Activation of CB1 cannabinoid receptors in the cerebellum acutely depresses excitatory synaptic transmission at parallel fibre-Purkinje cell synapses by decreasing the probability of glutamate release. This depression involves the activation of presynaptic 4-aminopyridine-sensitive K(+) channels by CB1 receptors, which in turn inhibits presynaptic Ca(2+) influx controlling glutamate release at these synapses. Using rat cerebellar frontal slices and fluorometric measures of presynaptic Ca(2+) influx evoked by stimulation of parallel fibres with the fluorescent dye fluo-4FF, we tested whether the CB1 receptor-mediated inhibition of this influx also involves a direct inhibition of presynaptic voltage-gated calcium channels. Since various physiological effects of CB1 receptors appear to be mediated through the activation of PTX-sensitive proteins, including inhibition of adenylate cyclases, activation of mitogen-activated protein kinases (MAPK) and activation of G protein-gated inwardly rectifying K(+) channels, we also studied the potential involvement of these intracellular signal transduction pathways in the cannabinoid-mediated depression of presynaptic Ca(2+) influx. The present study demonstrates that the molecular mechanisms underlying the CB1 inhibitory effect involve the activation of the PTX-sensitive G(i)/G(o) subclass of G proteins, independently of any direct effect on presynaptic Ca(2+) channels (N, P/Q and R (SNX-482-sensitive) types) or on adenylate cyclase or MAPK activity, but do require the activation of G protein-gated inwardly rectifying (Ba(2+)- and tertiapin Q-sensitive) K(+) channels, in addition to 4-aminopyridine-sensitive K(+) channels.