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(Circulation Research. 2002;91:232.)
© 2002 American Heart Association, Inc.

Cellular Biology

₁-Adrenoceptor-Mediated Breakdown of Phosphatidylinositol 4,5-Bisphosphate Inhibits Pinacidil-Activated ATP-Sensitive K⁺ Currents in Rat Ventricular Myocytes

Tetsuya Haruna, Hidetada Yoshida, Tomoe Y. Nakamura, Lai-Hua Xie, Hideo Otani, Tomonori Ninomiya, Makoto Takano, William A. Coetzee, Minoru Horie

From the Departments of Cardiovascular Medicine (T.H., H.Y., H.O., T.N., M.H.) and Physiology and Biophysics (L.-H.X., M.T.), Kyoto University Graduate School of Medicine, Kyoto, Japan, and Pediatric Cardiology (T.Y.N., W.A.C.), NYU School of Medicine, New York, NY.

Correspondence to Dr Minoru Horie, Department of Cardiovascular Medicine, Graduate School of Medicine, Kyoto University, 54 Shogoin Kawahara-cho Sakyo-ku, Kyoto 606-8507, Japan. E-mail horie{at}kuhp.kyoto-u.ac.jp

Phosphatidylinositol 4,5-bisphosphate (PIP₂) stimulates ATP-sensitive K⁺ (K_ATP) channel activity. Because phospholipase C (PLC) hydrolyzes membrane-bound PIP₂, which in turn may potentially decrease K_ATP channel activity, we investigated the effects of the ₁-adrenoceptor-G_q-PLC signal transduction axis on pinacidil-activated K_ATP channel activity in adult rat and neonatal mouse ventricular myocytes. The ₁-adrenoceptor agonist methoxamine (MTX) reversibly inhibited the pinacidil-activated K_ATP current in a concentration-dependent manner (IC₅₀ 20.9±6.6 µmol/L). This inhibition did not occur when the specific ₁-adrenoceptor antagonist, prazosin, was present. An involvement of G proteins is suggested by the ability of GDPßS to prevent this response. Blockade of PLC by U-73122 (2 µmol/L) or neomycin (2 mmol/L) attenuated the MTX-induced inhibition of K_ATP channel activity. In contrast, the MTX response was unaffected by protein kinase C inhibition or stimulation by H-7 (100 µmol/L) or phorbol 12,13-didecanoate. The MTX-induced inhibition became irreversible in the presence of wortmannin (20 µmol/L), an inhibitor of phosphatidylinositol-4 kinase, which is expected to prevent membrane PIP₂ replenishment. In excised inside-out patch membranes, pinacidil induced a significantly rightward shift of ATP sensitivity of the channel. This phenomenon was reversed by pretreatment of myocytes with MTX. Direct visualization of PIP₂ subcellular distribution using a PLC pleckstrin homology domain-green fluorescent protein fusion constructs revealed reversible translocation of green fluorescent protein fluorescence from the membrane to the cytosol after ₁-adrenoceptor stimulation. Our data demonstrate that ₁-adrenoceptor stimulation reduces the membrane PIP₂ level, which in turn inhibits pinacidil-activated K_ATP channels.

Key Words: ATP-sensitive K⁺ channels • phosphatidylinositol 4,5-bisphosphate • ₁-adrenoceptors

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