Intracellular mechanisms of cocaine-memory reconsolidation

描述(申请人提供)：环境导致的药物复发是可卡因使用者康复的主要障碍。与可卡因相关的环境通过必须在长期记忆中保持的上下文-反应-可卡因联系来维持对可卡因诱导行为的刺激控制。因此，有选择地削弱或消除这种可卡因记忆的治疗可能会降低曾经吸毒者的复发倾向。值得注意的是，可卡因相关的长期记忆在提取后变得不稳定(即记忆重新激活)，这些记忆需要重新稳定或重新整合到长期存储中才能维持。此外，有证据表明，杏仁基底外侧核(BLA)对于促进药物背景诱导的可卡因寻求的可卡因记忆的重新巩固是必需的(非强化的主动杠杆按压，它提供了可卡因的动机指数；Fuchs等人，2009年)。拟议的NRSA研究计划将通过探索BLA中可卡因记忆重新巩固的细胞内机制来扩展这一研究路线，重点放在两个信号分子上：细胞外信号相关激酶(ERK)和细胞周期蛋白依赖性蛋白激酶5(CDK5)。具体目标1将检验这一假说，即ERK介导的BLA信号有助于可卡因记忆的重新巩固，而可卡因记忆对于控制工具性可卡因寻找的上下文控制至关重要，并将探索这一现象的可能下游机制。初步数据显示，BLA U0126(MEK/ERK抑制剂)治疗以记忆重新激活依赖的方式损害可卡因记忆再巩固，并扰乱随后药物上下文诱导的可卡因寻找(Exp.(1A，正在进行中)。这些数据为Exp.1b，它将利用定量Western blotting来表征ELK-1、CREB和Zif268的激活变化，这些转录因子可能在记忆再巩固过程中被ERK在BLA中激活(目标1b)。作为后续，将使用反义寡核苷酸或小干扰RNA操作来评估ELK-1、CREB或Zif268的激活是否是可卡因记忆重新巩固和随后对可卡因寻找的上下文控制所必需的。具体目标2将检验这一假设，即BLA中的CDK5干扰可卡因记忆的重新巩固，而可卡因记忆对于控制器质性可卡因寻找的上下文控制至关重要。基于CDK5在体外负向调节ERK的证据，Exp.2将评估白血球内注射丁内酯(CDK5抑制剂)是否会以记忆重新激活的方式加强可卡因记忆的再巩固，并增加随后的药物背景诱导的可卡因寻找。综上所述，将结合复杂的行为程序和尖端的分子技术来探索可卡因记忆重新巩固的细胞内机制，这是一种维持不适应成瘾记忆的认知过程，从而刺激环境诱导的药物渴望和复发。这项研究努力有可能为成瘾治疗的发展提供新的见解。

DESCRIPTION (provided by applicant): Environmentally-induced drug relapse is a major obstacle in the rehabilitation of cocaine users. Cocaine- associated environments maintain stimulus control over cocaine-induced behavior via context-response- cocaine associations that must be maintained in long-term memory. Thus, treatments that selectively weaken or eliminate such cocaine memories may reduce the propensity to relapse in former drug addicts. Remarkably, cocaine-related long-term memories become labile following retrieval (i.e., memory reactivation), and these memories need to be re-stabilized, or reconsolidated, into long-term storage in order to be maintained. Furthermore, evidence suggests that the basolateral amygdala (BLA) is required for the reconsolidation of cocaine memories that promote drug context-induced cocaine seeking (non-reinforced active lever presses, which provided an index of motivation for cocaine; Fuchs et al., 2009). The proposed NRSA research plan will extend this line of inquiry by exploring the intracellular mechanisms of cocaine-memory reconsolidation in the BLA with a focus on two signaling molecules: extracellular signal-related kinase (ERK) and cyclin-dependent protein kinase 5 (Cdk5). Specific aim 1 will be to test the hypothesis that ERK-mediated signaling in the BLA facilitates the reconsolidation of cocaine memories that are critical for contextual control over instrumental cocaine seeking and will explore possible downstream mechanisms of this phenomenon. Preliminary data show that intra-BLA U0126 (MEK/ERK inhibitor) treatment impairs cocaine-memory reconsolidation in a memory reactivation-dependent manner and disrupts subsequent drug context-induced cocaine seeking (Exp. 1a, in progress). These data provide sound rationale for Exp. 1b, which will utilize quantitative Western blotting to characterize changes in the activation of Elk-1, CREB, and zif268, the transcription factors that are likely activated by ERK in the BLA during memory reconsolidation (Aim 1b). As follow-up, antisense oligonucleotide or small interfering RNA manipulation will be used to assess whether the activation of Elk-1, CREB, or zif268 is required for cocaine-memory reconsolidation and subsequent contextual control over cocaine seeking. Specific aim 2 will be to test the hypothesis that Cdk5 in the BLA interferes with the reconsolidation of cocaine memories that are critical for contextual control over instrumental cocaine seeking. Based on evidence that Cdk5 negatively regulates ERK in vitro, Exp. 2 will assess whether intra-BLA ¿-butyrolactone (Cdk5 inhibitor) treatment will enhance cocaine-memory reconsolidation in a memory reactivation-dependent manner and increase subsequent drug context-induced cocaine seeking. In summary, a combination of sophisticated behavioral procedures and cutting edge molecular techniques will be used to explore the intracellular mechanisms of cocaine-memory reconsolidation, a cognitive process that maintains maladaptive addictive memories and thus stimulates environmentally-induced drug craving and relapse. This research endeavor has the potential to provide novel insights for addiction treatment development.