CB 1 Receptor Antagonist SR141716A Inhibits Ca 2+ -Induced Relaxation in CB 1 Receptor-Deficient Mice

Abstract

Mesenteric branch arteries isolated from cannabinoid type 1 receptor knockout (CB 1 −/− ) mice, their wild-type littermates (CB 1 +/+ mice), and C57BL/J wild-type mice were studied to test the hypothesis that murine arteries undergo high sensitivity Ca 2+ -induced relaxation that is CB 1 receptor dependent. Confocal microscope analysis of mesenteric branch arteries from wild-type mice showed the presence of Ca 2+ receptor-positive periadventitial nerves. Arterial segments of C57 control mice mounted on wire myographs contracted in response to 5 μmol/L norepinephrine and responded to the cumulative addition of extracellular Ca 2+ with a concentration-dependent relaxation that reached a maximum of 72.0±6.3% of the prerelaxation tone and had an EC 50 for Ca 2+ of 2.90±0.54 mmol/L. The relaxation was antagonized by precontraction in buffer containing 100 mmol/L K + and by pretreatment with 10 mmol/L tetraethylammonium. Arteries from CB 1 −/− and CB 1 +/+ mice also relaxed in response to extracellular Ca 2+ with no differences being detected between the knockout and their littermate controls. SR141716A, a selective CB 1 antagonist, caused concentration-dependent inhibition of Ca 2+ -induced relaxation in both the knockout and wild-type strains (60% inhibition at 1 μmol/L). O-1918, a cannabidiol analog, had a similar blocking effect in arteries of both wild-type and CB 1 −/− mice at 10 μmol/L. In contrast, 1 μmol/L SR144538, a cannabinoid type 2 receptor antagonist, or 50 μmol/L 18α-glycyrrhetinic acid, a gap junction blocker, were without effect. SR141716A (1 to 30 μmol/L) was also assessed for nonspecific actions on whole-cell K + currents in isolated vascular smooth muscle cells. SR141716A inhibited macroscopic K + currents at concentrations higher than those required to inhibit Ca 2+ -induced relaxation, and appeared to have little effect on currents through large conductance Ca 2+ -activated K + channels. These data indicate that arteries of the mouse relax in response to cumulative addition of extracellular Ca 2+ in a hyperpolarization-dependent manner and rule out a role for CB 1 or CB 2 receptors in this effect. The possible role of a nonclassical cannabinoid receptor is discussed.