Expression of NUCB2/NEFASTIN-1 in the brain and gut of the teleostean model for aging research nothobranchius furzeri

The mechanisms that regulate food behavior are the result of a complex neuroendocrine integration. In 2006 a new regulating molecule of eating behavior called nesfatin–1 (Nesf-1) was identified. Nesf-1 and its precursor NEFA/nucleobindin 2 (NUCB2) showed a highly conserved structure between mammalian and non-mammalian species, where they were detected both in appetite-control hypothalamic nuclei and in peripheral tissues involved in energy metabolism. Although tissue distribution is known in Vertebrates, there are few reports on expression during ontogenesis. In this study we examine for the first time the age-related central and peripheral expression of NUCB2/Nesf-1 in a model organism for aging research the teleost Nothobranchius furzeri. Experiments were performed on fishes at 5 weeks post hatching (wph) (young-adult) and 27 wph (old) belonging to MZM 04/10 strain provided by the FLI Leibniz Institute. Our analysis were directed on brain areas involved in food intake and on rostral intestinal bulb, which is analogous of mammalian stomach. Sequence analysis evidenced that in N. furzeri gene structure was well conserved and protein sequence showed an overall identity of 78% with medaka, the closest related species. Real-Time PCR in old fishes compared to young revealed an increase in brain and a decrease in rostral intestinal bulb in NUCB2 expression levels. Either in young and aged brains both NUCB2 mRNA transcript and Nesf-1 immunoreactivity cells were detected not only in the hypothalamic area but also in non diencephalic regions, specifically telencephalon, optic tectum and semicircular tori. Western blot on brain revealed a band of about 40 kDa, confirming the specificity of antibody employed. Either in young and aged rostral intestinal bulb NUCB2 mRNA transcript was detected mainly in the lining epithelium while Nesf-1 immunoreactive cells were distributed in the submucosae. Our results represent a step for understanding the regulation of NUCB2/Nesf-1 during vertebrate aging processes