br Materials and methods br Results br Discussion

Materials and methods
Results
Discussion The current study validates a key role for GAL3 receptors in consummatory drive for sucrose, saccharin, and most notably ethanol intake in mice. We show that alcohol preferring mice drink less ethanol compared to wildtype mice when treated with the GAL3 antagonist, SNAP 37889, an effect that was also extended to morphine. This effect was independent of any sedative or motor deficit caused by the drug as illustrated by no difference between mice treated with SNAP 37889 or vehicle in both locomotor activity and rotarod performance. In addition, at the dose tested, mice did not develop a conditioned place preference to SNAP 37889 suggesting that the drug itself is not intrinsically rewarding. Liver assays revealed there was no difference in ethanol metabolising activity between SNAP 37889 or vehicle treated mice. Collectively, these data provide the first evidence that SNAP 37889 significantly decreases the intake of ethanol in mice in a binge drinking model and self-administration of morphine without affecting other important aspects of rodent behaviour. Despite evidence for galanin in the motivational aspects of eating (Crawley et al., 1990, Kyrkouli et al., 1986) and ethanol intake (Lewis et al., 2004, Rada et al., 2004), the specific subtype(s) of the galanin receptor mediating these effects is less established. Furthermore, given the calorific value of ethanol, it is unclear whether the role that GAL3 played in binge drinking occurred independently from other generalised effects upon feeding in mice. We show that antagonism of GAL3 reduces the short-term voluntary intake of ethanol, sucrose and saccharin suggesting that the motivation for binge drinking ethanol is not independent of other consummatory behaviour. However, it should be noted that SNAP 37889 had the most sustained effect on ethanol intake by significantly reducing consumption at all 4 hourly time points, compared to only the first 3 time points for sucrose and only the second and third time points for saccharin. SNAP 37889 when administered i.p. to rats has a 3254 of approximately 5 h, and is stable in plasma (ng/ml) for 4 h (Lundstrom et al., 2008). This pharmacokinetic profile of SNAP 37889 is consistent with our ethanol binge drinking data, where SNAP 37889 provided a significant effect over 4 h. The effect of this GAL3 antagonist highlights its potential to be co-administered with longer lasting drug therapies such as naltrexone, which has a half-life of 4–10 h and a slow terminal elimination-phase half-life of 96 h (Crabtree, 1983), compared to the SNAP 37889 terminal elimination-phase of approximately 25 h (Lundstrom et al., 2008). In humans, combination therapy of naltrexone and acamprosate showed an absence of contraindication in clinical practice (Mason et al., 2002). SNAP 37889 may therefore potentially be useful to help reduce ethanol intake in combination with other therapies that induce their actions via different mechanisms. This work provides new insights into the disruption by SNAP 37889 of the positive feedback loop that is present between ethanol and galanin (Barson et al., 2010, Chang et al., 2007, Karatayev et al., 2009, Picciotto, 2008). Galanin enhances DA in the NAc to augment the rewarding aspects of alcohol consumption (Rada et al., 1998), and increases NA in the PVN (Rada et al., 2004) which leads to a prompt feeding reaction. SNAP 37889 may exert its effects by reducing release of these monoamines through inhibiting galanin's stimulatory effect on both feeding and drug reward. Overall, SNAP 37889 appears to interfere with the mechanisms that drive consumption. This result is consistent with our previous findings showing SNAP 37889 (30 mg/kg) reduced operant response to ethanol, sucrose and saccharin in iP rats (Ash et al., 2011). It appears however that SNAP 37889 had the greatest effect on decreasing ethanol and sucrose intake, the two solutions containing calories, which suggests that antagonising GAL3 may be related to modifying energy balance, in addition to reward and/or palatability. While the evidence illustrating a role for GAL3 in feeding is limited, activation of GAL1 has been shown to increase food intake (Fang et al., 2012), while activation of GAL2 using the selective agonists M1153 and M1145 has shown no effect (Saar et al., 2011). The acute administration of M617, a GAL1 agonist, dramatically increases the consumption of high-fat milk (Saar et al., 2011) and when given intraventricularly, dose dependently augments consumption of cookie mash (14% fat, 79% carbohydrate and 7% protein) in Sprague Dawley rats (Lundström et al., 2005). Since GAL1 and GAL3 have comparable downstream signalling, it is likely activation of GAL3 may lead to similar effects on consummatory drive and energy balance.