Flumazenil on Multiple Stress and Withdrawal Anxiety

描述(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诱导的焦虑对长期接触乙醇的敏化，进一步支持Ballenger和Post(1978)的“点燃”假说。一个主要的发现是，苯二氮(BZD)拮抗剂氟马西尼可以最大限度地减少反复戒断方案后焦虑的敏化。根据氟马西尼的这些数据，推测一种对GABAA受体有负面影响的内源性物质是由反复应激敏化焦虑而引起的焦虑敏化。为了支持这一假设，给予两剂DMCM，一种BZD反向激动剂，而不是戒断，会导致在最后一次戒断期间焦虑的敏化。本次调查的目的是检验解释这些发现的具体假设。具体目标1将确定氟马西尼是否会拮抗戒断诱导的焦虑的反复应激敏感化，以及这种治疗是否导致内源性BZD反向激动剂安定结合抑制物(DBI)的mRNA表达增加，从而确定DBI的适应性变化可能对戒断诱导的焦虑起作用的大脑部位。特定目标2将测试反复应激或暴露于BZD反向激动剂是否会持续诱导戒断敏感化-在稍后再次暴露于慢性乙醇时诱发焦虑，就像反复戒断发生的那样。此外，我们将确定在多次戒断期间暴露于压力或BZD反向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。特定目标3将确定在特定目标1中确定的脑区微量注射氟马西尼是否会阻止多重应激后戒断诱导的焦虑的敏化，以及将BZD反向激动剂微量注射到选定的部位是否会敏化戒断诱导的焦虑。具体目标4将测试GABA(A)和BZD结合是否受到重复应激和单次戒断的影响，以及重复应激是否增加了对BZD反向激动剂的反应性。此外，还将进行实验，以确定对氟马西尼敏感的内源性化合物是否会随着反复停药而增加。明确病理性适应机制的基础(S)负责点燃与重复应激相关的戒断诱导的焦虑，可能会为更好地了解酒精滥用提供更好的见解，并提供新的治疗途径。

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.