Cloned δ-Opioid Receptors in GH3 Cells Inhibit Spontaneous Ca2+ Oscillations and Prolactin Release ThroughK IR Channel Activation

Abstract

Opioid receptors can couple to K+ and Ca2+ channels, adenylyl cyclase, and phosphatidyl inositol turnover. Any of these actions may be important in the regulation of neurotransmitter and hormone release from excitable cells. GH3 cells exhibit spontaneous oscillations of intracellular Ca2+concentration ([Ca2+]i) and prolactin release. Activation of cloned δ-opioid receptors stably expressed in GH3 cells inhibits both spontaneous Ca2+signaling and basal prolactin release. The objective of this study was to examine a possible role for K+ channels in these processes using the patch-clamp technique, fluorescence imaging, and a sensitive ELISA for prolactin. The selective δ receptor agonist [d-Pen2,d-Pen2]enkephalin (DPDPE) inhibited [Ca2+]i oscillations in GH3 cells expressing both μ and δ receptors (GH3MORDOR cells) but had no effect on control GH3 cells or cells expressing μ receptors alone (GH3MOR cells). The inhibition of [Ca2+]i oscillations by DPDPE was unaffected by thapsigargin pretreatment, suggesting that this effect is independent of inositol 1,4,5-triphosphate-sensitive Ca2+ stores. DPDPE caused a concentration-dependent inhibition of prolactin release from GH3MORDOR cells with an IC50 of 4 nM. DPDPE increased inward K+current recorded from GH3MORDOR cells but had no significant effect on K+ currents recorded from control GH3 cells or GH3MOR cells. The μ receptor agonist morphine also had no effect on currents recorded from control cells but activated inward K+ currents recorded from GH3MOR and GH3MORDOR cells. Somatostatin activated inward currents recorded from all three cell lines. The DPDPE-sensitive K+ current was inwardly rectifying and was inhibited by Ba2+ but not TEA. DPDPE had no effect on delayed rectifier-, Ca2+-, and voltage-activated or A-type K+ currents, recorded from GH3MORDOR cells. Ba2+ attenuated the inhibition of [Ca2+]i and prolactin release by DPDPE, whereas TEA had no effect, consistent with an involvement of K IR channels in these actions of the opioid.