Gulbronson Chloe I, Jahanian Sepideh, Gransee Heather M, Sieck Gary C, Mantilla Carlos B
Department of Anesthesiology & Perioperative Medicine, Mayo Clinic, 200 First Street SW, Rochester, MN 55905, USA.
Department of Physiology & Biomedical Engineering, Mayo Clinic, 200 First Street SW, Rochester, MN 55905, USA.
Cells. 2025 Mar 7;14(6):390. doi: 10.3390/cells14060390.
Autophagy impairments have been implicated in various aging conditions. Previous studies in cervical motor neurons show an age-dependent increase in the key autophagy proteins LC3 and p62, reflecting autophagy impairment and autophagosome accumulation. Chloroquine is commonly used to inhibit autophagy by preventing autophagosome-lysosome fusion and may thus emulate the effects of aging on the neuromuscular system. Indeed, acute chloroquine administration in old mice decreases maximal transdiaphragmatic pressure generation, consistent with aging effects. We hypothesized that chloroquine alters diaphragm muscle neuromuscular junction (NMJ) morphology and increases denervation. Adult male and female C57BL/6 × 129J mice between 5 and 8 months of age were used to examine diaphragm muscle NMJ morphology and denervation following daily intraperitoneal injections of chloroquine (10 mg/kg/d) or vehicle for 7 days. The motor end-plates and pre-synaptic terminals were fluorescently labeled with α-bungarotoxin and anti-synaptophysin, respectively. Confocal microscopy was used to assess pre- and post-synaptic morphology and denervation. At diaphragm NMJs, chloroquine treatment decreased pre-synaptic volume by 12% compared to the vehicle ( < 0.05), with no change in post-synaptic volume. Chloroquine treatment increased the proportion of partially denervated NMJs by 2.7-fold compared to vehicle treatment ( < 0.05). The morphological changes observed were similar to those previously reported in the diaphragm muscles of 18-month-old mice. These findings highlight the importance of autophagy in the maintenance of the structural properties at adult NMJs in vivo.
自噬功能障碍与多种衰老状况有关。先前对颈运动神经元的研究表明，关键自噬蛋白LC3和p62随年龄增长而增加，这反映了自噬功能障碍和自噬体积累。氯喹通常用于通过阻止自噬体与溶酶体融合来抑制自噬，因此可能模拟衰老对神经肌肉系统的影响。事实上，对老年小鼠急性给予氯喹会降低最大跨膈压的产生，这与衰老效应一致。我们假设氯喹会改变膈肌神经肌肉接头（NMJ）的形态并增加去神经支配。使用5至8个月大的成年雄性和雌性C57BL/6×129J小鼠，每日腹腔注射氯喹（10 mg/kg/d）或赋形剂7天，以检查膈肌NMJ的形态和去神经支配情况。运动终板和突触前末端分别用α-银环蛇毒素和抗突触素进行荧光标记。共聚焦显微镜用于评估突触前和突触后的形态以及去神经支配情况。在膈肌NMJ处，与赋形剂相比，氯喹处理使突触前体积减少了12%（P<0.05），而突触后体积没有变化。与赋形剂处理相比，氯喹处理使部分去神经支配的NMJ比例增加了2.7倍（P<0.05）。观察到的形态学变化与先前在18个月大的小鼠膈肌中报道的相似。这些发现突出了自噬在体内维持成年NMJ结构特性中的重要性。
