RIM genes differentially contribute to organizing presynaptic release sites

Tight coupling of Ca2+ channels to the presynaptic active zone is critical for fast synchronous neurotransmitter release. RIMs are multidomain proteins that tether Ca2+ channels to active zones, dock and prime synaptic vesicles for release, and mediate presynaptic plasticity. Here, we use conditional knockout mice targeting all RIM isoforms expressed by the Rims1 and Rims2 genes to examine the contributions and mechanism of action of different RIMs in neurotransmitter release. We show that acute single deletions of each Rims gene decreased release and impaired vesicle priming but did not alter the extracellular Ca2+-responsiveness of release (which for Rims gene mutants is a measure of presynaptic Ca2+ influx). Moreover, single deletions did not affect the synchronization of release (which depends on the close proximity of Ca2+ channels to release sites). In contrast, deletion of both Rims genes severely impaired the Ca2+ responsiveness and synchronization of release. RIM proteins may act on Ca2+ channels in two modes: They tether Ca2+ channels to active zones, and they directly modulate Ca2+-channel inactivation. The first mechanism is essential for localizing presynaptic Ca2+ influx to nerve terminals, but the role of the second mechanism remains unknown. Strikingly, we find that although the RIM2 C2B domain by itself significantly decreased Ca2+-channel inactivation in transfected HEK293 cells, it did not rescue any aspect of the RIM knockout phenotype in cultured neurons. Thus, RIMs primarily act in release as physical Ca2+-channel tethers and not as Ca2+-channel modulators. Different RIM proteins compensate for each other in recruiting Ca2+ channels to active zones, but contribute independently and incrementally to vesicle priming.