Abstract
The dopamine transporter (DAT) regulates dopamine (DA) neurotransmission by recapturing DA into the presynaptic terminals and is a principal target of the psychostimulant cocaine. The sigma-1 receptor (σ1R) is a molecular chaperone, and its ligands have been shown to modulate DA neuronal signaling, although their effects on DAT activity are unclear. Here, we report that the prototypical σ1R agonist (+)-pentazocine potentiated the dose response of cocaine self-administration in rats, consistent with the effects of the σR agonists PRE-084 and DTG (1,3-di-o-tolylguanidine) reported previously. These behavioral effects appeared to be correlated with functional changes of DAT. Preincubation with (+)-pentazocine or PRE-084 increased the Bmax values of [3H]WIN35428 binding to DAT in rat striatal synaptosomes and transfected cells. A specific interaction between σ1R and DAT was detected by co-immunoprecipitation and bioluminescence resonance energy transfer assays. Mutational analyses indicated that the transmembrane domain of σ1R likely mediated this interaction. Furthermore, cysteine accessibility assays showed that σ1R agonist preincubation potentiated cocaine-induced changes in DAT conformation, which were blocked by the specific σ1R antagonist CM304. Moreover, σ1R ligands had distinct effects on σ1R multimerization. CM304 increased the proportion of multimeric σ1Rs, whereas (+)-pentazocine increased monomeric σ1Rs. Together these results support the hypothesis that σ1R agonists promote dissociation of σ1R multimers into monomers, which then interact with DAT to stabilize an outward-facing DAT conformation and enhance cocaine binding. We propose that this novel molecular mechanism underlies the behavioral potentiation of cocaine self-administration by σ1R agonists in animal models.
Highlights
Neurotransmission by recapturing DA into the presynaptic terminals and is a principal target of the psychostimulant cocaine
Cysteine accessibility assays showed that 1R agonist preincubation potentiated cocaine-induced changes in dopamine transporter (DAT) conformation, which were blocked by the specific 1R antagonist CM304
To understand why preincubation with 1R agonists increased cocaine binding (Fig. 2), we examined whether 1R drugs could modulate the DAT conformation
Summary
A recent behavioral study showed that 1R agonists such as PRE-084 and 1,3-di-o-tolylguanidine (DTG), but not 1R antagonists, increased the potency of cocaine in a self-administration procedure [22]. In the present study we investigated the effects of 1R ligands on DAT function using behavioral, pharmacological, and biochemical methods. We present several lines of evidence supporting the hypothesis that interaction of 1R with DAT modulates the outward-facing conformation of DAT and facilitates cocaine binding to DAT. This molecular mechanism underscores a novel modulatory role of 1R on DA neurotransmission and suggests therapeutic potentials of 1R ligands in treating cocaine addiction
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