Actomyosin-mediated inhibition of synaptic vesicle release under CB1R activation.

Long-term synaptic plasticity is critical for adaptive function of the brain, but presynaptic mechanisms of functional plasticity remain poorly understood. Here, we show that changes in synaptic efficacy induced by activation of the cannabinoid type-1 receptor (CB1R), one of the most widespread G-protein coupled receptors in the brain, requires contractility of the neuronal actomyosin cytoskeleton. Specifically, using a synaptophysin-pHluorin probe (sypH2), we show that inhibitors of non-muscle myosin II (NMII) ATPase as well as one of its upstream effectors Rho-associated kinase (ROCK) prevent the reduction of synaptic vesicle release induced by CB1R activation. Using 3D STORM super-resolution microscopy, we find that activation of CB1R induces a redistribution of synaptic vesicles within presynaptic boutons in an actomyosin dependent manner, leading to vesicle clustering within the bouton and depletion of synaptic vesicles from the active zone. We further show, using sypH2, that inhibitors of NMII and ROCK specifically restore the release of the readily releasable pool of synaptic vesicles from the inhibition induced by CB1R activation. Finally, using slice electrophysiology, we find that activation of both NMII and ROCK is necessary for the long-term, but not the short-term, form of CB1R induced synaptic plasticity at excitatory cortico-striatal synapses. We thus propose a novel mechanism underlying CB1R-induced plasticity, whereby CB1R activation leads to a contraction of the actomyosin cytoskeleton inducing a reorganization of the functional presynaptic vesicle pool, preventing vesicle release and inducing long-term depression.

Feed-forward recruitment of electrical synapses enhances synchronous spiking in the mouse cerebellar cortex.

In the cerebellar cortex, molecular layer interneurons use chemical and electrical synapses to form subnetworks that fine-tune the spiking output of the cerebellum. Although electrical synapses can entrain activity within neuronal assemblies, their role in feed-forward circuits is less well explored. By combining whole-cell patch-clamp and 2-photon laser scanning microscopy of basket cells (BCs), we found that classical excitatory postsynaptic currents (EPSCs) are followed by GABAA receptor-independent outward currents, reflecting the hyperpolarization component of spikelets (a synapse-evoked action potential passively propagating from electrically coupled neighbors). FF recruitment of the spikelet-mediated inhibition curtails the integration time window of concomitant excitatory postsynaptic potentials (EPSPs) and dampens their temporal integration. In contrast with GABAergic-mediated feed-forward inhibition, the depolarizing component of spikelets transiently increases the peak amplitude of EPSPs, and thus postsynaptic spiking probability. Therefore, spikelet transmission can propagate within the BC network to generate synchronous inhibition of Purkinje cells, which can entrain cerebellar output for driving temporally precise behaviors.

Cannabinoid-induced actomyosin contractility shapes neuronal morphology and growth.

Endocannabinoids are recently recognized regulators of brain development, but molecular effectors downstream of type-1 cannabinoid receptor (CB1R)-activation remain incompletely understood. We report atypical coupling of neuronal CB1Rs, after activation by endo- or exocannabinoids such as the marijuana component ∆(9)-tetrahydrocannabinol, to heterotrimeric G12/G13 proteins that triggers rapid and reversible non-muscle myosin II (NM II) dependent contraction of the actomyosin cytoskeleton, through a Rho-GTPase and Rho-associated kinase (ROCK). This induces rapid neuronal remodeling, such as retraction of neurites and axonal growth cones, elevated neuronal rigidity, and reshaping of somatodendritic morphology. Chronic pharmacological inhibition of NM II prevents cannabinoid-induced reduction of dendritic development in vitro and leads, similarly to blockade of endocannabinoid action, to excessive growth of corticofugal axons into the sub-ventricular zone in vivo. Our results suggest that CB1R can rapidly transform the neuronal cytoskeleton through actomyosin contractility, resulting in cellular remodeling events ultimately able to affect the brain architecture and wiring.