The rate of uptake of sex steroids from water by Tinca tinca is influenced by their affinity for sex steroid binding protein in plasma

Abstract

Two experiments were carried out in which male and female tench Tinca tinca were placed in individual containers and tritiated steroids then added to the water. Water samples were collected over the next 6 or 7 h and the fish then sacrificed, bled and the gall bladder removed. Radioactivity was counted in all the samples. Over the course of the exposure period in the first experiment (7 h), radioactivity of 11‐ketotestosterone (11‐KT) in the water was depleted by 11%, 17,20β‐dihydroxypregn‐4‐en‐3‐one (17,20ß‐P) and 17,20α‐dihydroxypregn‐4‐en‐3‐one (17,20α‐P) by 28%, testosterone (T) by 56% and androstenedione (AD) by 68%. HPLC analysis of water samples at 3 h indicated that none of the steroids was extensively metabolized during the experiment. Females had a faster rate of uptake of AD than males. In the second experiment (6 h), radioactivity of cortisol in the water was depleted by 5%, 11‐KT by 7%, 17‐hydroxypregnen‐4‐ene (17‐P) by 17%, 17β‐oestradiol (E2) by 35%, T by 37% and AD by 44%. In both experiments, the amounts of radioactivity that were recovered from the gall bladder and plasma were positively correlated with the rate of disappearance of radioactivity from the water. The ability of the steroids to bind to sex steroid binding protein (SBP) of tench plasma was tested by incubating plasma with radioactive steroids and then separating bound and free with ice cold dextran‐coated charcoal. When plasma at a final dilution of 1 : 60 (v/v) was incubated with 5 nM of each steroid, the percentage of radiolabel bound to SBP was: T 48% AD 44%, E2 30%, 17‐P 17%, 11‐KT 13·2%, 17,20α‐P 10·3%, 17,20β‐P 4·5% and cortisol 0%. Saturation analysis established dissociation constants (Kd; mean ± s.e.) of 3·4 ± 0·4, 2·2 ± 0·2, 4·0 ± 0·3. 9·0 ± 2·8 and 51·8 nM and binding capacities (Bmax) of 201 ± 29, 201 ± 33, 165 ± 3, 187 ± 15 and 13·4 nM for T, AD, E2,17‐P and 17,20β‐P respectively. The ability of steroids to displace tritiated T and AD from SBP was in the rank order AD > T > E2 > 17,20αP = 17,20β‐P = 11‐KT = 17‐P > cortisol. Thus, the ability of tench plasma to bind certain steroids showed a relatively strong correlation with the ability of the fish to take up these steroids from water. Modelling of data for AD and 17,20β‐P helped to show why and how plasma binding had a strong influence on the rate of uptake (and hence release) of the steroids.