Heavy drinking and alcoholism are associated with decreases in the mass of brain grey and white matter, whereas abstinence in recovering alcoholics is associated with a progressive increase in brain mass (Pfefferbaum et al., 1995). Recent discoveries have indicated that the adult brain contains Neural Progenitor Cells (NPC) that form new neurons, gila and other nervous system cells. NPC actively proliferate under normal conditions in hippocampus and forebrain. NPC that exit cell cycle often die, although many migrate and differentiate over a period of weeks becoming mature neurons and other nervous system cells. Proliferation and differentiation-survival are independently regulated. We have found that acute ethanol administration reduces hippocampal NPC proliferation and chronic administration decreases both hippocampal proliferation and survival (Nixon and Crews, 2002). Withdrawal from chronic ethanol increases NPC proliferation and the formation of new neurons, e.g. neurogenesis. This proposal will test the overall hypothesis that ethanol administration alters adult NPC in a manner that contributes to the changes in brains of active alcoholics as well as recovering alcoholics. The specific hypotheses to be tested are: 1. Acute ethanol disrupts NPC proliferation. Complete ethanol dose and time courses will be determined for the entire brain. Immunocytochemistry using specific proliferation markers, such as BrdU, PCNA and Ki67, antigens marking different stages of cell cycle, as well as markers of cell death, e.g. TUNEL and antigens identifying apoptosis-necrosis will be determined. 2. Acute ethanol will disrupt NPC differentiation and survival. Various stages of NPC differentiation and cell death will be examined by prelabeling NPC with BrdU and immunocytochemistry for specific antigens to detect the changes in NPC differentiation with alcohol exposure. 3. Chronic Ethanol reduces NPC proliferation as well as differentiation and survival. We have found that 4-day binge ethanol administration induces ethanol neurotoxicity, tolerance and dependence as well as a reduction in hippocampal neurogenesis. Progressive changes in neurogenesis during chronic ethanol administration will be determined using immunocytochemical techniques to investigate clonal selection and altered differentiation. 4. Ethanol withdrawal increases NPC proliferation, and neurogenesis. We have discovered that withdrawal from 4-day binge ethanol administration results in increases in NPC proliferation in a variety of brain regions and in new neuron formation at 5 weeks of withdrawal. NPC will be identified by BrdU and other antigens. We will establish a time-course of ethanol withdrawal to distinguish proliferating and differentiating NPC as well as the consequent neurogenesis and gliogenesis from NPC. It is expected that ethanol withdrawal will result in the formation of new neural cells throughout brain that contribute to long term changes in brain. Understanding the effects of ethanol on NPC proliferation and differentiation-survival could further our comprehension of the CNS dysfunction that occurs in alcoholism and the subsequent regenerative processes in the brains of withdrawn abstinent recovering alcoholics.
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