Fluorescent Visualisation of the Hypothalamic Oxytocin Neurones Activated by Cholecystokinin-8 in Rats Expressing c- fos-Enhanced Green Fluorescent Protein and Oxytocin-Monomeric Red Fluorescent Protein 1 Fusion Transgenes.

The up-regulation of c- fos gene expression is widely used as a marker of neuronal activation elicited by various stimuli. Anatomically precise observation of c- fos gene products can be achieved at the RNA level by in situ hybridisation or at the protein level by immunocytochemistry. Both of these methods are time and labour intensive. We have developed a novel transgenic rat system that enables the trivial visualisation of c- fos expression using an enhanced green fluorescent protein (e GFP) tag. These rats express a transgene consisting of c- fos gene regulatory sequences that drive the expression of a c- fos-e GFP fusion protein. In c- fos-e GFP transgenic rats, robust nuclear e GFP fluorescence was observed in osmosensitive brain regions 90 min after i.p. administration of hypertonic saline. Nuclear e GFP fluorescence was also observed in the supraoptic nucleus ( SON) and paraventricular nucleus ( PVN) 90 min after i.p. administration of cholecystokinin ( CCK)-8, which selectively activates oxytocin ( OXT)-secreting neurones in the hypothalamus. In double transgenic rats that express c- fos-e GFP and an OXT-monomeric red fluorescent protein 1 (m RFP1) fusion gene, almost all m RFP1-positive neurones in the SON and PVN expressed nuclear e GFP fluorescence 90 min after i.p. administration of CCK-8. It is possible that not only a plane image, but also three-dimensional reconstruction image may identify cytoplasmic vesicles in an activated neurone at the same time. [ABSTRACT FROM AUTHOR]