Ahnert-Hilger G, John M, Kistner U, Wiedenmann B, Jarry H
Institut für Anatomie, Universitätsklinikum Charité, Humboldt-Universität zu Berlin, Germany.
Eur J Neurosci. 1998 Mar;10(3):1145-52. doi: 10.1046/j.1460-9568.1998.00129.x.
The immortalized hypothalamic neuronal cell lines GT1-1 and GT1-7 represent unique model systems to investigate the physiological control of gonadotropin-releasing hormone (GnRH) secretion. Using immunofluorescence microscopy, key proteins of regulated exocytosis, e.g. synaptotagmin, synaptobrevin and SNAP-25 (synaptosomal associated protein of 25 kDa) were found in GT1 neurons. In addition, GT1 neurons contained synaptophysin, a marker protein for small synaptic vesicles (SSVs) which are responsible for the storage of neurotransmitters such as gamma-aminobutyric acid (GABA). Upon subcellular fractionation, a lighter vesicle population characterized by synaptophysin separated from a denser vesicle population containing GnRH. Both vesicle populations contained synaptobrevin and synaptotagmin. Besides GnRH, GT1 neurons expressed glutamic acid decarboxylase at the mRNA-level and synthesized GABA. More importantly, GT1 neurons took up and stored 3H-GABA. The stored GABA was released after stimulation with increasing K+ concentrations and by alpha-latrotoxin. Reducing the extracellular Ca2+-concentration abolished stimulated secretion, indicating that GABA was released by regulated exocytosis. Hormone secretion from GT1 neurons is controlled by GABA via GABA(A) and GABA(B) receptors reflecting the situation in vivo. Our data provide the first evidence that GT1 neurons possess a second regulated secretory pathway sustained by SSVs storing and releasing GABA. The released GABA influences GnRH secretion by an auto- or paracrine loop.
永生化下丘脑神经元细胞系GT1-1和GT1-7是研究促性腺激素释放激素（GnRH）分泌生理调控的独特模型系统。利用免疫荧光显微镜技术，在GT1神经元中发现了调节性胞吐作用的关键蛋白，如突触结合蛋白、突触囊泡蛋白和SNAP-25（25 kDa的突触体相关蛋白）。此外，GT1神经元含有突触素，这是一种小突触囊泡（SSV）的标记蛋白，小突触囊泡负责储存神经递质，如γ-氨基丁酸（GABA）。经过亚细胞分级分离，以突触素为特征的较轻囊泡群体与含有GnRH的较密囊泡群体分离。两个囊泡群体都含有突触囊泡蛋白和突触结合蛋白。除了GnRH，GT1神经元在mRNA水平表达谷氨酸脱羧酶并合成GABA。更重要的是，GT1神经元摄取并储存3H-GABA。在增加K+浓度刺激和α- latrotoxin刺激后，储存的GABA被释放。降低细胞外Ca2+浓度可消除刺激后的分泌，表明GABA是通过调节性胞吐作用释放的。GT1神经元的激素分泌通过GABA(A)和GABA(B)受体受GABA控制，这反映了体内的情况。我们的数据首次证明GT1神经元拥有由储存和释放GABA的小突触囊泡维持的第二条调节性分泌途径。释放的GABA通过自分泌或旁分泌环影响GnRH分泌。
