Members of the three classes of opioid receptors (μ, δ and κ) have been cloned and characterized in unexcitable cell lines using biochemical techniques. However, an important function of these cloned receptors, their coupling to voltage-activated Ca2+ channels, remains untested. We stably transfected cloned rat μ-opioid receptor cDNAs into clonal pituitary GH3 cells. GH3 cells expressing μ-opioid receptors (GH3MOR cells) bound the receptor-specific ligands [D-Ala2,Me-Phe4,Gly-ol5]-enkephalin (DAMGO) and morphine with high affinity (K(i) = 1.0 and 7.2 nM, respectively), and these ligands also potently inhibited adenylyl cyclase activity (IC50 = 21.9 and 55.2 nM, respectively). Functional coupling of μ-opioid receptors to voltage-activated Ca2+ channels was compared with that of endogenous somatostatin (SRIF) receptors in GH3MOR cells, using the patch-clamp technique, with Ba2+ as the charge carrier. DAMGO (1 μM) and SRIF (1 μM) inhibited Ba2+ currents by 23.8 ± 1.0% and 22.9 ± 2.5%, respectively. DAMGO (0.1 nM to 10 μM) dose-dependently inhibited Ba2+ currents, with an IC50 of 105 nM. The μ-opioid receptor agonist morphine (1 μM) inhibited currents by 13.5 ± 1.1% and the δ-opioid receptor-selective ligand [D- Pen2.5]-enkephalin (1 μM) caused only 3.5 ± 2.1% inhibition. The inhibitory actions of DAMGO, morphine, and [D-Pen2,5]-enkephalin were reversed by naloxone. Ba2+ current inhibitions by DAMGO and SRIF were attenuated by pertussis toxin pretreatment. Nimodipine reduced the amplitude of Ba2+ current inhibition by DAMGO, suggesting that μ-opioid receptors couple to L-type Ca2+ channels in GH3MOR cells.
|Original language||English (US)|
|Number of pages||9|
|State||Published - 1995|