Department of Comparative Medicine, Stanford University, Stanford, California 94305, USA.
Epilepsia. 2011 Dec;52(12):2326-32. doi: 10.1111/j.1528-1167.2011.03315.x. Epub 2011 Nov 16.
Dentate granule cell axon (mossy fiber) sprouting creates an aberrant positive-feedback circuit that might be epileptogenic. Presumably, mossy fiber sprouting is initiated by molecular signals, but it is unclear whether they are expressed transiently or persistently. If transient, there might be a critical period when short preventative treatments could permanently block mossy fiber sprouting. Alternatively, if signals persist, continuous treatment would be necessary. The present study tested whether temporary treatment with rapamycin has long-term effects on mossy fiber sprouting.
Mice were treated daily with 1.5 mg/kg rapamycin or vehicle (i.p.) beginning 24 h after pilocarpine-induced status epilepticus. Mice were perfused for anatomic evaluation immediately after 2 months of treatment ("0 delay") or after an additional 6 months without treatment ("6-month delay"). One series of sections was Timm-stained, and an adjacent series was Nissl-stained. Stereologic methods were used to measure the volume of the granule cell layer plus molecular layer and the Timm-positive fraction. Numbers of Nissl-stained hilar neurons were estimated using the optical fractionator method.
At 0 delay, rapamycin-treated mice had significantly less black Timm staining in the granule cell layer plus molecular layer than vehicle-treated animals. However, by 6-month delay, Timm staining had increased significantly in mice that had been treated with rapamycin. Percentages of the granule cell layer plus molecular layer that were Timm-positive were high and similar in 0 delay vehicle-treated, 6-month delay vehicle-treated, and 6-month delay rapamycin-treated mice. Extent of hilar neuron loss was similar among all groups that experienced status epilepticus and, therefore, was not a confounding factor. Compared to naive controls, average volume of the granule cell layer plus molecular layer was larger in 0 delay vehicle-treated mice. The hypertrophy was partially suppressed in 0 delay rapamycin-treated mice. However, 6-month delay vehicle- and 6-month delay rapamycin-treated animals had similar average volumes of the granule cell layer plus molecular layer that were significantly larger than those of all other groups.
Status epilepticus-induced mossy fiber sprouting and dentate gyrus hypertrophy were suppressed by systemic treatment with rapamycin but resumed after treatment ceased. These findings suggest that molecular signals that drive mossy fiber sprouting and dentate gyrus hypertrophy might persist for >2 months after status epilepticus in mice. Therefore, prolonged or continuous treatment might be required to permanently suppress mossy fiber sprouting.
齿状回颗粒细胞轴突（苔藓纤维）发芽形成异常正反馈回路，可能引发癫痫。推测苔藓纤维发芽是由分子信号引发的，但尚不清楚这些信号是短暂表达还是持续表达。如果是短暂的，那么在短时间的预防性治疗可能会永久性阻断苔藓纤维发芽的关键时期。或者，如果信号持续存在，则需要持续治疗。本研究测试了雷帕霉素的短期治疗是否对苔藓纤维发芽有长期影响。
在匹罗卡品诱导的癫痫持续状态后 24 小时，每日用 1.5mg/kg 雷帕霉素或载体（腹腔内）处理小鼠。治疗 2 个月后立即（“0 延迟”）或不治疗 6 个月后（“6 个月延迟”）对小鼠进行解剖评估。一系列切片用 Timm 染色，相邻系列用尼氏染色。立体学方法用于测量颗粒细胞层加分子层的体积和 Timm 阳性分数。尼氏染色的颗粒细胞层加分子层的神经细胞数量用光学分选区法估计。
在 0 延迟时，雷帕霉素处理的小鼠颗粒细胞层加分子层的黑色 Timm 染色明显少于载体处理的动物。然而，到 6 个月延迟时，雷帕霉素处理的小鼠 Timm 染色显著增加。0 延迟载体处理、6 个月延迟载体处理和 6 个月延迟雷帕霉素处理的小鼠 Timm 阳性的颗粒细胞层加分子层的百分比均较高且相似。各组经历癫痫持续状态后的颗粒细胞层加分子层的神经细胞丢失程度相似，因此不是混杂因素。与对照相比，0 延迟载体处理的小鼠颗粒细胞层加分子层的平均体积更大。0 延迟雷帕霉素处理的小鼠的肥大部分受到抑制。然而，6 个月延迟载体和 6 个月延迟雷帕霉素处理的动物的颗粒细胞层加分子层的平均体积相似，明显大于其他所有组。
系统性雷帕霉素治疗抑制了癫痫持续状态诱导的苔藓纤维发芽和齿状回肥大，但在治疗停止后又恢复。这些发现表明，在小鼠癫痫持续状态后 >2 个月，驱动苔藓纤维发芽和齿状回肥大的分子信号可能持续存在。因此，可能需要延长或持续治疗才能永久抑制苔藓纤维发芽。
