Methamphetamine, mitochondria, and neurodegeneration

摘要 甲基苯丙胺(冰毒)是一种上瘾的精神刺激剂。我们的初步研究表明，慢性冰毒导致 黑质致密部(SNC)、多巴胺(DA)和蓝斑(LC)去甲肾上腺素的变性 (NE)但不包括腹侧被盖区(VTA)DA或中缝背侧(DR)5-羟色胺(5-HT)神经元。内华达州 和5-羟色胺是合成的单胺类神经递质，并被包装成轴突间隔中的囊泡。 METH增加SNC轴突的胞浆单胺和MAO依赖的线粒体应激，但不增加胞体。 初步研究表明，MAO抑制可防止冰毒诱导的SNC退化。这些数据表明 退变是由MAO依赖的轴突线粒体应激所致。然而，试点数据显示，冰毒也 Lc、VTA和DR轴突线粒体应激增加但慢性冰毒引起的SNC和LC但不能引起VTA或 博士退化症。我们在SNC中发现了L型钙通道(LCC)依赖的线粒体应激，但没有发现VTA 轴突。我们提出了两次打击假说，其中MAO和LCC依赖的轴突线粒体应激 是退化所必需的。变性的“触发器”是冰毒诱导的MAO依赖的轴突线粒体 应激和易损性是依赖于LCC的轴突线粒体应激的结果。因为毛-和LCC- 依赖应力在轴突中汇聚，我们提出轴突丢失先于躯体丢失的“死亡-回归”模式 堕落。实验数据表明，戒断过程中黑质核轴突线粒体应激增加。我们 假设这种压力是MAO依赖的，并在禁欲期间导致退化。最后，冰毒- 诱发的退化预计会损害新的物体识别和恐惧条件反射，它们是依赖的 分别在DA和NE上。假说将在大脑中使用双光子激光扫描显微镜进行验证 切片、遗传编码的生物传感器、MAO/LCC的药理和遗传操作，以及 免疫组织化学。目的1：确定单胺能轴突线粒体应激的机制， 慢性冰毒对线粒体的影响，以及线粒体应激在 禁欲。我们假设SNC、VTA、LC和DR轴突都有冰毒诱导的MAO依赖 而SNC和LC，而不是VTA或DR轴突，会产生LCC诱导的线粒体应激。 我们还假设，在慢性冰毒戒断过程中，轴突压力将保持较高水平，并将 依赖毛。目的2：确定MAO和LCCs在慢性冰毒诱导的变性中的作用 以及行为后果。我们假设从药物上抑制或从基因上删除MAO 或LCC将防止慢性冰毒诱导的易受伤害(SNC和LC)神经元的退化，并删除 来自抵抗(VTA和DR)神经元的CA2缓冲蛋白Calbindin将使它们容易受到 退化。我们进一步假设，在禁欲期间，堕落将是渐进的，毛- 依附的。最后，我们假设SNC和LC变性会导致认知障碍。

ABSTRACT Methamphetamine (meth) is an addictive psychostimulant. Our pilot studies show that chronic meth resulted in degeneration of substantia nigra pars compacta (SNc) dopamine (DA) and locus coeruleus (LC) norepinephrine (NE) but not ventral tegmental area (VTA) DA or dorsal raphe (DR) serotonin (5-HT) neurons in mice. DA, NE, and 5-HT are monoamine neurotransmitters synthesized and packaged into vesicles in axonal compartments. Meth increases cytosolic monoamines and MAO-dependent mitochondrial stress in SNc axons but not the soma. Pilot studies show that MAO inhibition prevented meth-induced SNc degeneration. These data suggest that degeneration is driven by MAO-dependent axonal mitochondrial stress. However, pilot data show meth also increased LC, VTA, and DR axonal mitochondrial stress yet chronic meth induced SNc and LC but not VTA or DR degeneration. We found L-type Ca2+ channel (LCC)-dependent mitochondrial stress in SNc but not VTA axons. We propose a `two-hit' hypothesis wherein both MAO- and LCC-dependent axonal mitochondrial stress are required for degeneration. The degeneration `trigger' is meth-induced MAO-dependent axonal mitochondrial stress and vulnerability is a consequence of axonal LCC-dependent mitochondrial stress. Since MAO- and LCC- dependent stress converge in axons, we propose that axonal loss precedes somatic loss in a `dying-back' pattern of degeneration. Pilot data indicate that SNc axonal mitochondrial stress was increased during abstinence. We hypothesize this stress is MAO-dependent and contributes to degeneration during abstinence. Lastly, meth- induced degeneration is expected to impair novel object recognition and fear conditioning which are dependent on DA and NE, respectively. Hypotheses will be tested using two-photon laser scanning microscopy in brain slices, genetically encoded biosensors, pharmacological and genetic manipulation of MAO/LCCs, and immunohistochemistry. Aim 1: Determine mechanisms of mitochondrial stress in monoaminergic axons, consequences of chronic meth on mitochondria, and whether mitochondrial stress continues during abstinence. We hypothesize that SNc, VTA, LC, and DR axons all have meth-induced MAO-dependent mitochondrial stress whereas SNc and LC but not VTA or DR axons will have LCC-induced mitochondrial stress. We also hypothesize that axonal stress will remain elevated throughout abstinence from chronic meth and will be MAO-dependent. Aim 2: Determine the roles of MAO and LCCs in chronic meth-induced degeneration and behavioral consequences. We hypothesize that pharmacologically inhibiting or genetically deleting MAO or LCCs will prevent chronic meth-induced degeneration of vulnerable (SNc and LC) neurons and deleting the Ca2+ buffering protein calbindin from resistant (VTA and DR) neurons will render them vulnerable to degeneration. We further hypothesize that degeneration will be progressive throughout abstinence and MAO- dependent. Lastly, we hypothesize that SNc and LC degeneration will result in cognitive impairments.