Genistein elicits biphasic effects on L-type Ca21 current in feline atrial myocytes

Wang, Yong G., and Stephen L. Lipsius. Genistein elicits biphasic effects on L-type Ca21 current in feline atrial myocytes. Am. J. Physiol. 275 (Heart Circ. Physiol. 44): H204–H212, 1998.—A perforated patch recording method was used to determine the effects of genistein (Gen), a protein tyrosine kinase (PTK) inhibitor, on basal L-type Ca21 current (ICa,L) in feline atrial myocytes. Gen (50 μM) elicited biphasic changes in ICa,L: an initial inhibition (255 6 4%; phase 1) followed by a secondary stimulation (34 6 9%; phase 2) of ICa,L. Withdrawal of Gen elicited a further potentiation of ICa,L (152 6 19%; phase 3) above control (n 5 46). In general, phase 1 inhibition and phase 3 potentiation varied directly with Gen concentration, and phase 2 stimulation exhibited biphasic concentration-dependent changes compared with control. When cells were dialyzed using a ruptured patch recording method, Gen elicited only inhibition of ICa,L; phases 2 and 3 were abolished. Vanadate (1 mM), an inhibitor of protein tyrosine phosphatase, abolished both Gen-induced inhibition and stimulation of ICa,L. Daidzein (50 μM), a weakly active analog of Gen, exerted no significant effects on ICa,L, and withdrawal of daidzein failed to potentiate ICa,L. In a few cells, Gen elicited a prominent vanadate-sensitive stimulation of ICa,L in the absence of any significant inhibition of ICa,L. Gen-induced changes in ICa,L were unaffected by either 100 μM 1,2-bis(2-aminophenoxy)ethane-N,N,N8,N8-tetraacetic acid (BAPTA)-acetoxymethyl ester (AM) or 1 μM ryanodine, agents that alter intracellular Ca21; 4 μM H-89 or 50 μM Rp diastereomer of adenosine 38,58-monophosphothioate (RP-cAMPS), inhibitors of protein kinase A (PKA); 0.1 μM calphostin C or 2 μM chelerythrine, inhibitors of protein kinase C (PKC); or 100 μM NG-monomethyl-L-arginine (LNMMA), an inhibitor of nitric oxide (NO) synthase. We conclude that in feline atrial myocytes, Gen acts via membrane-bound PTKs to inhibit ICa,L and via cytosolic PTKs to stimulate ICa,L. Gen-induced changes in ICa,L are not related to changes in intracellular Ca21 or to secondary interactions with either PKA, PKC, or NO signaling pathways. These results indicate that in atrial myocytes ICa,L is regulated by two independent and competing PTK signaling mechanisms.