behaviors. Further analysis revealed that SNI mice displayed a significant reduction in mature dendritic spines in the mPFC. After 4 weeks of housekeeping in EE, dendritic complexity, spine maturity, and the expression of AMPAR and NMDAR of SNI mice were restored, leading to a rescue of anxiety and depression-like behaviors. Moreover, pharmacological antagonism of AMPAR or NMDAR, by intraperitoneal injection of AP-5 or CNQX, effectively abolished the alleviating effect of EE on anxiety and depression-like behaviors in SNI mice.

Together, our findings uncovered a previously unrecognized AMPAR and NMDAR-dependent mechanism of EE in mitigates anxiety and depression-like behaviors in SNI mice via restoration of mPFC excitatory synaptic transmission.