树鼩脑缺血时神经元线粒体渗透性转导孔开放对线粒体呼吸的影响

目的：揭示光化学诱导树鼩脑缺血后不同时间内，缺血神经元线粒体呼吸功能的变化，观察血小板活化因子（PAF）受体拮抗剂银杏内酯B（GB）和免疫抑制剂环孢菌素A （CsA）对线粒体呼吸及神经元线粒体渗透性转导孔（MPT）开放的影响，探讨二者可能的神经保护机制及缺血时MPT开放与线粒体呼吸的相互关系。方法：建立光化学诱导树鼩脑缺血模型，分离缺血后4、24、72 h大脑皮层线粒体，用氧电极极谱法测定线粒体呼吸。于缺血6 h分别注射GB和CsA，24 h时观察相关指标。另取分离的线粒体，用CaCl₂诱导MPT开放，再分别加入CsA或GB，观察其对线粒体肿胀的影响。结果：缺血脑皮层神经元线粒体Ⅲ态呼吸速度、呼吸控制率（RCR）及磷氧比（P/O）逐渐下降，以缺血24 h的变化为著，与假手术组相比均有显著差异（P<0.01）。给予GB或CsA，Ⅲ态呼吸速度显著大于对照组（P<0.01），P/O比上升（P<0.05）。CaCl₂可诱导MPT开放，线粒体肿胀而透光率迅速下降，CsA有效阻滞了MPT开放，GB则不然。结论：光化学诱导树鼩局灶脑缺血后，缺血皮层线粒体呼吸明显障碍，GB或CsA可不同程度地改善缺血神经元线粒体代谢，可能与GB拮抗神经细胞膜上PAF受体活化，间接调节线粒体呼吸有关；而CsA主要通过抑制MPT开放而改善线粒体呼吸而具有神经保护作用。

The opening of neuronal mitochondria permeability transition pore affects respiratory function during thrombotic cerebral ischemia in tree shrews

AIM: The present study explored those changes in brain mitochondrial respiration in different times after thrombotic cerebral ischemia induced by photochemical reaction in primate's animal tree shrew and observed the effects of platelet-activating factor (PAF) receptor antagonist ginkgolide B (GB) and immunosuppressor cyclosporin A (CsA) on neuronal mitochondrial respiration and mitochondrial permeability transition pore(MPT) in twenty-four hours after occlusion and showed their neuronprotective mechanism and expatiate the relationship between mitochondrial respiration and MPT. METHODS: The focal thrombotic cerebral ischemia was formed by photochemistry-induced technology in tree shrews. At 4,24,72 h after focal cerebral ischemia, the neuronal mitochondria were centrifuged. Clark oxygen electrode was used to measure the changes in neuronal mitochondrial respiration.Pretreatment for experimental animals with GB and CsA at 6h after occlusion, we centrifuged the mitochondria and measured the changes in neuronal mitochondrial respiration at 24 h after occlusion. In addition, experiments were performed in a flurometer by measuring CaCl₂ 100 μmol/L induced centrifuged mitochondrial swelling and GB or CsA were added at the same time. RESULTS: All of respiration state Ⅲ, RCR and P/O decreased after cerebral ischemia. There were significant differences between every ischemic group and sham, especially at 24 h (P<0.01). Pretreatment for experimental animals with GB and CsA,the state Ⅲ and P/O recovered after ischemia for 24 h (P<0.01,P<0.05). CaCl₂ induced the opening of MPT and the light scattering decreased. When CsA or GB were added at the initiation of each experiment, studies on the swelling kinetics of isolated brain mitochondria showed the CsA but GB, inhibited calcium ions induced mitochondria swelling. CONCLUSIONS: Neuronal mitochondrial respiration and metabolism were inhibited after thrombotic cerebral ischemia induced by photochemical reaction. GB and CsA were shown to improve neuronal mitochondrial respiration. Probably, the neuronprotection of GB connects with it's antagonizing the activation of PAFreceptor in neuronal membrane and indirectly regulated the mitochondria respiration. Moreover, CsA improved neuronal mitochondrial respiration through inhibiting the opening of MPT.