MICROANALYSIS OF RAPIDLY-FROZEN TISSUE IN THE FIELD EMISSION STEM

扫描电子显微镜(STEM)，电子能量损失 能谱(EELS)和能量色散X射线能谱(EDXS)有 已开发并应用于钙和其他离子的测量 在神经元中。这些进展包括：玻璃化培养的能力 脑切片，改进的冷冻切片技术，能够表征 鳗鱼冷冻水化切片，低剂量数字暗视野切片 冷冻干燥切片的成像和高效X射线 微量分析。这些方法已被应用于鉴定钙 大鼠海马CA3区锥体细胞树突内的缓冲细胞器 紧随传入突触活动之后的几分钟。这个 结果显示，内质网的一个亚群是主要的， 这些树突中有高容量的钙缓冲液。自动减支活动 这些细胞器中的一个是分级的和可逆的。它也是非常 很健壮。在强直性刺激后，总钙离子 平均而言，这些细胞器内的浓度增加了20倍。 分析电子显微镜也被用来测量钙和 快速冷冻神经垂体神经中的其他元素浓度 最后，为了确定钙在细胞分泌中的作用 脑下垂体后叶激素。

Scanning transmission electron microscopy (STEM), electron energy-loss spectroscopy (EELS), and energy-dispersive X-ray spectroscopy (EDXS) have been developed and applied to the measurement of calcium and other ions in neurons. The developments include: capability to vitrify cultured brain slices, improved cryosectioning technique, ability to characterize frozen-hydrated sections by EELS, low-dose digital dark-field STEM imaging of the freeze-dried sections, and high-efficiency X-ray microanalysis. These methods have been applied to identify calcium buffering organelles in dendrites of CA3 hippocampal pyramidal cells in the minutes immediately following afferent synaptic activity. The results reveal a subset of endoplasmic reticulum as a major, high-capacity Ca++ buffer in these dendrites. The sequestration activity of these organelles was graded and reversible. It was also extremely robust. Following strong tetanic stimulation, the total Ca++ concentration within these organelles increased twentyfold, on average. Analytical electron microscopy has also been used to measure calcium and other elemental concentrations in rapidly frozen neurohypophyseal nerve endings, in order to determine the role of calcium in the secretion of posterior pituitary hormones.