Nicotinic-dopamine receptor interaction in a novel Parkinsonian mouse model.

描述（由申请人提供）：帕金森病是由于黑质多巴胺能神经元的进行性死亡导致基底神经节多巴胺释放中断而引起的。流行病学数据表明，帕金森病在吸烟者中发病率较低。此外，动物研究发现，尼古丁，烟草烟雾的成瘾成分，保护能神经元免受化学损伤，这种作用可能是由神经元尼古丁乙酰胆碱受体（nAChRs）介导的。nAChR如何调节DA神经传递？涉及哪些nAChR亚型？为了解决这些问题，我们设计了一种新的小鼠系，在烟碱受体α -4亚基的假定孔区表达单点突变Leu9'Ala。这种突变使得含有α -4（“α -4*”）的nAChRs对激动剂的敏感性提高了50倍，从而允许分离和扩增涉及α -4* nAChRs的行为和生理表型。最近的研究表明，α -4 β - 2* nachr可能与d2样受体直接作用；g蛋白偶联受体在中脑和纹状体神经元中表达，通常负调控活性。初步数据表明，在Leu9'Ala小鼠中d2样受体的激活会引起帕金森病症状，而这种症状在野生型动物中不会发生。特异性目的1验证了一种假设，即Leu9'Ala帕金森表型是由d2样多巴胺受体的激活引起的，并依赖于α -4* nAChR调节。这将通过给小鼠施用不同的多巴胺和烟碱受体拮抗剂并分析帕金森表型的严重程度来完成。具体目标2检验相互作用发生在黑质和/或地层中的假设。在特定目标3中，体内微透析将用于测定多巴胺和乙酰胆碱的释放。最后，特异性目的4验证了D2在Leu9'Ala小鼠中的激活，揭示了Gi/o偶联受体与中脑和/或纹状体神经元中的α -4* nachr之间的功能相互作用。这将通过测量D2激活前后神经元活动和尼古丁反应的变化来实现。本实验结果不仅将为帕金森病小鼠提供一种新的药理学、可逆性模型，而且还将增加对烟碱受体介导的神经递质调节的认识。预计本研究结果将为帕金森病提供一种新的药理学、可逆性小鼠模型。此外，该小鼠模型应该有助于阐明在正常和患病状态下自主运动的重要潜在神经元机制。

DESCRIPTION (provided by applicant): Parkinson's disease is caused by the disruption of dopamine release in basal ganglia due to the progressive death of dopaminergic neurons in substantia nigra. Epidemiological data indicate Parkinson's disease is less prevalent in smokers. In addition, animal studies have found that nicotine, the addictive component of tobacco smoke, protects DAergic neurons from chemical insult and that this effect is likely mediated by neuronal nicotinic acetylcholine receptors (nAChRs). How do nAChRs modulate DA neurotransmission and which nAChR subtypes are involved? To address these questions a novel mouse line was engineered expressing a single point mutation, Leu9'Ala, within the putative pore region of the nicotinic receptor alpha-4 subunit. This mutation renders alpha-4-containing ("alpha- 4*") nAChRs 50-fold more sensitive to agonist allowing for the isolation and amplification of behavioral and physiological phenotypes that involve alpha-4* nAChRs. Recent studies suggest that alpha-4 beta- 2* nAChRs may functionally interact directly with D2-like receptors; G-protein coupled receptors that are expressed in midbrain and striatal neurons and, normally, negatively regulate activity. Preliminary data indicate that activation of D2-like receptors in Leu9'Ala mice elicits Parkinsonian symptoms that do not occur in wild-type animals. Specific aim 1 tests the hypothesis that the Leu9'Ala Parkinsonian phenotype is caused by activation of a D2-like dopamine receptor and is dependent on alpha-4* nAChR modulation. This will be done by administering different dopamine and nicotinic receptor antagonists to mice and assaying the Parkinsonian phenotype severity. Specific aim 2 tests the hypothesis that the interaction takes place in substantia nigra and/or stratum. In specific aim 3, in vivo microdialysis will be utilized to assay dopamine and acetylcholine release. Finally, specific aim 4 tests the hypothesis that D2 activation in Leu9'Ala mice uncovers a functional interaction between Gi/o coupled receptors and alpha-4* nAChRs in midbrain and/or striatal neurons. This will be achieved by measuring changes in neuron activity and nicotinic responses before and after D2 activation. It is anticipated that the results from the proposed experiments will not only provide a new pharmacological, reversible, Parkinson's disease mouse model, but, also increase understanding of nicotinic receptor mediated modulation of DAergic neurotransmission. PUBLIC HEALTH RELEVANCE It is anticipated that the results from this study will provide a new pharmacological, reversible mouse model of Parkinson's disease. In addition, this mouse model should help elucidate underlying neuronal mechanisms important for voluntary movement in normal and diseased states.