Adolescent ethanol exposure and NMDA receptor maturation in cerebellum

描述（由申请人提供）：乙醇是青少年中最常见的滥用药物，但我们对其在青少年大脑中的作用知之甚少。我们试图了解乙醇对哺乳动物小脑皮层活动依赖性成熟和功能的影响，这主要是由兴奋性神经递质谷氨酸介导的。这很重要，因为小脑在运动和认知功能中起着至关重要的作用，并且在青少年时期继续经历实质性的增长。特别是，n -甲基- d -天冬氨酸受体（NR）的组成和功能会发生变化，而乙醇是包括小脑在内的许多脑区和神经元类型的NR拮抗剂。因此，预计乙醇会影响NRs本身以及位于NRs下游的信号级联，特别是细胞质和细胞核中的Ca2+和钙调素依赖性激酶。我们将采用大鼠模型，结合分子生物学、免疫化学和解剖学技术，重点研究内颗粒层中的NRs和CaMKs。我们假设，从青春期前开始长期使用乙醇将不可逆转地干扰谷氨酸能苔藓纤维颗粒神经元突触回路和信号分子的发育和功能。这将通过NR2A-C亚基mrna和免疫反应蛋白的特异性和持续性改变、NR1的外显子选择以及下游Ca2+依赖性激酶CaMKK和CaMKIV的成熟来反映。我们不预测大量的细胞损失对乙醇的反应，因为死亡通常是观察到当乙醇给予新生儿在出生后4 -6天。相反，乙醇可能会干扰突触的成熟，从而干扰青少年小脑的信息处理。我们之所以选择RO3支持机制，是因为这个试点项目将为未来研究乙醇对小脑生理和病理生理的功能影响奠定基础。这些未来的研究将与我们在上州医学大学和宾厄姆顿大学的合作者一起设计，他们是胎儿和青少年酒精研究领域的知名专家。在这个国家，青少年消费了至少四分之一的酒精，然而我们对酒精对青少年大脑的影响知之甚少。小脑皮层在青春期继续经历显著的生长和突触的完善，是乙醇最敏感的目标之一。我们将用大鼠模型验证乙醇不可逆地干扰兴奋性神经递质分子及其下游效应物成熟的假设。我们的结果将为今后的功能研究提供基础。

DESCRIPTION (provided by applicant): Ethanol is the most commonly abused drug in adolescents yet we know very little about its actions in adolescent brain. We seek to understand ethanol effects on the activity-dependent maturation and function of the mammalian cerebellar cortex, which is mediated primarily by the excitatory neurotransmitter glutamate. This is important because the cerebellum has critical roles in motor and cognitive functions, and continues to undergo substantial growth in the adolescent. In particular, N-methyl-D-aspartate receptor (NR) composition and function changes, and ethanol is an antagonist of NRs in many brain regions and neuronal types, including cerebellum. As such, ethanol is predicted to influence NRs themselves as well as signaling cascades that lie downstream of NRs, in particular Ca2+ and calmodulin-dependent kinases in the cytosol and nucleus. We will use a rat model and a combination of molecular biological, immunochemical and anatomical techniques, focusing on NRs and CaMKs in the internal granular layer. We hypothesize that chronic use of ethanol beginning in pre-adolescence will irreversibly interfere with the development and function of the glutamatergic mossy fiber-granule neuron synaptic circuitry and signaling molecules. This will be reflected by specific and persistent alterations in NR2A-C subunit mRNAs and immunoreactive proteins, exon selection in NR1, and maturation of the downstream Ca2+-dependent kinases, CaMKK and CaMKIV. We do not predict substantial cell loss in response to ethanol since death is typically observed when ethanol is administered to neonates at postnatal days 4 -6. Rather, ethanol is expected to interfere with synaptic maturation and therefore, information processing in the adolescent cerebellum. We chose the RO3 mechanism of support because this pilot project will serve as a foundation for future studies exploring the functional consequences of ethanol on cerebellar physiology and pathophysiology. These future studies will be designed with substantial input from our collaborators at Upstate Medical and Binghamton Universities who are established experts in the fields of fetal and adolescent alcohol research. Adolescents consume at least one-fourth of alcohol in this country, yet we understand little about ethanol effects in adolescent brain. The cerebellar cortex, which continues to undergo significant growth and synaptic refinement through adolescence is one of the most sensitive targets of ethanol. We will test the hypothesis that ethanol irreversibly interferes with maturation of excitatory neurotransmitter molecules and their downstream effectors using the rat model. Our results will serve as a basis for future functional studies.