A novel method for simultaneous anterograde and retrograde labeling of spinal cord motor tracts in the same animal
A novel method for simultaneous anterograde and retrograde labeling of spinal cord motor tracts in the same animal
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DOI：
10.1177/002215540104900905
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发表时间：
2001-09-01
影响因子：
3.2
通讯作者：
Tator, CH
中科院分区：
文献类型：
--
作者：
Tsai, EC;van Bendegem, RL;Tator, CH
由于组织固定等技术的不兼容，修复后的脊髓束的检查通常需要不同的动物组进行顺行和逆行追踪。然而，对接受相同修复的不同动物进行顺行和逆行标记可能无法准确检查该修复策略，因为在接受相同策略的动物中可能会出现广泛不同的结果。我们开发了一种可靠的方法，在同一动物中使用 Dil（一种亲脂性染料）顺行标记皮质脊髓束，并使用氟金（FG）逆行标记正常和受伤成年大鼠的多个脊髓运动束的皮质和脑干神经元。还研究了其他示踪剂组合（亲脂性染料或荧光葡聚糖），但效果较差。我们还开发了使用 Dil/FG 技术最小化自发荧光的方法，并发现 Dil/FG 技术与脊髓再生研究相关的几种标记物的脱钙和免疫组织化学兼容。因此，使用顺行 Dil 和逆行 FG 是一种用荧光示踪剂双向标记成人脊髓运动束的新技术，并且应该有助于在脊髓修复研究中证明神经突再生。
Examination of repaired spinal cord tracts has usually required separate groups of animals for anterograde and retrograde tracing owing to the incompatibility of techniques such as tissue fixation. However, anterograde and retrograde labeling of different animals subjected to the same repair may not allow accurate examination of that repair strategy because widely variable results can occur in animals subjected to the same strategy. We have developed a reliable method of labeling spinal cord motor tracts bidirectionally in the same animal using Dil, a lipophilic dye, to anterogradely label the corticospinal tract and Fluoro-Gold (FG) to retrogradely label cortical and brainstem neurons of several spinal cord motor tracts in normal and injured adult rats. Other tracer combinations (lipophilic dyes or fluorescent dextrans) were also investigated but were less effective. We also developed methods to minimize autofluorescence with the Dil/FG technique, and found that the Dil/FG technique is compatible with decalcification and immunohistochemistry for several markers relevant for studies of spinal cord regeneration. Thus, the use of anterograde Dil and retrograde FG is a novel technique for bidirectional labeling of the motor tracts of the adult spinal cord with fluorescent tracers and should be useful for demonstrating neurite regeneration in studies of spinal cord repair.