Low- and high-cocaine locomotor responding rats differ in reinstatement of cocaine seeking and striatal mGluR5 protein expression.

Behavioral responsiveness to initial cocaine use varies among individuals and may contribute to differential vulnerability to cocaine addiction. Rats also exhibit individual differences in cocaine's effects and can be classified as low or high cocaine responders (LCRs or HCRs, respectively), based on their initial cocaine-induced locomotor activity (10 mg/kg, i.p.). Here, we used the extinction/reinstatement model to address whether or not LCRs and HCRs differ in (i) extinction/reinstatement of cocaine self-administration behavior and (ii) levels of metabotropic glutamate receptors (mGluRs) following these behaviors. During the earliest acquisition sessions, LCRs exhibited significantly greater cocaine intake (0.8 mg/kg/infusion) and cocaine-paired lever responding than HCRs, but intake and lever responding converged by the end of the cocaine self-administration portion of the study. LCRs and HCRs did not differ in cocaine seeking during the first extinction session and extinguished cocaine seeking similarly. HCRs exhibited greater reinstatement than LCRs to lower (2.5 and 5 mg/kg), but not higher (10 mg/kg), i.p. priming doses of cocaine. The effect of drug-paired cues on reinstatement following extinction was complex, with HCRs and LCRs showing the greater effect of cue depending on the order in which cue- and drug-primed tests were given. Western blot analysis revealed that mGluR5 heteromers were significantly higher in the dorsal striatum of HCRs than LCRs following reinstatement testing. Although our previous findings with the LCR/HCR model have uniformly supported the idea that lower initial cocaine-induced activation predicts more ready development of cocaine addiction-like behaviors, here, we show a more complex relationship with cocaine reinstatement.

Striatal regulation of ΔFosB, FosB, and cFos during cocaine self-administration and withdrawal.

Chronic drug exposure induces alterations in gene expression profiles that are thought to underlie the development of drug addiction. The present study examined regulation of the Fos-family of transcription factors, specifically cFos, FosB, and ΔFosB, in striatal subregions during and after chronic intravenous cocaine administration in self-administering and yoked rats. We found that cFos, FosB, and ΔFosB exhibit regionally and temporally distinct expression patterns, with greater accumulation of ΔFosB protein in the nucleus accumbens (NAc) shell and core after chronic cocaine administration, whereas ΔFosB increases in the caudate-putamen (CPu) remained similar with either acute or chronic administration. In contrast, tolerance developed to cocaine-induced mRNA for ΔFosB in all three striatal subregions with chronic administration. Tolerance also developed to FosB expression, most notably in the NAc shell and CPu. Interestingly, tolerance to cocaine-induced cFos induction was dependent on volitional control of cocaine intake in ventral but not dorsal striatal regions, whereas regulation of FosB and ΔFosB was similar in cocaine self-administering and yoked animals. Thus, ΔFosB-mediated neuroadaptations in the CPu may occur earlier than previously thought with the initiation of intravenous cocaine use and, together with greater accumulation of ΔFosB in the NAc, could contribute to addiction-related increases in cocaine-seeking behavior.

Role of GluR1 expression in nucleus accumbens neurons in cocaine sensitization and cocaine-seeking behavior.

Chronic cocaine use reduces glutamate levels in the nucleus accumbens (NAc), and is associated with experience-dependent changes in (+/-)-alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) glutamate receptor membrane expression in NAc neurons. These changes accompany behavioral sensitization to cocaine and increased susceptibility to cocaine relapse. The functional relationship between neuroplasticity in AMPA receptors and the behavioral manifestation of cocaine addiction remains unclear. Thus, we examined the behavioral effects of up- and downregulating basal AMPA receptor function in the NAc core and shell using viral-mediated gene transfer of wild-type glutamate receptor 1 (wt-GluR1) or a dominant-negative pore-dead GluR1 (pd-GluR1), respectively. Transient increases in wt-GluR1 during or after cocaine treatments diminished the development of cocaine sensitization, while pd-GluR1 expression exacerbated cocaine sensitization. Parallel changes were found in D2, but not D1, receptor-mediated behavioral responses. As a correlate of the sensitization experiments, we overexpressed wt- or pd-GluR1 in the NAc core during cocaine self-administration, and tested the effects on subsequent drug-seeking behavior 3 weeks after overexpression declined. wt-GluR1 overexpression during self-administration had no effect on cocaine intake, but subsequently reduced cocaine seeking in extinction and cocaine-induced reinstatement, whereas pd-GluR1 facilitated cocaine-induced reinstatement. When overexpressed during reinstatement tests, wt-GluR1 directly attenuated cocaine- and D2 agonist-induced reinstatement, while pd-GluR1 enhanced reinstatement. In both experimental procedures, neither wt- nor pd-GluR1 expression affected cue-induced reinstatement. Together, these results suggest that degrading basal AMPA receptor function in NAc neurons is sufficient to facilitate relapse via sensitization in D2 receptor responses, whereas elevating basal AMPA receptor function attenuates these behaviors.

Local gene knockdown in the brain using viral-mediated RNA interference.

Conditional mutant techniques that allow spatial and temporal control over gene expression can be used to create mice with restricted genetic modifications. These mice serve as powerful disease models in which gene function in adult tissues can be specifically dissected. Current strategies for conditional genetic manipulation are inefficient, however, and often lack sufficient spatial control. Here we use viral-mediated RNA interference (RNAi) to generate a specific knockdown of Th, the gene encoding the dopamine synthesis enzyme tyrosine hydroxylase, within midbrain neurons of adult mice. This localized gene knockdown resulted in behavioral changes, including a motor performance deficit and reduced response to a psychostimulant. These results underscore the potential of using viral-mediated RNAi for the rapid production and testing of new genetic disease models. Similar strategies may be used in other model species, and may ultimately find applications in human gene therapy.

Antagonistic effects of dopaminergic signaling and ethanol on protein kinase A-mediated phosphorylation of DARPP-32 and the NR1 subunit of the NMDA receptor.

BACKGROUND: This article extends our initial investigation of the interactions between dopamine and glutamate receptor systems after acute exposure to ethanol. DARPP-32 (dopamine and cyclic adenosine monophosphate-regulated phosphoprotein of approximate molecular weight 32 kDa) is an important regulator of protein phosphatase-1 that in turn regulates a large number of effectors, including the NMDA receptor. METHODS: We measured the protein kinase A (PKA)-mediated phosphorylation of DARPP-32 and the NR1 subunit of the NMDA receptor. Initially, corpus striatum was assayed after intraperitoneal treatment of mice with the D1 agonist SKF82958, the D2 agonist and anticraving drug bromocriptine, or ethanol. In other experiments we blocked D1 receptors with the selective D1 antagonist SCH23390 or blocked D2 receptors with the selective D2 antagonist eticlopride. Finally, we examined combinations of some dopaminergic drugs with and without ethanol. RESULTS: SKF82958 alone significantly increased PKA-mediated phosphorylation of both DARPP-32 and NR1. Bromocriptine alone had no effect on pDARPP-32 or on pNR1. However, when D1 receptors were blocked, bromocriptine reduced the PKA-mediated phosphorylation of both DARPP-32 and NR1. Coincident treatment with bromocriptine and ethanol reversed both of these effects with D1 receptors blocked. The combination of eticlopride (D2 blocker) and SF82958 (D1 agonist) did not significantly alter either pDARPP-32 or pNR1. CONCLUSIONS: These data demonstrate antagonistic effects of acute ethanol exposure on D1 signaling in vivo and the potential of acute in vivo challenge protocols to help fill gaps in the understanding of ethanol's effects on protein phosphorylation.