VOLTAGE-SENSITIVE DYE RECORDING OF ACTION-POTENTIALS AND SYNAPTIC POTENTIALS FROM SYMPATHETIC MICROCULTURES
VOLTAGE-SENSITIVE DYE RECORDING OF ACTION-POTENTIALS AND SYNAPTIC POTENTIALS FROM SYMPATHETIC MICROCULTURES
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DOI：
10.1016/s0006-3495(91)82099-9
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发表时间：
1991-09-01
影响因子：
3.4
通讯作者：
PINE, J
中科院分区：
文献类型：
--
作者：
CHIEN, CB;PINE, J
考虑到适当的多细胞电生理技术，培养神经元的小网络（微培养）非常适合于突触可塑性的长期研究。为此，我们开发了一种利用电压敏感荧光染料从培养的脊椎动物神经元中进行光学记录的装置（Chien， C.-B.）。潘志强等，1991。j . >。方法。38:93 - 105)。我们在此评估该技术在记录大鼠颈上神经节（SCG）神经元微培养中的动作电位和突触电位的实用性。经过广泛的染料筛选和条件优化后，我们选择了苯乙烯基染料RH423，该染料对典型记录的线性荧光变化速度快，约为1%/100 mV。仪器的均方根噪声（受射击噪声限制）一般为0.03%，相当于3 mV的膜电位。照明至少100次，每次100毫秒，没有造成明显的光动力学损伤。结果表明，电压敏感染料可用于记录脊椎动物神经元的微培养，灵敏度高。细胞体和神经突均可检测到染料信号。来自推定树突的信号与细胞体的信号同时显示超极化和动作电位，而来自推定轴突的信号显示延迟传播的动作电位。胞体内的阈下突触电位偶有可见；然而，它们通常被通过同一像素的轴突信号所掩盖。这是由于SCG微培养的复杂解剖结构，其中有许多交叉的神经突，经常穿过细胞体。如果有一个更简单的微培养系统和更少的神经突，应该可以使用染料记录常规测量阈下突触强度。
Given the appropriate multicell electrophysiological techniques, small networks of cultured neurons (microcultures) are well suited to long-term studies of synaptic plasticity. To this end, we have developed an apparatus for optical recording from cultured vertebrate neurons using voltage-sensitive fluorescent dyes (Chien, C.-B., and J. Pine. 1991. J. Neurosci. Methods. 38:93-105). We evaluate here the usefulness of this technique for recording action potentials and synaptic potentials in microcultures of neurons from the rat superior cervical ganglion (SCG). After extensive dye screening and optimization of conditions, we chose the styryl dye RH423, which gave fast linear fluorescence changes of approximately 1%/100 mV for typical recordings. The root mean square noise of the apparatus (limited by shot noise) was typically 0.03%, equivalent to 3 mV of membrane potential. Illumination for at least 100 flashes of 100 ms each caused no noticeable photodynamic damage.Our results show that voltage-sensitive dyes can be used to record from microcultures of vertebrate neurons with high sensitivity. Dye signals were detected from both cell bodies and neurites. Signals from presumptive dendrites showed hyperpolarizations and action potentials simultaneous with those in the cell body, while those from presumptive axons showed delayed propagating action potentials. Subthreshold synaptic potentials in the cell body were occasionally detectable optically; however, they were usually masked by signals from axons passing through the same pixel. This is due to the complex anatomy of SCG microcultures, which have many crisscrossing neurites that often pass over cell bodies. Given a simpler microculture system with fewer neurites, it should be possible to use dye recording to routinely measure subthreshold synaptic strengths.