Neuroligin 1 (NLGN1) is a postsynaptic adhesion protein that binds presynaptic neurexins and undergoes activity dependent cleavage. We examined the time course of NLGN1 cleavage after photothrombotic stroke, its relationship to synaptic structural changes, and whether neuronal stimulation enhances cleavage in the peri-infarct cortex.We induced photothrombotic stroke in the mouse cortex and measured soluble NLGN1(sNLGN1) and full-length NLGN1(FL-NLGN1) by immunoblotting in peri-infarct tissue at 3, 7, 14, and 28 days. At day 7, we examined synaptic ultrastructure by electron microscopy and measured PSD-95 and VGLUT2 in synaptosome enriched fractions. We treated primary rat cortical neurons with bicuculine for 24 hours, with or without metalloprotease inhibitors. In a separate cohort, we optogenetically stimulated the peri-infarct cortex during the second week after stroke and collected tissue on day 14. Peri-infarct sNLGN1 levels were highest at day 7 and declined thereafter. At day 7, synaptic clefts were wider and postsynaptic densities were shorter than in sham mice. PSD-95 levels were lower in synaptosome-enriched fractions, whereas VGLUT2 levels did not differ significantly. In cultured neurons, bicuculline increased sNLGN1 in conditioned medium and decreased full-length NLGN1 in cell lysates. GM6001 and INCB3619 each significantly attenuated both bicuculline-induced changes. At day 14 after stroke, optogenetically stimulated mice had higher sNLGN1 and lower full-length NLGN1 levels in the peri-infarct cortex than controls. NLGN1 cleavage peaked when peri-infarct synapses showed structural changes. The culture experiments implicate metalloprotease activity in activity-dependent NLGN1 cleavage, and the optogenetic results suggest that neuronal stimulation can enhance cleavage during the subacute phase of stroke. Whether NLGN1 cleavage contributes to synaptic remodeling and functional recovery remains to be determined.