The Role of Cav1.2 L-type Ca2+ Channels in Cocaine-Induced Reinstatement

Cav1.2 L 型 Ca2 通道在可卡因诱导恢复中的作用

• 批准号: 8657167
• 负责人: Anjali M RAJADHYAKSHA
• 金额: $ 0.83万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-06-01 至 2017-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8657167
• 关键词: AMPA Receptors; Addictive Behavior; Behavior; Behavioral; C57BL/6 Mouse; Cell Fractionation; Cell surface; Cocaine; Cocaine Dependence; Data; Development; Diltiazem; Dopamine; Electron Microscopy; Extinction (Psychology); Face; Genetic; Illicit Drugs; Immunoblot Analysis; Immunoblotting; Investigation; Knock-out; Knowledge; Label; Laboratories; Learning; Measures; Mediating; Mediator of activation protein; Memory; Modeling; Molecular; Molecular Target; Mus; Mutant Strains Mice; Neurons; Nucleus Accumbens; Pathway interactions; Peptides; Pharmaceutical Preparations; Pharmacological Treatment; Pharmacology; Phosphorylation; Phosphorylation Site; Phosphotransferases; Property; Relapse; Research; Rodent; Rodent Model; Role; Surface; Synapses; Techniques; Technology; Testing; Therapeutic Intervention; Viral; Viral Vector; Withdrawal; Work; addiction; base; calmodulin-dependent protein kinase II; cocaine use; drug craving; drug relapse; genetic technology; inhibitor/antagonist; kinase inhibitor; mouse model; neurotoxic; novel; pre-clinical; preference; prevent; recombinase; response; therapeutic target; trafficking; transmission process; treatment strategy

DESCRIPTION (provided by applicant): Relapse to cocaine use, the highest among commonly abused illicit drugs, is a serious public problem and represents the primary challenge that exists for the treatment of cocaine addicts. Despite extensive investigation, molecular substrates that can serve as potential therapeutic targets to prevent relapse are limited. Thus, understanding the mechanisms of relapse and identifying new molecular targets for developing pharmacological treatments will greatly aid the field of addiction research. Recent preclinical rodent studies have suggested that craving and drug-induced relapse is mediated by enhanced synaptic AMPAR transmission in the nucleus accumbens (NAc) via GluA2-lacking-, GluA1-containinng-Ca2+-permeable AMPA receptors (CP-AMPARs). Work from our laboratory has identified the Cav1.2 L-type Ca2+ channel (LTCC) as a promising candidate for mediating cocaine-induced long-term behavioral responses and in regulating cell surface AMPARs. Using psychomotor sensitization, we find that Cav1.2 channels in the NAc, mediates cocaine-induced expression of sensitization following extended periods of withdrawal. Using the reinstatement of cocaine CPP model, we find that the LTCC antagonist, diltiazem delivered directly into the NAc blocks cocaine-induced reinstatement of cocaine CPP. Furthermore, we have found that Cav1.2-activated kinase pathways (CaMKII and ERK) regulate cocaine-induced increase in cell surface GluA1, but not GluA2 in the NAc. Thus, in this RO1 application we aim to capitalize on the knowledge we have gained to further explore molecular targets that mediate relapse to cocaine. We will test the central hypothesis that Cav1.2- activated kinase pathways in the NAc mediate cocaine-induced reinstatement of cocaine seeking via increase in NAc synaptic CP-AMPARs. To circumvent the challenge of the lack of Cav1.2-specific blockers, we propose to use the cutting edge Cre-lox P technology to generate local Cav1.2 knockout in the mouse NAc. Kinases will be manipulated by the use of viral vectors. In Aim 1.1, adenoassociated viral (AAV) vectors expressing Cre recombinase will be stereotaxically delivered into the NAc of Cav1.2 floxed mice. Mice will be behaviorally tested in cocaine-induced reinstatement of cocaine CPP. In Aim 1.2, molecular studies will be pursued to examine Cav1.2-induced AMPAR trafficking. In Aim 1.3, electron microscopy will be utilized to examine GluA1 trafficking in dopamine D1 neurons in the NAc shell. In Aim 2.1, viral vectors expressing kinase inhibitors will be stereotaxically delivered into the NAc of C57BL/6 mice. Mice will be behaviorally tested in cocaine-induced reinstatement of cocaine CPP. In Aim 2.2, role of kinases in AMPAR trafficking will be examined. In Aim 3, the functional significance of NAc CP-AMPARs, GluA1 trafficking and GluA1 phosphorylation in cocaine-induced reinstatement will be examined using pharmacology and genetic mutant mice. The results obtained from this study could greatly advance the field of cocaine addiction by identifying discrete molecular targets for developing pharmacological treatments for cocaine addicts.

描述（由申请人提供）：对可卡因使用的复发是普遍滥用的非法药物中最高的，是一个严重的公共问题，代表了治疗可卡因成瘾者的主要挑战。尽管进行了广泛的研究，但可以用作防止复发的潜在治疗靶标的分子底物受到限制。因此，了解复发的机制并确定开发药理治疗的新分子靶标将极大地帮助成瘾研究领域。最近的临床前啮齿动物研究表明，渴望和药物诱导的复发是通过通过GluA2-lacking-，Glua1-Containinng-Ca2+可渗透性AMPA受体（CP-pampars）通过GluA2-containinng-ca2+glua1-containinng-ca2+增强的突触AMPAR传播（NAC）介导的。我们实验室的工作确定CAV1.2 L型Ca2+通道（LTCC）是介导可卡因诱导的长期行为反应和调节细胞表面AMPAR的有前途的候选人。使用心理运动敏化，我们发现NAC中的CAV1.2通道介导可卡因诱导的延长戒断后敏化的表达。使用可卡因CPP模型的恢复原状，我们发现LTCC拮抗剂Diltiazem直接传递到NAC中可卡因诱导的可卡因CPP的恢复。此外，我们发现CAV1.2激活的激酶途径（CAMKII和ERK）调节可卡因诱导的细胞表面GLUA1的增加，而不是NAC中的GLUA2。因此，在此RO1应用中，我们旨在利用我们获得的知识，以进一步探索介导可卡因复发的分子靶标。我们将测试中心假设，即NAC中Cav1.2激活的激酶途径通过NAC突触CP-Ampars增加可卡因诱导的可卡因寻求可卡因。为了避免缺乏CAV1.2特异性阻滞剂的挑战，我们建议使用尖端CRE-LOX P技术在小鼠NAC中生成局部CAV1.2敲除。激酶将通过使用病毒载体来操纵。在AIM 1.1中，表达CRE重组酶的腺体相关病毒（AAV）载体将立体定义地递送到Cav1.2 Floxed小鼠的NAC中。小鼠将在可卡因CPP的可卡因诱导的恢复中进行行为测试。在AIM 1.2中，将进行分子研究以检查CAV1.2诱导的AMPAR运输。在AIM 1.3中，将利用电子显微镜检查NAC壳中多巴胺D1神经元中的GLUA1运输。在AIM 2.1中，表达激酶抑制剂的病毒载体将立体定位递送到C57BL/6小鼠的NAC中。小鼠将在可卡因CPP的可卡因诱导的恢复中进行行为测试。在AIM 2.2中，将检查激酶在AMPAR贩运中的作用。在AIM 3中，将使用药理学和遗传突变小鼠检查可卡因诱导的恢复中NAC CP-AMPAR，GLUA1运输和GLUA1磷酸化的功能意义。从这项研究中获得的结果可以通过确定开发可卡因成瘾药物治疗的离散分子靶标，可以大大推动可卡因成瘾领域。