Sullivan Megan R, Nimmerjahn Axel, Sarkisov Dmitry V, Helmchen Fritjof, Wang Samuel S-H
Department of Molecular Biology, Princeton University, Princeton, New Jersey, Princeton, NJ 08544, USA.
J Neurophysiol. 2005 Aug;94(2):1636-44. doi: 10.1152/jn.01013.2004. Epub 2005 Apr 20.
In vivo two-photon calcium imaging provides the opportunity to monitor activity in multiple components of neural circuitry at once. Here we report the use of bulk-loading of fluorescent calcium indicators to record from axons, dendrites, and neuronal cell bodies in cerebellar cortex in vivo. In cerebellar folium crus IIa of anesthetized rats, we imaged the labeled molecular layer and identified all major cellular structures: Purkinje cells, interneurons, parallel fibers, and Bergmann glia. Using extracellular stimuli we evoked calcium transients corresponding to parallel fiber beam activity. This beam activity triggered prolonged calcium transients in interneurons, consistent with in vitro evidence for synaptic activation of N-methyl-d-aspartate receptors via glutamate spillover. We also observed spontaneous calcium transients in Purkinje cell dendrites that were identified as climbing-fiber-evoked calcium spikes by their size, time course, and sensitivity to AMPA receptor antagonist. Two-photon calcium imaging of bulk-loaded cerebellar cortex is thus well suited to optically monitor synaptic processing in the intact cerebellum.
在体双光子钙成像提供了同时监测神经回路多个组成部分活动的机会。在此，我们报告了利用荧光钙指示剂的批量加载来记录麻醉大鼠小脑皮质轴突、树突和神经元细胞体的活动。在麻醉大鼠的小脑IIa小叶，我们对标记的分子层进行成像，并识别了所有主要细胞结构：浦肯野细胞、中间神经元、平行纤维和伯格曼胶质细胞。利用细胞外刺激，我们诱发了与平行纤维束活动相对应的钙瞬变。这种束状活动触发了中间神经元中长时间的钙瞬变，这与体外通过谷氨酸溢出对N-甲基-D-天冬氨酸受体进行突触激活的证据一致。我们还在浦肯野细胞树突中观察到自发钙瞬变，根据其大小、时间进程和对AMPA受体拮抗剂的敏感性，这些瞬变被确定为攀缘纤维诱发的钙峰。因此，批量加载的小脑皮质的双光子钙成像非常适合于光学监测完整小脑中的突触处理。
