REGULATION OF HEPATOCYTE PLASMA-MEMBRANE ALPHA(1)-ADRENERGIC RECEPTORS BY 4-BETA-PHORBOL 12-MYRISTATE 13-ACETATE

The effect of phorbol 12-myristate 13-acetate (PMA) on hepatocyte alpha(1)-adrenergic receptors was determined by [H-3]prazosin binding to plasma membranes from control and PMA-treated hepatocytes. Membranes from hepatocytes incubated with PMA (1 mu g/ml) for 1 h exhibited a 40% decrease in alpha(1)-adrenergic receptors (481+/-10 fmol/mg of protein; mean+/-S.E.M. for three separate experiments) relative to vehicle-treated (dimethylformamide) hepatocytes (802+/-91 fmol/mg of protein; n = 3), with no significant effect on the K-D. The PMA-induced decrease in alpha(1)-adrenergic receptors was maximal by 30 min and half-maximal inhibition of [H-3]prazosin binding occurred with a PMA concentration of approx. 15 ng/ml. Pretreatment of hepatocytes with staurosporine (5 mu M) blocked the effect of PMA, and 4 beta-phorbol 13-monoacetate was ineffective, suggesting the involvement of protein kinase C (PKC). Treatment of hepatocytes with primaquine (300 mu M) for 15 min decreased hepatocyte plasma membrane alpha(1)-adrenergic receptors by 34.0+/-2.4% (mean+/-S.E.M. of three experiments). Removal of primaquine allowed essentially complete recovery (98+/-4%; mean+/-S.E.M. for five separate experiments) of plasma membrane [H-3]prazosin binding within 20 min, suggesting that the alpha(1)-adrenergic receptor undergoes endocytotic recycling. Addition of PMA (1 pg/ml) to hepatocytes immediately after removal of primaquine, completely inhibited the increase in plasma membrane alpha(1)-adrenergic receptors relative to control cells, but had no effect on hepatocytes whose cell surface alpha(1)-receptors remaining after primaquine treatment had been inactivated by alkylation. These observations suggested that activation of PKC may facilitate the internalization of the alpha(1)-adrenergic receptor in hepatocytes.