Gleason E L, Spitzer N C
Department of Biology and Center for Molecular Genetics, University of California, San Diego, La Jolla,California 92093, USA.
J Neurophysiol. 1998 Jun;79(6):2986-98. doi: 10.1152/jn.1998.79.6.2986.
N-methyl--aspartate (NMDA) receptors are often the first ionotropic glutamate receptors expressed at early stages of development and appear to influence neuronal differentiation by mediating Ca2+ influx. Although less well studied, alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors also can generate Ca2+ elevations and may have developmental roles. We document the presence of AMPA and NMDA class receptors and the absence of kainate class receptors with whole cell voltage-clamp recordings from Xenopus embryonic spinal neurons differentiated in vitro. Reversal potential measurements indicate that AMPA receptors are permeable to Ca2+ both in differentiated neurons and at the time they first are expressed. The PCa/Pmonocation of 1.9 is close to that of cloned Ca2+-permeable AMPA receptors expressed in heterologous systems. Ca2+ imaging reveals that Ca2+ elevations are elicited by AMPA or NMDA in the absence of Mg2+. The amplitudes and durations of these agonist-induced Ca2+ elevations are similar to those of spontaneous Ca2+ transients known to act as differentiation signals in these cells. Two sources of Ca2+ amplify AMPA- and NMDA-induced Ca2+ elevations. Activation of voltage-gated Ca2+ channels by AMPA- or NMDA-mediated depolarization contributes approximately 15 or 30% of cytosolic Ca2+ elevations, respectively. Activation of either class of receptor produces elevations of Ca2+ that elicit further release of Ca2+ from thapsigargin-sensitive but ryanodine-insensitive stores, contributing an additional approximately 30% of Ca2+ elevations. Voltage-clamp recordings and Ca2+ imaging both show that these spinal neurons express functional AMPA receptors soon after neurite initiation and before expression of NMDA receptors. The Ca2+ permeability of AMPA receptors, their ability to generate significant elevations of [Ca2+]i, and their appearance before synapse formation position them to play roles in neural development. Spontaneous release of agonists from growth cones is detected with glutamate receptors in outside-out patches, suggesting that spinal neurons are early, nonsynaptic sources of glutamate that can influence neuronal differentiation in vivo.
N-甲基-D-天冬氨酸（NMDA）受体通常是在发育早期阶段表达的首批离子型谷氨酸受体，并且似乎通过介导Ca2+内流来影响神经元分化。虽然研究较少，但α-氨基-3-羟基-5-甲基-4-异恶唑丙酸（AMPA）受体也能引起Ca2+浓度升高，并且可能在发育过程中发挥作用。我们通过对体外分化的非洲爪蟾胚胎脊髓神经元进行全细胞电压钳记录，证明了AMPA和NMDA类受体的存在以及海人藻酸类受体的缺失。反转电位测量表明，AMPA受体在分化的神经元中以及首次表达时对Ca2+都是通透的。1.9的Ca2+通透率与异源系统中表达的克隆的Ca2+通透型AMPA受体相近。Ca2+成像显示，在不存在Mg2+的情况下，AMPA或NMDA能引起Ca2+浓度升高。这些激动剂诱导的Ca2+浓度升高的幅度和持续时间与已知在这些细胞中作为分化信号的自发Ca2+瞬变相似。有两种Ca2+来源会放大AMPA和NMDA诱导的Ca2+浓度升高。AMPA或NMDA介导的去极化激活电压门控Ca2+通道分别约占胞质Ca2+浓度升高的15%或30%。两类受体中的任何一类被激活都会使Ca2+浓度升高，从而引起从毒胡萝卜素敏感但对兰尼碱不敏感的储存库中进一步释放Ca2+，这又额外贡献了约30%的Ca2+浓度升高。电压钳记录和Ca2+成像均显示，这些脊髓神经元在神经突起始后不久且在NMDA受体表达之前就表达功能性AMPA受体。AMPA受体的Ca2+通透性、它们产生显著的[Ca2+]i升高的能力以及它们在突触形成之前的出现表明它们在神经发育中发挥作用。通过外向膜片上的谷氨酸受体检测到生长锥自发释放激动剂，这表明脊髓神经元是谷氨酸的早期非突触来源，能够在体内影响神经元分化。
