Rose C R, Kovalchuk Y, Eilers J, Konnerth A
I. Physiologisches Institut, Universität des Saarlandes, Homburg, Germany.
Pflugers Arch. 1999 Dec;439(1-2):201-7. doi: 10.1007/s004249900123.
Dendritic spines are assumed to be the smallest units of neuronal integration. Because of their miniature size, however, many of their functional properties are still unclear. New insights in spine physiology have been provided by two-photon laser-scanning microscopy which allows fluorescence imaging with high spatial resolution and minimal photodamage. For example, two-photon imaging has been employed successfully for the measurement of activity-induced calcium transients in individual spines. Here, we describe the first application of two-photon imaging to measure Na+ transients in spines and dendrites of CA1 pyramidal neurons in hippocampal slices. Whole-cell patch-clamped neurons were loaded with the Na(+)-indicator dye SBFI (sodium-binding benzofuran-isophthalate). In situ calibration of SBFI fluorescence with ionophores enabled the determination of the actual magnitude of the [Na+]i changes. We found that back-propagating action potentials (APs) evoked Na+ transients throughout the proximal part of the dendritic tree and adjacent spines. The action-potential-induced [Na+]i transients reached values of 4 mM for a train of 20 APs and monotonically decayed with a time constant of several seconds. These results represent the first demonstration of activity-induced Na+ accumulation in spines. Our results demonstrate that two-photon Na+ imaging represents a powerful tool for extending our knowledge on Na+ signaling in fine cellular subcompartments.
树突棘被认为是神经元整合的最小单位。然而，由于其微小的尺寸，其许多功能特性仍不清楚。双光子激光扫描显微镜为棘突生理学提供了新的见解，它能够进行高空间分辨率且光损伤最小的荧光成像。例如，双光子成像已成功用于测量单个棘突中活性诱导的钙瞬变。在此，我们描述了双光子成像在测量海马切片中CA1锥体神经元的棘突和树突中Na+瞬变方面的首次应用。全细胞膜片钳记录的神经元被加载Na(+)-指示剂染料SBFI（钠结合苯并呋喃-间苯二甲酸酯）。使用离子载体对SBFI荧光进行原位校准，能够确定[Na+]i变化的实际幅度。我们发现，反向传播动作电位（APs）在整个树突树近端部分和相邻棘突中诱发Na+瞬变。对于一串20个APs，动作电位诱导的[Na+]i瞬变达到4 mM的值，并以几秒的时间常数单调衰减。这些结果首次证明了活性诱导的Na+在棘突中的积累。我们的结果表明，双光子Na+成像代表了一种强大的工具，可扩展我们对精细细胞亚区室中Na+信号传导的认识。
