We found that sex chromosome complement and not gonadal phenotype determined alcohol habit formation (): XY mice (XYM and XYF) were not sensitive to reinforcer devaluation and responded at levels comparable to those prior to devaluation, indicating habitual behavior. In contrast, XX mice (XXM and XXF) were sensitive to devaluation, demonstrating goal-directed behavior. Neither gonadal phenotype nor gonadectomy predicted whether mice responded in a habitual or goal-directed manner during the habit test, confirming that sex chromosome complement rather than gonadal status influenced behavior. Conversely, in an ad libitum
context, gonadal females consumed more alcohol than gonadal males () with no effect sex chromosome complement. A trend toward a gonadectomy by gonadal sex interaction suggests an organizing and activating influence of hormones on free alcohol drinking. These data are consistent with several reports indicating greater free alcohol consumption in female rodents (Middaugh et al., 1999
; Lancaster et al., 1996
). The consumption test also confirmed that the alcohol reinforcer was successfully devalued only in mice that underwent alcohol CTA. These data suggest that the habitual responding observed in the XY (XYM/XYF) mice could not be attributed to deficits in learning/memory of alcohol devaluation.
Research has demonstrated that reinforcer exposure, rather than the number of responses made, predicts habit formation (Adams, 1982
). Importantly, there were no differences in the number of alcohol reinforcers earned during acquisition (), though chromosomal males (XYF/XYM) had higher response rates than chromosomal females (XXF/XXM) and mice with testes (XYM/XXM) had higher response rates than those without (XXF/XYF), as shown in . This difference in response efficiency, whereby some groups make more responses but receive equal access to reinforcer, may reflect differences in motivation to work for alcohol reinforcer or disparity in the ability to track reinforcer availability. Additionally, the chromosome-mediated differences in habitual responding did not appear to be explained by differences in locomotor behavior. Locomotor activity at baseline was determined by an interaction between gonadal and chromosomal sex such that XXM mice made more beam breaks than other genotypes. While we did not observe an increase in activity in response to ethanol in this strain of mouse (MF1), after both acute and chronic exposure to alcohol, mice that developed testes (XYM/XXM) made more beam breaks during the session than those without. A non-significant trend towards an interaction between testing condition, gonadal sex and gonadectomy, indicates that the locomotor effects of alcohol are not mediated by sex chromosome complement, but rather a potential interaction between organizational and activational effects of gonadal hormone in which testicular hormones predict increased locomotor activity ().
Together, these data suggest that sex chromosome complement influences habit formation for alcohol, independently of gonadal phenotype and hormone circulation, in such a way that male chromosomal status predicts rapid alcohol habit formation. These findings stand in contrast to our data with “natural” reinforcers (Quinn et al., 2007
) where habit formation for sucrose was more rapid in XX than XY mice. Though our data do not distinguish between the pharmacological effects and caloric value of alcohol, the direction of the sex differences in the rate of habit formation of alcohol stands in direct contrast to that of habit for a sucrose reinforcer similar in caloric value, indicating that caloric value alone cannot explain the observed effect of sex chromosome complement on behavior. Sex chromosome complement, therefore, differentially affects habit formation depending on the reinforcer, possibly explaining differences in rates of dependence on various drugs and foods in men and women.
It has been reported that alcohol habits form more rapidly than habits for natural reinforcers in male animals (Dickinson et al., 2002
), but the rate of alcohol habit formation in female rodents has not been previously studied. The shift in response strategy from goal-directed action to stimulus-response habit is thought to result from a transition from ventral striatal circuitry including prefrontal cortex to a more dorsal circuit involving dorsolateral striatum (e.g. Everitt & Robbins, 2005
), and alcohol may interact with this corticostriatal circuitry to influence the rate of progression from goal-directed to habitual responding. For example, alcohol exposure can produce impaired prefrontal cortical function (Sullivan & Pfefferbaum, 2005
) and possibly disrupt input to the ventral striatum (Szumlinski et al., 2007
). Additionally, alcohol exposure has been shown to increase activation of the NR2B subunit of NMDA receptors in the dorsolateral striatum, producing enhanced signaling consistent with habit formation (Wang et al., 2007
). Furthermore, bath application of alcohol on striatal slices results in decreased long-term potentiation in the dorsomedial striatum, where decreased activity is associated with habit formation (Yin et al., 2007
). Work from Devaud et al. (2000)
also found that gonadectomy did not influence sex-specific effects of alcohol on NMDA or GABAA
receptors in cortex indicating that activational influences of hormones were not a factor; however, their study cannot distinguish whether genetic or organizational hormonal factors were involved. Our data suggest that sex chromosomes, independent of gonadal hormones, mediate sex differences in habit formation for alcohol and may underlie sex differences in alcohol-induced neuroadaptations.
How genetic factors influence the shift from goal-directed behaviors to habit is only beginning to be understood. A recent study from Yu et al. (2009)
demonstrated that the absence of A2A
adenosine receptors in the striatum impaired the development of habitual responding for food. Additionally, work from Hilario et al. (2007)
has indicated a role for endocannabinoid signaling in habit development through the use of a cannabinoid receptor type 1 (CB1) knockout mouse. Both A2A
and CB1 receptors are thought to interact with dopamine receptors as receptor heteromers to modulate neurotransmission in the striatum (Ferre et al., 2009
) and may influence dopamine receptor-mediated behaviors (Martin et al., 2008
; Hsu et al., 2010
). Notably, no studies have examined genetic factors involved in alcohol habits, and how alcohol may act at these synapses to influence long-term plasticity.
It is becoming evident that sex chromosomes play a key role in the regulation of plasticity-associated signaling cascades (Jazin & Cahill, 2010
), which may underlie shifts in cortico-striatal function. Sex differences in striatal circuitry have been shown to result from a combination of gonadal and chromosomal influences. Sex chromosome complement has been shown to influence TH expression in dopaminergic neurons in vitro
(Carruth et al., 2002
). It has also been shown that sex chromosome complement can cause differential gene expression in the adult striatum in vivo
(Chen et al., 2009
). Collectively, these studies suggest that sex chromosome-mediated differences in genes that regulate synaptic plasticity within the striatum may underlie functional sex differences in the development of habits. Study of Y chromosome genes, genes that escape X chromosome inactivation and their paralogues on the Y chromosome may provide a starting point for exploration of genetic sex differences. Further investigation into the precise neurobiological factors that mediate the divergence in habit formation caused by male and female chromosomal status, and the influence of alcohol consumption, is critical for developing gender-specific treatment and early intervention methods for individuals suffering from and at-risk for developing alcohol dependence.