初级传入神经中的 NMDA 受体需要 Src 家族激酶的磷酸化作用才能在大鼠脊髓中诱导 P 物质释放。
NMDA receptors in primary afferents require phosphorylation by Src family kinases to induce substance P release in the rat spinal cord.
机构信息
Center for Neurobiology of Stress, CURE: Digestive Diseases Research Center, Division of Digestive Diseases, Department of Medicine, David Geffen School of Medicine at the University of California, Los Angeles, Los Angeles, CA, USA.
出版信息
Neuroscience. 2010 Mar 31;166(3):924-34. doi: 10.1016/j.neuroscience.2010.01.009. Epub 2010 Jan 13.
The function of N-methyl-d-aspartate (NMDA) receptors in primary afferents remains controversial, in particular regarding their ability to evoke substance P release in the spinal cord. The objective of this study was, first, to confirm that substance P release evoked by NMDA is mediated by NMDA receptors in primary afferent terminals. Second, we investigated whether these NMDA receptors are inactivated in some conditions, which would explain why their effect on substance P release was not observed in some studies. Substance P release was induced in spinal cord slices and measured as neurokinin 1 (NK1) receptor internalization in lamina I neurons. NMDA (combined with d-serine) induced NK1 receptor internalization with a half of the effective concentration (EC50) of 258 nM. NMDA-induced NK1 receptor internalization was abolished by the NK1 receptor antagonist L-703,606, confirming that is was caused by substance P release, by NMDA receptor antagonists (MK1801 and ifenprodil), showing that it was mediated by NMDA receptors containing the NR2B subunit, and by preincubating the slices with capsaicin, showing that the substance P release was from primary afferents. However, it was not affected by lidocaine and omega-conotoxin MVIIA, which block Na+ channels and voltage-dependent Ca2+ channels, respectively. Therefore, NMDA-induced substance P release does not require firing of primary afferents or the opening of Ca2+ channels, which is consistent with the idea that NMDA receptors induce substance P directly by letting Ca2+ into primary afferent terminals. Importantly, NMDA-induced substance P release was eliminated by preincubating the slices for 1 h with the Src family kinase inhibitors PP1 and dasatinib, and was substantially increased by the protein tyrosine phosphatase inhibitor BVT948. In contrast, PP1 did not affect NK1 receptor internalization induced by capsaicin. These results show that tyrosine-phosphorylation of these NMDA receptors is regulated by the opposite actions of Src family kinases and protein tyrosine phosphatases, and is required to induce substance P release.
N-甲基-D-天冬氨酸（NMDA）受体在初级传入神经中的功能仍然存在争议，特别是关于它们在脊髓中引发 P 物质释放的能力。本研究的目的是，首先，证实 NMDA 引发的 P 物质释放是由初级传入末梢的 NMDA 受体介导的。其次，我们研究了这些 NMDA 受体在某些条件下是否失活，这可以解释为什么它们对 P 物质释放的影响在某些研究中没有观察到。P 物质释放是在脊髓切片中诱导的，并作为 I 层神经元中神经激肽 1（NK1）受体内化来测量。NMDA（与 D-丝氨酸联合使用）诱导 NK1 受体内化的有效浓度（EC50）为 258 nM。NMDA 诱导的 NK1 受体内化被 NK1 受体拮抗剂 L-703,606 所阻断，证实其是由 P 物质释放引起的，被 NMDA 受体拮抗剂（MK1801 和ifenprodil）所阻断，表明其是由含有 NR2B 亚基的 NMDA 受体介导的，并且通过预先用辣椒素孵育切片来证实，其 P 物质释放来自初级传入神经。然而，它不受利多卡因和ω-芋螺毒素 MVIIA 的影响，后者分别阻断钠通道和电压依赖性钙通道。因此，NMDA 诱导的 P 物质释放不需要初级传入神经的放电或钙通道的开放，这与 NMDA 受体通过让钙进入初级传入末梢直接诱导 P 物质的观点一致。重要的是，用 Src 家族激酶抑制剂 PP1 和 dasatinib 预先孵育切片 1 小时可消除 NMDA 诱导的 P 物质释放，而蛋白酪氨酸磷酸酶抑制剂 BVT948 则大大增加了 P 物质释放。相比之下，PP1 不影响辣椒素诱导的 NK1 受体内化。这些结果表明，这些 NMDA 受体的酪氨酸磷酸化受到 Src 家族激酶和蛋白酪氨酸磷酸酶的相反作用的调节，并且是诱导 P 物质释放所必需的。
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