Desensitization of the human D(1) dopamine receptor: Evidence for Involvement of both cyclic AMP-dependent and receptor-specific protein kinases.
Human SK-N-MC neurotumor cells express D(1) dopamine receptors coupled to adenylyl cyclase. Following exposure of the cells to dopamine, there was a time- and dose-dependent loss of dopamine responsiveness that involved both a reduction in the maximum response (V(max)) and a shift to less sensitivity in the dose response (K(act)). Although the shift in K(act) occurred more rapidly than the reduction in V(max), both effects were completed within a 20-min exposure of the cells to dopamine. During this rapid period of desensitization, there was no loss of D(1) receptors from membranes prepared from dopamine-treated cells, but there was a reduction in the proportion of agonist high-affinity binding sites. More prolonged exposure to D(1) agonists led to a progressive loss of binding activity. The desensitization was homologous as exposure of the cells, which have beta(1)-adrenergic receptors, to isoproterenol did not alter their response to dopamine. A shift in K(act) but not in V(max) was observed when membranes from control cells were incubated with dopamine, ATP, and the catalytic subunit of cyclic AMP-dependent protein kinase (PKA). To pursue the role of protein kinases in the desensitization process, cells were transiently made permeable, loaded with a PKA inhibitor or with heparin, an inhibitor of the beta-adrenergic receptor kinase, and exposed to dopamine. The PKA inhibitor blocked the shift in K(act), whereas heparin inhibited the reduction in V(max). Our results suggest that desensitization of human D(1) dopamine receptors involves both PKA and a receptor-specific kinase and that the action of both kinases in intact cells requires agonist-occupied receptors.
Zhou
X M
XM
Membrane Biochemistry Section, Laboratory of Molecular and Cellular Neurobiology, National Institute of Neurological Disorders and Stroke, The National Institutes of Health, Bethesda, Maryland 20892, USA.
Sidhu
A
A
Fishman
P H
PH