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

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 micromol/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 micromol/L). O-1918, a cannabidiol analog, had a similar blocking effect in arteries of both wild-type and CB(1)(-/-) mice at 10 micromol/L. In contrast, 1 micromol/L SR144538, a cannabinoid type 2 receptor antagonist, or 50 micromol/L 18alpha-glycyrrhetinic acid, a gap junction blocker, were without effect. SR141716A (1 to 30 micromol/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.