Jackson Joshua G, Robinson Michael B
Children's Hospital of Philadelphia Research Institute, and Departments of Pediatrics and.
Children's Hospital of Philadelphia Research Institute, and Departments of Pediatrics and Systems Pharmacology and Experimental Therapeutics, University of Pennsylvania, Pennsylvania 19104
J Neurosci. 2015 Nov 11;35(45):15199-213. doi: 10.1523/JNEUROSCI.2049-15.2015.
We recently showed that inhibition of neuronal activity, glutamate uptake, or reversed-Na(+)/Ca(2+)-exchange with TTX, TFB-TBOA, or YM-244769, respectively, increases mitochondrial mobility in astrocytic processes. In the present study, we examined the interrelationships between mitochondrial mobility and Ca(2+) signaling in astrocyte processes in organotypic cultures of rat hippocampus. All of the treatments that increase mitochondrial mobility decreased basal Ca(2+). As recently reported, we observed spontaneous Ca(2+) spikes with half-lives of ∼1 s that spread ∼6 μm and are almost abolished by a TRPA1 channel antagonist. Virtually all of these Ca(2+) spikes overlap mitochondria (98%), and 62% of mitochondria are overlapped by these spikes. Although tetrodotoxin, TFB-TBOA, or YM-244769 increased Ca(2+) signaling, the specific effects on peak, decay time, and/or frequency were different. To more specifically manipulate mitochondrial mobility, we explored the effects of Miro motor adaptor proteins. We show that Miro1 and Miro2 are both expressed in astrocytes and that exogenous expression of Ca(2+)-insensitive Miro mutants (KK) nearly doubles the percentage of mobile mitochondria. Expression of Miro1(KK) had a modest effect on the frequency of these Ca(2+) spikes but nearly doubled the decay half-life. The mitochondrial proton ionophore, FCCP, caused a large, prolonged increase in cytosolic Ca(2+) followed by an increase in the decay time and the spread of the spontaneous Ca(2+) spikes. Photo-ablation of mitochondria in individual astrocyte processes has similar effects on Ca(2+). Together, these studies show that Ca(2+) regulates mitochondrial mobility, and mitochondria in turn regulate Ca(2+) signals in astrocyte processes.
In neurons, the movement and positioning of mitochondria at sites of elevated activity are important for matching local energy and Ca(2+) buffering capacity. Previously, we demonstrated that mitochondria are immobilized in astrocytes in response to neuronal activity and glutamate uptake. Here, we demonstrate a mechanism by which mitochondria are immobilized in astrocytes subsequent to increases in intracellular [Ca(2+)] and provide evidence that mitochondria contribute to the compartmentalization of spontaneous Ca(2+) signals in astrocyte processes. Immobilization of mitochondria at sites of glutamate uptake in astrocyte processes provides a mechanism to coordinate increases in activity with increases in mitochondrial metabolism.
我们最近发现，分别用河豚毒素（TTX）、TFB - TBOA或YM - 244769抑制神经元活动、谷氨酸摄取或反向钠/钙交换，可增加星形胶质细胞突起中线粒体的移动性。在本研究中，我们检测了大鼠海马器官型培养物中星形胶质细胞突起中线粒体移动性与钙信号之间的相互关系。所有增加线粒体移动性的处理均降低了基础钙水平。正如最近报道的那样，我们观察到自发钙峰，其半衰期约为1秒，传播距离约为6μm，并且几乎被TRPA1通道拮抗剂消除。实际上，所有这些钙峰都与线粒体重叠（98%），62%的线粒体被这些钙峰重叠。尽管河豚毒素、TFB - TBOA或YM - 244769增加了钙信号，但对峰值、衰减时间和/或频率的具体影响有所不同。为了更具体地操纵线粒体移动性，我们研究了米罗（Miro）运动衔接蛋白的作用。我们发现Miro1和Miro2均在星形胶质细胞中表达，并且钙不敏感的Miro突变体（KK）的外源性表达使移动线粒体的百分比几乎增加了一倍。Miro1（KK）的表达对这些钙峰的频率有适度影响，但使衰减半衰期几乎增加了一倍。线粒体质子离子载体羰基氰化物4 - （三氟甲氧基）苯腙（FCCP）导致胞质钙大幅、持续增加，随后自发钙峰的衰减时间和传播增加。对单个星形胶质细胞突起中的线粒体进行光消融对钙有类似影响。总之，这些研究表明钙调节线粒体移动性，而线粒体反过来调节星形胶质细胞突起中的钙信号。
在神经元中，线粒体在高活性位点的移动和定位对于匹配局部能量和钙缓冲能力很重要。此前，我们证明线粒体在星形胶质细胞中因神经元活动和谷氨酸摄取而固定。在这里，我们展示了一种机制，通过该机制线粒体在细胞内钙浓度升高后在星形胶质细胞中固定，并提供证据表明线粒体有助于星形胶质细胞突起中自发钙信号的区室化。线粒体在星形胶质细胞突起中谷氨酸摄取位点的固定提供了一种机制，以协调活动增加与线粒体代谢增加。
