NEUROHYPOPHYSEAL AND FLUID HOMEOSTASIS IN TRANSGENIC RATS EXPRESSING A TAGGED RAT VASOPRESSIN PREPROPEPTIDE IN HYPOTHALAMIC NEURONS

We have developed a transgenic system that, for the first time, facili tates monitoring of the regulatory dynamics of a central peptidergic s ystem from transcription of a neuropeptide gene to the storage and rel ease of the mature secretory product. A rat vasopressin (VP) transgene (5-VCAT-3), the expression of which is restricted to hypothalamic vas opressinergic magnocellular neurons in rats, contains a sequence that, if translated, would place a unique hexadecapeptide (DRSAGYYGLFKDRKEK , abbreviated to DR-12-EK) at the C-terminus of the VP precursor. We h ave raised an antibody against this ''tag'' and, using immunohistochem istry, electron microscopy, RIA, and HPLC, have shown for the first ti me that a VP transgene RNA is translated into a protein product found, in a processed form, in secretory granules in the posterior pituitari es of transgenic rats. Disruption of the C-terminus of the VP precurso r by the peptide tag is well tolerated and does not disrupt VP product ion or disturb salt and Rater balance. An osmotic stimulus increased h ypothalamic DR-12-EK levels, but changes in posterior pituitary DR-12- EK levels were more complex. After 5 days of salt-loading, DR-12-EK le vels fell, as would be expected if its release was coordinate with tha t of VP. However, after 10 days of salt-loading, posterior pituitary D R-12-EK levels increased, despite the lower level of VP. This probably reflects the greater response of the transgene to osmotic challenge a t the RNA level, increasing the proportion of DR-12-EK-containing tran slation products transported to the posterior pituitary relative to th ose derived from the endogenous gene. The exaggerated response of the tagged transgene to osmotic challenge at both RNA and protein levels a ffords a new opportunity to study the regulatory dynamics of the VP sy stem at the molecular level, but within the physiologically advantageo us context of the intact animal.