An amylin analogue attenuates alcohol-related behaviours in various animal models of alcohol use disorder

Abstract

Recent findings have identified salmon calcitonin (sCT), an amylin receptor agonist and analogue of endogenous amylin, as a potential regulator of alcohol-induced activation of the mesolimbic dopamine system and alcohol consumption. Providing that the role of amylin signalling in alcohol-related behaviours remains unknown, the present experiments investigate the effect of sCT on these behaviours and the mechanisms involved. We showed that repeated sCT administration decreased alcohol and food intake in outbred rats. Moreover, single administration of the potent amylin receptor antagonist, AC187, increased short-term alcohol intake in outbred alcohol-consuming rats, but did not affect food intake. Acute administration of sCT prevented relapse-like drinking in the "alcohol deprivation effect" model in outbred alcohol-experienced rats. Additionally, acute sCT administration reduced operant oral alcohol self-administration (under the fixed ratio 4 schedule of reinforcement) in selectively bred Sardinian alcohol-preferring rats, while it did not alter operant self-administration (under the progressive ratio schedule of reinforcement) of a highly palatable chocolate-flavoured beverage in outbred rats. Lastly, we identified differential amylin receptor expression in high compared to low alcohol-consuming rats, as reflected by decreased calcitonin receptor and increased receptor activity modifying protein 1 expression in the nucleus accumbens (NAc) of high consumers. Collectively, our data suggest that amylin signalling, especially in the NAc, may contribute to reduction of various alcohol-related behaviours.

Fig. 1

Repeated sCT administration attenuates voluntary alcohol intake, alcohol preference, and food intake and decreases body weight in outbred rats. a Repeated sCT treatment (5 μg/kg, IP) reduced alcohol intake in rats (N = 25) in the intermittent access 20% alcohol two-bottle-choice drinking paradigm at the time point of 1 h on the first and second treatment days compared to vehicle (N = 25). b Repeated sCT administration reduced alcohol intake at the time point of 24 h on the first treatment day. sCT reduced alcohol preference at the time point of c 1 h on treatment days 1 and 2 and at the time point of d 24 h on treatment day 1. sCT increased water intake at the time point of e 1 h on treatment day 1, but reduced water intake on treatment day 3 when compared to vehicle. At f 24 h, the sCT group showed increased water intake only on the first treatment day. Repeated sCT injections decreased total fluid intake at the time point of g 1 h on the first treatment day, but decreased water intake on the and last treatment day when compared to vehicle. sCT administration had no effect on water at the time point of h 24 h on any treatment day. sCT increased i 24-hour water intake during the untreated days when compared to vehicle. Repeated sCT injections (5 μg/kg, IP) decreased food intake in rats at the time point of j 1 h and k 24 h on all treatment days when compared to vehicle. l Repeated sCT administration increased the 24-hour values of percentage of body weight change. (Data are presented as mean ± SEM; *P < 0.05, **P < 0.001, ***P < 0.001, ****P < 0.0001; white circle indicates vehicle, black circle indicates sCT)

Fig. 2

AC187 increases short-term alcohol intake, but does not affect food intake in outbred rats. Single administration of AC187 (250 μg/kg) did not affect a alcohol intake in outbred rats (N = 9) compared to vehicle (Veh) (N = 9) at the time point of 1 h, but significantly increased alcohol consumption at the time point of 4 h. There was no difference in long-term alcohol intake 24 h after AC187 administration. AC187 did not have an effect on b alcohol preference scores at any time point of 1 h, 4 h, or 24 h. No effect was noted on c water intake at any measured time of 1 h, 4 h, or 24 h, similarly to d total fluid intake scores for 1, 4, and 24 h, respectively. AC187 did not affect e food intake at the time point of 1 h, 4 h, or 24 h. (Data are presented as mean ± SEM; *P < 0.05, n.s.: P > 0.05)

Fig. 3

sCT prevents ADE in outbred rats. A single injection of sCT (5 μg/kg) prevents the alcohol deprivation effect (ADE) in rats (N = 18) abstained from alcohol for 10 days. There was an increase in alcohol intake post abstinence in the vehicle (Veh) group, but not in the sCT group when compared to baseline values. (Data are presented as mean ± SEM; ***P < 0.001 for baseline vs sCT within the Veh group, ^#P < 0.05 for ADE vs ADE between the Veh-sCT groups, n.s.: P > 0.05)

Fig. 4

sCT administration decreases lever-responding for alcohol and amount of self-administered alcohol in sP rats. a Acute sCT administration in the dose of 1 μg/kg (sCT1) (N = 12) and 5 μg/kg (sCT5) (N = 12) decreased the number of lever-responses for alcohol in a dose-response manner when compared to vehicle (Veh) (N = 12) in Sardinian alcohol-preferring (sP) rats exposed to the FR4 schedule of reinforcement. b Acute administration of sCT in the same doses reduced the amount of self-administered alcohol in a dose-response manner. c In the same experiment, body weight change (in grams) was increased by acute administration of sCT in two doses in a dose response-like effect. (Data are presented as mean ± SEM;⁺P < 0.05,⁺⁺P < 0.01 for Veh vs sCT1, ****P < 0.0001 for Veh vs sCT5 and ^###P < 0.001, ^####P < 0.0001 for sCT1 vs sCT5)

Fig. 5

The expression of the CALCR gene in NAc is lower in high alcohol-consuming rats, whereas the expression of RAMP1 is higher in the same brain area. a The expression of CALCR in NAc is decreased in high (N = 19) compared to low alcohol-consuming (N = 25) rats. Conversely, b the expression of RAMP1 in the same area is higher in high compared to low alcohol-consuming rats. (Data are presented as mean ± SEM of the ΔC_T values; ⁺P < 0.05)