Ghrelin receptor (GHS-R1A) antagonism suppresses both alcohol consumption and the alcohol deprivation effect in rats following long-term voluntary alcohol consumption

Abstract

Alcohol dependence is a heterogeneous disorder where several signalling systems play important roles. Recent studies implicate that the gut-brain hormone ghrelin, an orexigenic peptide, is a potential mediator of alcohol related behaviours. Ghrelin increases whereas a ghrelin receptor (GHS-R1A) antagonist decreases alcohol consumption as well as operant self-administration of alcohol in rodents that have consumed alcohol for twelve weeks. In the present study we aimed at investigating the effect of acute and repeated treatment with the GHS-R1A antagonist JMV2959 on alcohol intake in a group of rats following voluntarily alcohol consumption for two, five and eight months. After approximately ten months of voluntary alcohol consumption the expression of the GHS-R1A gene (Ghsr) as well as the degree of methylation of a CpG island found in Ghsr was examined in reward related brain areas. In a separate group of rats, we examined the effect of the JMV2959 on alcohol relapse using the alcohol deprivation paradigm. Acute JMV2959 treatment was found to decrease alcohol intake and the effect was more pronounced after five, compared to two months of alcohol exposure. In addition, repeated JMV2959 treatment decreased alcohol intake without inducing tolerance or rebound increase in alcohol intake after the treatment. The GHS-R1A antagonist prevented the alcohol deprivation effect in rats. There was a significant down-regulation of the Ghsr expression in the ventral tegmental area (VTA) in high- compared to low-alcohol consuming rats after approximately ten months of voluntary alcohol consumption. Further analysis revealed a negative correlation between Ghsr expression in the VTA and alcohol intake. No differences in methylation degree were found between high- compared to low-alcohol consuming rats. These findings support previous studies showing that the ghrelin signalling system may constitute a potential target for development of novel treatment strategies for alcohol dependence.

Figure 1. Acute JMV2959 treatment decreases alcohol intake in rats.

(A) The acute effects of the GHS-R1A antagonist JMV2959 (1 or 3 mg/kg) administration on alcohol intake was evaluated at 1 hour time interval after both two and five months of voluntary alcohol consumption. (B) The acute effects of the GHS-R1A antagonist JMV2959 (1 or 3 mg/kg) administration on alcohol intake was evaluated at 4 hour time interval after both two and five months of voluntary alcohol consumption. JMV2959 significantly decreases voluntary alcohol consumption compared to vehicle and the effect was more pronounced after five compared to two months of alcohol consumption. All values represent mean alcohol intake (g/kg) ± SEM (n = 19, *P<0.05, **P<0.01, ***P<0.001 compared to corresponding vehicle or as indicated).

Figure 2. Acute JMV2959 treatment increases water intake in rats.

(A) Acute administration of the GHS-R1A antagonist JMV2959 (3 mg/kg) significantly increased water consumption compared to vehicle at 1 hour time interval in rats voluntarily consuming alcohol for two and five months before the treatment. (B) Acute administration of the GHS-R1A antagonist JMV2959 (3 mg/kg) significantly increased water consumption compared to vehicle at 4 hour time interval in rats voluntarily consuming alcohol for two and five months before the treatment. All values represent mean water intake (ml) ± SEM (n = 19, ***P<0.001 compared to corresponding vehicle).

Figure 3. Acute JMV2959 treatment decreases alcohol preference in rats.

(A) Acute administration of the GHS-R1A antagonist JMV2959 (1 or 3 mg/kg) decreases the alcohol preference compared to vehicle at 1 hour time interval in rats voluntarily consuming alcohol for two and five months before the treatment. Acute administration of the GHS-R1A antagonist JMV2959 (1 or 3 mg/kg) decreases the alcohol preference compared to vehicle at 4 hour time interval in rats voluntarily consuming alcohol for two and five months before the treatment. All values represent mean alcohol preference (%) ± SEM (n = 19, *P<0.05, **P<0.01, ***P<0.001 compared to corresponding vehicle or as indicated).

Figure 4. Repeated JMV2959 treatment decreases alcohol intake in rats.

(A) Repeated administration of the GHS-R1A antagonist JMV2959 (3 mg/kg) significantly decreased voluntary alcohol consumption compared to vehicle at 1 hour time interval in rats voluntarily consuming alcohol for eight months before the treatment. (B) Repeated administration of the GHS-R1A antagonist JMV2959 (3 mg/kg) significantly decreased voluntary alcohol consumption compared to vehicle at 24 hour time interval in rats voluntarily consuming alcohol for eight months before the treatment. No tolerance to treatment was observed and no rebound drinking following discontinuation of treatment was observed. All values represent mean alcohol intake (g/kg) ± SEM (n = 19, *P<0.05, **P<0.01, ***P<0.001 compared to vehicle on the corresponding treatment day).

Figure 5. Repeated JMV2959 treatment increases water intake in rats.

(A) Repeated administration of the GHS-R1A antagonist JMV2959 (3 mg/kg) significantly increased water intake compared to vehicle at the 1 hour time interval in rats voluntarily consuming alcohol for eight months before the treatment. (B) Repeated administration of the GHS-R1A antagonist JMV2959 (3 mg/kg) significantly decreased alcohol preference compared to vehicle at the 1 hour time interval in rats voluntarily consuming alcohol for eight months before the treatment. All values represent mean ± SEM (n = 19, *P<0.05, **P<0.01, ***P<0.001 compared to vehicle on the corresponding treatment day).

Figure 6. Acute JMV2959 treatment prevents the alcohol deprivation effect in rats.

Following seven weeks of alcohol consumption (baseline, white bars) the rats were exposed to ten days of alcohol abstinence. Before alcohol was reintroduced (black bars) the rats were treated with wither JMV2959 or vehicle. An alcohol deprivation effect was observed in vehicle treated rats but not in rats treated with the GHS-R1A antagonist, JMV2959. Data are presented as mean alcohol intake (g/kg/1 hr) ± SEM. (n = 15 in each group, *P<0.05 compared to corresponding baseline).

Figure 7. Scatter plot of Ghsr expression in the VTA.

The present figure shows a significant negative correlation between the expression of Ghsr in the VTA (data presented as arbitrary units 2^−ΔΔC _T where the mean ΔC_T of the group of low alcohol consuming rats was set as the calibrator) and mean alcohol consumption (g/kg/24 hrs) in rats that have voluntarily been drinking alcohol for ten months.