Inhibition of human recombinant T-type calcium channels by phytocannabinoids in vitro

Background and Purpose: T-type Ca channels (I _Ca) regulate neuronal excitability and contribute to neurotransmitter release. The phytocannabinoids Δ ⁹-tetrahydrocannabinol and cannabidiol effectively modulate T-type I _Ca, but effects of other biologically active phytocannabinoids on these channels are unknown. We thus investigated the modulation of T-type I _Ca by low abundance phytocannabinoids. Experimental Approach: A fluorometric (fluorescence imaging plate reader [FLIPR]) assay was used to investigate modulation of human T-type I _Ca (Ca _V3.1, 3.2 and 3.3) stably expressed in FlpIn-TREx HEK293 cells. The biophysical effects of some compounds were examined using whole-cell patch clamp recordings. Key Results: In the FLIPR assay, all 11 phytocannabinoids tested modulated T-type I _Ca, with most inhibiting Ca _V3.1 and Ca _V3.2 more effectively than Ca _V3.3. Cannabigerolic acid was the most potent inhibitor of Ca _V3.1 (pIC ₅₀ 6.1 ± 0.6) and Ca _V3.2 (pIC ₅₀ 6.4 ± 0.4); in all cases, phytocannabinoid acids were more potent than their corresponding neutral forms. In patch clamp recordings, cannabigerolic acid inhibited Ca _V3.1 and 3.2 with similar potency to the FLIPR assay; the inhibition was associated with significant hyperpolarizing shift in activation and steady-state inactivation of these channels. In contrast, cannabidiol, cannabidivarin, and cannabigerol only affected channel inactivation. Conclusion and Implications: Modulation of T-type calcium channels is a common property of phytocannabinoids, which all increase steady-state inactivation at physiological membrane potentials, with some also affecting channel activation. Thus, T-type I _Ca may be a common site of action for phytocannabinoids, and the diverse actions of phytocannabinoids on channel gating may provide insight into structural requirement for selective T-type I _Ca modulators.