EPHB2 FORWARD SIGNALING REGULATES PARVALBUMIN INTERNEURON INHIBITORY STRENGTH, NETWORK EXCITABILITY, AND BEHAVIOR

Disruption of excitatory–inhibitory balance contributes to hyperexcitable neuronal networks in neurodevelopmental disorders. Although the receptor tyrosine kinase EphB2 is a major regulator of excitatory synapse development, its role in inhibitory circuit function remains unclear. Parvalbumin interneurons provide powerful perisomatic inhibition onto CA1 pyramidal cells, and changes in their synaptic efficacy strongly influence hippocampal network excitability. We previously showed that deletion of EphB2 from parvalbumin cells increases presynaptic parvalbumin/vGAT sites and strengthens parvalbumin-to-pyramidal cell connectivity in CA1, suggesting that EphB2 negatively regulates inhibitory output. Here, we directly examined how EphB2 forward signaling controls inhibitory synaptic function and behavior using genetic gain- and loss-of-function approaches. EphB2 kinase activity was manipulated with point mutants F620D (kinase-hyperactive) and K661R (kinase-dead). Whole-cell patch-clamp recordings from CA1 pyramidal cells revealed that kinase-dead K661R mice exhibit enhanced inhibitory input, reflected by increased spontaneous inhibitory postsynaptic currents, whereas kinase-hyperactive F620D mice show reduced inhibitory drive. These functional changes were accompanied by corresponding structural alterations: K661R mice displayed increased parvalbumin/vGAT and vGAT/gephyrin colocalization, higher puncta density in the stratum pyramidale, and tighter presynaptic–postsynaptic alignment, while F620D mice showed reduced overlap and more dispersed inhibitory synapses. Importantly, these synaptic and electrophysiological effects translated to behavior. Loss of EphB2 signaling in parvalbumin cells conferred protection against pentylenetetrazole-induced seizures, indicating reduced network hyperexcitability in vivo. Together, our findings demonstrate that EphB2 kinase activity in parvalbumin interneurons negatively regulates inhibitory synaptic strength, hippocampal excitability, and seizure susceptibility, highlighting EphB2 as a key modulator of inhibitory circuit function and excitatory–inhibitory balance.