Elimination of zinc from synaptic vesicles in the intact mouse brain by disruption of the ZnT3 gene.

The mammalian protein ZnT3 resides on synaptic vesicle membranes of zinc-containing neurons, suggesting its possible role in vesicular zinc transport. We show here that histochemically reactive zinc, corresponding to the zinc found within synaptic vesicles, was undetectable in the brains of mice with targeted disruption of the ZnT3 gene. Total zinc levels in the hippocampus and cortex of these mice were reduced by about 20%. The ultrastructure of mossy fiber boutons, which normally store the highest levels of vesicular zinc, was unaffected. Mice with one normal ZnT3 allele had reduced levels of ZnT3 protein on synaptic vesicle membranes and had intermediate amounts of vesicular zinc. These results demonstrate that ZnT3 is required for transport of zinc into synaptic vesicles and suggest that vesicular zinc concentration is determined by the abundance of ZnT3.

Ganglioneuromas and renal anomalies are induced by activated RET(MEN2B) in transgenic mice.

Multiple endocrine neoplasia type 2B (MEN2B) is an autosomal dominant syndrome characterized by the development of medullary thyroid carcinoma, pheochromocytomas, musculoskeletal anomalies and mucosal ganglioneuromas. MEN2B is caused by a specific mutation (Met918-->Thr) in the RET receptor tyrosine kinase. Different mutations of RET lead to other conditions including MEN2A, familial medullary thyroid carcinoma and intestinal aganglionosis (Hirschsprung disease). Transgenic mice were created using the dopamine beta-hydroxylase promoter to direct expression of RET(MEN2B) in the developing sympathetic and enteric nervous systems and the adrenal medulla. DbetaH-RET(MEN2B) transgenic mice developed benign neuroglial tumors, histologically identical to human ganglioneuromas, in their sympathetic nervous systems and adrenal glands. The enteric nervous system was not affected. The neoplasms in DbetaH-RET(MEN2B) mice were similar to benign neuroglial tumors induced in transgenic mice by activated Ras expression under control of the same promoter. Levels of phosphorylated MAP kinase were not increased in the RET(MEN2B)-induced neurolglial proliferations, suggesting that alternative pathways may play a role in the pathogenesis of these lesions. Transgenic mice with the highest levels of DbetaH-RET(MEN2B) expression, unexpectedly developed renal malformations analogous to those reported with loss of function mutations in the Ret gene.

Role of the Y5 neuropeptide Y receptor in feeding and obesity.

Neuropeptide Y (NPY), a 36-amino-acid neuromodulator abundantly expressed in the brain, has been implicated in the regulation of food intake and body weight. Pharmacological data suggest that NPY's stimulatory effect on appetite is transduced by the G-protein-coupled NPY Y5 receptor (Y5R). We have inactivated the Y5R gene in mice and report that younger Y5R-null mice feed and grow normally; however, they develop mild late-onset obesity characterized by increased body weight, food intake and adiposity. Fasting-induced refeeding is unchanged in younger Y5R-null mice and they exhibit normal sensitivity to leptin. Their response to intracerebroventricular (i.c.v.) administration of NPY and related peptides is either reduced or absent. NPY deficiency attenuates the obesity syndrome of mice deficient for leptin (ob/ob), but these effects are not mediated by NPY signaling through the Y5R because Y5R-null ob/ob mice are equally obese. These results demonstrate that the Y5R contributes to feeding induced by centrally administered NPY and its analogs, but is not a critical physiological feeding receptor in mice.

Oncogenesis and altered differentiation induced by activated Ras in neuroblasts of transgenic mice.

Sympathetic neurons, enteric neurons and adrenal chromaffin cells all derive from the neural crest. During development these cells migrate, proliferate, survive and differentiate in a highly controlled fashion influenced by local signals encountered during their migration. Aberrations of these processes are responsible for a variety of developmental defects and malignancies. Many of the environmental signals influencing these precursor cells activate receptor tyrosine kinases that can signal, at least in part, via Ras pathways. To assess the extent to which Ras can alter neuroblast cell number and fate in vivo, we expressed activated H-Ras in transgenic mice using the dopamine-beta-hydroxylase promoter, which directs expression to these cells prior to and after their differentiation. Ganglioneuromas and occasional neuroblastomas formed in the adrenal gland and preaortic sympathetic ganglia. Curiously, neurons of the superior cervical ganglia and the gut were largely unaffected despite demonstrated expression of activated Ras. The sensitivity of preaortic sympathetic neurons and adrenal chromaffin cells to the effects of oncogenes such as Ras may explain the predilection of neuroblastomas in humans to these sites. The ability to analyse neuroblastoma development in these mice may shed light on the molecular basis of certain types of human neuroblastoma.