Rapid stimulation of cyclic AMP production by aldosterone in rat inner medullary collecting ducts E A Sheader, E T Wargent, N Ashton and R J Balment School of Biological Sciences, University of Manchester, G.38 Stopford Building, Oxford Road, Manchester M13 9PT, UK (Requests for offprints should be addressed to N Ashton; Email: nick.ashton@man.ac.uk) (E T Wargent is now at Clore Laboratory, University of Buckingham, Hunter Street, Buckingham MK18 1EG, UK) Abstract Aldosterone stimulates sodium transport in the inner medullary collecting duct (IMCD) via the classic genomic pathway, but it is not known whether it also acts via a rapid, non-conventional pathway in this part of the nephron. The IMCD regulates the final sodium content of urine and expresses vasopressin receptors coupled to adenylate cyclase. The recently reported rapid, non- genomic actions of aldosterone have been associated mainly with an increase in intracellular Ca 2+ ; however, it has also been shown to stimulate cAMP generation. Thus the aim of this study was to determine whether aldoster- one stimulates rapid generation of cAMP in isolated IMCD segments. IMCD segments were microdissected from Sprague–Dawley rat kidneys and incubated at 37 C for 4 min with aldosterone (10 12 to 10 6 M), vaso- pressin (10 12 to 10 6 M), or a combination of hormones in the presence of a phosphodiesterase inhibitor. cAMP was measured by radioimmunoassay. While corticosterone and dexamethasone were ineffective, aldosterone stimu- lated a dose-dependent increase in cAMP within 4 min (P < 0·05). This action of aldosterone was not inhibited by the MR antagonist spironolactone. Co-incubation of aldosterone with vasopressin resulted in a further increase in cAMP generation above that induced by the neuro- hypophysial hormone alone. Aldosterone-mediated cAMP generation was not inhibited by a vasopressin V 1 or V 2 receptor antagonist. These data support a novel and rapid, non-genomic effect of aldosterone in IMCD. Aldosterone does not apparently interact with the vasopressin receptor to stimulate cAMP generation. Journal of Endocrinology (2002) 175, 343–347 Introduction In recent years aldosterone has been shown to exert rapid, non-genomic effects in a number of tissues, including the kidney (Harvey & Higgins 2000), colon (Maguire et al. 1999) and vascular smooth muscle (Christ et al. 1995), as well as human mononuclear leukocytes (Wehling et al. 1987). These effects do not appear to be mediated via the classical type 1 mineralocorticoid receptor (MR), but rather by a distinct membrane-associated receptor whose identity has yet to be fully described. While the structure of this receptor remains obscure, there is a growing body of work describing the signal transduction pathways which link receptor activation to the rapid actions of aldosterone. To date, the pathway most commonly reported involves intracellular Ca 2+ ; however, aldosterone has also been shown to stimulate rapid generation of cyclic AMP (cAMP) in porcine coronary vascular smooth muscle cells (Christ et al. 1999). Aldosterone’s classic target site is the principal cells of the cortical collecting duct; however, it has also been shown to stimulate sodium transport in the inner medul- lary collecting duct (IMCD) (Stokes 2000). The IMCD comprises three morphologically distinct regions: the outer segment, IMCD 1 , which consists of principal and inter- calated cells similar to those in the outer medullary collecting duct, the middle segment, IMCD 2 , which is a mixture of principal, intercalated and IMCD cells, and the inner segment, IMCD 3 , which consists solely of IMCD cells (Tisher & Madesen 1996). In vivo micropuncture studies have demonstrated that aldosterone is able to stimulate sodium reabsorption via the type 1 MR in IMCD (Uhlich et al. 1969), but it is not known whether it also exerts a rapid, non-genomic effect in this part of the nephron. The major aim of this study was to determine whether aldosterone can stimulate rapid generation of cAMP in IMCD segments. We have also investigated the potential interaction between aldosterone and vasopressin in view of the sensitivity of IMCD to vasopressin via its V 2 receptor coupled to adenylate cyclase (Wargent et al. 1999). Materials and Methods Male Sprague–Dawley rats (200–250 g body weight; Charles Rivers Ltd, Margate, Kent, UK) were maintained 343 Journal of Endocrinology (2002) 175, 343–347 0022–0795/02/0175–343  2002 Society for Endocrinology Printed in Great Britain Online version via http://www.endocrinology.org