Separate receptors mediate oxytocin and vasopressin stimulation of cAMP in rat inner medullary collecting duct cells

Abstract

SUMMARYThe two neurohypophysial hormones arginine vasopressin (AVP) and oxytocin have actions in the inner medullary collecting duct (IMCD) where both peptides induce an increas8e in cAMP accumulation. The present study has employed a novel IMCD cell line to determine whether these two hormones induce cAMP accumulation via common or separate receptors, and to characterize the potential receptors responsible. Equal volumes of vehicle (150 mM NaCI) or hormone/antagonist solutions were added to aliquots of lo4 IMCD cells in the presence of M 3-isobutylmethylxanthine (IBMX) and incubated at 37OC for 4min. cAMP levels were determined by radioimmunoassay and protein concentration by Bradford assay. Both AVP and oxytocin elicited dose-dependent increases in cAMP generation, though oxytocin was less potenlt than AVP (EC,, = 1.6 x lo-' M vs. 7.4 x lo-'' M). AVP at lo-' M and oxytocin at M, concentrations sufficient to elicit near-maximal cAMP accumulation, resulted in cAMP levels of 73.4 k 1.7 and 69.0 k 3.3 pmol (mg protein)-' (4 min)-', respectively (n = lo), compared with the vehicle-treated basal value of 37.7 f 2.2 pmol (mg protein)-' (4 min)-' ( P c 0.001, n = 10). Combined AVP (lo-' M) and oxytocin M) resulted in cAMP accumulation of 63.8 f 3.1 pmol (mg protein)-' (4 min)-' (n = lo), which was not significantly different from the effect of oxytocin alone, but slightly less than that for AVP alone ( P c 0.05). A submaximal concentration of AVP (lo-" M) induced cAMP accumulation of 48.6 f 2.5 pmol (mg protein)-' (4 min)-' ( P c 0.01 compared with basal level of 34.9 f 2.4 pmol (mg protein)-' (4 min)-', n = lo), which was blocked in the presence of a vasopressin V, receptor antagonist M OPC-31260) but not by the oxytocin receptor antagonist ( low6 M [Pen',pMePhe2,Thr4,0rns]oxytocin) ( 3 6 3 k 6.1 and 45.1 f 1.3 pmol (mg protein)-' (4 min)-' respectively, P c 0.05, n = 10). A submaximal concentration of oxytocin M) induced a cAMP accumulation of 45.8 f 1.8 pmol (mg protein)-' (4 min)-' (n = lo), which was reduced by addition of M oxytocin antagonist (36.3 & 2.1 pmol (mg protein)-' (4 min)-', P c 0.05, n = lo), whereas co-incubation with M of the V, receptor antagonist had no effect (43.2 f 1.3 pmol (mg protein)-' (4 min)-', n = 10).These results indicate that AVP and oxytocin induce cAMP accumulation from a common ATP pool in IMCD cells, and that separate vasopressin V, and oxytocin receptor systems are involved, perhaps coupled to a common adenylate cyclase system. INTRODUCTIONThe antidiuretic osmotic water transport (P,) in the inner medullary collecting duct of most mammals, including humans and rats, is regulated by arginine vasopressin (A'VP) via CAMPmediated translocation of aquaporin-2-containing vesicles to the luminal membrane. The other neurohypophysial hormone, oxytocin, has also been demonstrated to have an antit Corresponding author: rnopleetw@stud.rnan.ac.uk , 1984;Lyness et al. 1985) and has also been found to stimulate an increase in osmotic water transport (P,) in microdissected inner medullary colle...

African lungfish,Protopterus annectens, possess an arginine vasotocin receptor homologous to the tetrapod V2-type receptor

“…Molecular phylogenetic analysis also showed that the putative lungfish V2-type receptor belongs to the V2-type receptor group of tetrapods and was distinguishable from the V1-type receptor or other neurohypophysial hormone receptor groups. It has been known that the V1-and V2-type receptors in tetrapods are coupled to the PLC/PKC pathway and AC/PKA pathway, respectively (Wargent et al, 1999;Kohno et al, 2003). In the present study, administration of AVT to CHO cells transfected with lungfish V1a-type receptor or V2-type receptor resulted in intracellular Ca 2+ mobilization and the accumulation of cAMP in a concentration-dependent manner, respectively.…”

Nitrergic modulation of vasopressin, oxytocin and atrial natriuretic peptide secretion in response to sodium intake and hypertonic blood volume expansion

“…OT binds to the V2 AVP receptor because of its structural similarity to AVP. Vasopressin and OT induce cAMP accumulation from a common ATP pool in inner medullary collecting duct cells, and the separate AVP V2 and OT receptor systems may be coupled to a common adenylate cyclase system (30). In addition to their peripheral effects, these peptides may also have other effects that could complement their physiological actions.…”

Vasotocin and isotocin regulate aquaporin 1 function in the sea bream

“…3), the effect mediated by AVT V2-type receptors on both AQP1a and AQP1b function would involve stimulation of PKA and perhaps PKC. It is unsurprising that AQP1a is regulated by AVTRV2 given that PKA is the preferential signaling pathway associated with this receptor, and PKC signaling seems to play a minor role (Wargent et al, 1999;Warne, 2001). However, previous studies in frog demonstrated the V2-type receptor could also function via stimulation of PLC pathways, one of the alternative pathways for PKC (Acharjee et al, 2004;Zhu et al, 1994).…”

“…The regulation of AQP function can be achieved via different signaling pathways (Dibas et al, 1998). AVTR/AVPR V1a-type and ITR were shown to act via PKC, which uses phospholipase C (PLC), inositol-1,4,5-trisphosphate (IP3) or Ca 2+ intracellular signaling pathways (Wargent et al, 1999). In contrast, PKA, whose activity is dependent on cellular levels of cyclic AMP (cAMP), is preferentially used by AVT/AVP V2-type receptors (Wargent et al, 1999;Warne, 2001).…”

“…AVTR/AVPR V1a-type and ITR were shown to act via PKC, which uses phospholipase C (PLC), inositol-1,4,5-trisphosphate (IP3) or Ca 2+ intracellular signaling pathways (Wargent et al, 1999). In contrast, PKA, whose activity is dependent on cellular levels of cyclic AMP (cAMP), is preferentially used by AVT/AVP V2-type receptors (Wargent et al, 1999;Warne, 2001). Although direct phosphorylation of AQPs was not demonstrated in this study, we have shown that sea bream Aqp1a-mediated water transport is positively regulated by PKC but not PKA signaling, whereas the opposite holds true for Aqp1b (Fig.…”