Effect of membrane fluidity on brain receptors and transporters

该神经药理学研究项目的目的是研究膜成分/流动性对大脑神经递质功能的影响。假设是膜成分和/或流动性的变化将对某些受体和转运蛋白以及某些大脑区域产生特异性影响。我们建议在体外研究牛脑膜中硬化膜剂（胆固醇半琥珀酸酯和硬脂酸）与流化膜剂（亚油酸、L-α-磷脂酰胆碱和 DHA、omega-3（一种广泛使用的营养补充剂））的剂量反应。我们将研究与多巴胺转运蛋白 DAT 相比，流动性变化如何影响多巴胺 (D2)、腺苷 (A2A)、GABAA、血清素（5-HT1A、5-HT2A）谷氨酸（NMDA、mGluR5）受体。将研究额叶皮层、海马体和纹状体，这些脑膜代表从流动性更大到刚性更大的脑膜。流动性将用荧光探针测量。将通过蛋白质印迹法测量总膜受体和转运蛋白及其与特定放射性配体的结合的表面水平。功能变化将通过与第二信使和关键信号蛋白的偶联来测量。在大鼠体内，将用老年大鼠研究呈现更坚硬的膜的病症，而将通过用慢性他汀类药物治疗、DHA 治疗或缩醛磷脂（以增加 DHA 水平）治疗降低胆固醇来研究更多的流体膜。对于体内实验，我们还将通过原位杂交测量这些受体的 mRNA，以评估它们的合成。我们预计微小的膜变化将显着影响所研究的膜受体，并且这具有重要的生理后果。可以施用影响膜的化合物（或设计饮食）来增强针对这些受体和转运蛋白的药物治疗的功效，从而成功衰老。

The objective of this research program in neuropharmacology is to investigate the effect of membrane composition/fluidity on neurotransmitter function in the brain. The hypothesis is that changes of membrane composition and/or fluidity will have effects specific for certain receptors and transporters and specific for certain brain regions. We propose to investigate in vitro a dose-response of agents that rigidify membranes (cholesteryl hemisuccinate and stearic acid) compared to agents that fluidify membranes (linoleic acid, L-alpha-phosphatidylcholine and DHA, omega-3, a widely used nutritional supplement) in bovine brain membranes. We will investigate how fluidity changes affect dopamine (D2), adenosine (A2A), GABAA, serotonin (5-HT1A, 5-HT2A) glutamate (NMDA, mGluR5) receptors compared to the dopamine transporter DAT. The frontal cortex, the hippocampus and the striatum representing a range from more fluid to more rigid brain membranes will be studied. Fluidity will be measured with fluorescent probes. Total membrane receptors and transporters will be measured by Western blot and their surface levels with binding of specific radioligands. The functional changes will be measured with coupling to second messengers and key signaling proteins. In vivo in rats a condition presenting a more rigid membrane will be investigated with old rat whereas more fluid membranes will be investigated by reducing cholesterol with a chronic statin drug treatment, DHA treatment or plasmalogen (to increase DHA levels) treatment. For in vivo experiments we will also measure by in situ hydridization mRNA of these receptors to assess their synthesis. We expect that small membrane changes will significantly affect the membrane receptor investigated and that this has important physiological consequences. Compounds affecting membranes could be administered (or a diet designed) to enhance efficacy of drug treatments targeting these receptors and transporters for successful aging.