5a-reductase, an enzyme regulating glucocorticoid action in the testis of Rhinella arenarum (Amphibia: Anura).

TESONE AJ; REGUEIRA E; CANOSA LF; CEBALLOS NR

GENERAL AND COMPARATIVE ENDOCRINOLOGY

ACADEMIC PRESS INC ELSEVIER SCIENCE

Lugar: Amsterdam; Año: 2012 vol. 176 p. 500 - 506

0016-6480

The reduction of A-ring of glucocorticoids to produce 5a-dihydro-derivatives by 5a-reductases has been considered as a pathway of irreversible inactivation. However, 5a-reduced metabolites of corticosterone and testosterone have significant biological activity. In this paper, we investigated whether toad testicular 5a-reductase (5a-Red) is able to transform corticosterone into 5a-dihydrocorticosterone. Furthermore, we studied the role of 5a-reduced metabolite of corticosterone as a glucocorticoid receptor (GR) agonist. The activity of 5a-Red was assayed in subcellular fractions with [3H]corticosterone or [3H]testosterone as substrate. The enzyme localizes in microsomes and its optimal pH is between 7 and 8. The activity is not inhibited by finasteride. These results support the conclusion that toad 5a-Red resembles mammalian type 1 isoenzyme. Kinetic studies indicate that neither Km nor Vmax for both corticosterone and testosterone were significantly different among reproductive periods. The Km value for testosterone was significantly higher than that for corticosterone, indicating that the C-21 steroid is the preferred substrate for the enzyme. Studies of the binding capacity of 5a-dihydrocorticosterone (5a-DHB) to the testicular GR show that 5a-DHB is able to displace the binding of [3H]dexamethasone to testicular cytosol with a similar potency than corticosterone. The inhibition constant (Ki) values for corticosterone and 5a-DHB were similar, 31.33 ± 2.9 nM and 35.24 ± 2.3 nM, respectively. In vitro experiments suggest that 5a-DHB is an agonist of toad testicular GR, decreasing the activity of the key enzyme for androgen synthesis, the cytochrome P450 17-hydroxylase, C17,20-lyase.