Voltage-Gated R-Type Calcium Channel Inhibition via Human μ-, δ-, and κ-opioid Receptors Is Voltage-Independently Mediated by Gβγ Protein Subunits

Elucidating the mechanisms that modulate calcium channels via opioid receptor activation is fundamental to our understanding of both pain perception and how opioids modulate pain. Neuronal voltage-gated N-type calcium channels (Ca(v)2.2) are inhibited by activation of G protein-coupled opioid receptors (ORs). However, inhibition of R-type (Ca(v)2.3) channels by mu- or kappa-ORs is poorly defined and has not been reported for delta-ORs. To investigate such interactions, we coexpressed human mu-, delta-, or kappa-ORs with human Ca(v)2.3 or Ca(v)2.2 in human embryonic kidney 293 cells and measured depolarization-activated Ba2+ currents (I-Ba). Selective agonists of mu-, delta-, and kappa-ORs inhibited I-Ba through Ca(v)2.3 channels by 35%. Ca(v)2.2 channels were inhibited to a similar extent by kappa-ORs, but more potently (60%) via mu- and delta-ORs. Antagonists of delta- and kappa-ORs potentiated I-Ba amplitude mediated by Ca(v)2.3 and Ca(v)2.2 channels. Consistent with G protein beta gamma (G beta gamma) interaction, modulation of Ca(v)2.2 was primarily voltage-dependent and transiently relieved by depolarizing prepulses. In contrast, Ca(v)2.3 modulation was voltage-independent and unaffected by depolarizing prepulses. However, Ca(v)2.3 inhibition was sensitive to pertussis toxin and to intracellular application of guanosine 5'-[beta-thio] diphosphate trilithium salt and guanosine 5'-[gamma-thio] triphosphate tetralithium salt. Coexpression of G beta gamma-specific scavengers-namely, the carboxyl terminus of the G protein-coupled receptor kinase 2 or membrane-targeted myristoylated-phosducin-attenuated or abolished Ca(v)2.3 modulation. Our study reveals the diversity of OR-mediated signaling at Ca(v)2 channels and identifies neuronal Ca(v)2.3 channels as potential targets for opioid analgesics. Their novel modulation is dependent on pre-existing OR activity and mediated by membrane-delimited G beta gamma subunits in a voltage-independent manner.