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CNP-53 predominates in all tissue and fluids (hypothalamus pituitary CSF and plasma).proCNP is increased by dexamethasone in all tissues/fluids except CSF.CNP-22 is present in hypothalamus, posterior pituitary gland and CSF.ProCNP is present, and CNP-22 is absent, from anterior pituitary gland and plasma.Similar profiles in both tissues suggest the gland contributes to plasma CNP.C-type natriuretic peptide (CNP) is a paracrine growth factor widely expressed within tissues of the central nervous system. Consistent with this is the high concentration of CNP in cerebrospinal fluid (CSF), exceeding levels in the systemic circulation. CNP abundance is high in hypothalamus and especially enriched in pituitary tissue where – in contrast to hypothalamus – processing to CNP-22 is minimal. Recently we have shown that dexamethasone acutely raises CNP peptides throughout the brain as well as in CSF and plasma. Postulating that molecular forms of CNP would differ in central tissues compared to forms in pituitary and plasma, we have characterized the molecular forms of CNP in tissues (hypothalamus, anterior and posterior pituitary gland) and associated fluids (CSF and plasma) using size-exclusion high performance liquid chromatography (SE-HPLC) and radioimmunoassay in control (saline-treated) and dexamethasone-treated adult sheep. Three immunoreactive-CNP components were identified which were consistent with proCNP (1-103), CNP-53 and CNP-22, but the presence and proportions of these different fragments differed among tissues. Peaks consistent with CNP-53 were the dominant form in all tissues and fluids. Peaks consistent with proCNP, conspicuous in hypothalamic extracts, were negligible in CSF whereas proportions of low molecular weight immunoreactivity (IR) consistent with CNP-22 were similar in hypothalamus, posterior pituitary gland and CSF. In contrast, in both plasma and the anterior pituitary gland, proportions of higher molecular weight IR, consistent with CNP-53 and proCNP, predominated, and low molecular weight IR consistent with CNP-22 was very low. After dexamethasone, proCNP like material – but not other forms – was increased in all samples except CSF, consistent with increased synthesis and secretion. In conclusion, immunoreactive forms of CNP in central tissues differ from those identified in anterior pituitary tissue and plasma – suggesting that the anterior pituitary gland may contribute to systemic levels of CNP in some physiological settings.