Methamphetamine Addiction Vulnerability: The Glutamate, the Bad, and the Ugly - PubMed (original) (raw)

. 2017 Jun 1;81(11):959-970.

doi: 10.1016/j.biopsych.2016.10.005. Epub 2016 Oct 13.

Kevin D Lominac 2, Rianne R Campbell 2, Matan Cohen 2, Elissa K Fultz 2, Chelsea N Brown 2, Bailey W Miller 2, Sema G Quadir 2, Douglas Martin 2, Andrew B Thompson 2, Georg von Jonquieres 3, Matthias Klugmann 3, Tamara J Phillips 4, Tod E Kippin 5

Affiliations

Methamphetamine Addiction Vulnerability: The Glutamate, the Bad, and the Ugly

Karen K Szumlinski et al. Biol Psychiatry. 2017.

Abstract

Background: The high prevalence and severity of methamphetamine (MA) abuse demands greater neurobiological understanding of its etiology.

Methods: We conducted immunoblotting and in vivo microdialysis procedures in MA high/low drinking mice, as well as in isogenic C57BL/6J mice that varied in their MA preference/taking, to examine the glutamate underpinnings of MA abuse vulnerability. Neuropharmacological and Homer2 knockdown approaches were also used in C57BL/6J mice to confirm the role for nucleus accumbens (NAC) glutamate/Homer2 expression in MA preference/aversion.

Results: We identified a hyperglutamatergic state within the NAC as a biochemical trait corresponding with both genetic and idiopathic vulnerability for high MA preference and taking. We also confirmed that subchronic subtoxic MA experience elicits a hyperglutamatergic state within the NAC during protracted withdrawal, characterized by elevated metabotropic glutamate 1/5 receptor function and Homer2 receptor-scaffolding protein expression. A high MA-preferring phenotype was recapitulated by elevating endogenous glutamate within the NAC shell of mice and we reversed MA preference/taking by lowering endogenous glutamate and/or Homer2 expression within this subregion.

Conclusions: Our data point to an idiopathic, genetic, or drug-induced hyperglutamatergic state within the NAC as a mediator of MA addiction vulnerability.

Keywords: Conditioned place preference; Glutamate; Homer proteins; MAHDR; Metabotropic glutamate receptor; NMDA receptor; Nucleus accumbens.

Copyright © 2016 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.

PubMed Disclaimer

Figures

Figure 1

Figure 1. NAC GLU correlates of MA addiction vulnerability/resiliency in a genetic model

(A) No net-flux in vivo microdialysis procedures were employed to estimate basal GLU content within the shell-core interface of the NAC of MA-naïve mice selectively bred for Methamphetamine High Drinking (MAHDR) and Methamphetamine Low Drinking (MALDR) and (B) an examination of y=0 confirmed higher GLUEC in MAHDR vs. MALDR mice [t(13)=8.93, p<0.0001]. **_(C)_** Conventional microdialysis detected marked line differences also in the capacity of acute MA (2 mg/kg, IP) to elevate GLUEC [Genotype X Time interaction [F(11,143)=4.37, p<0.0001], with MAHDR mice exhibiting a large rise in GLUEC [one-way ANOVA, F(11,77)=4.74, p<0.0001], but no change in GLUEC in MALDR animals (one-way ANOVA, p=0.36). Immunoblotting conducted on NACc tissue of MALDR and MAHDR mice revealed line differences in the total protein expression of **_(D)_** EAAT3 [t(37)=2.32, p=0.03], **_(E)_** mGlu5 monomer [t(37)=2.78, p=0009] and **_(F)_** both Homer1b/c [t(37)=2.75, p=0.007] and Homer2a/b [t(37)=3.01, p=0.005]. **_(G)_** Although Homer1b/c levels within the NACs did not vary with line (t-test, p=0.27), MAHDR mice also exhibited greater mGlu5 [t(35)=2.28, p=0.03] and Homer2a/b [t(35)=2.08, p=0.04] expression within this subregion. **_(H)_** Line differences were not apparent regarding the expression of NMDA receptor subunits within either subregion (t-tests, all p’s>0.20). Samples sizes indicated in parentheses.

Figure 2

Figure 2. Initial subjective response to MA predicts MA addiction vulnerability in an inbred murine model

The predicative validity of responses in our MA place-conditioning procedure for addiction vulnerability/resiliency was determined in (A) a cohort of B6 mice expressing divergent subjective responses to the MA-paired compartment [F(2,54)=170.42, p<0.0001; *p<0.05 vs. Neutral, LSD post-hoc tests; one-sample t-tests, for CPA: t(8)=7.30, p<0.0001; for CPP: t(30)=20.46, p<0.0001; for Neutral, p=0.29; +p<0.05 vs. CPP Score = 0]. (B) Early in operant-conditioning (FI20 reinforcement schedule), all mice progressively increased their proportion of responses towards the aperture that delivered 20 mg/L MA; however CPP mice consistently directed a greater proportion of their total responses towards the active hole than the other phenotypes [Phenotype effect: F(2,53)=4.90, p=0.01; FRI20 Day effect: F(4,212)=13.87, p<0.0001; interaction: p=0.79; ** p<0.03 vs. CPA/Neutral, LSD post-hoc tests]. (C) CPP Score correlated positively with the average response allocation during the first 5 days of training (from panel b) [r=0.32, p=0.02; N=56]. (D) For those mice that successfully completed training, a dose-response analysis of MA intake revealed a shift upwards in CPP mice, relative to the other phenotypes [MA dose: F(3,144)=15.85, p<0.0001; Phenotype: F(2,48)=7.02, p=0.002; interaction, p=0.57; ** p<0.05 vs. CPA/Neutral, LSD post-hoc test]. (E) CPP Score predicted the intake of 10 mg/L MA (as well as 40 mg/L, not shown) under an FR5/FI20 reinforcement schedule. (F) A demand-intake analysis of behavior when 10 mg/L MA served as the reinforcer revealed greater MA intake under low demand in CPP mice, relative to the other phenotypes [Phenotype X Schedule: F(5,240)=5.82, p<0.0001; ** p<0.05 vs. CPA/Neutral, LSD post-hoc tests]. Sample sizes are indicated in parentheses and the results of the correlational analyses are presented in their corresponding panels.

Figure 3

Figure 3. NAC glutamate correlates of MA addiction vulnerability/resiliency in an inbred murine model

(A) Immunoblotting was employed on MA-induced CPP, CPA and Neutral-B6 mice to assay for indices of protein levels of glutamate transmission within NAC subregions [CPP Score: F(1,37)=121.00, p<0.0001; *p<0.05 vs. Neutral, LSD post-hoc tests; +p<0.05 vs. CPP Score = 0 sec, one-sample t-tests]. (B) Within the NACc, mGlu1 and mGlu5 levels were highest in CPP mice [for mGlu1: [F(3,49)=21.88, p<0.0001, ***p<0.05 vs. all other groups, LSD post-hoc tests; for mGlu5: [F(3,49)=2.70, p=0.06; for mGlu2/3, p=0.34] and the expression of both mGlu1 and mGlu5 was predicted by CPP Score (from panel a). (C) Homer2a/b expression was also elevated within the NACc of CPP mice [for Homer2a/b: F(3,49)=8.92, p<0.0001; ***p<0.05 vs. all other groups, LSD post-hoc tests; for Homer1b/c: p=0.93] with expression also predicted by CPP score. (D) Within the NACs, Homer2a/b levels were also the most highly expressed in CPP mice [F(3,41)=8.10, p<0.0001; *p<0.05 Neutral vs. SAL; ***p<0.05 CPP vs. other groups, LSD post-hoc tests]. Samples sizes are indicated in parentheses.

Figure 4

Figure 4. Endogenous glutamate within the NAC bi-directionally regulates MA-preference

(A) No net-flux in vivo microdialysis procedures revealed that repeated MA (10 daily injections of 2 mg/kg, IP) increased basal extracellular glutamate content (y=0), but only at the 21-day withdrawal time-point (21 WD) [Treatment X Withdrawal: F(1,30)=7.80, p=0.009]. No group difference in the Ed was apparent when the slopes of the linear regressions for each plot were compared (Treatment X Withdrawal ANOVA, all p’s>0.08). *p<0.05 vs. SAL; +p<0.05 vs. 1 WD (t-tests). **_(B)_** A significant Treatment X Withdrawal X Time interaction was observed for the change in extracellular glutamate elicited by a challenge injection of 1 mg/kg MA, administered at either 1 or 21 WD [F(11,319)=3.12, p=0.001]. This interaction reflected the fact that the MA challenge injection lowered glutamate below baseline in MA-treated animals at 1 WD, but these same mice exhibited a robust glutamate response to the same challenge at 21 WD [Withdrawal X Time interaction: F(11,176)=2.93, p=0.001]. In contrast, the MA challenge elicited a modest glutamate rise in SAL animals that was comparable between microdialysis sessions [Withdrawal X Time ANOVA, all p’s>0.05]. *p<0.05 vs. SAL (tests for simple effects). **_(C)_** Repeated MA treatment sensitized mice to the capacity of the EAAT inhibitor TBOA to increase NAC glutamate, but this effect was apparent only in early withdrawal [for 1 WD, Dose X Treatment: F(2,30)=6.62, p=0.004; for 21 WD, Dose X Treatment: p=0.39; *p<0.05 for MA vs. SAL, t-tests]. **_(D)_** Repeated MA treatment also sensitized mice to the capacity of the mGlu2/3 autoreceptor agonist APDC to reduce NAC glutamate, irrespective of withdrawal [Dose X Treatment: F(2,48)=4.64, p=0.004; 3-way interaction: p=0.94], with SAL-MA differences in the magnitude of the glutamate reduction observed at both 1 WD [50 μM: t(12)=2.59, p=0.02] and 21 WD [5μM: t(14)=2.95, p=0.01; 50μM: t(13)=3.38, p=0.005; *p<0.05 for MA vs. SAL]. **_(E)_** In a separate cohort of B6 mice, the repeated pairing of MA (4 × 2 mg/kg) elicited a place-preference when mice were tested in a drug-free state (no infusion; open bars; Drug X Side, p>0.75]. However, raising and lowering endogenous glutamate via infusion of 300 μM TBOA or 50 μM APDC into the NACs, respectively, potentiated and reversed the place-preference, relative to vehicle (VEH)-infused animals [Drug X Side: F(1,21)=17.84, p<0.0001; *p<0.05 Paired vs. Unpaired or conditioning; +p<0.05 vs. VEH, t-tests]. Samples sizes are indicated in the data bars.

Figure 5

Figure 5. Repeated MA experience elevates NACs Homer2 expression to promote MA-seeking and – taking

(A) Immunoblotting conducted on the NACs of B6 mice treated repeatedly with MA (10 IP injections of 2 mg/kg) revealed a moderate, albeit significant, reduction in Homer1b/c expression relative to SAL controls, irrespective of withdrawal [Treatment effect: F(1,36)=4.61, p=0.04; n’s=11–12]. Interestingly, these same animals exhibited a polar opposite increase in Homer2a/b expression [Treatment effect: F(1,31)=5.47, p=0.03; n’s=11–12]. (B) No changes in Homer protein expression were observed within the adjacent NACc (Treatment X Withdrawal ANOVAs, all p’s>0.20; n’s=10–12). For both panels a & b, representative immunoblots are provided, corresponding with their respective datasets above and *p<0.05 denotes a main Treatment effect (p<0.05). **_(C)** When tested in a MA-free state, an intra-NACs infusion of AAV-shRNA-Homer2b did not impact the magnitude of a MA-conditioned place-preference induced by 4 pairings of 2 mg/kg MA. However, only GFP animals exhibited a relative increase in place-preference magnitude when tested in the presence of a 2 mg/kg MA interoceptive cue, however the group difference in this regard was shy of statistical significance [AAV X Side X Test: F(1,23)=3.04, p=0.09]. (D) During operant conditioning for reinforcement by 10 mg/L MA, shRNA-infused mice emitted fewer nose-pokes than green fluorescent protein (GFP) controls with increasing response requirement [AAV X Schedule: F(2,44)=5.92, p=0.005; *p<0.05 vs. GFP, t-tests]. **_(E)_** In parallel, shRNA-infused mice consumed less MA with increasing response requirement during this training phase [AAV X Schedule: F(2,44)=7.84, p=0.001; *p<0.05 vs. GFP, t-tests]. **_(F)_** shRNA-Homer2b infusion also flattened the MA dose-nose-poke response function under an FR1/FI20 schedule of reinforcement [AAV X Dose: F(4,88)=4.75, p=0.002; *p<0.05 vs. GFP, t-tests]. **_(G)_** Although it appeared that shRNA-Homer2b infusion reduced the intake of 10 mg/L MA, there was no systematic effect of knock-down on the MA dose-intake function [AAV X Dose, p>0.3]. **(H)_** Visualization of the GFP reporter by fluorescent microscopy indicating neuronal transduction within the NACs at 10X magnification. For panels c–g, sample sizes are indicated in parentheses.

Comment in

References

    1. United Nations Office on Drugs and Crime. World Drug Report 2015. United Nations publication, Sales No. E.15.XI.6; 2015.
    1. Cruickshank CC, Dyer KR. A review of the clinical pharmacology of methamphetamine. Addiction. 2009;104:1085–1099. - PubMed
    1. Sheridan J, Butler R, Wheeler A. Initiation into methamphetamine use: qualitative findings from an exploration of first time use among a group of New Zealand users. J Psychoactive Drugs. 2009;41:11–17. - PubMed
    1. Chait LD. Factors influencing the reinforcing and subjective effects of d-amphetamine in humans. Behav Pharmacol. 1993;4:191–199. - PubMed
    1. de Wit H, Uhlenhuth EH, Johanson CE. Individual differences in the reinforcing and subjective effects of amphetamine and diazepam. Drug Alcohol Depend. 1986;16:341–60. - PubMed

Publication types

MeSH terms

Substances

Grants and funding

LinkOut - more resources