STUDIES ON THE EFFECTS OF ANANDAMIDE IN RAT HEPATIC-ARTERY

1 The effects of anandamide on K+ currents and membrane potential have been examined in freshly-isolated smooth muscle cells from rat hepati c artery and the results compared with the effects of this arachidonic acid derivative on tension and membrane potential changes in segments of whole artery. 2 In the presence of 0.3 mM L-NOARG and 10 mu M indo methacin, anandamide (0.1-100 mu M) and endothelium-derived hyperpolar izing factor (EDHF; liberated by acetylcholine, 0.01-10 mu M) each rel axed endothelium-intact segments of hepatic artery precontracted with phenylephrine. These effects of anandamide, but not those of EDHF, wer e antagonized by the cannabinoid receptor antagonist, SR141716A (3 mu M). 3 The relaxant effects of anandamide were unaffected by a toxin co mbination (apamin plus charybdotoxin, each 0.3 mu M) which abolishes E DHF relaxations and were essentially unchanged in endothelium-denuded arteries. The relaxant effects of anandamide in endothelium-intact art eries were significantly reduced in a physiological salt solution cont aining 30 mM KCl and abolished when the K+ concentration was raised to 60 mM. 4 Anandamide (10 mu M), acetylcholine (1 mu M, via release of EDHF) and levcromakalim (10 mu M) each markedly hyperpolarized the mem brane potential of the smooth muscle cells of endothelium-intact arter ies. However, when the endothelium was removed, the hyperpolarizing ef fects of both anandamide (10 mu M) and acetylcholine were essentially abolished whereas those of levcromakalim (10 mu M) were unaffected. 5 Under voltage-clamp conditions, anandamide (10 mu M) abolished spontan eous transient outward currents (STOCs) in freshly-isolated single hep atic artery cells held at 0 mV but had no effect on the holding curren t at this potential. In current-clamp mode, the spontaneous hyperpolar izing potentials which corresponded to the STOCs were abolished with n o significant change in basal membrane potential. 6 Anandamide (10 mu M) abolished the iberiotoxin-sensitive K+ current (I-BK(Ca)) produced by caffeine and the corresponding hyperpolarizations generated by this xanthine derivative in current-clamp mode. In contrast, anandamide ha d no effect on I-BK(Ca) generated on exposure to NS1619 (30 mu M). 7 I t was concluded that anandamide is not EDHF in the rat hepatic artery. Anandamide-induced hyperpolarization is exerted indirectly and requir es the presence of the endothelium. Anandamide also acts on the smooth muscle cells to inhibit processes which require functional intracellu lar calcium stores. This direct action seems more important than membr ane hyperpolarization in relaxing phenylephrine-contracted vessels.