NMDA受體的二價陽離子通透性及通道阻斷之分子機制

Abstract

NMDA受體（N-methyl-D-aspartate receptor, NMDAR）是一種麩胺酸受體（Glutamate Receptor），與其他麩胺酸受體最大的不同處在於其高度的鈣離子通透性、以及鎂離子對其賦予的電壓依賴性。也因此，除了突觸訊息的傳遞外，NMDA受體也參與了突觸可塑性、學習記憶整合等高層級神經生理上的調控。NMDAR由四個次單元所構成，每個次單元皆有四個穿膜區段，其中第二穿膜區段為構成孔洞內壁最主要的部份。過去的研究指出，位於第二穿膜區段的尖端上，NR1的N616或是NR2B的N615、N616這幾個結構上較為相近的胺基酸，若將其突變則會對鈣離子的通透性、以及鎂離子的抑制性造成很大的影響。本實驗藉由patch clamp的技術來記錄表現在非洲爪蟾蛙卵上的重組NR1-NR2B電流，測量外側的鎂離子、鈣離子在不同電壓下通透的情形。發現 NR1與NR2B的N site其功能分別對鎂離子和鈣離子的通透性影響是有差異的。在極負電壓下，鎂離子可以穿過NMDAR，NR1 N616突變後會僅會些微增加極負電壓下鎂離子的通透性，而NR2B N615的突變會明顯改變鎂離子與NMDAR之間的親和力。NR1 N616的突變則也會對鈣離子和鈉離子的通透電流造成影響。此外鎂離子和鈣離子都會對NMDAR的門閥性質有影響，特別是作用在NR1 N616上。綜合以上幾點可以發現，NMDAR次單元具有功能性的不對稱性。NR1與NR2B的N site分別與鈣離子和鎂離子的通道阻斷作用較為有關。

N-methyl-D-aspartate receptors(NMDARs) are subtypes of ionotropic glutamate receptors that have distinct characteristics including high permeability to calcium ions and voltage-dependant block by magnesium ion. Thus, NMDARs play an important role in neural transmission, neuronal plasticity and long-term potentiation. NMDAR is composed of four subunits, and each subunits have four transmembrane domains(M1-4). The re-entrant loop (M2) forms part of the channel pore. The amino acid on the tip of M2 loop, like NR1-N616 and NR2B-N615, play an important role in Mg2+ block and divalent cation permeability. In this study, we examined the mechanism of extracellular calcium permeability and Mg2+ block in recombinant NR1-NR2B expressed on Xenopus oocytes. The NMDAR currents were measured with excised outside-out macropatchs. We found that Mg2+ could permeate through NMDAR in extreme low voltage. The mutant NR2B containing N615Q and N615C may increase external Mg2+ permeability and decrease binding affinity ,which indicate that the NR2B N615 may be the energy barrier for Mg2+ permeation in the pore. NR1 N616Q increase the permeability of Mg2+ in extreme low voltage, and also has a strong influence on calcium permeability. We conclude that the subunits of NMDAR do not have a completely symmetric alignment in tense of permeation of divalent ions.