Galanin can modulate neuronal activity in brain regions related to drug dependence and withdrawal. The neuropeptide is expressed in most of the noradrenergic neurons of the LC (Xu et al., 1998
), and is increased after chronic morphine administration and withdrawal (McClung et al., 2005
). Studies in vitro
have shown that galanin decreases the firing of LC neurons by hyperpolarizing LC noradrenergic neurons (Rasmussen et al., 1990
;Seutin et al., 1989
;Sevcik et al., 1993
), an effect hypothesized to decrease signs of opiate withdrawal. This has been proposed to be due to a GalR1-mediated increase in membrane permeability to potassium ions (Pieribone et al., 1995
The current dataset demonstrates that galanin expression is regulated in the LC under conditions leading to morphine dependence and withdrawal, and shows that GalR1 is critical for mediating the effects of galanin to reduce opiate withdrawal signs. Therefore, these studies identify GalR1 as a novel target for the treatment of opiate dependence and withdrawal. This study complements earlier work on the ability of the galanin system to oppose the behavioral correlates of morphine dependence and withdrawal (Zachariou et al., 2003
Opiate dependence and withdrawal produce adaptive changes in the cAMP pathway, and one of the most robust changes is increased adenylyl cyclase levels, and thus cAMP production, in the noradrenergic LC neurons (Kogan et al., 1992
). In vitro
studies show that galanin inhibits cAMP formation following chronic morphine administration and withdrawal (Hawes et al., 2006
). The first evidence for a potential role of the GalR1 subtype in the actions of galanin on opiate withdrawal came from in situ
hybridization studies showing up-regulation of this receptor in LC neurons following chronic morphine administration and withdrawal (Zachariou et al., 2000
). Furthermore, GalR1 expression in the LC is also modulated by galanin signaling. Both the opiate administration/withdrawal-induced up-regulation of GalR1 and the modulation of GalR1 levels by galanin occur through a cAMP-dependent mechanism involving phosphorylation of the transcription factor cAMP regulatory element-binding protein (CREB) that regulates the GalR1 promoter (Zachariou et al., 2001
;Hawes et al., 2005
). Here, we show that transcription of the galanin peptide is also increased in LC neurons following opiate dependence and withdrawal. Consistent with the opponent process theory (Koob and Le, 2008
;Koob and Volkow, 2010
) this up-regulation, along with an increase in GalR1, may be part of a compensatory mechanism to oppose both the increased firing, and cAMP production, of the LC neurons following chronic morphine administration and withdrawal (Hawes et al., 2005
;Zachariou et al., 2000
). Consistent with previous findings that application of the galanin agonist galnon alleviates several signs of opiate withdrawal (Zachariou et al., 2003
), we show that galanin over-expression in the LC leads to a milder withdrawal syndrome, most likely mediated via GalR1. Interestingly, in naïve animals, galanin appears to have a different effect on stress mediated responses, as galanin over-expression leads to augmented stress evoked noradrenaline release (Kehr et al., 2001
). We speculate that galanin has distinct roles in stress responses in naïve versus morphine dependent mice. Also, as mentioned above, galanin-mu opioid receptor interactions in the LC are critical for the modulation of LC firing (Sevick et al., 1993). It is possible that GalR-mu opioid receptor dimers can be formed and might have distinct signaling properties that would be abolished in GalR1 knockout mice, resulting in increased LC firing in response to noradrenaline, and a more severe withdrawal phenotype, but this remains to be explored at the molecular level.
Our behavioral data using genetically modified mice, lacking GalR1 or GalR2, show that GalR1 is selectively involved in regulating in opiate withdrawal behaviors, as mice lacking this receptor subtype show significantly more exaggerated symptoms of morphine withdrawal than their WT controls, whereas the GalR2 knockout mice show no overall significant withdrawal phenotype. However, although not statistically significant, there is some reduction in jumps, wet dog shakes and ptosis, which might be attributed to compensatory changes in GalR1 receptor expression/function in GalR2 mutants. These behavioral studies were conducted on several genetically modified mouse lines and their respective WT controls. Transgenic or mutant founder animals were paired with WTs of the same strain to generated heterozygotes, from which were bred homozygous transgenic or mutant mice and their respective WTs. Therefore, although these studies did not use littermates derived from continuous heterozygous breeding (which can be a limitation when interpreting behavioral data, but avoids the generation of unwanted mice) the WTs used were strain, age and sex-matched and housed in an identical environment. Our findings are in agreement with previous knowledge and are coherent between the various genetically modified mice used in this study. These studies help clarify the function of galanin receptor subtypes in morphine dependence and withdrawal and provide important information for new pharmacological strategies for the treatment of morphine dependence and addiction.
Several lines of evidence including in situ
hybridization, gene array analysis, pharmacological and behavioral studies indicate an important role for the galanin peptide and GalR1 receptor subtype in opiate reward, dependence and withdrawal (Picciotto, 2008
). In spite of the importance of alleviating withdrawal to help addicts achieve abstinence, very few drugs to date are available to treat opiate withdrawal symptoms. Compounds targeting α2 adrenergic receptors, such as clonidine, show limited success and have significant side effects. Thus, GalR1 agonists may provide a novel approach to alleviate opiate withdrawal.
In summary, the current set of studies provides novel information about the mechanisms underlying the actions of galanin in opiate dependence and withdrawal and point to GalR1 as a potential new pharmacological target.