AJP - Renal Physiology, Vol 261, Issue 5 760-F766, Copyright © 1991 by American Physiological Society

ARTICLES

Characterization of prazosin-sensitive alpha 2 B-adrenoceptors expressed by cultured rat IMCD cells

G. Yasuda, L. Sun, S. Umemura, W. A. Pettinger and W. B. Jeffries
Department of Medicine, Creighton University School of Medicine, Omaha, Nebraska 68131.

alpha 2-Adrenoceptor subtype expression was investigated in cultured rat inner medullary collecting duct (IMCD) cells using radioligand binding studies, Northern blot analysis, and adenosine 3',5'-cyclic monophosphate (cAMP) assays. [3H]rauwolscine bound to a single class of alpha 2-adrenoceptors with high affinity [Kd = 1.7 +/- 0.3 nM, maximum binding (Bmax) = 45.2 +/- 10.8 fmol/mg protein]. alpha 2-Adrenoceptor ligands inhibited [3H]rauwolscine binding with a rank order of potency characteristic of interaction with the alpha 2B-adrenoceptor [inhibitory constant (Ki) values (in nM) rauwolscine (1.95) greater than ARC-239 (8.52) greater than prazosin (237) greater than oxymetazoline (30,000)]. Northern blot analysis was performed using poly(A)+ RNA isolated from 90% confluent rat IMCD cells and probes derived from alpha 2-adrenoceptor DNA sequences from the rat nonglycosylated alpha 2B-adrenoceptor and the human alpha 2A-adrenoceptor. The alpha 2B probe hybridized to a 4.2-kb band under high stringency conditions, but the alpha 2A-adrenoceptor probe did not hybridize to this band. In functional studies, the full alpha 2-adrenoceptor agonists epinephrine and UK-14,304 potently inhibited vasopressin-stimulated cAMP accumulation by 50 to 70% [half-maximal response (EC50) (in nM) epinephrine = 11.2, UK-14,304 = 6.4]. Guanabenz and clonidine were partial agonists, inhibiting cAMP accumulation by 30 to 40% and were less potent than the full agonists [EC50 (in nM) 56.0 guanabenz and 94.5 clonidine]. Epinephrine-induced inhibition of cAMP accumulation was blocked by rauwolscine, prazosin, and ARC-239 but not by the alpha 1-adrenoceptor antagonist corynanthine. We conclude that rat IMCD cells in primary culture express functional alpha 2-adrenoceptors of the alpha 2B-subtype.

This article has been cited by other articles:

				D. P. Wallace, G. Reif, A.-M. Hedge, J. B. Thrasher, and P. Pietrow Adrenergic regulation of salt and fluid secretion in human medullary collecting duct cells Am J Physiol Renal Physiol, October 1, 2004; 287(4): F639 - F648. [Abstract] [Full Text] [PDF]

				R. Nasrallah, J. Zimpelmann, S. Singh, and R. L. Hebert Molecular and biochemical characterization of prostacyclin receptors in rat kidney Am J Physiol Renal Physiol, February 1, 2001; 280(2): F266 - F277. [Abstract] [Full Text] [PDF]

				A. J. Rouch and L. H. Kudo Role of PGE2 in alpha 2-induced inhibition of AVP- and cAMP-stimulated H2O, Na+, and urea transport in rat IMCD Am J Physiol Renal Physiol, August 1, 2000; 279(2): F294 - F301. [Abstract] [Full Text] [PDF]

				D. Cussac, S. Schaak, C. Gales, C. Flordellis, C. Denis, and H. Paris alpha 2B-Adrenergic receptors activate MAPK and modulate proliferation of primary cultured proximal tubule cells Am J Physiol Renal Physiol, May 1, 2002; 282(5): F943 - F952. [Abstract] [Full Text] [PDF]