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

Abstract

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 (Cav2.2) are inhibited by activation of G protein-coupled opioid receptors (ORs). However, inhibition of R-Type (Cav2.3) channels by μ-or κ-ORs is poorly defined and has not been reported for δ-ORs. To investigate such interactions, we coexpressed human μ-, δ-, or κ-ORs with human Cav2.3 or Cav2.2 in human embryonic kidney 293 cells and measured depolarizationactivated Ba21 currents (IBa). Selective agonists of μ-, δ-, and κ-ORs inhibited IBa through Cav2.3 channels by 35%. Cav2.2 channels were inhibited to a similar extent by κ-ORs, but more potently (60%) via μ-and δ-ORs. Antagonists of δ-and κ-ORs potentiated IBa amplitude mediated by Cav2.3 and Cav2.2 channels. Consistent with G protein bg (Gbg) interaction, modulation of Cav2.2 was primarily voltage-dependent and transiently relieved by depolarizing prepulses. In contrast, Cav2.3 modulation was voltage-independent and unaffected by depolarizing prepulses. However, Cav2.3 inhibition was sensitive to pertussis toxin and to intracellular application of guanosine 59-[b-Thio]diphosphate trilithium salt and guanosine 59-[g-Thio]triphosphate tetralithium salt. Coexpression of Gbgspecific scavengers-namely, the carboxyl terminus of the G protein-coupled receptor kinase 2 or membrane-Targeted myristoylateδ-phosducin-Attenuated or abolished Cav2.3 modulation. Our study reveals the diversity of OR-mediated signaling at Cav2 channels and identifies neuronal Cav2.3 channels as potential targets for opioid analgesics. Their novel modulation is dependent on pre-existing OR activity and mediated by membrane-delimited Gbg subunits in a voltageindependent manner.