Adenovirus-mediated gene delivery to hypothalamic magnocellular neurons in mice.
Adenovirus-mediated gene delivery to hypothalamic magnocellular neurons in mice.
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腺病毒介导的基因递送至小鼠下丘脑大细胞神经元。
DOI：
10.1161/01.hyp.34.4.756
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发表时间：
1999
期刊：
影响因子：
--
通讯作者：
Johnson,AK
中科院分区：
文献类型：
--
作者：
Vasquez,EC;Beltz,TG;Meyrelles,SS;Johnson,AK
-加压素是由下丘脑视上核(SON)和室旁核(PVN)的大细胞神经元合成的，并由其在神经垂体(NH)的轴突终末释放。加压素作为一种抗利尿激素和血管活性物质，在控制体液和心血管内环境稳定方面发挥着重要作用。由于其明确的神经生物学和功能重要性，SON/PVN-NH系统是建立遗传物质基因转移到小鼠中枢神经系统特定途径的理想方法。在这些研究中，我们使用复制缺陷型腺病毒(Ad)载体在成年小鼠中比较了编码β-半乳糖苷酶(β-GAL)的LacZ基因和编码绿色荧光蛋白的基因LacZ的效率。在分离的培养细胞中，当NH、PVN和SON的病毒浓度达到每张盖片上2×107个空斑形成单位时，可获得有效的转染率，而不会产生细胞毒性。然而，随着时间的延长，与绿色荧光蛋白(5%至50%的细胞)相比，检测到更高水平的β-GAL表达(50%至75%的细胞)。采用立体定位的方法，在小鼠脑垂体腺注射4×106个空斑形成单位。在这些动物的材料中，我们能够观察到β-Gal基因在NH以及下丘脑室旁核和SON的大细胞神经元中的表达。这些实验结果表明，Ad-Rous肉瘤病毒启动子-β-Gal被注射部位(NH)的神经末梢摄取，并逆行运输到投射到NH的神经元胞体。我们的结论是，这些实验方法的应用将为生理学研究提供强大的工具，并为治疗疾病提供潜在的治疗基因。
—Vasopressin is synthesized by magnocellular neurons in supraoptic (SON) and paraventricular (PVN) hypothalamic nuclei and released by their axon terminals in the neurohypophysis (NH). With its actions as an antidiuretic hormone and vasoactive agent, vasopressin plays a pivotal role in the control of body fluids and cardiovascular homeostasis. Because of its well-defined neurobiology and functional importance, the SON/PVN-NH system is ideal to establish methods for gene transfer of genetic material into specific pathways in the mouse central nervous system. In these studies, we compared the efficiency of transferring the gene lacZ, encoding for β-galactosidase (β-gal), versus a gene encoding for green fluorescent protein by using replication-deficient adenovirus (Ad) vectors in adult mice. Transfection with viral concentrations up to 2×107plaque-forming units per coverslip of NH, PVN, and SON in dissociated, cultured cells caused efficient transfection without cytotoxicity. However, over an extended period of time, higher levels (50% to 75% of the cells) of β-gal expression were detected in comparison with green fluorescent protein (5% to 50% of the cells). With the use of a stereotaxic approach, the pituitary glands of mice were injected with Ad (4×106plaque-forming units). In material from these animals, we were able to visualize the expression of the β-gal gene in the NH and in magnocellular neurons of both the PVN and SON. The results of these experiments indicate that Ad–Rous sarcoma virus promoter–β-gal is taken up by nerve terminals at the injection site (NH) and retrogradely transported to the soma of the neurons projecting to the NH. We conclude that the application of these experimental approaches will provide powerful tools for physiological studies and potential approaches to deliver therapeutic genes to treat diseases.