Effects of Age on Ethanol Withdrawal Toxicity: Mechanisms and Therapy

描述(申请人提供)：酒精中毒是长期酒精滥用的结果，与认知和精神运动功能的老化有关。由于酗酒者会经历反复的酒精戒断(EW)，我们建议使用重复EW来研究衰老是否通过氧化侮辱线粒体来加剧EW对认知和精神运动功能的影响。我们还建议调查雌激素的丢失是否会导致这个问题。这项工作对女性酗酒者可能有特别的意义，她们在晚年同时经历了EW和雌激素缺乏症。我们的初步研究表明，雌激素对年轻和老年雌性大鼠的EW毒性都有保护作用。在拟议的研究中，我们将使用三个年龄组的卵巢完整的雌性大鼠：5个月大、12个月大和16个月大，开始酒精饮食。我们建议使用25天的酒精饮食周期，然后5天停药，重复5个周期。这些周期的时间安排将使大鼠处于发情期(高雌激素水平)，此时停止饮食，新的体征达到顶峰。在目标1中，我们将确定电子战在衰老过程中对认知和精神运动功能的影响。处于发情期的大鼠将从间歇性酒精饮食中撤出，并使用既定的行为方法测试精神运动功能。在目标2中，我们将描述AGE和EW之间有害相互作用背后的线粒体氧化机制。我们将测量氧化标记物(活性氧物种、脂质过氧化)和随之而来的线粒体功能障碍(线粒体膜电位崩溃和ATP丢失)。在目标3中，我们将表征氧化修饰的线粒体酶细胞色素c氧化酶(COX)，我们假设它介导了AGE和EW之间的有害相互作用。我们将用液相色谱-质谱法和串联质谱法对COX和过氧化脂质修饰的COX进行定量。在目标4中，我们将确定衰老过程中雌激素缺乏是否会导致电子政务导致的线粒体氧化损伤和精神运动障碍。同样的精神运动和氧化标记物将在卵巢完整的大鼠和去卵巢的大鼠身上被测量，无论是否使用雌激素替代。这些研究最终可能有助于更好地理解和应对女性酗酒者在大脑老化过程中接受电击的情况。

DESCRIPTION (provided by applicant): Alcoholism is a consequence of prolonged ethanol abuse and has been associated with the aging of cognition and psychomotor function. Because alcoholics go through repeated rounds of ethanol withdrawal (EW), we propose using repeated EW to investigate whether aging with exacerbates the effects of EW on cognition and psychomotor function through oxidative insults to mitochondria. We also propose to investigate whether a loss of estrogen contributes to this problem. This work may have particular significance for female alcoholics, who simultaneously undergo EW and estrogen deficiency in their advanced age. Our pilot study has demonstrated that estrogen protects against EW toxicity in both young and old female rats. In the proposed study, we will use three age groups of ovary-intact female rats: 5 months old, 12 months old, and 16 months old, when the ethanol diet begins. We propose to use a diet cycle of 25 days of ethanol followed by 5 days of withdrawal repeated for 5 cycles. These cycles will be timed such that rats will be at the estrus phase (high estrogen levels) when diet is removed and EW signs peak. In Aim 1, we will determine the effects of EW on cognition and psychomotor functions during aging. Rats at the estrus phase will be withdrawn from an intermittent ethanol diet and tested for psychomotor function using established behavioral methods. In Aim 2, we will characterize the mitochondrial oxidative mechanisms underlying the deleterious interaction between age and EW. We will measure oxidative markers (reactive oxygen species, lipid peroxidation) and consequent mitochondrial dysfunction (collapse of mitochondrial membrane potential and ATP loss). In Aim 3, we will characterize oxidatively modified mitochondrial enzyme cytochrome c oxidase (COX) which we hypothesize mediates the deleterious interaction between age and EW. We will quantify carbonylation of COX and lipid-peroxide-modified COX using liquid chromatography-mass spectrometry and tandem mass spectrometry. In Aim 4, we will determine whether estrogen deficiency during aging contributes to EW-induced oxidative damage to mitochondria and psychomotor deficit. The same psychomotor and oxidative markers will be measured in ovary-intact and ovariectomized rats with or without estrogen replacement. These studies may ultimately contribute to a better understanding of and strategy for female alcoholics undergoing EW during brain aging.