Reduced alcohol intake and reward associated with impaired endocannabinoid signaling in mice with a deletion of the glutamate transporter GLAST

Abstract

A hyperglutamatergic state has been hypothesized to drive escalation of alcohol intake. This hypothesis predicts that an impairment of glutamate clearance through inactivation of the astrocytic glutamate transporter, GLAST (EAAT1), will result in escalation of alcohol consumption. Here, we used mice with a deletion of GLAST to test this prediction. WT and GLAST KO mice were tested for alcohol consumption using two-bottle free-choice drinking. Alcohol reward was evaluated using conditioned place preference (CPP). Sensitivity to depressant alcohol effects was tested using the accelerating rotarod, alcohol-induced hypothermia, and loss of righting reflex. Extracellular glutamate was measured using microdialysis, and striatal slice electrophysiology was carried out to examine plasticity of the cortico-striatal pathway as a model system in which adaptations to the constitutive GLAST deletion can be studied. Contrary to our hypothesis, GLAST KO mice showed markedly decreased alcohol consumption, and lacked CPP for alcohol, despite a higher locomotor response to this drug. Alcohol-induced ataxia, hypothermia, and sedation were unaffected. In striatal slices from GLAST KO mice, long-term depression (LTD) induced by high frequency stimulation, or by post-synaptic depolarization combined with the l-type calcium channel activator FPL 64176 was absent. In contrast, normal synaptic depression was observed after application of the cannabinoid 1 (CB1) receptor agonist WIN55,212-2. Constitutive deletion of GLAST unexpectedly results in markedly reduced alcohol consumption and preference, associated with markedly reduced alcohol reward. Endocannabinoid signaling appears to be down-regulated upstream of the CB1 receptor as a result of the GLAST deletion, and is a candidate mechanism behind the reduction of alcohol reward observed.

Figure 1

Decreased 2-bottle free choice consumption of alcohol in GLAST KO mice. GLAST KO mice showed decreased alcohol consumption at the highest alcohol concentration tested, 16% (A), and showed a decreased preference for alcohol vs water both at 12 and 16% (B). Data are mean±SEM. n=28-46/genotype for consumption. For detailed statistics, see Results.

Figure 2

Loss of alcohol reward in GLAST KO mice. There was no significant difference between genotypes in preference for side going to become alcohol-paired in the initial habituation session (A). WT and GLAST HET mice showed robust place preference for the alcohol-paired side, while GLAST KO mice did not show any side preference, and consequently differed from WT mice. (B). Data are mean±SEM, n=17-18 for condition place preference, **P<0.01 compared to WT.

Figure 3

Extracellular glutamate in the Nc. Accumbens was not affected by the GLAST deletion, neither at baseline, nor following systemic administration of alcohol (2g/kg) n=7-8 / genotype / time point. Data are mean±SEM μM.

Figure 4

LTD-induction but not CB1R activation is impaired in slices from GLAST KO mice. HFS was insufficient to induce LTD in slices from GLAST KO mice (A), even during blocked NMDA receptors (B). Application of the L-type calcium channel activator FPL 64176 (500 nM) paired with slight depolarization of the postsynaptic cell (-50 mV) induced LTD in control slices, but not in slices from GLAST KO mice (C). WIN55,212-2 induced a robust depression, showing that activation of CB1R and downstream mechanism from this receptor is functional in slices from GLAST KO mice (D). EPSC values from MSNs located in the dorsolateral striatum are mean±SEM. Example traces show evoked EPSCs at baseline (black) and after HFS stimulation (gray) in KO (A), and WT (B), and in KO following 20 min of FPL or WIN 55,212-2 treatment (C, D). Arrows mark time point for HFS. n=5-11. Field potential recordings reviled that LTP could be induced by HFS in slices from GLAST KO mice, while both LTP and LTD was observed in WT (E, F). n=12-14. Scale bar is 0.025 ms and 100 pA.