/ 2x14 2323 Mp 495 Friday May 30 11:31 AM EL–PEP (v.18#4) 2323 495 Peptides, Vol. 18, No. 4, pp. 495–504, 1997 Copyright  1997 Elsevier Science Inc. Printed in the USA. All rights reserved 0196-9781/97 $17.00 / .00 PII S0196-9781( 96 ) 00329-4 Angiotensinogen and Natriuretic Peptide mRNAs in Rat Brain: Localization and Differential Regulation by Adrenal Steroids in Hypothalamus MARY C. RYAN, PEI-JUAN SHEN AND ANDREW L. GUNDLACH 1 University of Melbourne, Clinical Pharmacology and Therapeutics Unit, Department of Medicine, Austin and Repatriation Medical Centre, Heidelberg, 3084, Australia Received 15 October 1996; Accepted 11 December 1996 RYAN, M. C., P.-J. SHEN AND A. L. GUNDLACH. Angiotensinogen and natriuretic peptide mRNAs in rat brain: localization and differential regulation by adrenal steroids in hypothalamus. PEPTIDES 18 ( 4 ) 495 – 504, 1997. — Adrenal steroids have been shown to modulate angiotensin II and natriuretic peptide systems—peptide synthesis and metabolism—in vitro. In the present study the effects of adrenal steroids on mRNA encoding the angiotensin II precursor, angiotensinogen (AOGEN), and the natri- uretic peptides, atrial natriuretic peptide (ANP) and C-type natriuretic peptide (CNP) in the rat hypothalamus were investigated using quantitative in situ hybridization histochemistry of [ 35 S ] - and [ 33 P ] -labeled oligonucleotide probes. Adrenalectomy produced an apparent overall decrease in preproAOGEN (ppAOGEN) mRNA in presumed astrocytes in the anterior hypothalamus with significant decreases (ANOVA) measured in the medial preoptic area, the ventral region of the medial preoptic area, the paraven- tricular, suprachiasmatic, supraoptic, and periventricular nuclei. ppAOGEN mRNA levels were restored by both glucocorticoid ( dexamethasone; 2 mg /ml in drinking water ) and mineralocorticoid ( aldosterone; 50 mg / kg, SC ) replacement. Treatment of intact animals with dexamethasone ( 2 mg /ml in drinking water for 5 days ) and aldosterone ( 100 mg / kg, SC, daily for 10 days ) produced a significant increase in ppAOGEN mRNA in those hypothalamic regions affected by adrenalectomy. ppANP and ppCNP mRNA- positive neurons were successfully detected using [ 35 S ] - and [ 33 P ] -labeled probes, respectively, and were abundant in the anterior hypothalamus, particularly in the anteromedial preoptic nucleus of the medial preoptic area. In contrast to the effects on ppAOGEN mRNA, however, alterations in adrenal steroid levels did not significantly change ppANP or ppCNP mRNA levels in neurons of the anteromedial preoptic nucleus or in the arcuate nucleus. These results indicate that adrenal steroids modulate AOGEN gene transcription in vivo, consistent with previous reports of increased levels of ppAOGEN mRNA in a number of brain regions in response to acute dexamethasone treatment and reports of decreased AOGEN immunoreactivity in brain regions of adrenalectom- ized rats. In contrast, despite reports of modulation of hypothalamic ANP immunoreactivity following adrenalectomy and dexa- methasone treatment, it would appear that adrenal steroids do not alter the transcription or stability of hypothalamic natriuretic peptides mRNA in vivo.  1997 Elsevier Science Inc. Atrial natriuretic peptide C-type natriuretic peptide Angiotensin II Adrenalectomy Corticotrophin-releasing factor Glucocorticoid Mineralocorticoid In situ hybridization histochemistry 1 Requests for reprints should be addressed to Dr. A. L. Gundlach, Clinical Pharmacology and Therapeutics Unit, Austin and Repatriation Medical Centre, Heidelberg, 3084 Australia. THE mammalian natriuretic peptide system consists of at least three structurally homologous peptides which are differentially ex- pressed throughout the periphery and central nervous system (CNS ) and thought to be involved in the regulation of blood fluid homeo- stasis and cardiovascular and endocrine function (19,43). Atrial na- triuretic peptide (ANP) and C-type natriuretic peptide (CNP) im- munoreactivity (IR) and the mRNA encoding their peptide precursors, preproANP (ppANP) and preproCNP (ppCNP) have been localized to discrete neuronal populations in the CNS with largely unique distributions, although some overlap is apparent, par- ticularly in the hypothalamus including the anteromedial preoptic nucleus and the arcuate nucleus (15,27,32,44). Conversely, brain natriuretic peptide, although originally isolated from porcine brain, has not been detected in the CNS of human or rat (19). Although ANP and CNP show highly conserved structure among different species, the relative abundance of ANP-IR and CNP-IR and mRNA in the CNS varies considerably with CNP being the most abundant natriuretic peptide in human and porcine brain, and ANP more abun- dant in the rat CNS (19,32,36,53). Studies of natriuretic peptide receptor (NPR) mRNA in rat brain have found the NPR-B sub- type —which exhibits a higher affinity for CNP than ANP, the pref- erential ligand for the NPR-A subtype —to be the predominant NR in the hypothalamus, suggesting an important role for CNP in this region (26). In this respect both ANP and CNP have been shown