The GABAB Positive Allosteric Modulator ADX71441 Attenuates Alcohol Self-Administration and Relapse to Alcohol Seeking in Rats

Abstract

GABAergic signaling is involved in modulating the reinforcing properties of alcohol, and GABA_B receptors have been proposed as a potential target for clinical treatment of alcoholism. The orthosteric GABA_B receptor agonist baclofen has been shown to suppress operant self-administration of alcohol in animals and alcohol use in alcohol-dependent patients, but its utility is limited by a narrow therapeutic index. We tested the effects of ADX71441, a novel GABA_B receptor positive allosteric modulator, on alcohol-related behaviors in rats. We first assessed the effects of ADX71441 (1, 3, 10 and 30 mg/kg, I.P.) on both non-dependent and dependent male Wistar rats trained to self-administer 20% alcohol. We then determined the effects of ADX71441 on stress-induced as well as cue-induced relapse-like behavior. Finally, we sought to identify the brain regions through which ADX71441 may act to prevent relapse-like behavior by mapping the neuronal activation induced by stress-induced reinstatement of alcohol-seeking using c-Fos immunohistochemistry. ADX71441 dose-dependently decreased alcohol self-administration of both dependent and non-dependent animals, but its potency was higher in alcohol-dependent rats. Furthermore, both cue- and stress-induced alcohol seeking were blocked by the GABA_B receptor positive allosteric modulator. Finally, pretreatment with 3 mg/kg of ADX71441 before stress-induced reinstatement significantly decreased c-Fos expression in a network of brain regions implicated in stress-induced relapse, comprising the nucleus accumbens shell, the dorsal raphe nucleus and the medial prefrontal cortex. Our findings support a causal role of GABA_B receptors in alcohol reinforcement and relapse to alcohol seeking. These effects are observed in the absence of significant sedative side effects. Jointly, these observations indicate that GABA_B receptor positive allosteric modulators merit being tested clinically for the treatment of alcoholism. Our data also point to a potential biomarker of target engagement for early clinical studies.

Figure 1

ADX71441 strongly decreases 20% alcohol self-administration. (a) Mean reinforcers (±SEM) earned during a 30-min FR2 self-administration session of 20% EtOH following either vehicle or ADX71441 treatment (1, 3, 10 or 30 mg/kg; n=32; *p<0.001). (b) Mean active lever presses (±SEM) completed during a 30-min FR2 self-administration session of 20% EtOH following either vehicle or ADX71441 treatment (1, 3, 10 or 30 mg/kg; n=32; *p<0.001). (c) Mean distance traveled (±SEM) following either vehicle or ADX71441 treatment (1, 3, 10 or 30 mg/kg; n=18; *p<0.001). (d) Mean breakpoint (±SEM) reached during a progressive ratio session of 20% EtOH following either vehicle or ADX71441 treatment (3 or 10 mg/kg; n=10, 11 by groups; *p<0.001).

Figure 2

Rats with a history of dependence show an enhanced sensitivity to suppression of alcohol self-administration by ADX71441. (a) Schematic representation of the timeline of the experiment. (b) Mean reinforcers (±SEM) earned during eleven consecutive 30-min FR3 self-administration sessions of 20% EtOH following vapor exposure (n=16 and 14 by groups; *p<0.05). (c) Mean active lever presses (±SEM) completed during 11 consecutive 30-min FR3 self-administration sessions of 20% EtOH following vapor exposure (n=16 and 14 by groups; *p<0.05). (d) Mean reinforcers (±SEM) earned during a 30-min FR3 self-administration session of 20% EtOH following either vehicle or ADX71441 treatment (1 or 3 mg/kg; n=16 and 14 by groups; *p<0.05). (e) Mean active lever presses (±SEM) completed during a 30-min FR3 self-administration session of 20% EtOH following either vehicle or ADX71441 treatment (1 or 3 mg/kg; n=16 and 14 by groups; ^#p<0.05 compared to baseline, *p<0.05 compared to respective vehicle).

Figure 3

ADX71441 blocks both cue-induced and stress-induced alcohol-seeking (a) mean number of non-reinforced lever presses (±SEM) during the 30-min test for cue-induced reinstatement following either saline or ADX71441 treatment (3 or 10 mg/kg; n=10 by groups; *p<0.001 compared to vehicle, #p<0.001 compared to extinction). (b) Mean number of non-reinforced lever presses (±SEM) during the 30-min test for stress-induced reinstatement following either saline or ADX71441 treatment (3 or 10 mg/kg; n=10, 11 by groups; *p<0.01 compared to vehicle, ^#p<0.01 compared to extinction).

Figure 4

ADX71441 attenuates stress-induced neuronal activity in the nucleus accumbens shell, central amygdala and dorsal raphe nucleus. Mean c-Fos-positive cells (±SEM) following either vehicle or ADX71441 treatment (3 mg/kg) in the (a) nucleus accumbens shell (NAcSh), (b) central amygdala (CeA), (c) dorsal raphe nucleus (DRN), (d) medial prefrontal cortex (mPFC), (e) bed nucleus of stria terminalis (BNST) and (f) basolateral amygdala (BLA; n=9–11 by groups; *p<0.05). Representative images are shown at × 40 magnification.

Figure 5

Stress-induced neuronal activity in an interconnected network of brain structures is strongly correlated with stress-induced relapse-like behavior, and is suppressed by ADX71441. Principal component extraction followed by varimax normalized rotation of c-Fos data identified two independent networks that showed high measures of within-network connectivity. Network 1 consisted of the NAcSh, DRN and mPFC, and its activity was highly correlated with relapse-like behavior (a). In contrast, activity of Network 2, which consisted of BLA, CeA and BNST did not show any correlation with behavior (b). Activity of Network 1, but not Network 2, was suppressed by ADX71441 (c; ***p<.0001; for detailed statistics, see Results section).