The effects of different culture media, glucose, pyridine nucleotides and adenosine on the activity of 11 beta-hydroxysteroid dehydrogenase in rat Leydig cells

11 beta-Hydroxysteroid dehydrogenase (11 beta HSD) reversibly converts gluc ocorticoids into inert 11-ketosteroids. The direction of the reaction has b een found to vary with the cell type and sub-cellular reparation used. We h ave investigated if the directionality of 11 beta HSD can be influenced by the nature of the culture medium and compounds added during incubation of r at testis Leydig cells.:We found that when the cells were cultured in Dulbe cco's Modified Eagle Medium (DMEM) that the dehydrogenase (11 beta DH) acti vity was higher than the reductase (11KSR) activity (11 beta DH:11KSR ratio similar to 2:1). When glucose was omitted from the DMEM a higher 11 beta D H:11KSR ratio (similar to 33:1) was obtained. However, when the cells were cultured in a combination of DMEM/Ham's F12 (1:1, v/v), a ninefold increase in 11KSR activity was obtained whereas 11 beta DH activity was inhibited b y 64% compared with cells incubated in DMEM alone. Consequently, the predom inant activity changed from a dehydrogenase to a reductase (11 beta DH: 11K SR ratio 1:15). Addition of the individual components of the Ham's F12 medi um to DMEM showed that only pyruvate and/or the amino acids were able to mi mic the effects of DMEM/Ham's F12. Similar differential effects were found when NAD(+), NADH or adenosine were added to the Leydig cells incubated in DMEM (three to fivefold increases and 20-50% decreases in 11KSR and 11 beta DH activities, respectively). In contrast, NADP(+) was found to increase 1 1 beta DH activity (up to threefold) but NADPH had no effect on 11KSR activ ity. Cells incubated with DMEM/Ham's F12, NAD(+), NADP(+) and adenosine wer e found to have higher ATP levels (four to sixfold) than those incubated in DMEM alone. These results illustrate that the relative 11 beta DH and 11KS R activities of 11 beta HSD in Leydig cells are markedly and differentially altered by the nature of the incubation medium and compounds added. (C) 19 99 Elsevier Science Ireland Ltd. All rights reserved.