Brain and Mind Research Institute and Department of Neurology, Weill Medical College of Cornell University, New York, New York 10065, and Department of Neuroscience and Sackler School of Biomedical Sciences, Tufts University, Boston, Massachusetts 02111.
J Neurosci. 2014 Feb 5;34(6):2331-48. doi: 10.1523/JNEUROSCI.2689-13.2014.
Motor neurons are progressively and predominantly degenerated in ALS, which is not only induced by multiple intrinsic pathways but also significantly influenced by the neighboring glial cells. In particular, astrocytes derived from the SOD1 mutant mouse model of ALS or from human familial or sporadic ALS patient brain tissue directly induce motor neuron death in culture; however, the mechanisms of pathological astroglial secretion remain unclear. Here we investigated abnormal calcium homeostasis and altered exocytosis in SOD1G93A astrocytes. We found that purinergic stimulation induces excess calcium release from the ER stores in SOD1G93A astrocytes, which results from the abnormal ER calcium accumulation and is independent of clearance mechanisms. Furthermore, pharmacological studies suggested that store-operated calcium entry (SOCE), a calcium refilling mechanism responsive to ER calcium depletion, is enhanced in SOD1G93A astrocytes. We found that oxidant-induced increased S-glutathionylation and calcium-independent puncta formation of the ER calcium sensor STIM1 underlies the abnormal SOCE response in SOD1G93A astrocytes. Enhanced SOCE contributes to ER calcium overload in SOD1G93A astrocytes and excess calcium release from the ER during ATP stimulation. In addition, ER calcium release induces elevated ATP release from SOD1G93A astrocytes, which can be inhibited by the overexpression of dominant-negative SNARE. Selective inhibition of exocytosis in SOD1G93A astrocytes significantly prevents astrocyte-mediated toxicity to motor neurons and delays disease onset in SOD1G93A mice. Our results characterize a novel mechanism responsible for calcium dysregulation in SOD1G93A astrocytes and provide the first in vivo evidence that astrocyte exocytosis contributes to the pathogenesis of ALS.
运动神经元在 ALS 中逐渐且主要退化，这不仅是由多种内在途径引起的，而且还受到邻近神经胶质细胞的显著影响。特别是，源自 ALS 突变 SOD1 小鼠模型或源自人类家族性或散发性 ALS 患者脑组织的星形胶质细胞在培养物中直接诱导运动神经元死亡；然而，病理性星形胶质细胞分泌的机制仍不清楚。在这里，我们研究了 SOD1G93A 星形胶质细胞中异常的钙稳态和改变的胞吐作用。我们发现嘌呤能刺激诱导 SOD1G93A 星形胶质细胞中内质网（ER）库的过量钙释放，这是由于 ER 钙积累异常，并且独立于清除机制。此外，药理学研究表明，储存操纵的钙内流（SOCE）是一种响应 ER 钙耗竭的钙填充机制，在 SOD1G93A 星形胶质细胞中增强。我们发现，氧化应激诱导的 ER 钙传感器 STIM1 的 S-谷胱甘肽化增加和钙独立性斑点形成是 SOD1G93A 星形胶质细胞中异常 SOCE 反应的基础。增强的 SOCE 导致 SOD1G93A 星形胶质细胞中 ER 钙超载和 ATP 刺激时 ER 钙释放增加。此外，ER 钙释放诱导 SOD1G93A 星形胶质细胞中 ATP 释放增加，这可以通过过表达显性负 SNARE 来抑制。在 SOD1G93A 星形胶质细胞中选择性抑制胞吐作用可显著防止星形胶质细胞介导的运动神经元毒性，并延迟 SOD1G93A 小鼠疾病的发作。我们的结果描述了 SOD1G93A 星形胶质细胞中钙失调的新机制，并提供了体内证据表明星形胶质细胞胞吐作用有助于 ALS 的发病机制。
