IDENTIFICATION AND CHARACTERIZATION OF FUNCTIONAL ANGIOTENSIN-II RECEPTORS ON CULTURED HEART MYOCYTES

  • 1 February 1986
    • journal article
    • research article
    • Vol. 236  (2) , 438-444
Abstract
Cultured neonatal rat myocytes have been investigated as a potential system to study the molecular mechanism of angiotensin II (AII) stimulation of heart contractile behavior. Intact cultured cells and the membranes from these cells bind [125I]AII in a biphasic manner. The data were analyzed as two classes of binding sites on membrane preparations: Kd1 = 0.65 nM; maximum binding (Bmax1) = 245 fmol/mg of protein and Kd2 = 5.57 nM, Bmax2 = 720 fmol/mg protein. The receptor on intact cells is specific as AII peptide analogs were potent at inhibiting the binding of [125I]AII. An AII antagonist, [125I]Sar1,Leu8-AII, recognized a single class of high affinity receptors on intact cells: Kd = 0.63 nM, Bmax = 318 fmol/mg of protein, or 45,300 sites per cell. Guanine nucleotides regulated the binding of AII to membranes by shifting the receptor from a high affinity to a low affinity form without changing Kd2. Conversely, these nucleotides altered the high affinity binding of [125I]Sar1,Leu8-AII by increasing Kd without changing Bmax. All was found to stimulate the contractile frequency of spontaneously beating myocytes with maximal effects (a 50% stimulation) observed at 5 nM hormone. The responses were reversible with half-maximal effects observed at doses around 1 nM. The antagonist, Sar1,Ala8-AII, could inhibit the chronotropic stimulatory responses. It is concluded that these cultured myocytes express an AII receptor system with properties conistent with a true hormone receptor system. Furthermore, studies on the pharmacology of the chronotropic responses of these cells to AII demonstrate that the receptors are functional.