Dopamine (DA)-producing neurons in the ventral midbrain are generated from a specified neuronal lineage and form selective axonal pathways that mediate multiple CNS functions. Expression of the gene encoding tyrosine hydroxylase (TH), which is a key enzyme of catecholamine biosynthesis, is regulated during the development of midbrain DA neurons. In the present study, we report the developmental regulation and cell type specificity of TH gene promoter in the ventral midbrain by using a green fluorescent protein (GFP) reporter system. Transgenic mice were generated that express GFP in the majority of midbrain DA neurons under the control of the 9-kb upstream region of the rat TH gene. At an early embryonic stage, GFP expression was induced in the developing DA neurons, and the expression was then markedly down-regulated at later embryonic stages. However, the expression was reactivated and approached the adult levels during early postnatal development. These developmental changes in GFP expression patterns suggest the presence of multistep regulatory mechanisms for TH gene expression during DA neuron development. The TH promoter appears to possess transcriptional elements at least necessary for the induction of TH expression at the early embryonic stage and its reactivation during the post-natal development.

Na(v)2/NaG is a putative sodium channel, whose physiological role has long been an enigma. We generated Na(v)2 gene-deficient mice by inserting the lacZ gene. Analysis of the targeted mice allowed us to identify Na(v)2-producing cells by examining the lacZ expression. Besides in the lung, heart, dorsal root ganglia, and Schwann cells in the peripheral nervous system, Na(v)2 was expressed in neurons and ependymal cells in restricted areas of the CNS, particularly in the circumventricular organs, which are involved in body-fluid homeostasis. Under water-depleted conditions, c-fos expression was markedly elevated in neurons in the subfornical organ and organum vasculosum laminae terminalis compared with wild-type animals, suggesting a hyperactive state in the Na(v)2-null mice. Moreover, the null mutants showed abnormal intakes of hypertonic saline under both water- and salt-depleted conditions. These findings suggest that the Na(v)2 channel plays an important role in the central sensing of body-fluid sodium level and regulation of salt intake behavior.

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antisense oligodeoxynucleotide ͉ c-fos knockout mice ͉ amygdala ͉ conditioned taste aversion ͉ zif268

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Dopamine (DA) exerts synaptic organization of basal ganglia circuitry through a variety of neuronal populations in the striatum. We performed conditional ablation of striatal neuronal types containing DA D2 receptor (D2R) by using immunotoxin-mediated cell targeting. Mutant mice were generated that express the human interleukin-2 receptor alpha-subunit under the control of the D2R gene. Intrastriatal immunotoxin treatment of the mutants eliminated the majority of the striatopallidal medium spiny neurons and cholinergic interneurons. The elimination of these neurons caused hyperactivity of spontaneous movement and reduced motor activation in response to DA stimulation. The elimination also induced upregulation of GAD gene expression in the globus pallidus (GP) and downregulation of cytochrome oxidase activity in the subthalamic nucleus (STN), whereas it attenuated DA-induced expression of the immediate-early genes (IEGs) in the striatonigral neurons. In addition, chemical lesion of cholinergic interneurons did not alter spontaneous movement but caused a moderate enhancement in DA-induced motor activation. This enhancement of the behavior was accompanied by an increase in the IEG expression in the striatonigral neurons. These data suggest that ablation of the striatopallidal neurons causes spontaneous hyperactivity through modulation of the GP and STN activity and that the ablation leads to the reduction in DA-induced behavior at least partly through attenuation of the striatonigral activity as opposed to the influence of cholinergic cell lesion. We propose a possible model in which the striatopallidal neurons dually regulate motor behavior dependent on the state of DA transmission through coordination of the basal ganglia circuitry.

The primate lentiviral vector system based on human immunodeficiency virus type 1 (HIV-1) has been used for a wide range of gene therapy trials in animal models. Axonal transport in the retrograde direction, which is observed with some viral vectors, confers a considerable advantage to gene transfer into neuronal cell bodies that are localized in regions remote from the injection site of the vectors. However, retrograde axonal transport of the HIV-1-based lentiviral vector pseudotyped with vesicular stomatitis virus glycoprotein is reported to be inefficient. In the present study, we developed an efficient gene transfer system through retrograde transport in the brain with the HIV-1-based vector pseudotyped with rabies virus glycoprotein (RV-G). Injection of the RV-G-pseudotyped HIV-1 vector into the dorsal striatum of mice yielded an increase in gene transfer into neuronal populations in the cerebral cortex, thalamus, and ventral midbrain, each of which innervates the striatum. In addition, injection of the RV-G-pseudotyped vector into the monkey striatum (putamen) resulted in highly efficient transfer into neurons in the ventral midbrain (nigrostriatal dopamine neurons). Our results indicate that pseudotyping of the HIV-1 vector with RV-G enhances the efficiency of gene transfer through retrograde axonal transport in both mouse and monkey brains. This primate lentiviral vector system will provide a powerful approach to gene therapy for neurological and neurodegenerative diseases by means of enhanced retrograde transport.

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