Pan Ping-Yue, Cai Qian, Lin Lin, Lu Pei-Hua, Duan Shumin, Sheng Zu-Hang
Department of Neurobiology, Shanghai Second Medical University, 280 Chong Qing Nan Road, 200025 Shanghai, China.
J Biol Chem. 2005 Jul 8;280(27):25769-79. doi: 10.1074/jbc.M502356200. Epub 2005 May 12.
Identifying the molecules that regulate both the recycling of synaptic vesicles and the SNARE components required for fusion is critical for elucidating the molecular mechanisms underlying synaptic plasticity. SNAP-29 was initially isolated as a syntaxin-binding and ubiquitously expressed protein. Previous studies have suggested that SNAP-29 inhibits SNARE complex disassembly, thereby reducing synaptic transmission in cultured superior cervical ganglion neurons in an activity-dependent manner. However, the role of SNAP-29 in regulating synaptic vesicle recycling and short-term plasticity in the central nervous system remains unclear. In the present study, we examined the effect of SNAP-29 on synaptic transmission in cultured hippocampal neurons by dual patch clamp whole-cell recording, FM dye imaging, and immunocytochemistry. Our results demonstrated that exogenous expression of SNAP-29 in presynaptic neurons significantly decreased the efficiency of synaptic transmission after repetitive firing within a few minutes under low and moderate frequency stimulations (0.1 and 1 Hz). In contrast, SNAP-29 did not affect the density of synapses and basal synaptic transmission. Whereas neurotransmitter release was unaffected during intensive stimulation, recovery after synaptic depression was impaired by SNAP-29. Furthermore, knockdown of SNAP-29 expression in neurons by small interfering RNA increased the efficiency of synaptic transmission during repetitive firing. These findings suggest that SNAP-29 acts as a negative modulator for neurotransmitter release, probably by slowing recycling of the SNARE-based fusion machinery and synaptic vesicle turnover.
识别调控突触小泡循环以及融合所需的SNARE组件的分子，对于阐明突触可塑性背后的分子机制至关重要。SNAP-29最初是作为一种与 syntaxin 结合且广泛表达的蛋白质被分离出来的。先前的研究表明，SNAP-29抑制SNARE复合体的拆解，从而以一种活动依赖的方式减少培养的颈上神经节神经元中的突触传递。然而，SNAP-29在调节中枢神经系统中突触小泡循环和短期可塑性方面的作用仍不清楚。在本研究中，我们通过双膜片钳全细胞记录、FM染料成像和免疫细胞化学方法，研究了SNAP-29对培养的海马神经元突触传递的影响。我们的结果表明，在低频率和中等频率刺激（0.1和1Hz）下，几分钟内重复放电后，突触前神经元中外源表达的SNAP-29显著降低了突触传递效率。相比之下，SNAP-29不影响突触密度和基础突触传递。虽然在强烈刺激期间神经递质释放不受影响，但SNAP-29损害了突触抑制后的恢复。此外，通过小干扰RNA敲低神经元中SNAP-29的表达，增加了重复放电期间的突触传递效率。这些发现表明，SNAP-29可能通过减缓基于SNARE的融合机制和突触小泡周转的循环，作为神经递质释放的负调节剂。
