Gγ7 proteins contribute to coupling of nociceptin/orphanin FQ peptide (NOP) opioid receptors and voltage-gated Ca²⁺ channels in rat stellate ganglion neurons

The nociceptin/orphanin FQ peptide (NOP) opioid receptors regulate neurotransmitter release via inhibition of voltage-gated Ca²⁺ channels (Ca_V2.2) in sympathetic and sensory neurons. Stimulation of NOP receptors by its endogenous agonist, nociception (Noc), leads to membrane-delimited, voltage-dependent (VD) block of Ca_V2.2 channel currents mediated by Gβγ protein subunits. Previously we reported that the pertussis toxin-sensitive Gαi1 and Gβ2/β4 isoforms mediate the functional coupling of NOP opioid receptors with Ca_V channels in rat stellate ganglion (SG) sympathetic neurons. In the present report we extended our studies by identifying the Gγ subunit that forms the heterotrimer within this signaling pathway. Small interference RNA (or siRNA) was employed to silence the expression of the natively expressed Gγ subunits. Initial PCR assays indicated that SG neurons expressed seven Gγ subunits. Silencing Gγ3 subunits did not alter signaling between NOP receptors and Ca²⁺ channels. However, after Gγ7 isoforms were silenced, the Noc-mediated inhibition of Ca_V channels was significantly decreased when compared to SG neurons transfected with scrambled siRNA. We observed that Gγ10 and Gγ11 mRNA levels increased 2.5- and 2.7-fold, respectively, after Gγ7 subunits were silenced. However, this compensatory increase in mRNA expression did not appear to fully rescue the NOP receptor coupling efficiency. Additionally, both Gγ2 and Gγ5 levels increased 50 and 75%, respectively, while Gγ3 and Gγ4 expression levels remained relatively unchanged. Taken together, our findings suggest that the Gαi1/Gβ2(β4)/Gγ7 heterotrimeric G protein complex determines the NOP receptor-mediated modulation of Ca_V channels in SG neurons.