Received for publication May 10, 2007.
Revised August 1, 2007.
Accepted for publication August 1, 2007.

THE ENDOGENOUS CANNABINOID ANANDAMIDE INHIBITS CROMAKALIM-ACTIVATED K⁺ CURRENTS IN FOLLICLE-ENCLOSED XENOPUS OOCYTES

Abstract

The effect of the endogenous cannabinoid anandamide on K⁺ currents activated by the K_ATP channel opener cromakalim was investigated in follicle-enclosed Xenopus oocytes using the two-electrode voltage-clamp technique. Anandamide (1-90 µM) reversibly inhibited cromakalim-induced K⁺ currents with an IC₅₀ value of 8.1 ± 2 µM. Inhibition was noncompetitive and independent of membrane potential. Pretreatment of oocytes with the CB₁ receptor antagonist SR141716A (1 µM), the CB₂ receptor antagonist SR144528 (1 µM), or pertussis toxin (5 µg/ml) did not alter the inhibitory effect of anandamide, suggesting that known cannabinoid receptors are not involved in anandamide inhibition of K⁺ currents. Similarly, neither the amidohydrolase inhibitor, phenylmethylsulfonyl fluoride (0.2 mM) nor the cyclooxygenase inhibitor, indomethacin (5 µM) affected anandamide inhibition of K⁺ currents suggesting that the effects of anandamide are not mediated by its metabolic products. In radioligand binding studies, anandamide inhibited the specific binding of the K_ATP ligand [³H]glibenclamide in the oocyte microsomal fractions with an IC₅₀ value of 6.3 ± 0.4 µM. Gonadotropin-induced oocyte maturation and the cromakalim-acceleration of progesterone-induced oocyte maturation were significantly inhibited in the presence of 10 µM anandamide. Collectively, these results indicate that cromakalim-activated K⁺ currents in follicular cells of Xenopus oocytes are modulated by anandamide via a cannabinoid receptor-independent mechanism and the inhibition of these channels by anandamide alters the responsiveness of oocytes to gonadotropin and progesterone.

Key words: KATP channels, Xenopus oocytes, anandamide, cromakalim, electrophysiology, oocyte development