Differential effects of norepinephrine and phorbol ester on alpha-1 adrenergic receptor number and surface-accessibility in DDT1 MF-2 cells

Abstract

The short (5–60 min) and long (24 hrs) term effects of norephrine (10 uM) and the phorbol ester, 12-O-tetradecanoyl phorbol-13-acetate (10 nM), on total cellular and surface-accessible alpha-1 adrenergic receptor number were determined in DDT1 MF-2 smooth muscle cells. The density of alpha-1 adrenergic receptors was determined with [3H]-prazosin in a crude cellular homogenate (total cellular receptors) and in intact cells at 4°C (surface-accessible receptors). Under basal conditions, all receptors were accessible to the cell surface at 4°C. Short term norepinephrine exposure caused an approximately 40% decrease in surface-accessible binding without a change in total receptor number. Long term norepinephrine exposure caused a further decrease in surface-accessible binding, and an approximately 30 % decrease in total receptor number. In contrast, phorbol ester had no effect on surface-accessible or total receptor number with either short or long term exposure. These data suggest that sequestration of cell surface alpha-1 adrenergic receptors is an early step in the process of agorist-mediated down-regulation. In DDT1 MF-2 cells, phorbol ester, alone, does not mimmick the effect of agonist on receptor sequestration or number.