Viral-Mediated Alterations in CaMKII Alter Behavioral Responding to Amphetamine

描述（由申请人提供）：反复将大鼠暴露于安非他明（AMPH）等药物可导致运动反应增强、脑桥核（NAcc）多巴胺（DA）溢出和大鼠在数周至数月后再次暴露于药物时服药。这些现象是敏感化的表现，可能在从偶然使用药物向滥用药物过渡的过程中发挥重要作用。揭开神经适应的基础表达敏化增强的行为和神经化学反应的药物-可能有助于确定治疗干预的目标。在介导敏化表达的信号级联中起重要作用的许多蛋白质之一是钙/钙调蛋白依赖性蛋白激酶II（CaMKII）。而急性AMPH诱导的运动反应和NAcc DA释放是钙依赖性的，敏化运动和DA释放是钙和CaMK II依赖性的。此外，暴露于反复，间歇注射AMPH的大鼠纹状体切片显示钙调素水平增强和CaMKII活性增加相比，从盐水暴露的控制。我们假设，使用病毒介导的基因转移，以修改CaMKII水平的NAcc将改变对AMPH的行为反应。在这个资助申请中提出的实验将测试两个主要假设。假设1：增加NAcc中CaMKII的水平将增强对低阈值剂量AMPH的运动反应，并增强药物的自我给药，其作用类似于在暴露于致敏药物方案的大鼠中观察到的作用。大鼠将通过手术植入双侧引导插管和静脉内导管。然后进行病毒载体的显微注射，并进行运动和自我给药实验以确定这种操作如何改变对AMPH的敏感性。将使用免疫印迹法测定病毒介导的基因转移的效力。假设二：在致敏大鼠的NAcc中增加CaMKII的无活性突变形式的水平将减少在这些大鼠中观察到的增强的AMPH自我给药。将大鼠暴露于AMPH的重复、间歇注射，随后是停药期，在此期间将对动物进行手术准备。在药物自我给药一段时间后，将进行病毒载体的显微注射，随后评估这种操作对药物自我给药的影响。使用成瘾动物模型是研究从偶然到强迫性药物使用转变的有效方法。这项研究将有助于预防药物成瘾的最终目标，通过调查如何防止在先前暴露于AMPH的大鼠中观察到的增强药物摄入。

DESCRIPTION (provided by applicant): Repeatedly exposing rats to drugs such as amphetamine (AMPH) can lead to enhanced locomotor responding, nucleus accumbens (NAcc) dopamine (DA) overflow and drug taking when rats are re-exposed to the drug weeks to months later. These phenomena, manifestations of sensitization, may play an important role in the transition from casual drug use to abuse. Unraveling the neuroadaptations that underlie the expression of sensitization-enhanced behavioral and neurochemical responding to the drug-- may help identify targets for therapeutic intervention. One of many proteins that play an important role in mediating signaling cascades underlying the expression of sensitization is calcium/calmodulin-dependent protein kinase II (CaMKII). While acute AMPH-induced locomotor responding and NAcc DA release are calcium-independent, sensitized locomotion and DA release are calcium and CaMKII-dependent. In addition, striatal slices from rats exposed to repeated, intermittent injections of AMPH show enhanced calmodulin levels and increased CaMKII activity compared to those obtained from saline-exposed controls. We hypothesize that using viral-mediated gene transfer to modify CaMKII levels in the NAcc will alter behavioral responding to AMPH. The experiments proposed in this grant application will test two main hypotheses. Hypothesis 1: Increasing levels of CaMKII in the NAcc will enhance locomotor responding to a low, threshold dose of AMPH and enhance self-administration of the drug, effects similar to those observed in rats exposed to a sensitizing drug regimen. Rats will be surgically implanted with bilateral guide cannulae and intravenous catheters. Microinjections of the viral vectors will then be made and locomotor and self- administration experiments will be conducted to determine how this manipulation alters sensitivity to AMPH. The efficacy of viral-mediated gene transfer will be determined using immunoblotting. Hypothesis 2: Increasing levels of an inactive mutant form of CaMKII in the NAcc of sensitized rats will diminish enhanced AMPH self-administration observed in these rats. Rats will be exposed to repeated, intermittent injections of AMPH followed by a withdrawal period, during which the animals will be surgically prepared. Following a period of drug self-administration, microinjections of the viral vectors will be made and the effect of this manipulation on drug self-administration subsequently assessed. Using an animal model of addiction is an effective way to study the transition form casual to compulsive drug use. This research will contribute to the ultimate goal of preventing drug addiction by investigating ways to prevent the enhanced drug taking observed in rats that have been previously exposed to AMPH.