Mouse genetic studies on the functions of the mammalian catecholaminergic nervous system

(1)儿茶酚胺特异性的遗传改变和肾上腺素能受体亚型的调节表达-产生在DBH启动子控制下表达PNMT的转基因小鼠以切换儿茶酚胺特异性。在转基因小鼠的交感神经元支配的靶组织中，β 2-AR结合位点的数量显著减少，并且β 1-AR结合位点在组织中以不同的方式调节。我们的数据表明，在转基因小鼠中，儿茶酚胺特异性的改变导致靶组织中β-AR亚型的表达受到调节。(2)多巴胺β-羟化酶的高水平表达和对儿茶酚胺代谢的调节-用多拷贝的人DBH小基因构建体产生转基因小鼠以实现DBH的过表达。转基因产物被正确地加工成糖基化的成熟多肽。在转基因小鼠中表达人DBH导致该酶的显著增加 ...更多信息 活性在各种含儿茶酚胺的组织中，但去甲肾上腺素和肾上腺素的稳态水平通常保持无儿茶酚胺周转率的加速，这表明一些调节机制，以保持恒定的去甲肾上腺素合成率，尽管DBH蛋白的量增加。(3)酪氨酸羟化酶突变小鼠中的儿茶酚胺耗竭和围产期致死性-通过基因靶向产生的缺乏TH基因的小鼠在胚胎发育后期或出生后不久显示出致死性。TH突变导致三种儿茶酚胺严重耗竭，但不影响正常产生儿茶酚胺的细胞的总体形态发育。对存活突变体的心电图分析显示心动过缓，表明纯合子小鼠的心脏功能发生了改变。此外，将人TH转基因转移到纯合子小鼠中纠正了突变体表型，表明通过表达人酶恢复了TH活性。我们的研究结果表明，TH是必不可少的动物在妊娠后期的发展和出生后的生存。(4)通过免疫毒素介导的细胞靶向对特定神经元类型的条件性破坏-我们已经开发了一种新的转基因方法，利用免疫毒素的细胞毒性活性消融脑中的诱导选择性神经元。产生在DBH启动子控制下表达人IL-2 Ra的转基因小鼠。用免疫毒素抗Tac（Fv）-PE 40 i. c. v.处理动物，该免疫毒素选择性杀死携带人IL-2 R α的动物细胞。该免疫毒素仅在转基因小鼠中引起特征性行为异常，伴随着含DBH神经元的急剧丧失以及DBH活性和去甲肾上腺素水平的显著降低。这种方法提供了一种通用的技术，通过靶向人类神经退行性疾病的动物模型，

(1) Genetic alteration of catecholamine specificity and regulated expression of adrenergic receptor subtypes-Transgenic mice expressing PNMT under the control of the DBH promoter were generated to switch catecholamine specificity. In the targeted tissues innervated by sympathetic neurons of transgenic mice, the number of beta2-AR binding sites was dramatically decreased, and beta1-AR binding sites were regulated in a different fashion among the tissues. Our data indicate that alteration of catecholamine specificity in transgenic mice leads to regulated expression of the beta-AR subtypes in the target tissues. (2) High level expression of dopamine beta-hydroxylase and regulation of catecholamine metabolism-Transgenic mice were generated with multiple copies of a human DBH minigene construct to achieve overexpression of DBH.The transgene products were correctly processed to a glycosylated mature polypeptide. Expression of human DBH in transgenic mice led to marked increase in the enzyme … More activity in various catecholamine-containing tissues, but the steady-state levels of noradrenaline and adrenaline were normally maintained without the acceleration of the catecholamine turnover rate, suggesting some regulatory mechanisms to preserve a constant rate of noradrenaline synthesis in spite of the increased amount of DBH protein. (3) Catecholamine depletion and perinatal lethality in tyrosine hydroxylase-mutant mice-Mice lacking the TH gene generated by gene targeting showed the lethality at a late stage of embryonic development or shortly after birth. The TH mutation resulted in severe depletion of three kinds of catecholamine, but did not affect gross morphological development of the cells that normally produce catecholamines. Analysis of electrocardiograms of surviving mutants showed bradycardia, suggesting an alteration of cardiac funcitons in the homozygous mice. In addition, tranfer of a human TH transgene into the homozygous mice corrected the mutant phenotype, showing recovery of TH activity by expression of the human enzyme. Our results represent that TH is essential for survival of the animals during the late gestational development and after birth. (4) Conditional disruption of specific neuronal types by immunotoxin-mediated cell targeting-We have developed a novel transgenic approach to ablate inducibly selective neurons in the brain with the cytotoxic activity of immunotoxins. Transgenic mice were created that express the human IL-2Ralpha under the control of the DBH promoter. The animals were treated i.c.v. with an immunotoxin anti-Tac (Fv) -PE40, which selectively kills animal cells bearing human IL-2Ralpha. The immunotoxin caused a characteristic behavioral abnormality only in transgenic mice, which was accompanied by a dramatic loss of DBH-containing neurons and a significant decrease in DBH activity and noradrenaline levels. This approach provides a general technique to create animal models of human neurodegenerative disorders by targetin Less