Garbelli Rita, Inverardi Francesca, Medici Valentina, Amadeo Alida, Verderio Claudia, Matteoli Michela, Frassoni Carolina
Clinical Epileptology and Experimental Neurophysiology Unit, Fondazione IRCCS, Istituto Neurologico C Besta, via Celoria 11, Milano, Italy.
J Comp Neurol. 2008 Jan 20;506(3):373-86. doi: 10.1002/cne.21505.
Synaptosomal associated protein of 25 kDa (SNAP-25) is a SNARE component of the exocytotic apparatus involved in the release of neurotransmitter. We used multiple-labeling immunofluorescence, confocal microscopy, and ultrastructural immunocytochemistry to examine the expression of SNAP-25 in excitatory and inhibitory terminals from different rat and human brain areas. Glutamatergic and GABAergic terminals were identified by staining for the vesicular glutamate transporter (vGLUT1), glutamic acid decarboxylase (GAD67), or the vesicular GABA transporter (vGAT). In all examined areas GABAergic terminals did not display detectable levels of SNAP-25, whereas glutamatergic terminals expressed the protein to a variable extent. Codistribution analysis revealed a high colocalization between pixels detecting SNAP-25 labeling and pixels detecting vGLUT1 immunoreactivity. On the contrary, a low degree of pixel colocalization, comparable to that between two unrelated antigens, was detected between SNAP-25 and vGAT, thus suggesting a random overlap of immunofluorescence signals. Our immunofluorescence evidence was supported by ultrastructural data, which clearly confirmed that SNAP-25 was undetectable in GABAergic terminals identified by both their typical morphology and specific staining for GABA. Interestingly, our ultrastructural results confirmed that a subset of glutamatergic synapses do not contain detectable levels of SNAP-25. The present study extends our previous findings obtained in rodent hippocampus and provides evidence that SNAP-25 expression is highly variable between different axon terminals both in rat and human brain. The heterogeneous distribution of SNAP-25 may have important implications not only in relation to the function of the protein as a SNARE but also in the control of network excitability.
25 kDa突触体相关蛋白（SNAP - 25）是参与神经递质释放的胞吐装置的一种可溶性N - 乙基马来酰胺敏感因子附着蛋白受体（SNARE）成分。我们使用多重标记免疫荧光、共聚焦显微镜和超微结构免疫细胞化学方法，来检测不同大鼠和人类脑区兴奋性和抑制性终末中SNAP - 25的表达。通过对囊泡型谷氨酸转运体（vGLUT1）、谷氨酸脱羧酶（GAD67）或囊泡型GABA转运体（vGAT）进行染色来鉴定谷氨酸能和GABA能终末。在所有检测区域中，GABA能终末未显示出可检测到的SNAP - 25水平，而谷氨酸能终末则在不同程度上表达该蛋白。共分布分析显示，检测SNAP - 25标记的像素与检测vGLUT1免疫反应性的像素之间存在高度共定位。相反，在SNAP - 25和vGAT之间检测到的像素共定位程度较低，与两种不相关抗原之间的共定位程度相当，因此表明免疫荧光信号存在随机重叠。我们的免疫荧光证据得到了超微结构数据的支持，这些数据清楚地证实，在通过典型形态和GABA特异性染色鉴定的GABA能终末中未检测到SNAP - 25。有趣的是，我们的超微结构结果证实，一部分谷氨酸能突触不含有可检测到的SNAP - 25水平。本研究扩展了我们先前在啮齿动物海马体中获得的发现，并提供证据表明，在大鼠和人类大脑中，不同轴突终末之间SNAP - 25的表达高度可变。SNAP - 25的异质性分布不仅可能对该蛋白作为SNARE的功能具有重要意义，而且可能对网络兴奋性的控制也具有重要意义。
