Stressed Avoider rats show blunted sensitivity to alcohol's aversive effects: Potential contributions of the lateral habenula and lateral hypothalamus

Abstract

Avoidance coping following stress exposure predicts heightened alcohol drinking. Similarly, blunted sensitivity to the aversive effects of alcohol facilitates increased drinking. However, the relationship between stress exposure, coping mechanism, and sensitivity to alcohol's aversive effects is unknown. In rats, predator odor stress increases alcohol intake in animals that show persistent avoidance of stress-paired stimuli, termed "Avoiders". Here, we tested the hypothesis that Avoider rats have blunted sensitivity to alcohol's aversive effects using an alcohol-induced conditioned taste aversion (CTA) paradigm. After a single conditioning session, Non-Avoider rats acquired alcohol-induced CTA while Avoiders did not. Male rats across all groups eventually acquired alcohol CTA after four conditioning sessions. However, in females, only Non-Avoiders acquired alcohol-induced CTA. In male Non-Avoider rats, a single CTA-inducing dose of alcohol increased cFos expression in the lateral habenula (LHb), an important nucleus in aversion signaling. In male Avoiders, the same dose of alcohol decreased LHb cFos expression. cFos expression in the lateral hypothalamus (LH), which provides glutamatergic inputs to the LHb, was also diminished by alcohol in male Avoider rats. In females, alcohol had no effect on cFos cell counts in the LHb. However, in the LH, alcohol diminished cFos expression in female Non-Avoiders. Collectively, these findings suggest that stressed Avoider rats are hyposensitive to alcohol's aversive effects, which may facilitate their heightened alcohol drinking after stress. Sex- and stress group-specific differences in LH and LHb recruitment highlight these regions as candidates for mediating stress-induced changes in alcohol behaviors.

Keywords: alcohol; aversion; avoidance; central amygdala; lateral habenula; lateral hypothalamus; predator odor.

Figure 1:. Non-Avoider rats develop CTA to 1.5 g/kg alcohol injection, some Avoiders do not.

A. Experiment timeline. B. Pre-conditioning baseline saccharin intake (in Session 1) in Avoiders and Non-Avoiders. C. Change in saccharin intake after one conditioning session in which an alcohol or saline injection was paired with saccharin access. D. Saccharin intake in each of all 4 conditioning sessions in male and E. female rats. F. Percent decrease in saccharin consumption from the first to the last conditioning session. G. Correlations between time spent in odor-paired chamber (avoidance scores) and percent decrease in saccharin consumption in males and H. females. Created in BioRender. Bonauto, S. (2025) https://BioRender.com/klnmq1f

Figure 2:. Alcohol blunts activity in the LHb of Male Avoiders and increases activity in the CeA after stress and 1.5 g/kg ethanol i.p. injection.

A, C, E. cFos density (count of positive cFos expressing cells per area measured in pixels) of males separated by stress group (Control, Non-Avoider, and Avoider) and treatment with each stress group (Saline or Alcohol). B, D, F. cFos density (count of positive cFos expressing cells per area measured in pixels) of females separated by stress group (Control, Non-Avoider, and Avoider) and treatment with each stress group (Saline or Alcohol).

Figure 3:. Correlations of cFos density between key regions

Heat maps of cFos density (measured by count per area in pixels) of the LH, LHb, and CeA divided by sex, stress group, and treatment of alcohol or saline. In each sex, the cFos densities between brain areas were compared within stress group and treatment. Correlations between cFos density in the LH and LHb were seen in the Control Saline, Non-Avoider Alcohol, and Avoider Alcohol groups.

Figure 4:. Representative images of regions quantified for cFos.

Sample images of the LHb, LH, and CeA from male rats reflecting all experimental conditions: Control, Non-Avoider, or Avoider treated with Saline or Alcohol. Abbreviations: MHb: medial habenula, sm: stria medullaris of the thalamus, PVT: paraventricular thalamic nucleus, fr: fasciculus retroflexus, IC: internal capsule, f: fornix, ZI: zona incerta, Opt: optic tract, EGP: external globus pallidus, MeA: medial amygdala, LA: lateral amygdala, STIA: striatum, CPu: caudate putamen (striatum).

Figure 5:. Blood Alcohol Concentrations

A. The concentration of ethanol (mg/dl) in serum of males before injection and 30 and 90 minutes after injection. B. The concentration of ethanol (mg/dl) in serum of females before injection and at 30 and 90 minutes after injection.

Figure 6.. Hyperactive exploration behavior in Avoider females during predator odor CPA

A. Representative heat maps for one rat in each stress group and sex. Color scale represents the proportion of total duration (5 minutes) in designated location, red is greater time. B. Stress group outcomes based on 4-day predator odor CPA for males and females (male Avoiders = 20; female Avoiders = 16). C. Latency to enter the odor-associated context during Posttest for all stressed rats. D. Number of entries into odor-paired context for stressed rats. E. Average velocity within the odor-paired context. F. Total distance traveled within the odor-paired chamber. G. Percentage of time outside of odor-paired context that was spent in the center zone (proximal to odor-paired context). H. Total distance traveled within the entire apparatus during avoidance Posttest.