Synaptic remodeling of GluA1 and GluA2 expression in the nucleus accumbens promotes susceptibility to cognitive deficits concomitant with downstream GSK3 β mediated neurotoxicity in female mice during abstinence from voluntary oral methamphetamine

Abstract

Stimulant-use disorders can present with long-term cognitive and mental health deficits. Little is known about the underlying molecular mechanisms perpetuating sex differences in cognitive and behavioral deficits in preclinical models of addiction to stimulants such as methamphetamine (MA). The current study investigated the neurochemical shifts underlying sex disparities in MA-induced working memory deficits and an addictive phenotype following abstinence from chronic MA abuse. We used our previously reported mouse model of voluntary oral methamphetamine administration (VOMA) consisting of an acquisition phase (days 1-14) characterized by escalating doses of MA and a binge phase (days 14-28) characterized by static doses. Female VOMA mice exhibited sustained MA consumption during the binge phase, demonstrating sex-specific vulnerabilities to the maintenance of MA addiction. The 8-arm radial maze was used to test spatial working memory performance following abstinence from VOMA. Results indicate working memory deficits correlated to higher MA consumption in females only. Hippocampal and accumbal tissue were collected and analyzed by immunoblotting. Female VOMA mice had decreased GluA1, but not GluA2, in the hippocampus, which may perpetuate synaptic destabilization and working memory deficits. Female-specific increases in GluA1 and p-GSK3β expression in accumbal tissue suggest vulnerability toward abstinence-induced drug craving and heightened downstream neurotoxicity. Our study reveals female-specific neurochemical shifts in hippocampal and accumbal AMPA receptor signaling following abstinence from chronic MA consumption that may perpetuate female susceptibility to MA-induced cognitive deficits. These data demonstrate a novel molecular pathway that would exacerbate memory deficits and perpetuate an addictive phenotype in female populations following MA abuse.

Keywords: AMPA receptors; Hippocampus; Methamphetamine addiction; Nucleus accumbens; Sex differences.

Fig. 1.

The escalation voluntary oral methamphetamine administration (VOMA) model produces increased consumption in female VOMA mice accompanied by increased total WM errors on the RAM. The paradigm lasted 8 weeks, with acclimation and shaping during weeks 1–2; VOMA during weeks 3–6; behavioral assessments during weeks 7–8 (a). Body weights were tracked across all eight weeks, and there were no significant differences between drug and control mice (b). The acquisition phase of VOMA was characterized from days 1–14 with the average MA consumption over days 11–14 designated as baseline consumption per animal. There was a no significant effect of sex (male vs female; F_1,26=0.20, p = 0.66), no significant sex by day interaction (F_13,338=0.32, p = 0.99) but a significant effect of day (days 1–14; F_13,338=125.20, p<0.0001) during the acquisition phase of VOMA with no significant post-hoc differences (c). The binge phase of VOMA was characterized from days 15–28. Difference scores of MA intake per day from baseline consumption per animal (Δ consumption) were calculated for each day from days 15–28. There was a significant effect of sex (male vs female; F_1,26=14.70, p<0.001), a significant effect of day (days 15–28; F_13,338=4.07, p<0.001), and a significant interaction effect of sex by day (F_13,338=3.78, p<0.001) during the binge phase of VOMA. Females consumed significantly more MA compared to baseline consumption compared to males on days 18, 21–24, and 26–28, p<0.05 (d). Working-memory performance on the RAM was assessed across baits 1–4 and baits 5–8. WM performance is shown as a percent of sex-specific control groups. There was no significant effect of bait (F_1,26=0.66, p = 0.42), no effect of sex (F_1,26=1.79, p = 0.19), and no bait by sex interaction (F_1,26=0.48, p = 0.49) (e). Analyses of total number of working memory errors on the RAM correlated to average MA consumption during the binge phase for male VOMA mice revealed no significant correlation between total WM errors and binge consumption rate (r₁₂=−0.21, p = 0.47) (f). Analyses of total number of working memory errors on the RAM correlated to average MA consumption during the binge phase for female VOMA mice revealed a significant correlation between total WM errors and binge consumption rate (r₁₂=0.54, p<0.05, *) (g).

Fig. 2.

AMPAR signaling in the hippocampus demonstrates baseline sex differences. Hippocampal GluA1 (a) and GluA2 expression (b) is significantly lower in female mice compared to male mice. There was no difference in the hippocampal ratio of GluA1/GluA2 (c). There was no significant correlation between total working memory errors and the ratio of hippocampal GluA1/GluA2 expression in males (r₉=0.48, p = 0.14), (d) nor in females (r₁₀=0.09, p = 0.79) (e). VOMA: voluntary oral methamphetamine administration. * S=overall effect of sex, * D=overall effect of drug, *S x * D= interaction effect of sex by drug. ** p<0.01, *** p < 0.001.

Fig. 3.

Abstinence from VOMA produces female specific decreases in GluA2 signaling in the nucleus accumbens compared to males and a correlation to working memory errors. Female mice demonstrate decreased GluA1 expression (a) compared to males with abstinence from VOMA producing overall increases in GluA1 expression in the nucleus accumbens. Female VOMA mice display decreased GluA2 (b) expression in the nucleus accumbens compared to male VOMA mice with no differences in the ratio of GluA1/GluA2 (c). There was no significant correlation between total working memory errors and the ratio of accumbal GluA1/GluA2 expression in males (r₁₁=0.29, p = 0.34) (d), however, there was a significant correlation found in females (r₁₁=−0.66, * p<0.05) (e). Female VOMA mice displayed more working memory errors correlated with a lower accumbal GluA1/GluA2 ratio. VOMA: voluntary oral methamphetamine administration. * S=overall effect of sex, * D=overall effect of drug, *S x * D= interaction effect of sex by drug. ** p<0.01, *** p < 0.001.

Fig. 4.

p-GSK3β signaling is dysregulated following abstinence from chronic VOMA. Female mice are more vulnerable to Hippocampal GSK3β expression (a) does not significantly change whereas mice display decreased p-GSK3β expression (b) with female VOMA mice displaying less p-GSK3β compared to male mice following abstinence and no differences in the ratio of p-GSK3β/GSK3β expression. There was no significant correlation between total working memory errors and the ratio of hippocampal p-GSK3β/GSK3β expression in males (r₁₂=0.04, p = 0.89), nor in females (r₁₁=−0.11, p = 0.71). VOMA: voluntary oral methamphetamine administration. * S=overall effect of sex, * D= overall effect of drug, *S x * D= interaction effect of sex by drug. ** p<0.01, *** p < 0.001.

Fig. 5.

Abstinence from VOMA produces female specific decreases in p-GSK3β signaling in the nucleus accumbens. Female mice demonstrate decreased GSK3β (a) expression in the nucleus accumbens compared to males. Female VOMA mice display decreased p-GSK3β (b) expression in the nucleus accumbens compared to male mice, with both sexes decreasing the ratio of p-GSK3β/GSK3β following abstinence (c). There was no significant correlation between total working memory errors and the ratio of accumbal p-GSK3β/GSK3β expression in males (r₁₀=−0.20, p = 0.53), nor in females (r₁₂=−0.16, p = 0.59). VOMA: voluntary oral methamphetamine administration. * S=overall effect of sex, * D= overall effect of drug, *S x * D= interaction effect of sex by drug. ** p<0.01, *** p < 0.001.