α-Neurexins Together with α2δ-1 Auxiliary Subunits Regulate Ca2+ Influx through Cav2.1 Channels

Action potential-evoked neurotransmitter release is impaired in knock-out neurons lacking synaptic cell-adhesion molecules alpha-neurexins (alpha Nrxns), the extracellularly longer variants of the three vertebrate Nrxn genes. Ca2+ influx through presynaptic high-voltage gated calcium channels like the ubiquitous P/Q-type (Ca(V)2.1) triggers release of fusion-ready vesicles at many boutons. alpha 2 delta Auxiliary subunits regulate trafficking and kinetic properties of Ca(V)2.1 pore-forming subunits but it has remained unclear if this involves alpha Nrxns. Using live cell imaging with Ca2+ indicators, we report here that the total presynaptic Ca2+ influx in primary hippocampal neurons of alpha Nrxn triple knock-out mice of both sexes is reduced and involved lower Ca(V)2.1-mediated transients. This defect is accompanied by lower vesicle release, reduced synaptic abundance of Ca(V)2.1 pore-forming subunits, and elevated surface mobility of alpha 2 delta-1 on axons. Overexpression of Nrxn1 alpha in alpha Nrxn triple knock-out neurons is sufficient to restore normal presynaptic Ca2+ influx and synaptic vesicle release. Moreover, coexpression of Nrxn1 alpha together with alpha 2 delta-1 subunits facilitates Ca2+ influx further but causes little augmentation together with a different subunit, alpha 2 delta-3, suggesting remarkable specificity. Expression of defined recombinant Ca(V)2.1 channels in heterologous cells validates and extends the findings from neurons. Whole-cell patch-clamp recordings show that Nrxn1 alpha in combination with alpha 2 delta-1, but not with alpha 2 delta-3, facilitates Ca2+ currents of recombinant Ca(V)2.1 without altering channel kinetics. These results suggest that presynaptic Nrxn1 alpha acts as a positive regulator of Ca2+ influx through Ca(V)2.1 channels containing alpha 2 delta-1 subunits. We propose that this regulation represents an important way for neurons to adjust synaptic strength.