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

描述(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节多巴胺释放中断所致。流行病学数据表明，帕金森氏症在吸烟者中较少流行。此外，动物研究发现，烟草烟雾中的成瘾成分尼古丁可以保护DA能神经元免受化学伤害，这种作用可能是由神经元烟碱型乙酰胆碱受体(NAChRs)介导的。NAChRs如何调节DA神经传递，涉及哪些nAChR亚型？为了解决这些问题，设计了一种新的小鼠品系，在尼古丁受体α-4亚单位的假定孔区内表达单点突变Leu9‘Ala。这种突变使含有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-4β-2*nAChRs可能在功能上直接与D2样受体相互作用；G蛋白偶联受体在中脑和纹状体神经元中表达，正常情况下负向调节活动。初步数据表明，Leu9‘Ala小鼠体内D2样受体的激活会引发野生动物不会出现的帕金森症状。特定目的1验证了Leu9‘Ala帕金森病表型是由D2样多巴胺受体激活引起的假设，并依赖于α-4*nAChR调节。这将通过给小鼠注射不同的多巴胺和尼古丁受体拮抗剂并分析帕金森氏症的表型严重程度来完成。特定目的2验证了这种相互作用发生在黑质和/或层中的假设。在具体目标3中，将利用体内微透析来测定多巴胺和乙酰胆碱的释放。最后，特定目标4测试了这一假设，即Leu9‘Ala小鼠的D2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*nAChRs之间的功能相互作用。这将通过测量D2激活前后神经元活性和尼古丁反应的变化来实现。预计这些实验结果不仅将提供一种新的药理学的、可逆的帕金森病小鼠模型，而且还将增加对烟碱受体介导的DAR能神经传递调节的理解。公共卫生相关性预计这项研究的结果将提供一种新的药理学的、可逆的帕金森氏病小鼠模型。此外，这个小鼠模型应该有助于阐明在正常和疾病状态下对自主运动至关重要的潜在神经机制。

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.