A Pertussis Toxin Substrate Regulates α1-Adrenergic Dependent Phosphatidylinositol Hydrolysis in Cultured Rat Myocytes*

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Abstract
The chronotropic response of the heart to axadrenergic catecholamines is influenced by pertussis toxin under certain conditions. In view of the fact that α1-adrenergic action is mediated by the phosphatidylinositol pathway of hormone action in many cells, we examined the hypothesis that α-adrenergic agonists stimulate phosphatidylinositol hydrolysis in cardiomyocytes and that this effect is sensitive to pertussis toxin. Addition of norepinephrine to cultured rat ventricular myocytes prelabeled with myo-[2-3H]inositol resulted in rapid and significant accumulation of inositol phosphate (IP1) and inositol biphosphate. Norepinephrine-stimulated IP1 formation was not inhibited by propranolol, but was inhibited by α-adrenergic antagonists with an order of potency indicating α1-adrenergic receptor subselectivity: prazosin (α1i; 3 nM) > yohimbine (α2; 10 μM). The effect of norepinephrine to enhance IP1 formation was markedly attenuated in cells pretreated with pertussis toxin. Pertussis toxin also induced the transfer of ADP-ribose from NAD to a 41,000-dalton membrane protein in these cells. The concentration of pertussis toxin resulting in maximal inhibition of norepinephrine-stimulated IP1 formation correlated well with the concentration of pertussis toxin necessary to completely ADP-ribosylate a 41,000-dalton membrane protein (1 ng/ml). The range over which pertussis toxin inhibited norepinephrinedependent IP1 formation and ADP-ribosylated the 41,000-dalton substrate was virtually identical. These observations establish a role for a 41,000-dalton pertussis toxin substrate in coupling the α1-adrenergic receptor to phosphoinositol hydrolysis in myocardial cells. (Endocrinology120: 1889–1895, 1987)