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ERK-Directed Phosphorylation of mGlu5 Gates Methamphetamine Reward and Reinforcement in Mouse - PubMed

  • ️Fri Jan 01 2021

ERK-Directed Phosphorylation of mGlu5 Gates Methamphetamine Reward and Reinforcement in Mouse

Elissa K Fultz et al. Int J Mol Sci. 2021.

Abstract

Methamphetamine (MA) is a highly addictive psychomotor stimulant drug. In recent years, MA use has increased exponentially on a global scale, with the number of MA-involved deaths reaching epidemic proportions. There is no approved pharmacotherapy for treating MA use disorder, and we know relatively little regarding the neurobiological determinants of vulnerability to this disease. Extracellular signal-regulated kinase (ERK) is an important signaling molecule implicated in the long-lasting neuroadaptations purported to underlie the development of substance use disorders, but the role for this kinase in the propensity to develop addiction, particularly MA use disorder, is uncharacterized. In a previous MA-induced place-conditioning study of C57BL/6J mice, we characterized mice as MA-preferring, -neutral, or -avoiding and collected tissue from the medial prefrontal cortex (mPFC). Using immunoblotting, we determined that elevated phosphorylated ERK expression within the medial prefrontal cortex (mPFC) is a biochemical correlate of the affective valence of MA in a population of C57BL/6J mice. We confirmed the functional relevance for mPFC ERK activation for MA-induced place-preference via site-directed infusion of the MEK inhibitor U0126. By contrast, ERK inhibition did not have any effect upon MA-induced locomotion or its sensitization upon repeated MA treatment. Through studies of transgenic mice with alanine point mutations on T1123/S1126 of mGlu5 that disrupt ERK-dependent phosphorylation of the receptor, we discovered that ERK-dependent mGlu5 phosphorylation normally suppresses MA-induced conditioned place-preference (MA-CPP), but is necessary for this drug's reinforcing properties. If relevant to humans, the present results implicate individual differences in the capacity of MA-associated cues/contexts to hyper-activate ERK signaling within mPFC in MA Use Disorder vulnerability and pose mGlu5 as one ERK-directed target contributing to the propensity to seek out and take MA.

Keywords: addiction; metabotropic glutamate receptor 5; methamphetamine; place-conditioning; reinforcement; self-administration; vulnerability.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1

Elevated phospho-ERK expression within mPFC is a biochemical correlate of the affective valence of methamphetamine. (Inset): Outline of the procedural timeline for the immunoblotting study of ERK correlates of the place-conditioning elicited by the repeated pairing of 2 mg/kg methamphetamine (MA) in male C57BL/6J mice. Mice were tested for the expression of their conditioned response in a MA-free state (15 min session) and brains removed (~2–5 min later) for immunoblotting. (A) Summary of the differences in the time spent in the methamphetamine (MA)—versus saline (SAL)—paired compartment of the place-conditioning apparatus between mice spontaneously exhibiting a MA-conditioned place-preference (CPP), a MA-conditioned place-aversion (CPA), and no conditioned response (Neutral). Sample sizes are indicated in parentheses. Note data in Panel A is from Ref. [26]. (B) Cartoon depicting the tissue dissection of the mPFC. (C) Summary of the mPFC levels of total ERK, p(Tyr204)-ERK, and their ratio exhibited by CPP, Neutral, and CPA mice, as well as mice conditioned with SAL. The raw immunoblots are provided in the unpublished materials. (D) Results of the correlational analysis conducted between the CPP Score and the ratio of phosphorylated to total ERK within the mPFC. The data in Panels (A,C) represent the mean ± SEMs of the number of mice indicated in parentheses. * p < 0.05 vs. all other groups; ** p < 0.05 vs. CPA and Neutral.

Figure 2
Figure 2

ERK inhibition within the mPFC blocks an established MA-conditioned place-preference, without affecting locomotor activity. (Inset) Procedural time-line for the neuropharmacological study of ERK inhibition upon the expression of a MA-conditioned place-preference. (A) Intra-mPFC infusion of the MEK inhibitor U0126 lowered the expression of a place-preference induced by the repeated pairing of 2 mg/kg methamphetamine. In contrast, no effect of repeated vehicle (VEH) infusion was detected. (B) U0126 infusion did not alter the locomotor activity of the mice during place-preference testing. (C) Cartoons depicting the locations of the microinjector tips within the mPFC. The data represent the means ± SEMs of the number of mice indicated in parentheses. * p < 0.05 CPP Score vs. 0 (i.e., presence of a conditioned response; 1-sample t-tests).

Figure 3
Figure 3

Disruption of ERK-dependent phosphorylation of mGlu5 augments methamphetamine-induced place-preference without affecting methamphetamine-induced psychomotor activity. (Inset) Procedural timeline of the study characterizing the effects of the Grm5AA/AA mutation upon MA-induced changes in behavior under place-conditioning procedures. Comparison of the dose–response functions for (A) methamphetamine (MA)-induced place-preference, (B) acute drug-induced locomotor activity, and (C) locomotor sensitization (defined as the difference in distance traveled from injection 1 to 4 of MA-conditioning) between WT Grm5TS/TS and mutant Grm5AA/AA mice. The data represent the means ± SEMs of the number of mice indicated in parentheses. * p < 0.05 (main effect of Genotype).

Figure 4
Figure 4

Disruption of ERK-dependent phosphorylation of mGlu5 blunts methamphetamine reinforcement and intake under operant-conditioning procedures. (Inset) Procedural timeline of the operant-conditioning studies conducted in Grm5AA/AA and Grm5TS/TS mice. When the first five days of self-administration training were considered, Grm5AA/AA mutants exhibited less (A) methamphetamine (MA)-directed responding for reinforcement by 20 mg/L MA and (B) less MA intake than WT Grm5TS/TS mice. When the response requirement for reinforcement by 20 mg/L MA progressively increased, mutant mice continued to exhibit (C) less MA-directed responding and (D) less intake. When the dose of the MA reinforcer varied under an FR1 schedule of reinforcement, the MA-directed responding (E) and MA intake (F) continued to be lower in Grm5AA/AA mice versus Grm5TS/TS controls. No genotypic differences were detected for responding in the inactive hole during (A’) early training, (C’) demand testing, or (E’) dose–response testing. The data represent the means ± SEMs of the number of mice indicated in parentheses. Note: not all mice tested for reinforcement by 5–40 mg/L MA advanced to testing under the 80 mg/L dose. * p < 0.05 (main Genotype effect); * p < 0.05 vs. Grm5TS/TS (t-test).

Figure 5
Figure 5

Disruption of ERK-dependent phosphorylation of mGlu5 only modestly blunts MA intake in the home cage. When presented simultaneously with four sipper tubes containing, 5, 10, 20, and 40 mg/L MA, (A) Grm5AA/AA mice tended to consume less 40 mg/L MA and (B) exhibited lower overall MA intake than their WT Grm5TS/TS controls. Neither of the genotypic differences were statistically reliable. The data represent the means ± SEMs of the number of mice indicated in parentheses.

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