水杨酸钠的螺旋神经节神经元损害中DR介导NMDAR与GABAaR相互作用的研究

Interaction between NMDAR and GABAaR involves multiple neural physiological and pathological process. Our previous studies showed that sodium salicylate(SS) induced spiral ganglion neurons’(SGNs) excitoxicity was mediated by interaction between NMDAR and GABAaR. However, mechanisms of interaction between the two receptors remain unclear. It has been proved that DR has reciprocal effects on the interaction between NMDAR and GABAaR respectively in neurons. Our recent studies also showed that DRs’ activity affects the interaction between NMDAR and GABAaR induced by SS in SGNs. Therefore, we hypothesize that the SS-induced ototoxicity associated with interaction between NMDAR and GABAaR is intermediated by DR in SGNs. In order to verify this hypothesis, we proposed to give observations both in the physiological and in SS conditions by using the patch clamp recording, immunofluorescence with laser confocal microscopy, real-time fluorescent PCR and gene recombination technique. Molecular mechanisms of DR mediating the two receptors, such as receptors’ phosphorylation, receptors’ trafficking, [Ca2+]i changes, will also be considered at the same time, to provide a deep research on receptors’ function ,cell membrane state and molecular level，which will lead to a new train of thought for underlying mechanisms of ototoxicity of drug-induced hearing impairment.

NMDAR与GABAaR相互作用涉及多种神经病理生理过程，我们的研究证实，水杨酸钠(SS)介导了NMDAR与GABAaR的相互作用是SS致螺旋神经节神经元(SGN)损害的途径之一，然而这两受体相互作用的机制尚不完全清楚。多巴胺受体(DR)能与神经元中的NMDAR和GABAaR有相互作用已被证实，我们的研究也显示，DR活动可影响SS对SGN中NMDAR与GABAaR的相互作用。据此推测：DR介导NMDAR与GABAaR之间的相互作用并参与了SS导致SGN损害的过程。为验证这一假说，我们拟用膜片钳记录、免疫荧光激光共聚焦显微镜、实时荧光PCR、基因重组等技术，观察SGN在生理状态下、及水杨酸钠作用下，受体磷酸化状态、受体转运、[Ca2+]i变化时，DR对NMDAR与GABAaR相互作用在受体功能、受体在胞膜和分子水平的影响，以及DR对这两个受体的作用位点。为研究耳聋的深层机制提供新的思路。

NMDAR与GABAaR相互作用涉及多种神经病理生理过程，我们的研究证实，水杨酸钠(SS)介导NMDAR与GABAaR的相互作用是SS致螺旋神经节神经元(SGN)损害的途径之一，然而这两受体相互作用的机制尚不完全清楚。多巴胺受体(DR)能与神经元中的NMDAR和GABAaR有相互作用已被证实，我们的研究也显示，DR活动可影响SS对SGN中NMDAR与GABAaR的相互作用。据此推测：DR介导NMDAR与GABAaR之间的相互作用并参与了SS导致SGN损害的过程。为验证这一假说，我们采用膜片钳记录、免疫荧光、实时荧光PCR、Western Blot等技术，发现一般情况下SGN的NMDAR可轻度抑制GABAaR电流，但差异无统计学意义。当SS作用时，越高浓度的SS作用下使NMDAR对GABAaR电流的抑制作用越强。SS或NMDA对GABAAaR电流有抑制用,Ca2+/CaMKII介导了NMDAR对GABAaR 的抑制作用。SS、DR1、DR2介导NMDAR与GABAaR的受体内吞，并抑制GABAaR应答，这是通过DR1、DR2-PKC途径调节，作用位点包括GABRβ3-S383/S408/409，GABRɣ2-S327及NR2B-Y1472。