Flumazenil on Multiple Stress and Withdrawal Anxiety

描述（由申请人提供）：我们实验室的工作已经证明，反复的压力和戒断会导致戒断引起的焦虑对短暂暴露于慢性乙醇的敏感化，进一步支持Ballenger和Post（1978）的“点火”假说。一个主要的发现是氟马西尼，一种苯二氮卓类药物（BZD）拮抗剂，在重复戒断方案后最小化焦虑的敏感性。基于氟马西尼的这些数据，我们推测一种内源性物质对GABAA受体有负作用，是焦虑反复应激致敏引起焦虑致敏的原因。为了支持这一假设，两剂DMCM（一种bzd逆激动剂）代替戒断会导致最终戒断期间焦虑的敏感化。本研究的目的是检验具体的假设，以解释这些发现。特异性目的1将确定氟马西尼是否会拮抗戒断性焦虑的反复应激致敏，以及这种治疗是否会导致地西泮结合抑制剂（DBI） mRNA表达的增加，DBI是一种内源性bzd逆激动剂，从而确定DBI的适应性变化可能导致戒断性焦虑的大脑部位。特异性目标2将测试反复应激或暴露于bzd逆激动剂是否会在随后再次暴露于慢性乙醇后持续诱导戒断诱导焦虑的敏化，正如反复戒断所发生的那样。此外，我们将确定在多次停药期间暴露于压力或bzd逆激动剂是否会延长持续时间，在此期间，再次暴露于慢性乙醇将继续导致焦虑敏化。特异性目标3将确定在特异性目标1中确定的大脑区域中微量注射氟马西尼是否会阻断多重应激后戒断性焦虑的致敏，以及在选定的部位微量注射bzd逆激动剂是否会使戒断性焦虑致敏。特异性目标4将测试GABA(A)和BZD的结合是否受到重复应激和单次停药的影响，以及对BZD逆激动剂的反应性是否因重复应激而增加。这将通过实验加以补充，以确定对氟马西尼敏感的内源性化合物是否随着反复停药而增加。定义病理适应机制的基础，负责“点燃”与反复压力相关的戒断性焦虑，可能会为酒精滥用提供更深入的了解，并为治疗提供新的途径。

DESCRIPTION (provided by applicant): Work in our laboratory has demonstrated that repeated stresses and withdrawals leads to sensitization of the withdrawal-induced anxiety to a brief exposure to chronic ethanol, in further support of the Ballenger and Post (1978)"kindling" hypothesis. A major finding is that flumazenil, a benzodiazepine (BZD) antagonist, minimizes the sensitization of anxiety following the repeated withdrawal protocol. Based upon these data with flumazenil, it is presumed that an endogenous substance having a negative effect on GABAA receptors is responsible for the sensitization of anxiety induced by the repeated stress sensitization of anxiety. In support of this hypothesis, two doses of DMCM, a BZD-inverse agonist, given instead of withdrawal results in sensitization of anxiety during a final withdrawal. The purpose of the present investigation is to test specific hypotheses to account for these findings. Specific Aim 1 will determine whether repeated stress sensitization of withdrawal-induced anxiety will be antagonized by flumazenil and whether this treatment results in an increase in mRNA expression for diazepam binding inhibitor (DBI), an endogenous BZD-inverse agonist, allowing identification of brain sites where adaptive change in DBI could be contributing to withdrawal-induced anxiety. Specific Aim 2 will test whether the repeated stresses or exposure to a BZD-inverse agonist will persist in inducing sensitization of withdrawal-induce anxiety upon later re-exposure to chronic ethanol, as occurs with repeated withdrawals. Additionally, we will determine if exposure to stress or a BZD-inverse agonist during multiple withdrawals will extend the duration during which later re-exposure to chronic ethanol will continue result in sensitization of anxiety. Specific Aim 3 will determine if microinjection of flumazenil into brain regions identified in Specific Aim 1 will block sensitization of withdrawal-induced anxiety following multiple stresses and whether microinjection of a BZD-inverse agonist into the selected sites will sensitize withdrawal-induced anxiety. Specific Aim 4 will test whether GABA(A) and BZD binding is affected by repeated stresses and a single withdrawal and whether responsiveness to a BZD-inverse agonist is increased by the repeated stresses. This will be complemented by experiments to identify whether an endogenous compound sensitive to flumazenil increases with repeated withdrawals. Defining the basis of the pathological adaptive mechanism(s) responsible for "kindling" of withdrawal-induced anxiety associated with repeated stresses may provide greater insight into alcohol abuse and provide new avenues for treatment.