The studies described in this revised proposal investigate the functional implications and the mechanistic underpinnings of a novel form of glucocorticoid feedback regulation in the hypothalamo- neurohypophyseal system. We have identified a neuroendocrine paradigm in which negative glucocorticoid feedback on vasopressin expression is established by cell-type specific, de novo, induction of glucocorticoid receptors during hypoosmolality. Magnocellular vasopressin gene expression is generally insensitive to circulating glucocorticoid levels, and this insensitivity can be ascribed to the absence of glucocorticoid receptors in magnocelluar vasopressin neurons. With the establishment of hypoosmolality (plasma osmolality less than 275 mEq/liter), however, the complete inhibition of vasopressin transcription and the first-order decay of cytoplasmic vasopressin mRNA levels is accompanied by the induction of glucocorticoid receptor expression. In the hypoosmolar rat, the inhibition of adrenal steroid synthesis, the antagonism of glucocorticoid receptors with RU486, or adrenalectomy, all result in the re-establishment of basal vasopressin transcriptional activity; thus, the induction of glucocorticoid receptor expression in magnocellular vasopressin neurons represents a novel form of physiologically-specific feedback control that has a significant impact on central nervous system models of hormone regulated feedback in general, on proposed mechanisms of vasopressin regulation in particular, and possibly on the clinical management of patients with hypoosmolality and adrenal insufficiency. Within Aim 1, studies propose to examine the synaptic pharmacology and osmotic afferent pathways involved in the hypoosmolar induction of glucocorticoid receptor expression. We will also examine the temporal association of magnocellular phosphoCREB (pCREB) and CREMalpha expression in this process. In addition, we propose to test the functional role of pCREB in this process by utilizing mice deficient in CREB expression via gene targeting. Within Aim 2, studies propose to study the physiological significance of this novel glucocorticoid feedback system on neurohypophyseal vasopressin expression, and to investigate the apparent uncoupling of vasopressin transcription and secretion under hypoosmolar conditions and/or adrenal insufficiency. Finally, we will examine the new hypothalamic peptide, orphanin FQ, and the magnocellular expression of its receptor, LC132, as negative regulators of magnocellular function during hypoosmolality.
