CHARACTERIZATION OF PUTATIVE STEROID-RECEPTORS IN THE MEMBRANE, CYTOSOL AND NUCLEAR FRACTIONS FROM THE OLFACTORY TISSUE OF BROWN AND RAINBOW-TROUT

Specific binding sites for testosterone have been detected in three co mpartments of olfactory tissue from brown and rainbow trout. Binding o f H-3-testosterone to the membrane fraction of olfactory tissue is of high affinity (K-d = 0.5 - 1.9 nM) and limited capacity (N-max = 30 - 60 fmol mg(-1) protein). Binding is reversible, and is eliminated by p rotease treatment. The membrane binding site exhibits a high degree of ligand specificity; 11 beta-hydroxytestosterone, 11-ketotestosterone, 17 alpha-hydroxyprogesterone, 17 alpha,20 beta-dihydroxy-4-pregnen-3- one, cortisol, and estradiol-17 beta all fail to displace testosterone at 20-fold excess while testosterone itself competes successfully. Th ese attributes are consistent with the presence of specific steroid re ceptor proteins. Binding of testosterone within the cytosol is of mode rate affinity (K-d = 9.0 - 23.0 nM) and high capacity (N-max = 0.5 - 2 .9 pmol mg(-1) protein) and is more readily displaced by a number of s teroid competitors than is the case for the membrane site. The rate of association and dissociation of testosterone from the cytosolic bindi ng site is markedly more rapid than the equivalent processes in the me mbrane fraction. Binding of testosterone to the nuclear extract is of high affinity (K-d similar to 3.0 nM) and limited capacity (N-max simi lar to 50 fmol mg(-1) protein). There are no substantial differences b etween species or between sexes in the affinity or capacity of testost erone-binding sites in nuclear extract or membrane fraction. However, cytosolic testosterone-binding sites are three-to four-fold more abund ant in rainbow trout than in brown trout, and female rainbow trout hav e more cytosolic binding sites than male rainbow trout, but a lower af finity for testosterone than male sites, Preliminary evidence supports the involvement of the membrane-associated testosterone-binding site in olfactory processes. Rainbow trout display an EOG response to testo sterone at a concentration (greater than or equal to 10(-9) M) which i s consistent with the equilibrium dissociation constant (K-d) of the m embrane-associated testosterone-binding site. Binding of H-3-testoster one to the membrane-associated site shows a pH dependency which is com parable to the effects of pH on the EOG response to testosterone in in tact fish. The attributes of the intracellular testosterone-binding si tes are common to testosterone receptors in other fish tissues which a re known androgen target tissues. This suggests that the development a nd/or function of salmonid olfactory tissue may be susceptible to infl uence by endogenous testosterone.