Effect of membrane fluidity on brain receptors and transporters

这项神经药理学研究计划的目的是探讨膜成分/流动性对脑内神经递质功能的影响。假设膜成分和/或流动性的变化将对某些受体和转运体产生特定的影响，并对特定的大脑区域产生特定的影响。我们建议在体外研究牛脑膜硬化剂(琥珀酸甘油酯和硬脂酸)与流动性药剂(亚油酸、L-α-磷脂酰胆碱和DHA，一种广泛使用的营养补充剂)之间的剂量-反应关系。我们将与多巴胺转运体DAT相比，研究流动性变化如何影响多巴胺(D2)、腺苷(A2A)、GABAA、5-羟色胺(5-HT1A，5-HT2A)谷氨酸(NMDA，mGluR5)受体。将对额叶皮质、海马体和纹状体进行研究，它们代表了从更多的液体到更坚硬的脑膜。流动性将用荧光探针测量。总的膜受体和转运体将通过蛋白质印迹和结合特定放射性配基的表面水平来测量。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，将在老年大鼠身上研究出现更坚硬的细胞膜的情况，而通过慢性他汀类药物治疗、DHA治疗或纤溶酶原(以增加DHA水平)降低胆固醇来研究更多的液膜。对于体内实验，我们还将通过原位氢化来测量这些受体的mRNA，以评估它们的合成。我们预计，微小的膜变化将显著影响所研究的膜受体，这将产生重要的生理后果。可以使用影响膜的化合物(或设计的饮食)来提高针对这些受体和转运体的药物治疗的有效性，从而成功地老化。

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.