Characterisation of Glucocorticoid Receptors in Peripheral Blood Leukocytes of Carp, Cyprinus carpio L. F. A. A. Weyts,* , † B. M. L. Verburg-van Kemenade,* and G. Flik† * Cell Biology and Immunology Laboratory, Wageningen Institute of Animal Sciences, Agricultural University, P.O. Box 338, 6700 AH Wageningen, The Netherlands; and † Department of Animal Physiology, University of Nijmegen, Toernooiveld, 6524 ED Nijmegen, The Netherlands Accepted February 6, 1998 Binding studies with [ 3 H]cortisol revealed the presence of a single class of cortisol-binding sites on carp periph- eral blood leukocytes (PBL). These binding sites showed high affinity ( K d of 3.8 nM) and low capacity (490 binding sites per cell), indicative of receptor binding. Affinity for cortisone was 254-fold lower than for corti- sol. Affinity for the two synthetic glucocorticoids dexa- methasone and triamcinolone acetonide (TA) was 4- and 10-fold higher than for cortisol, respectively. Further evidence for the GR character of the receptor came from results showing that cortisol induced apoptosis, which could be blocked by the glucocorticoid analogue RU486. A single meal of cortisol-containing food elevated plasma cortisol concentrations and decreased GR density in PBL, as measured 3 h later. The percentage of circulating B lymphocytes also decreased. Cortisol-induced redistri- bution of B lymphocytes from the blood, due to cortisol treatment, may explain the decrease of GR numbers in PBL, although downregulation of available GR remains possible.  1998 Academic Press In all teleosts so far studied, including the common carp Cyprinus carpio L., cortisol is the major corticoste- roid. Cortisol has widespread effects on osmoregula- tion, on cellular energy metabolism, and on the im- mune system (Wendelaar Bonga, 1997). Effects on the fish immune system include reduced antibody titers (Wechsler et al., 1986), redistribution of leukocytes (Maule and Schreck, 1990b), and leukocyte apoptosis (Weyts et al., 1997). In rainbow trout, cortisol acts via the glucocorticoid receptor (GR; Ducouret et al., 1995). Glucocorticoid actions on the mammalian immune system are mediated by binding to a specific cytoplas- matic GR (Bamberger et al., 1996). Receptor-like bind- ing to glucocorticoids has been detected in several fish organs (Chakraborti et al., 1987; DiBattista et al., 1983; Lee et al., 1992; Knoebl et al., 1996; Maule and Schreck, 1990a). However, there is no direct evidence for recep- tor-mediated effects on a fish immune system. Parameters that define steroid sensitivity of a cell or tissue are the number of receptors and the affinity and specificity of the receptor for that particular steroid (Bamberger, 1996). In mammalian systems downregu- lation of GR numbers in the cell counteracts potential detrimental effects of high glucocorticoid concentra- tions (Burnstein et al., 1991; Silva et al., 1994). In vivo cortisol (or cortisol analogue) treatment of salmonids downregulates receptor numbers in liver (Pottinger, 1990; Lee et al., 1992), gill (Shrimpton and Randall, 1994), and brain (Lee et al., 1992). However, in leuko- cytes isolated from coho salmon spleen and headkid- ney, receptor numbers increased following chronic stress or in vivo cortisol treatment (Maule and Schreck, 1991). Redistribution of leukocytes, caused by cortisol treatment, may have contributed to this effect. Cortisol induces apoptosis in carp PBL (Weyts et al., 1997). The present study characterises cortisol recep- tors in carp PBLfor cortisol-binding capacity, affinity, and specificity. To further characterise the PBLcortisol receptor, a specific GR blocker (RU486) was used to General and Comparative Endocrinology 111, 1–8 (1998) Article No. GC987080 1 0016-6480/98 $25.00 Copyright  1998 by Academic Press All rights of reproduction in any form reserved.