Axonal CB1 Receptors Mediate Inhibitory Bouton Formation via cAMP Increase and PKA

Experience-dependent formation and removal of inhibitory synapses are essential throughout life. For instance, GABAergic synapses are removed to facilitate learning, and strong excitatory activity is accompanied by the formation of inhibitory syn-apses to maintain coordination between excitation and inhibition. We recently discovered that active dendrites trigger the growth of inhibitory synapses via CB1 receptor-mediated endocannabinoid signaling, but the underlying mechanism remained unclear. Using two-photon microscopy to monitor the formation of individual inhibitory boutons in hippocampal organotypic slices from mice (both sexes), we found that CB1 receptor activation mediated the formation of inhibitory boutons and pro-moted their subsequent stabilization. Inhibitory bouton formation did not require neuronal activity and was independent of Gi/o-protein signaling, but was directly induced by elevating cAMP levels using forskolin and by activating Gs-proteins using DREADDs. Blocking PKA activity prevented CB1 receptor-mediated inhibitory bouton formation. Our findings reveal that axonal CB1 receptors signal via unconventional downstream pathways and that inhibitory bouton formation is triggered by an increase in axonal cAMP levels. Our results demonstrate an unexpected role for axonal CB1 receptors in axon-specific, and context-dependent, inhibitory synapse formation.

Automated summary

The study demonstrates that inhibitory synapse formation and stabilization in hippocampal organotypic slices is mediated by CB1 receptor activation and involves unconventional downstream pathways. Neuronal activity is not required for inhibitory bouton formation, which can be induced by elevating cAMP levels and activating Gs-proteins. Inhibition of PKA activity prevents the CB1 receptor-mediated formation of inhibitory boutons.