Regulation and functional characterization of a rat recombinant dopamine D3 receptor

Barbara A. Cox, Mary P. Rosser, Michal R. Kozlowski, Kevin M. Duwe, Rachael L. Neve, Kim A. Neve

Research output: Contribution to journalArticle

40 Scopus citations

Abstract

We stably expressed a rat D3 receptor cDNA in C6 glioma cells (C6‐D3 cells), quantifying receptor expression with the radioligands [125I]epidepride (KD = 0.1 nM) and [3H]spiperone (KD = 0.7 nM). As reported previously for D2 receptors, quinpirole induced a 9–16% increase in the rate of extracellular acidification by C6‐D3 cells. The acidification was inhibited by epidepride and by the Na+/H+ antiporter inhibitors, amiloride and methylisobutylamiloride, but pertussis toxin treatment had no effect on quinpirole‐induced extracellular acidification. These data suggest that D3 receptor stimulation of Na+/H+ exchange in C6 glioma cells is not mediated by the pertussis toxinsensitive G proteins, Gi or Go. Overnight treatment of C6‐D3 cells with N‐propylnorapomorphine, dopamine, or quinpirole resulted in large concentration‐dependent increases (up to 500%) in the density of D3 receptors on membranes prepared from the cells. Antagonists had smaller, variable effects on the density of D3 receptors in C6‐D3 cells, except for domperidone, which significantly increased the density of D3 receptors. Treatment with pertussis toxin had no effect on the agonist‐induced receptor up‐regulation, indicating that an interaction with pertussis toxin‐sensitive G proteins was not required. Densitometry analysis of Northern blots of RNA prepared from C6‐D3 cells showed no significant N‐propylnorapomorphine‐induced increase in D3 receptor message. Treatment with cycloheximide, however, completely prevented receptor up‐regulation by N‐propylnorapomorphine. Pretreatment of C6‐D2 cells with 1'0 pM DA resulted in a substantial heterologous sensitization, in which isoproterenol‐stimulated adenylyl cyclase activity was enhanced more than twofold. In contrast, isoproterenol‐stimulated enzyme activity was inhibited by greater than 50% in C6‐D3 cells pretreated with dopamine. These results confirm one functional response to activation of D3 receptors and demonstrate that the density of D3 receptors, like D2 receptors, is increased after incubation of intact cells with agonists. © 1995 Wiley‐Liss, Inc.

Original languageEnglish (US)
Pages (from-to)1-9
Number of pages9
JournalSynapse
Volume21
Issue number1
DOIs
StatePublished - Sep 1995

Keywords

  • Antiporter
  • Signal transduction
  • cDNA

ASJC Scopus subject areas

  • Cellular and Molecular Neuroscience

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