Depression of synaptic outputs by reduction of Ca2+ influx, but not RRP vesicles, becomes evident upon CB2R hyper-expression at cerebellar Purkinje cell terminals.

Synaptic transmission is dynamically regulated by neuromodulators. One well-studied example is cannabinoid receptor type 1 (CB1R)-mediated suppression of neurotransmitter release by reducing presynaptic Ca2+ influx. Recently, GPR55 was shown to regulate transmitter release through a mechanism distinct from that of CB1R. However, the presynaptic role of cannabinoid receptor type 2 (CB2R) remains unclear. Here we studied this issue using cerebellar Purkinje cells (PCs), which are known to express both CB2R and GPR55 and amenable to direct patch-clamp recordings from axon terminals. At naïve PC synapses onto target neurons, application of a CB2R agonist did not affect synaptic transmission. The lack of effect was ascribed to minimal endogenous CB2R at distal axon terminals of PCs. Exogenous expression of CB2R enabled suppression of synaptic transmission upon pharmacological activation. Direct voltage-clamp recordings of presynaptic Ca2+ current and membrane capacitance changes at boutons demonstrated that CB2R activation reduced presynaptic Ca2+ influx without affecting the total amount of readily releasable vesicles, leading to less vesicle exocytosis through lowered release probability. Notably, replacement of two intracellular loops of CB2R with those of GPR55 converted the site of action for synaptic suppression from Ca2+ channel inhibition to the reduction of the readily releasable vesicles, as GPR55 does. These findings indicate that, when sufficiently present at axon terminals, CB2R suppresses transmitter release through reduction of Ca2+ influx like CB1R, and suggest that intracellular loops of cannabinoid receptors determine whether transmitter release is suppressed through inhibition of presynaptic Ca2+ channels or reduction of the readily releasable pool of vesicles.