Effects of isolation stress and voluntary ethanol exposure during adolescence on ethanol and nicotine co-use in adulthood using male rats

Abstract

Background: Alcohol use in adolescence may increase susceptibility to substance use disorders (SUDs) in adulthood. This study determined if voluntary ethanol (EtOH) consumption during adolescence, combined with social isolation, alters the trajectory of EtOH and nicotine intake during adulthood, as well as activating brain neuroinflammation.

Methods: Adolescent male isolate- and group-housed rats were given 0.2 % saccharin/20 % EtOH (Sacc/EtOH) or water using intermittent 2-bottle choice; controls were given water in both bottles (n=17-20 per group). Some rats from each group (n=5-6) were euthanized one week later to measure autoradiographic [³H]PK-11195 binding, an indicator of microglial reactivity, and the remainder (n=11-14 per group) were tested in adulthood in 2-bottle choice, followed by nicotine self-administration using an incremental fixed ratio (FR) schedule with Sacc/EtOH and water concurrently available.

Results: Isolation housing increased adolescent intake of Sacc/EtOH, but the increase did not produce an observable neuroimmunological response in brain. Adolescent EtOH exposure decreased adult intake of both Sacc/EtOH and unsweetened EtOH, with isolate-housed rats showing a greater effect than group-housed rats. In the co-use model, a cross-price economic demand analysis revealed a substitutional relationship between Sacc/EtOH and nicotine, but no effect of adolescent Sacc/EtOH exposure. Compared to group-housed rats, isolate-housed rats were more sensitive to the changing price of nicotine and showed greater substitutability of Sacc/EtOH for nicotine.

Conclusion: The current results suggest that adolescent EtOH exposure per se, with or without isolation stress, does not likely explain the enhanced risk for either alcohol or nicotine use later in life.

Keywords: Adolescence; Economic Demand; Ethanol; Isolation; Microglia; Nicotine.

Fig. 1

Adolescent Sacc/EtOH consumption and preference ratio in 2-bottle choice across sessions in group- and isolate-housed male rats. A) Adolescent Sacc/EtOH consumption (g/kg) in group- (open circles) and isolate- (filled circles) housed rats over the 15 intermittent 2-bottle choice sessions. B) Adolescent Sacc/EtOH consumption (g/kg). C) Water consumption (g/kg) collapsed across the 15 2-bottle choice sessions. D) Sacc/EtOH preference ratio displayed as % relative to total fluid consumption over the 15 intermittent 2-bottle choice sessions. E) Sacc/EtOH preference ratio displayed as % relative to total fluid consumption collapsed across the 15 2-bottle choice sessions. All data presented as mean ± SEM. # indicates p < 0.05, ## indicates p < 0.01 main effect of session relative to mixed-effects analysis. * Indicates p < 0.05, *** indicates p < 0.001 relative to t-test.

Fig. 2

Representative pseudocolored autoradiograms containing region-specific outlines. A) Anterior section (6.63 (+1.70)) in which [³H]-PK11195 binding was analyzed in the cingulate cortex, white matter area, somatosensory cortex, striatum, nucleus accumbens shell, and nucleus accumbens core. B) Posterior section (12.05 (-5.00)) in which [³H]-PK11195 binding was analyzed in the thalamus, dorsal hippocampus, ventral hippocampus, visual cortex, auditory cortex, and entorhinal cortex. The bar represents the pseudocolored Green Fire Blue Look Up Table (LUT) from ImageJ on a scale from 1 (dark)-256 (light).

Fig. 3

Adult 20 % EtOH consumption and preference ratio in 2-bottle choice across sessions in group- and isolate-housed male rats exposed to water or EtOH during adolescence. A) Adult 20 % EtOH consumption (g/kg) in group- (open symbols) and isolate- (filled symbols) housed rats exposed to water (circles) or Sacc/EtOH (squares) as adolescents over the seven 2-bottle choice sessions. B) Adult 20 % EtOH intake collapsed across housing over the seven 2-bottle choice sessions. C) 20 % EtOH preference ratio displayed as percent relative to total fluid consumption over the seven 2-bottle choice sessions. D) 20 % EtOH preference ratio displayed as percent relative to total fluid consumption collapsed across the seven 2-bottle choice sessions. All data presented as mean ± SEM. # indicates main effect of housing, p < 0.05; & indicates session x adolescent exposure interaction relative to the mixed-effects analysis, p < 0.05. *p < 0.05, **p < 0.01, ***p < 0.001, relative to Bonferroni multiple comparisons test.

Fig. 4

Adult Sacc/EtOH consumption and preference ratio in 2-bottle choice across sessions in group- and isolate-housed male rats exposed to water or EtOH during adolescence. A) Adult Sacc/EtOH consumption (g/kg) in group- (open symbols) and isolated- (closed symbols) housed rats exposed to water (circles) or Sacc/EtOH (squares) as adolescents over the seven 2-bottle choice sessions. B) Adult Sacc/EtOH consumption collapsed across the seven 2-bottle choice sessions. C) Sacc/EtOH preference ratio displayed as percent relative to total fluid consumption over the seven 2-bottle choice sessions. D) Sacc/EtOH preference ratio displayed as percent relative to total fluid consumption collapsed across the seven 2-bottle choice sessions. All data presented as mean ± SEM. # indicates main effect of adolescent exposure relative to the mixed-effects analysis, p < 0.05. **p < 0.01, relative to Bonferroni multiple comparisons test.

Fig. 5

EtOH-Nicotine Co-Use: A) Nicotine intake (mg/kg) across conditions over each FR. B) Mean nicotine intake at each FR, collapsed across adolescent exposure groups. C) Nicotine intake for each condition collapsed across FR. D) Sacc/EtOH consumption (g/kg) across conditions at each FR. All data presented as mean ± SEM. &,&,&, indicates housing x FR interaction relative to mixed-effects model, p < 0.001; # indicates housing x adolescent exposure interaction relative to mixed-effects model, p < 0.05. *p < 0.05, **p < 0.01, ***p < 0.001, relative to Bonferroni multiple comparisons test.

Fig. 6

Demand elasticity for nicotine and Sacc/EtOH: A-D) Nicotine (mg/kg) and Sacc/EtOH intake (g/kg) for group-housed EtOH treated (filled symbols) and controls (open symbols) as a function of the changing price in nicotine. E & F) Normalized intake for Sacc/EtOH (squares) and nicotine (circles) for EtOH treatment (filled line) and controls (dotted line) as a function of the changing price in nicotine in group- (E) and isolate- (F) housed animals. Values are expressed as percent of maximal intake for individual subjects and curves were derived from nicotine own-price and Sacc/EtOH cross-price equations. All data presented as mean ± SEM.