A PUTATIVE CORTISOL RECEPTOR IN THE RAINBOW-TROUT ERYTHROCYTE - STRESS PREVENTS STARVATION-INDUCED INCREASES IN SPECIFIC BINDING OF CORTISOL

Binding sites for the steroid hormone cortisol, with characteristics t ypical of a steroid receptor, were detected in the rainbow trout (Onco rhynchus mykiss) erythrocyte. Binding of [H-3]cortisol to a washed and purified erythrocyte suspension was saturable (B-max=0.33+/-0.06 fmol per 2x10(6) cells; approximately 100+/-18 sites per cell; mean +/- S. E.M., N=6), of high affinity (K-d=3.7+/-0.4 nmol l(-1)) and reversible in the presence of an excess of unlabelled ligand. Maximum levels of specific binding were observed within 60 min of the addition of [H-3]c ortisol at 4 degrees C and were stable for 2-3 h. Within 20 min of the addition of excess unlabelled ligand, 60 % of specifically bound [H-3 ]cortisol had dissociated. Both dexamethasone and cortisol completely displaced specifically bound [H-3]cortisol at 100-fold excess, whereas a 1000-fold excess of unlabelled cortisone, 11-ketotestosterone, oest radiol-17 beta, testosterone and 17 alpha,20 beta-dihydroxy-4-pregnen- 3-one failed to displace specifically bound [H-3]cortisol completely. Specific binding sites for [H-3]cortisol were located predominantly (9 2 %) within the cytosolic fraction of the erythrocyte, with a trace am ount of specific binding (8%) detectable in the membrane fraction. No specific binding of [3H]cortisol was apparent in the erythrocyte nucle ar fraction. A 7 day period of confinement stress resulted in no signi ficant change in the number of erythrocyte cortisol-binding sites in r ainbow trout, although plasma cortisol levels were significantly eleva ted in the stressed fish. However, in control unconfined fish, there w as a progressive and significant increase in the amount of specificall y bound cortisol per cell during the course of the experiment (from 0. 097+/-0.030 to 0.260+/-0.070 fmol per 2x10(6) cells). A similar result was obtained when the experiment was repeated for confirmation. In bo th experiments, food was withheld from control and confined fish becau se of the negative impact of stress on appetite. The possibility that the increase in the number of erythrocyte cortisol-binding sites was r elated to the withdrawal of food was tested by quantifying the amount of specifically bound cortisol in erythrocytes over a 14 day period in unstressed rainbow trout maintained on normal rations and in unstress ed fish from which food was withheld. A significant increase in the am ount of specifically bound cortisol was observed with time in the fast ed fish (from 0.33+/-0.07 to 0.53+/-0.03 fmol per 2x10(6) cells). Thes e data suggest that the abundance of erythrocyte cortisol-binding site s in trout is a function of nutritional status and that stress opposes a fasting-induced increase in the number of binding sites.