High alcohol/sucrose consumption during dark circadian phase in C57Bl/6J mice: Involvement of hippocampus, lateral septum and urocortin-positive cells of the Edinger-Westphal nucleus

Rationale: Identification of the neuroanatomical substrates regulating alcohol consumption is important for the understanding of alcoholism. Previous studies mapping changes in brain activity used rodent models of alcohol drinking with relatively low alcohol intakes. Objectives: This study was aimed to identify brain regions changing activity after high voluntary intake of alcohol-containing solutions. Methods: Adult male C57BL/6J mice were trained to drink a 10% ethanol/10% sucrose solution in daily 30-min limited-access sessions during the dark phase of the circadian cycle. Control groups of animals consumed 10% sucrose or water. Analysis of c-Fos immunohistochemistry (as a marker for neuronal activity) was performed at 90 min after the last alcohol drinking session. Results: The limited access procedure led to high intakes (2.9±0.3 g/kg) and blood alcohol concentrations of 251±46 mg%. Expression of c-Fos was significantly higher in the alcohol/sucrose group than both the water and sucrose groups in the Edinger-Westphal nucleus, and significantly lower in the alcohol/sucrose group than two control groups in hippocampal subregions, posterior hypothalamus and dorsal lateral septum. Double immunohistochemistry showed that alcohol-induced c-Fos-positive cells in the Edinger-Westphal nucleus colocalized with the neuropeptide urocortin. In addition, intake and/or blood alcohol concentrations correlated with c-Fos expression in specific subregions of the hippocampus, hypothalamus, prefrontal cortex, lateral septum and midbrain. Conclusions: The dark phase voluntary limited-access procedure in mice leads to intakes of alcohol-containing solutions that are considered highly intoxicating. Brain regions showing alcohol-specific changes in c-Fos expression after this procedure can be connected into a novel neurocircuit, including lateral septum, hippocampus, hypothalamus, and the Edinger-Westphal nucleus.