Gene Delivery and Addiction

重组腺相关病毒（rAAV）载体经常用于向中枢神经系统的基因递送，并且能够转导神经元。 使用表达胶质源性神经营养因子（GDNF）的AAV载体，我们能够与我们的内部合作者Yavin Shaham博士一起参与研究。 我们发现，注射腺相关病毒（AAV）病毒载体含有大鼠GDNF cDNA的可卡因戒断后的第一天增加线索诱导的可卡因寻求停药后第11和31天，这种效果没有观察到使用对照病毒。 完整的研究探讨了GDNF在可卡因渴望孵化中的作用，并于最近发表。 我们部门多年来一直在研究的蛋白质之一是骨形态发生蛋白7，特别是它的神经营养/神经再生特性。 我们以前证明，外源性应用骨形态发生蛋白7（BMP 7）减少6-羟基多巴胺介导的神经退行性疾病的帕金森氏病的啮齿动物模型。我们在过去的一年中发表了一项研究，表明在黑质纹状体通路的多巴胺能神经元中表达BMP受体II的截短形式的转基因小鼠更容易受到甲基苯丙胺的攻击，如通过黑质中的TUNEL标记所确定的。 我们的数据表明，BMP信号的缺陷增加了对高剂量MA诱导的损伤的脆弱性。 在第二项研究中，我们发现甲基苯丙胺抑制了小鼠黑质中BMP 7的表达。 通过黑质网状组织中酪氨酸羟化酶（TH）免疫染色和自发活动的变化，仅携带一个BMP 7等位基因的小鼠更容易受到甲基苯丙胺毒性的影响。 通过小鼠脑侧脑室外源性递送BMP 7蛋白降低甲基苯丙胺毒性，如通过自发活动和TH免疫染色所测量的。 最近，我们正在研究BMP 7神经再生作用的细胞和分子机制，重点是影响细胞外基质（ECM）或更普遍的细胞外环境的能力。 使用原代大鼠神经元，我们正在研究BMP 7影响ECM修饰酶的能力。 总的来说，我们的数据表明，BMP 7和BMP受体信号传导在各种神经退行性变模型中具有神经保护和神经再生作用。 我们最近产生了一种表达人μ阿片受体（huMOR）的AAV载体，并正在评估huMOR在小鼠甲基苯丙胺致敏中的作用。 我们的初步研究结果表明，通过AAV载体在特定脑区域中表达huMOR改变了甲基苯丙胺的致敏性。 这项工作正在进行中。 我们还产生了表达谷氨酸转运蛋白（GLT-1）的AAV载体，并证明其功能。 我们正在使用大鼠中风模型研究GLT-1减少缺血引起的损伤的能力。 此外，我们正在与Roy Wise博士合作，研究GLT-1改变大鼠可卡因寻求行为的能力。 用来调节细胞外谷氨酸的水平 我们已经开始实验检查通过AAV过量GLT-1过表达降低谷氨酸兴奋毒性的能力。 我们也开始研究特定脑区GLT-1过表达对甲基苯丙胺致敏的影响。 这些实验正在进行中。

Recombinant adeno-associated viral (rAAV) vectors are frequently used for gene delivery to the central nervous system and are capable of transducing neurons. Using an AAV vector expressing glial derived neurotrophic factor (GDNF), we were able to contribute to a study with our intramural collaborator, Dr. Yavin Shaham. We show that injections of an adeno-associated virus (AAV) viral vector containing rat GDNF cDNA on the first day after cocaine withdrawal increased cue-induced cocaine-seeking on withdrawal days 11 and 31; this effect was not observed using a control virus. The complete study examines the role of GDNF in the incubation of cocaine craving and has recently been published. One of the proteins that our section has been studying for many years is bone morphogenetic protein 7, specifically, its neurotrophic/neuroregenerative properties. We previously demonstrated that exogenous application of bone morphogenetic protein 7 (BMP7) reduced 6-hydroxydopamine-mediated neurodegeneration in a rodent model of Parkinson's disease. We published a study this past year showing that transgenic mice expressing a truncated form of the BMP receptor II in dopaminergic neurons of the nigrostriatal pathway were more vulnerable to methamphetamine challenge as determined by TUNEL labeling in the substantia nigra. Our data suggest that a deficiency in BMP signaling increases vulnerability to insults induced by high doses of MA. In a second study, we show that methamphetamine suppressed BMP7 expression in the substantia nigra of mice. Mice that carry only one allele of BMP7 were more vulnerable to methamphetamine toxicity as measured by tyrosine hydroxylase (TH) immunostaining in the nigra reticulate and changes in locomotor activity. Exogenous delivery of BMP7 protein via the lateral ventricle of mouse brain reduced methamphetamine toxicity as measured by locomotor activity and TH immunostaining. Recently, we are examining the cellular and molecular mechanisms of BMP7 neuroregenerative effects by focusing on the ability to influence the extracellular matrix (ECM) or more generally the extracellular environment. Using a primary rat neurons, we are examining BMP7's ability to influence ECM-modifiying enzymes. Collectively, our data demostrate that BMP7 and BMP receptor signaling have neuroprotective and neuroregenerative effects in various models of neurodegeneration. We recently generated an AAV vector expressing the human mu opioid receptor (huMOR) and are evaluating the role of huMOR in methamphetamine sensitization in mice. Our preliminary findings show that huMOR expression by an AAV vector in specific brain regions alters methamphetamine sensitization. This work is ongoing. We have also generated an AAV vector expressing the glutamate transporter (GLT-1) and demonstrated that it functional. We have ongoing work examining the ability of GLT-1 to reduce damage caused by ischemia using a rat model of stroke. Additionally, we are collaborating with Dr. Roy Wise to examine the ability of GLT-1 to alter cocaine seeking behavior in rats. was created to modulated the levels of extracellular glutamate. We have begun experiments examining the ability of excess GLT-1 overexpression by AAV to reduce excitoxicity by glutamate. We have also begun examining the GLT-1 overexpression in specfic brain regions for alterations to methamphetamine sensitization. These experiments are ongoing.