Arginine vasopressin and oxytocin in the bovine adrenal gland

Abstract

ABSTRACT The concentrations of immunoreactive oxytocin and arginine vasopressin (AVP) and their respective neurophysins (NpI and NpII) were compared in bovine adrenal cortex and medulla. While the concentration of AVP was similar in both tissues there was more NpII in the medulla. The medulla also contained much more oxytocin and NpI than the cortex. The extracted AVP and oxytocin had identical retention times to those of the synthetic peptides on high-performance liquid chromatography (HPLC) and were biologically active in assays for antidiuretic and milk-ejection activity (with potencies of 310 units/mg and 340 units/mg respectively). Adrenal NpI and NpII behaved identically to commercially available neurohypophysial proteins on HPLC. Oxytocin, NpI and AVP were assayed in five subcellular fractions of bovine adrenal medulla prepared on discontinuous sucrose gradients. A high proportion of each co-localized with noradrenaline and adrenaline in the chromaffin granule fraction. Binding of [3H]AVP and [3H]oxytocin to crude bovine adrenal medulla membranes was dependent upon both time and temperature. The binding sites were specific and saturable: studies with the V1 AVP antagonist d(CH2)5Tyr(Me)AVP and the V2 agonist 1-deamino-8-d-AVP indicated that the AVP receptor was V1 in specificity. Scatchard plots showed that each ligand interacted with a single high-affinity, low-capacity binding site: oxytocin dissociation constant (Kd) 3·1 ±0·29 nmol/l, maximum binding capacity (Bmax) 89·6 ±18·4 fmol/mg protein (n = 3); AVP Kd 0·73 ±0·02 nmol/l, Bmax 26·5 ±8·3 fmol/mg protein (n = 3). Oxytocin and AVP had no effect on basal catecholamine release from bovine chromaffin cells in primary monolayer culture. However, both peptides inhibited acetylcholine- or nicotine-stimulated noradrenaline and adrenaline release in a dose-related manner. Neither inhibited noradrenaline or adrenaline secretion stimulated by veratridine- or potassium-induced depolarization. We conclude that the bovine adrenal cortex and medulla contain authentic AVP and oxytocin. In the medulla the peptides are packaged in secretory granules. The presence of the related neurophysins and high-affinity receptors in the medulla suggests that the peptides are both synthesized and have their site of action within this tissue. The function of AVP and oxytocin in the medulla may be indicated by the inhibition of acetylcholine-stimulated catecholamine secretion in vitro, although the effect requires high concentrations of either peptide. J. Endocr. (1987) 115, 141–149