STEADY-STATE STEROID 5-ALPHA-REDUCTASE MESSENGER-RIBONUCLEIC-ACID LEVELS AND IMMUNOCYTOCHEMICAL LOCALIZATION OF THE TYPE-1 PROTEIN IN THE RAT TESTIS DURING POSTNATAL-DEVELOPMENT

Steroid 5 alpha-reductase is the rate-limiting enzyme in the productio n of 5 alpha-reduced steroids in many tissues. Developmental changes i n 5 alpha-reductase activity play an important role in regulating the amount of testosterone that is secreted by the testis. To date, the re gulation of testicular 5 alpha-reductase has been studied extensively at the level of enzyme activity. Regulation at the messenger RNA (mRNA ) and protein levels, however, has not been investigated. The objectiv es of the present study were to determine the steady state mRNA levels for the 5 alpha-reductase isozymes, types 1 and 2, and to immunolocal ize the 5 alpha-reductase type 1 protein in the developing rat testis (7-91 days postpartum). Consistent with previously reported enzyme act ivity studies, type 1 5 alpha-reductase mRNA levels were most abundant in the immature animal (days 21-28). Unlike 5 alpha-reductase activit y, however, type 1 mRNA levels did not decline thereafter to reach nea rly undetectable levels in the adult (day 91). In contrast, 5 alpha-re ductase type 1 mRNA levels remained relatively constant between days 4 2-91. The 5 alpha-reductase type 1 transcript size did not remain cons tant during post natal testicular development. The characteristic 2.5- kilobase type 1 transcript size was detected in immature rats (days 21 -28), whereas in the adult (day 91), a slower migrating 2.7-kilobase t ype 1 mRNA species was observed. An antipeptide antiserum specific to rat 5 alpha-reductase type 1 was used to immunolocalize the 5 alpha-re ductase type 1 protein. At all ages examined, the immunoperoxidase rea ction was localized predominantly to the interstitial tissue of the te stis. On postnatal day 7, clusters of interstitial cells resembling fe tal Leydig cells were clearly immunoreactive. The staining intensity i ncreased steadily from day 7 onwards, so that by days 21 and 28, inter stitial cells with the appearance of immature Leydig cells were intens ely immunoreactive (peak expression). This was followed by a progressi ve decrease in staining intensity between days 28-91, so that by day 9 1 (adult) Leydig cell immunoreactivity was barely detectable. Immunocy tochemical staining revealed a predominantly cytoplasmic localization; significant nuclear staining was not evident. We conclude that the ex pression of the 5 alpha-reductase type 1 protein is primarily found in the cytoplasm of Leydig cells, is dependent on age, and that this exp ression closely parallels 5 alpha-reductase enzyme activity.