δ-Opioid-Induced Liberation of Gβγ Mobilizes Ca²⁺ Stores in NG108-15 Cells

Abstract

Activation of δ-opioid receptors in NG108-15 cells releases Ca²⁺ from an intracellular store through activation of a pertussis toxin-sensitive G protein. We tested the hypothesis that activation of δ-opioid receptors mobilizes inositol 1,4,5-trisphosphate (IP₃)-sensitive Ca²⁺stores via liberation of Gβγ. Fura-2-based digital imaging was used to study the mechanism of opioid-induced increases in [Ca²⁺]_i in NG108-15 cells. Exposure tod-Ala²-d-Leu⁵enkephalin (100 nM) for 90 s induced increases in [Ca²⁺]_i that were blocked by microinjection of the IP₃ receptor antagonist heparin (pipette concentration = 100 mg/ml) but not by sham injection. Microinjection of a peptide that binds Gβγ (QEHA, 1 mM) decreased the d-Ala²-d-Leu⁵enkephalin-evoked response. Microinjection of an inactive peptide (SKEE, 1 mM) that does not bind to Gβγ failed to inhibit the opioid-induced increase in [Ca²⁺]_i. Microinjection of a peptide (QLKK, 15 mM) that binds to free Gα_q blocked the increase evoked by 3 nM bradykinin, but microinjection of an inactive peptide (ADRK, 15 mM) did not. Microinjection of QLKK did not significantly affect the opioid-induced increase in [Ca²⁺]_i. Collectively, these data demonstrate that activation of δ-opioid receptors induces the release of Ca²⁺ from IP₃-sensitive stores in NG108-15 cells through activation of the βγ subunits of inhibitory G proteins.