Colin I, Blondeau C, Baude A
ITIS, CNRS, UMR 6150, Batiment N', 31 chemin Joseph Aiguier, 13402 Cedex 20, Marseille, France.
Neuroscience. 2002;115(4):1023-33. doi: 10.1016/s0306-4522(02)00541-9.
Neurokinins (substance P, neurokinin A and neurokinin B) and the neurokinin receptors, the NK1 and NK3 receptors, are largely expressed in the nucleus of the solitary tract (NST) where they are involved in the central regulation of visceral function. Studying the mechanisms that control neurokinin release can provide valuable information concerning the control of autonomic functions subserved by the NST. Glutamate is the principal excitatory neurotransmitter in the NST and the main neurotransmitter of afferent vagal fibers. Neurokinins and glutamate may interact within the NST. In the present study, we have examined the contribution of the N-methyl-D-aspartate (NMDA) and alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) subtypes of glutamate receptors on the release of the endogenous neurokinins in the NST. We used internalization of the NK1 or NK3 receptor as an index of endogenous neurokinin release assessed by immunocytochemical visualization of the NK1 or NK3 receptor endocytosis. Experiments were performed in vitro using rat brainstem slices. A first series of experiments were done in order to validate our in vitro preparation. Application of substance P, neurokinin A or neurokinin B induced dose-dependent internalization of NK1 and NK3 receptor. This was blocked by the endocytosis inhibitor, phenylarzine oxide. The NK1 receptor antagonist SR140333 blocked internalization of NK1 receptor induced by the three neurokinins. In addition, the internalization NK1 or NK3 receptor was reversible. These results demonstrate that internalization and recycling mechanisms of NK1 or NK3 receptor were preserved in in vitro brainstem slices. Application of NMDA or AMPA induced internalization of NK1 receptor. This was blocked by the application of SR140333 suggesting that NK1 receptor internalization is due to the binding of endogenous neurokinin released under the effects of NMDA and AMPA. Application of NMDA or AMPA had no effect on NK3 receptor. Application of tetrodotoxin blocked NK1 receptor internalization induced by NMDA, demonstrating that the release of neurokinins is dependent of axon potential propagation. This result excludes the hypothesis of a release on neurokinins via pre-synaptic NMDA receptors located on neurokinin-containing axon terminals. NMDA or AMPA may directly induce neurokinin release in the NST by acting on receptors located on the cell bodies and dendrites of neurokinin-containing neurons. Release of neurokinins may also be the result of a general activation of neuron networks of the NST by NMDA or AMPA. To conclude, our results suggest that glutamate, through activation of post-synaptic NMDA and AMPA receptors, contributes to neurokinin signaling in the NST.
神经激肽（P物质、神经激肽A和神经激肽B）以及神经激肽受体，即NK1和NK3受体，主要表达于孤束核（NST），它们参与内脏功能的中枢调节。研究控制神经激肽释放的机制可为了解由NST所维持的自主功能的控制提供有价值的信息。谷氨酸是NST中的主要兴奋性神经递质以及传入迷走神经纤维的主要神经递质。神经激肽和谷氨酸可能在NST内相互作用。在本研究中，我们研究了N-甲基-D-天冬氨酸（NMDA）和α-氨基-3-羟基-5-甲基-4-异恶唑丙酸（AMPA）亚型的谷氨酸受体对NST中内源性神经激肽释放的作用。我们将NK1或NK3受体的内化作为内源性神经激肽释放的指标，通过对NK1或NK3受体内吞作用的免疫细胞化学可视化进行评估。实验在体外使用大鼠脑干切片进行。进行了第一系列实验以验证我们的体外制备方法。应用P物质、神经激肽A或神经激肽B可诱导NK1和NK3受体的剂量依赖性内化。这被内吞抑制剂氧化苯胂阻断。NK1受体拮抗剂SR140333可阻断这三种神经激肽诱导的NK1受体内化。此外，NK1或NK3受体的内化是可逆的。这些结果表明，NK1或NK3受体的内化和再循环机制在体外脑干切片中得以保留。应用NMDA或AMPA可诱导NK1受体内化。这可被SR140333的应用所阻断，表明NK1受体内化是由于在NMDA和AMPA作用下释放的内源性神经激肽的结合。应用NMDA或AMPA对NK3受体无影响。应用河豚毒素可阻断NMDA诱导的NK1受体内化，表明神经激肽的释放依赖于轴突电位的传播。这一结果排除了通过位于含神经激肽轴突终末上的突触前NMDA受体释放神经激肽的假说。NMDA或AMPA可能通过作用于含神经激肽神经元的细胞体和树突上的受体直接诱导NST中的神经激肽释放。神经激肽的释放也可能是NMDA或AMPA对NST神经元网络的普遍激活的结果。总之，我们的结果表明，谷氨酸通过激活突触后NMDA和AMPA受体，在NST的神经激肽信号传导中发挥作用。
