A modified KOH-collagenase method applied to scanning electron microscopic observations of peripheral nerves.
A modified KOH-collagenase method applied to scanning electron microscopic observations of peripheral nerves.
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一种改良的 KOH-胶原酶方法，应用于周围神经的扫描电子显微镜观察。
DOI：
10.1679/aohc.51.223
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发表时间：
1988
影响因子：
--
通讯作者：
C. Idé
中科院分区：
文献类型：
--
作者：
T. Ushiki;C. Idé
通过米勒等人（1982）改良的KOH-胶原酶法观察周围神经系统的某些部分。将小鼠坐骨神经、大鼠背根神经节、舌肌和空肠在室温下用含有多聚甲醛和戊二醛的固定剂固定一天以上。固定后，在60 ℃下用5 N KOH溶液处理标本5-10 min，在37 ℃下浸入胶原酶溶液中3-5 h，并进行扫描电子显微镜（SEM）处理。在充分处理的标本中，结缔组织基质和基底层被完全去除，不会造成任何严重的组织损伤。周围神经细胞成分的三维可视化实现了以下结果：1）坐骨神经的单个神经纤维被清楚地暴露。有髓纤维为无分支索状，在郎维氏结处有局部环状收缩，无髓纤维则相互分支、交叉，形成松散的网状结构。雪旺细胞胞质的聚集部位在有髓纤维表面肿胀。中轴突被可视化为纵向沟。2)背根神经节内可见被卫星细胞覆盖的神经节细胞。神经节细胞被其自身的卷曲的树突-轴突突起覆盖，从而形成最初的Cajal肾小球。卫星细胞在肾小球延伸出许多指状突起，围绕着树突-轴突突起。3)舌肌内可见运动终板。终板处的终末雪旺细胞（终末细胞）清晰可见：圆形的核周体沿着终板内的轴突分支延伸出胞质突起。这些突起从边缘发出细小的手指状突起。4)舌肌内血管自主神经丛清晰可见。无髓神经分支和闭合，在血管周围形成复杂的神经网络。高倍镜下可见神经突起和相关的雪旺细胞。5)空肠粘膜下神经丛由大量神经节和相互连接的纤维束组成，构成复杂的神经网络。这些观察结果表明，这种改进的KOH消化方法是有用的扫描电镜研究的神经元在各种组织中的三维细胞组织。
Certain parts of the peripheral nervous system were observed by a modification of the KOH-collagenase method by Miller et al. (1982). The sciatic nerve of the mouse, and dorsal root ganglion, lingual muscle and jejunum of the rat were fixed over one day at room temperature in a fixative containing paraformaldehyde and glutaraldehyde. After fixation, specimens were treated with 5N KOH solution for 5-10 min at 60 degrees C, immersed in a collagenase solution for 3-5 h at 37 degrees C and processed for scanning electron microscopy (SEM). In adequately treated specimens, connective tissue matrices and basal laminae were completely removed without causing any severe tissue damage. The three-dimensional visualization of cellular elements of peripheral nerves was enabled with the following results: 1) Individual nerve fibers of the sciatic nerve were clearly exposed. Myelinated fibers were non-branching cords with local annular constrictions at the node of Ranvier, while unmyelinated ones branched and anastomosed with one another to form loose networks. The sites of accumulation of the Schwann cell cytoplasm swelled on the surface of the myelinated fibers. Mesaxons were visualized as longitudinal furrows. 2) Ganglion cells covered by satellite cells were observed in the dorsal root ganglion. The ganglion cells were covered by their own convoluted dendro-axonal processes, thus forming the initial glomeruli of Cajal. Satellite cells at the glomeruli extended many finger-like projections surrounding the dendro-axonal processes. 3) Motor endplates were observed in lingual muscles. Terminal Schwann cells (teloglia) at the endplate were clearly visualized: the round perikaryon extended cytoplasmic processes along the axonal branches within the endplate. The processes issued fine finger-like projections from their margins. 4) Vascular autonomic plexuses were clearly demonstrated in lingual muscles. Unmyelinated nerves branched and anastomosed to form elaborate nerve networks around the vessels. Neuronal processes and associated Schwann cells were identifiable at high magnification. 5) Submucous nerve plexuses in the jejunum consisted of numbers of ganglia and interconnecting strands of fibers which formed very complicated networks. These observations indicate that this modified KOH-digestion method is useful for the SEM study of the three-dimensional cellular organization of nervous elements in various tissues.