Uptake of norepinephrine and related catecholamines by cultured chromaffin cells: characterization of cocaine-sensitive and -insensitive plasma membrane transport sites.

Abstract

Norepinephrine and its closely related analogues, dopamine and epinephrine, are transported into chromaffin cells in culture by two distinct types of sites on the plasma membrane: one is sensitive to cocaine while the other is not. The cocaine-sensitive site has a high affinity for catecholamines and depends on sodium in the medium. The apparent Km for norepinephrine uptake by the cocaine-sensitive site is 5.8 microM when determined in the presence of 118 mM NaCl, obtained using nonlinear least-square curve fitting. Detailed kinetic analysis has also shown cocaine to be a competitive inhibitor of norepinephrine uptake with an apparent Ki of ca. 1 microM. This site is blocked by a series of tricyclic antidepressant drugs with relative potencies characteristic of norepinephrine transport sites in neurons. In contrast, the cocaine-insensitive site(s) have a low affinity for norepinephrine (apparent Km, approximately 88 microM) and are also able to transport catecholamine analogues such as dimethyl-epinephrine and isoproterenol, which have bulky groups attached to the amine moiety. Transport of norepinephrine at both sites is blocked by low temperature, by mitochondrial uncouplers, and by other metabolic inhibitors. Both of these transport sites in the chromaffin cell plasma membrane, therefore, appear to be different from the well-characterized catecholamine transport sites in the chromaffin granule membrane on the basis of substrate specificity and their sensitivity to inhibitors.