Molecular mechanism of pregnenolone sulfate action at NR1/NR2B receptors
Molecular mechanism of pregnenolone sulfate action at NR1/NR2B receptors
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DOI：
10.1523/jneurosci.2099-04.2004
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发表时间：
2004-11-17
影响因子：
5.3
通讯作者：
Vyklicky, L
中科院分区：
文献类型：
--
作者：
Horak, M;Vlcek, K;Vyklicky, L
NMDA受体在CNS中高度表达，并且参与兴奋性突触传递和突触可塑性以及兴奋性毒性。它们有几个别构调节剂的结合位点，包括神经类固醇，由神经组织合成的内源性化合物，并预期在局部起作用。从表达NR 1 -1a/NR 2B受体的人胚肾293细胞的全细胞膜片钳记录显示，神经甾体硫酸双烯醇酮（PS）（300 μ M），当应用于静息NMDA受体时，增强随后对1 mM谷氨酸的响应幅度五倍，并减缓其失活两倍。相同浓度的PS，当施加在NMDA受体激活1 mM谷氨酸，只有一个小的影响。PS与静息NMDA受体结合和不结合的结合和解离速率常数估计分别为3.3+/-2.0 mM（-1）sec（-1）和0.12+/-0.02 sec（-1），对应于37 μ M的表观亲和力Kd。实验结果表明，PS增强NMDA受体反应的分子机制是由于峰通道开放概率（P-o）的增加。在PS的连续存在下记录谷氨酸的反应表现出显着的时间依赖性下降。我们的研究结果表明，这种下降是由受体激活后NMDA受体对PS亲和力的变化引起的，这些结果表明PS是NMDA受体P-o的调节剂，其有效性被谷氨酸结合所降低。这种调节可能对神经元的兴奋性有重要影响。
NMDA receptors are highly expressed in the CNS and are involved in excitatory synaptic transmission and synaptic plasticity as well as excitotoxicity. They have several binding sites for allosteric modulators, including neurosteroids, endogenous compounds synthesized by the nervous tissue and expected to act locally. Whole-cell patch-clamp recording from human embryonic kidney 293 cells expressing NR1-1a/NR2B receptors revealed that neurosteroid pregnenolone sulfate (PS) (300 muM), when applied to resting NMDA receptors, potentiates the amplitude of subsequent responses to 1 mM glutamate fivefold and slows their deactivation twofold. The same concentration of PS, when applied during NMDA receptor activation by 1 mM glutamate, has only a small effect. The association and dissociation rate constants of PS binding and unbinding from resting NMDA receptors are estimated to be 3.3+/-2.0 mM(-1) sec(-1) and 0.12+/-0.02 sec(-1), respectively, corresponding to an apparent affinity K-d of 37 muM. The results of experiments indicate that the molecular mechanism of PS potentiation of NMDA receptor responses is attributable to an increase in the peak channel open probability (P-o). Responses to glutamate recorded in the continuous presence of PS exhibit marked time-dependent decline. Our results indicate that the decline is induced by a change of the NMDA receptor affinity for PS after receptor activation.These results suggest that the PS is a modulator of NMDA receptor P-o, the effectiveness of which is lowered by glutamate binding. This modulation may have important consequences for the neuronal excitability.