G protein activation by endomorphins in the mouse periaqueductal gray matter

The midbrain periaqueductal gray matter (PAG) is an important brain region for the coordination of mu-opioid-induced pharmacological actions. The present study was designed to determine whether newly isolated mu-opioid peptide endomorphins can activate G proteins through mu-opioid receptors in the PAG by monitoring the binding to membranes of the non-hydrolyzable analog of GTP, guanosine-5'-O-(3-[S-35]thio]triphosphate ([S-35]GTP gamma S). An autoradiographic [S-35]GTP gamma S binding study showed that both endomorphin-1 and -2 produced similar anatomical distributions of activated G proteins in the mouse midbrain region. in the mouse PAG, endomorphin-1 and -2 at concentrations from 0.001 to 10 mu M increased [S-35]GTP gamma S binding in a concentration-dependent manner and reached a maximal stimulation of 74.6 +/- 3.8 and 72.3 +/- 4.0%, respectively, at 10 mu M In contrast, the synthetic selective mu-opioid receptor agonist [D-Ala(2),NHPhe(4),Gly-ol]enkephalin (DAMGO) had a much greater efficacy and produced a 112.6 +/- 5.1% increase of the maximal stimulation. The receptor specificity of endomorphin-stimulated [S-35]GTP gamma S binding was verified by coincubating membranes with endomorphins in the presence of specific mu-, delta- or kappa-opioid receptor antagonists. Coincubation with selective mu-opioid receptor antagonists beta-funaltrexamine or D-Phe-Cys-Tyr-D-Trp-orn-Thr-Phe-Thr-NH2 (CTOP) blocked both endomorphin-1 and-2-stimulated [S-35]GTP gamma S binding. In contrast, neither delta- nor kappa-opioid receptor antagonist had any effect on the [S-35]GTP gamma S binding stimulated by either endomorphin-1 or -2. These findings indicate that both endomorphin-1 and -2 increase [S-35]GTP gamma S binding by selectively stimulating mu-opioid receptors with intrinsic activity less than that of DAMGO and suggest that these new endogenous ligands might be partial agonists for mu-opioid receptors in the mouse PAG. Copyright (C) 2000 National Science Council. ROC and S. Karger AG, Basel.