Doi K, Ohno K, Iwakura S, Takahashi Y, Kubo T
Department of Otolaryngology, Osaka University Medical School, Japan.
Acta Otolaryngol Suppl. 1995;520 Pt 2:334-7. doi: 10.3109/00016489509125264.
The expression of glutamate receptor gene family in the vestibular Scarpa's ganglion (VG) of rat was determined using molecular biological techniques including reverse transcription (RT), polymerase chain reaction (PCR), amplification, DNA sequencing, and immunocytochemistry. Oligonucleotide primers were designed from the previously cloned sequences of the AMPA-selective glutamate receptors (GluR1-4), KA-selective glutamate receptors (GluR5-7, KA1&2), NMDA-selective glutamate receptors (NMDAR1&2), and metabotropic glutamate receptors (mGluR1-4). The cDNAs synthesized from the VG mRNAs were amplified with the specific primers for the subunits of each glutamate receptor gene family. Analysis of the nucleotide sequences of the amplified cDNAs identified the expression of GluR1-4, GluR5, and NMDAR1 in the VG. Sequence analysis identified the expression of all the four subunits of GluR1-4 in two alternative spliced versions named "flip" and "flop" in the VG. In an immunocytochemical study, a large number of VG neurons showed only GluR2/3-like immunoreactivity (IR) while GluR1- or GluR4-IR could not be determined in the VG. The present study suggested that the subunits of GluR1-4, GluR5, and NMDAR1 may be assembled into the hetero-oligomeric glutamate receptor complex in the VG and that the VG neuron-specific stoichiometry of the receptor complex may be functionally significant for the afferent signal transduction in the rat peripheral vestibular system.
采用包括逆转录(RT)、聚合酶链反应(PCR)、扩增、DNA测序及免疫细胞化学等分子生物学技术,测定大鼠前庭斯卡帕神经节(VG)中谷氨酸受体基因家族的表达。根据先前克隆的AMPA选择性谷氨酸受体(GluR1 - 4)、KA选择性谷氨酸受体(GluR5 - 7、KA1&2)、NMDA选择性谷氨酸受体(NMDAR1&2)及代谢型谷氨酸受体(mGluR1 - 4)的序列设计寡核苷酸引物。用各谷氨酸受体基因家族亚基的特异性引物扩增从VG mRNA合成的cDNA。对扩增的cDNA核苷酸序列分析确定了VG中GluR1 - 4、GluR5和NMDAR1的表达。序列分析确定了VG中GluR1 - 4的所有四个亚基以名为“flip”和“flop”的两种可变剪接形式表达。在免疫细胞化学研究中,大量VG神经元仅显示GluR2/3样免疫反应性(IR),而在VG中无法确定GluR1或GluR4的IR。本研究提示,GluR1 - 4、GluR5和NMDAR1的亚基可能在VG中组装成异源寡聚体谷氨酸受体复合物,并且该受体复合物的VG神经元特异性化学计量学可能对大鼠外周前庭系统的传入信号转导具有功能意义。