Takeda Atsushi, Fuke Sayuri, Tsutsumi Wataru, Oku Naoto
Department of Medical Biochemistry, School of Pharmaceutical Sciences, University of Shizuoka, 52-1 Yada, Suruga-ku, Shizuoka, Japan.
J Neurosci Res. 2007 Dec;85(16):3666-72. doi: 10.1002/jnr.21449.
The role of zinc in excitation of Schaffer collateral-CA1 pyramidal cell synapses is poorly understood. Schaffer collaterals stained with ZnAF-2 or ZnAF-2DA, a membrane-impermeable or a membrane-permeable zinc indicator, respectively, were treated by tetanic stimulation (200 Hz, 1 sec). Extracellular and intracellular ZnAF-2 signals were increased in the stratum radiatum of the CA1, in which Schaffer collateral synapses exist. Both the increases were completely blocked in the presence of 1 mM CaEDAT, a membrane-impermeable zinc chelator, suggesting that 1 mM CaEDTA is effective for chelating zinc released from Schaffer collaterals. The role of Schaffer collateral zinc in presynaptic activity was examined by using FM4-64, a fluorescent indicator for vesicular exocytosis. The decrease in FM4-64 signal during tetanic stimulation (10 Hz, 180 sec) was enhanced in Schaffer collaterals in the presence of 1 mM CaEDTA but suppressed in the presence of 5 microM ZnC1(2), suggesting that zinc released from Schaffer collaterals suppresses presynaptic activity during tetanic stimulation. When Schaffer collateral synapses stained with calcium orange AM, a membrane-permeable calcium indicator, were regionally stimulated with 1 mM glutamate, calcium orange signal was increased in the CA1 pyramidal cell layer. This increase was enhanced in the presence of CaEDTA and attenuated in the presence of zinc. These results suggest that zinc attenuates excitation of Schaffer collateral synapses elicited with glutamate via suppression of presynaptic activity.
锌在海马体联合纤维-海马体CA1区锥体细胞突触兴奋过程中的作用目前仍知之甚少。分别用膜不通透性锌指示剂ZnAF-2或膜通透性锌指示剂ZnAF-2DA对海马体联合纤维进行染色，然后施加强直刺激（200Hz，1秒）。在存在海马体联合纤维突触的CA1区辐射层中，细胞外和细胞内的ZnAF-2信号均增强。在加入1mM CaEDAT（一种膜不通透性锌螯合剂）后，这两种增强均被完全阻断，这表明1mM CaEDTA能有效螯合从海马体联合纤维释放的锌。通过使用FM4-64（一种用于囊泡胞吐作用的荧光指示剂）来研究海马体联合纤维锌在突触前活动中的作用。在1mM CaEDTA存在的情况下，强直刺激（10Hz，180秒）期间FM4-64信号的减少在海马体联合纤维中增强，但在5μM ZnC1₂存在的情况下受到抑制，这表明从海马体联合纤维释放的锌在强直刺激期间抑制突触前活动。当用膜通透性钙指示剂钙橙AM染色的海马体联合纤维突触用1mM谷氨酸进行局部刺激时，CA1区锥体细胞层中的钙橙信号增强。在CaEDTA存在时这种增强更为明显，而在锌存在时则减弱。这些结果表明，锌通过抑制突触前活动减弱了由谷氨酸引发的海马体联合纤维突触的兴奋。
