Investigation of Mitochondrial Function in Bipolar Disorder

锂和丙戊酸盐（VPA）是双相情感障碍的常见情绪稳定剂，并显示出间接调节细胞内途径，使用分子如cAMP反应元件结合蛋白（CREB），脑源性神经营养因子（BDNF），Bcl-2和丝裂原活化蛋白激酶（MAPK），其活性可能产生一些情绪稳定药物的有益作用。具体而言，锂和丙戊酸盐在体内和体外上调Bcl-2，通过阻断线粒体凋亡程序防止神经元细胞死亡。越来越多的证据表明，线粒体功能障碍在各种神经退行性疾病和精神疾病的进展中起着关键作用。因此，增强线粒体功能可能有助于改善与各种神经精神疾病相关的神经可塑性和细胞弹性的损伤。本研究旨在探讨情绪稳定剂对线粒体功能的影响及对神经毒性的影响。我们发现，锂和VPA的长期处理增强了细胞呼吸速率。此外，锂或VPA的慢性处理增强了线粒体功能，如通过SH-SY 5 Y细胞中的线粒体膜电位和线粒体氧化所确定的。体内研究表明，锂或VPA长期治疗可在线粒体水平上防止甲基苯丙胺诱导的毒性。此外，这些药物防止甲基苯丙胺诱导的线粒体细胞色素c的减少，线粒体抗凋亡Bcl-2/Bax的比例，和线粒体细胞色素氧化酶（考克斯）的活动。DNA微阵列分析表明，与凋亡途径和线粒体功能相关的几种蛋白质的基因表达水平被甲基苯丙胺改变，这些变化被锂或VPA处理减弱。其中一个基因Bcl-2是锂和VPA的共同靶点。用特异性Bcl-2 siRNA敲低Bcl-2降低了锂和VPA诱导的线粒体氧化增加。这些发现表明，锂和VPA增强线粒体功能，并防止线粒体介导的毒性。这些药物可能在治疗与线粒体功能受损相关的其他疾病，如神经退行性疾病和精神分裂症中具有潜在的临床实用性。 此外，双相情感障碍中的钙功能障碍被认为是该疾病中描述的最可重复的生物学异常，最近的数据支持双相情感障碍个体线粒体和内质网（ER）中钙调节异常的作用。最近的人类遗传数据显示，Bcl-2 geners 956572（变体AA）中的单核苷酸多态性（SNP）是双相情感障碍的风险等位基因，其特征还在于低Bcl-2水平和等位基因携带者中细胞凋亡发生率增加。由于Bcl-2通过其对ER和线粒体的作用调节细胞内Ca++信号传导，我们已经研究了Bcl-2在双相情感障碍患者的淋巴母细胞中的细胞内Ca++调节中的功能作用，这些患者呈现该Bcl-2 SNP的三种变体（AA、AG或GG）。我们发现，Bcl-2缺陷的变体AA提出了升高的基础胞质Ca++水平，虽然有没有定量差异之间的SNP变体在管腔ER和线粒体钙浓度，也不是在Ca++进入。然而，在变体AA中特异性观察到1，4，5-三磷酸肌醇（InsP 3）介导的Ca++动员后细胞溶质Ca++水平增加，同时ER钙池减少增强，并在变体GG中被Bcl-2抑制剂模拟，表明Bcl-2水平降低与通过InsP 3受体增强的ER Ca++释放相关。Bcl-2和InsP 3受体之间的功能相互作用似乎与双相情感障碍的治疗相关，因为长期给予携带Bcl-2风险等位基因AA的人细胞锂诱导异常高的基础胞质Ca++水平和InsP 3介导的ER Ca++释放程度降低。结果表明，双相情感障碍中Bcl-2 SNP的变化通过调节InsP 3依赖的ER Ca++释放促进了钙信号的紊乱。

Lithium and valproate (VPA) are common mood stabilizers for bipolar disorder and are shown to indirectly regulate intracellular pathways, using molecules like cAMP response element-binding protein (CREB), brain derived neurotrophic factor (BDNF), Bcl-2, and mitogen-activated protein kinases (MAPK), whose activities may produce some of the beneficial effects of the mood stabilizing drugs. Specifically, lithium and valproate upregulate Bcl-2 in vivo and in vitro, preventing neuronal cell death through blocking the mitochondrial program for apoptosis. Accumulating evidence suggests that mitochondrial dysfunction plays a critical role in the progression of a variety of neurodegenerative and psychiatric disorders. Thus, enhancing mitochondrial function could potentially help ameliorate the impairments of neural plasticity and cellular resilience associated with a variety of neuropsychiatric disorders. A series of studies was undertaken to investigate the effects of mood stabilizers on mitochondrial function and mitochondria-mediated neurotoxicity. We found that long-term treatment with lithium and VPA enhanced cell respiration rate. Furthermore, chronic treatment with lithium or VPA enhanced mitochondrial function as determined by mitochondrial membrane potential and mitochondrial oxidation in SH-SY5Y cells. In vivo studies showed that long-term treatment with lithium or VPA protected against methamphetamine-induced toxicity at the mitochondrial level. Furthermore, these agents prevented the methamphetamine-induced reduction of mitochondrial cytochrome c, the mitochondrial anti-apoptotic Bcl-2/Bax ratio, and mitochondrial cytochrome oxidase (COX) activity. DNA microarray analysis demonstrated that the gene expression levels of several proteins related to the apoptotic pathway and mitochondrial functions were altered by methamphetamine, and these changes were attenuated by treatment with lithium or VPA. One of the genes, Bcl-2, is a common target for lithium and VPA. Knockdown of Bcl-2 with specific Bcl-2 siRNA reduced the lithium- and VPA-induced increases in mitochondrial oxidation. These findings illustrate that lithium and VPA enhance mitochondrial function and protect against mitochondria-mediated toxicity. These agents may have potential clinical utility in the treatment of other diseases associated with impaired mitochondrial function, such as neurodegenerative diseases and schizophrenia. In addition, calcium dysfunction in bipolar disorder is considered the most reproducible biological abnormality described in the illness, and recent data support a role for calcium dysregulation in the mitochondrial and endoplasmic reticulum (ER) of individuals with bipolar disorder. Recent human genetic data show that a single nucleotide polymorphism (SNP) in the Bcl-2 geners956572 (variant AA) is a risk allele for bipolar disorder that is also characterized by low Bcl-2 levels and an increased incidence of apoptosis in carriers of the allele. Since Bcl-2 regulates intracellular Ca++ signaling via its effects on the ER and mitochondria, we have examined the functional role of Bcl-2 in intracellular Ca++ regulation in lymphoblasts from bipolar disorder patients presenting three variants (AA, AG or GG) of this Bcl-2 SNP. We found that the Bcl-2-deficient variant AA presents elevated basal cytosolic Ca++ levels, although there were no quantitative differences between SNP variants in luminal ER and mitochondrial calcium concentrations, nor in Ca++ entry. However, increased cytosolic Ca++ levels after inositol 1, 4, 5-trisphosphate (InsP3)-mediated Ca++ mobilization, in parallel with enhanced reduction of the ER calcium pool, were observed specifically in variant AA and mimicked by Bcl-2 inhibitors in variant GG, showing that decreased Bcl-2 levels are associated with enhanced ER Ca++ release via InsP3 receptors. Functional interaction between Bcl-2 and InsP3 receptors appears relevant to the therapeutics of bipolar disorder, since chronic lithium given to human cells carrying the Bcl-2 risk allele AA induced a decrease in the abnormally high basal cytosolic Ca++ levels and in the extent of InsP3-mediated ER Ca++ release. The results demonstrate that Bcl-2 SNP variations in bipolar disorder facilitate deranged calcium signaling through modulation of InsP3-dependent ER Ca++ release.