Accumulating evidence indicates that the adolescent hippocampus is highly susceptible to alcoholinduced structural damage and behavioral deficits. Microglia are vitally important brain constituents needed to support and maintain proper neural function; however, alcohol's effects on microglia have only recently gained attention. The microglial response to alcohol during adolescence has yet to be studied; therefore, we examined hippocampal microglial activation in an adolescence binge alcohol exposure model. Adolescent male Sprague-Dawley rats were administered ethanol 3 times/day for 4 days and were sacrificed 2, 7, and 30 days later. Bromodeoxy-Uridine was injected 2 days after ethanol exposure to label dividing cells. Microglia morphology was scored using the microglia marker Iba-1, while the extent of microglial activation was examined with ED-1, major histocompatability complex-II (MHC-II), and tumor necrosis factor (TNF)-α expression. Ethanol induced significant morphological change in hippocampal microglia, consistent with activation. In addition, ethanol increased the number of BrdU+ cells throughout all regions of the hippocampus 2 days after the last dose. Confocal microscopy showed that the proliferating BrdU+ cells in each region were Iba-1+ microglia. Importantly, newly born microglia survived and retained their morphological characteristics 30 days after ethanol exposure. Ethanol did not alter hippocampal ED-1, MHC-II, or TNF-α expression, suggesting that a single period of binge ethanol exposure does not induce a full microglial-driven neuroinflammatory response. These results establish that ethanol triggers partial microglial activation in the adolescent hippocampus that persists through early adulthood, suggesting that alcohol exposure during this unique developmental time period has long-lasting consequences.

Binge alcohol exposure in adolescent rats potently inhibits adult hippocampal neurogenesis by altering neural progenitor cell (NPC) proliferation and survival; however, it is not clear whether alcohol results in an increase or decrease in net proliferation. Thus, the effects of alcohol on hippocampal NPC cell cycle phase distribution and kinetics were assessed in an adolescent rat model of an alcohol use disorder. Cell cycle distribution was measured using a combination of markers (Ki-67, bromo-deoxy-uridine incorporation, and phospho-histone H3) to determine the proportion of NPCs within G1, S, and G2/M phases of the cell cycle. Cell cycle kinetics were calculated using a cumulative bromo-deoxy-uridine injection protocol to determine the effect of alcohol on cell cycle length and S-phase duration. Binge alcohol exposure reduced the proportion of NPCs in S-phase, but had no effect on G1 or G2/M phases, indicating that alcohol specifically targets S-phase of the cell cycle. Cell cycle kinetics studies revealed that alcohol reduced NPC cell cycle duration by 36% and shortened S-phase by 62%, suggesting that binge alcohol exposure accelerates progression through the cell cycle. This effect would be expected to increase NPC proliferation, which was supported by a slight, but significant increase in the number of Sox-2+ NPCs residing in the hippocampal subgranular zone following binge alcohol exposure. These studies suggest the mechanism of alcohol inhibition of neurogenesis but also reveal the earliest evidence of the compensatory neurogenesis reaction that has been observed a week after binge alcohol exposure.

Adult neurogenesis is now widely accepted as an important contributor to hippocampal integrity and function but also dysfunction when adult neurogenesis is affected in neuropsychiatric diseases such as alcohol use disorders. Excessive alcohol consumption, the defining characteristic of alcohol use disorders, results in a variety of cognitive and behavioral impairments related wholly or in part to hippocampal structure and function. Recent preclinical work has shown that adult neurogenesis may be one route by which alcohol produces hippocampal neuropathology. Alcohol is a pharmacologically promiscuous drug capable of interfering with adult neurogenesis through multiple mechanisms. This review will discuss the primary mechanisms underlying alcohol-induced changes in adult hippocampal neurogenesis including alcohol's effects on neurotransmitters, CREB and its downstream effectors, and the neurogenic niche.

Background Chronic intake of ethanol has been linked to serious health consequences such as cardiac and liver problems, cognitive impairments, and brain damage. Alcohol’s detrimental effects depend upon the dose, duration and pattern of exposure with binge drinking as one of the most common, but most damaging, patterns of intake. Little is known about the threshold of the damaging effects of alcohol. Therefore, these experiments sought to determine a threshold for brain damage using various markers of neurodegeneration. Methods Adult male Sprague-Dawley rats were administered nutritionally complete liquid diet containing either ethanol (25% w/v) or isocaloric dextrose every 8 hours for either 1 (mean dose: 13.4 ± 0.3 g/kg/day, mean BEC: 336.2 ± 18.8 mg/dl) or 2 days (mean dose: 10.9 ± 0.3 g/kg/day, mean BEC: 369.8 ± 18.1 mg/dl). Based on a known time course of various neurodegeneration-associated events, rats were perfused transcardially immediately following, 2 days after, or 7 days post ethanol exposure. To label actively dividing cells, some animals were injected with BromodeoxyUridine (BrdU) two hours prior to perfusion. Tissue was then analyzed for the presence of BrdU (cell proliferation), FluoroJade B (degenerative neurons), and vimentin (reactive astrogliosis) immunoreactivity. Results One or two days of ethanol exposure failed to alter cell proliferation at any of the time points analyzed. However, significant 2 to 9-fold increases in neuronal degeneration in limbic cortex and clear evidence of reactive gliosis as indicated by a 2 to 8-fold upregulation in vimentin immunoreactivity in the hippocampus were observed following as little as one day of binge ethanol exposure. Conclusions These results indicate that as little as one day (24 hours) of high BEC, binge-like ethanol exposure is enough to elicit signs of alcohol-induced brain damage in adult rats. Further, reactive gliosis may be a more sensitive marker of alcohol-induced damage in the hippocampus.

Neural stem cell-driven adult neurogenesis contributes to the integrity of the hippocampus. Excessive alcohol consumption in alcoholism results in hippocampal degeneration that may recover with abstinence. Reactive, increased adult neurogenesis during abstinence following alcohol dependence may contribute to recovery, but the mechanism driving reactive neurogenesis is not known. Therefore, adult, male rats were exposed to alcohol for four days and various markers were used to examine cell cycle dynamics, the percentage and number of neural progenitor cell subtypes, and the percentage of quiescent versus activated progenitors. Using a screen for cell cycle perturbation, we showed that the cell cycle is not likely altered at 7 days in abstinence. As the vast majority of Bromodeoxyuridine-positive (+) cells were co-labeled with progenitor cell marker, Sox2, we then developed a quadruple fluorescent labeling scheme to examine Type-1, -2a, -2b and -3 progenitor cells simultaneously. Prior alcohol dependence indiscriminately increased all subtypes at 7 days, the peak of the reactive proliferation. An evaluation of the time course of reactive cell proliferation revealed that cells begin proliferating at 5 days post alcohol, where only actively dividing Type 2 progenitors were increased by alcohol. Furthermore, prior alcohol increased the percentage of actively dividing Sox2+ progenitors, which supported that reactive neurogenesis is likely due to the activation of progenitors out of quiescence. These observations were associated with granule cell number returning to normal at 28 days. Therefore, activating stem and progenitor cells out of quiescence may be the mechanism underlying hippocampal recovery in abstinence following alcohol dependence.

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These data indicate that acute blockade of the NPY Y1 receptor with a systemically bioavailable NPY Y1RA reduces voluntary ethanol consumption by C57BL/6J mice. These results are consistent with observations that hypothalamic infusion of NPY increases ethanol drinking by rats.

These data suggest that low NPY levels in the amygdala and/or the shell of the NAc, which are not compensated for by similar changes in CRF levels, may contribute to the high ethanol consumption characteristic of C57BL/6J mice.