Neurexin-2 restricts synapse numbers and restrains the presynaptic release probability by an alternative splicing-dependent mechanism.

α- and β-neurexins are extensively alternatively spliced, presynaptic cell-adhesion molecules that are thought to organize synapse assembly. However, recent data revealed that, in the hippocampus in vivo, the deletion of one neurexin isoform, , surprisingly increased excitatory synapse numbers and enhanced their presynaptic release probability, suggesting that restricts, instead of enabling, synapse assembly. To delineate the synaptic function and mechanism of action of , we examined cultured hippocampal neurons as a reduced system. In heterologous synapse formation assays, different alternatively spliced Nrxn2β isoforms robustly promoted synapse assembly similar to Nrxn1β and Nrxn3β, consistent with a general synaptogenic function of neurexins. Deletion of from cultured hippocampal neurons, however, caused a significant increase in synapse density and release probability, replicating the in vivo data that suggested a synapse-restricting function. Rescue experiments revealed that two of the four Nrxn2β splice variants (Nrxn2β-SS4+/SS5- and Nrxn2β-SS4+/SS5+) reversed the increase in synapse density in -deficient neurons, whereas only one of the four Nrxn2β splice variants (Nrxn2β-SS4+/SS5+) normalized the increase in release probability in -deficient neurons. Thus, a subset of splice variants restricts synapse numbers and restrains their release probability in cultured neurons.