Hou Dongmei, Suzuki Kazuhiro, Wolfgang William J, Clay Catherine, Forte Michael, Kidokoro Yoshiaki
Gunma University School of Medicine, Maebashi 371-8511, Japan.
J Neurosci. 2003 Jul 2;23(13):5897-905. doi: 10.1523/JNEUROSCI.23-13-05897.2003.
Gs(alpha) is a subunit of the heterotrimeric G-protein complex, expressed ubiquitously in all types of cells, including neurons. Drosophila larvae, which have mutations in the Gs(alpha) gene, are lethargic, suggesting an impairment of neuronal functions. In this study, we examined synaptic transmission at the neuromuscular synapse in Gs(alpha)-null (dgsR60) embryos shortly before they hatched. At low-frequency nerve stimulation, synaptic transmission in mutant embryos was not very different from that in controls. In contrast, facilitation during tetanic stimulation was minimal in dgsR60, and no post-tetanic potentiation was observed. Miniature synaptic currents (mSCs) were slightly smaller in amplitude and less frequent in dgsR60 embryos in normal-K+ saline. In high-K+ saline, mSCs with distinctly large amplitude occurred frequently in controls at late embryonic stages, whereas those mSCs were rarely observed in dgsR60 embryos, suggesting a developmental defect in the mutant. Using the Gal4-UAS expression system, we found that these phenotypes in dgsR60 were caused predominantly by lack of Gs(alpha) in presynaptic neurons and not in postsynaptic muscles. To test whether Gs(alpha) couples presynaptic modulator receptors to adenylyl cyclase (AC), we examined the responses of two known G-protein-coupled receptors in dgsR60 embryos. Both metabotropic glutamate and octopamine receptor responses were indistinguishable from those of controls, indicating that these receptors are not linked to AC by Gs(alpha). We therefore suggest that synaptic transmission is compromised in dgsR60 embryos because of presynaptic defects in two distinct processes; one is uncoupling between the yet-to-be-known modulator receptor and AC activation, and the other is a defect in synapse formation.
Gs(α)是异源三聚体G蛋白复合物的一个亚基,在包括神经元在内的所有类型细胞中普遍表达。Gs(α)基因发生突变的果蝇幼虫行动迟缓,提示神经元功能受损。在本研究中,我们在Gs(α)基因缺失(dgsR60)的胚胎孵化前不久,检测了其神经肌肉突触处的突触传递。在低频神经刺激下,突变胚胎中的突触传递与对照胚胎并无太大差异。相比之下,dgsR60胚胎在强直刺激期间的易化作用极小,且未观察到强直后增强。在正常钾离子盐溶液中,dgsR60胚胎中的微小突触电流(mSCs)幅度略小且频率较低。在高钾离子盐溶液中,对照胚胎在胚胎后期阶段经常出现幅度明显较大的mSCs,而在dgsR60胚胎中很少观察到这些mSCs,这表明突变体存在发育缺陷。使用Gal4-UAS表达系统,我们发现dgsR60中的这些表型主要是由突触前神经元而非突触后肌肉中缺乏Gs(α)引起的。为了测试Gs(α)是否将突触前调制受体与腺苷酸环化酶(AC)偶联,我们检测了dgsR60胚胎中两种已知的G蛋白偶联受体的反应。代谢型谷氨酸受体和章鱼胺受体的反应与对照无明显差异,表明这些受体不通过Gs(α)与AC相连。因此,我们认为dgsR60胚胎中的突触传递受损是由于两个不同过程中的突触前缺陷;一个是未知的调制受体与AC激活之间的解偶联,另一个是突触形成缺陷。