One hundred and ten male Swiss white mice were killed in batches of ten weekly between birth and the end of the 10th week of postnatal life. To demonstrate 3β-hydroxysteroid dehydrogenase activity histochemically, sections of testis from every animal were incubated with the following steroid substrates: (1) sodium 3β-sulphoxypregn-5-en-20-one (pregnenolone sulphate), (2) sodium 3β-sulphoxy-17α-hydroxypregn-5-en-20-one (17α-hydroxypregnenolone sulphate), (3) sodium 3β-sulphoxyandrost-5-en-17-one (DHA sulphate), (4) 3β,16α-dihydroxypregn-5-en-20-one (16α-hydroxypregnenolone), (5) pregn-5-ene-3β,20β-diol (pregnenediol), (6) androst-5-ene-3β, 17β-diol (androstenediol).
Pregnenolone sulphate was rapidly used by the entire interstitium at all ages. 17α-Hydroxypregnenolone sulphate was metabolized by some Leydig cells of all age groups. DHA sulphate was not utilized histochemically by the Leydig cells of the various age groups, but formazan deposition occurred in the mature seminiferous epithelium. This is the only steroid so far investigated to give an histochemical reaction with the germinal epithelium, and 3β-hydroxysteroid dehydrogenase activity has not previously been described in the seminiferous tubules. The utilization of steroid sulphates differently from the free steroids in the histochemical demonstration of 3β-hydroxysteroid dehydrogenase activity suggests that the presence of a sulphate group may affect enzyme-substrate binding.
With 16α-hydroxypregnenolone and pregnenediol as substrates, 3β-hydroxysteroid dehydrogenase activity was demonstrable at birth, increased progressively until the 6th week of postnatal life, and subsequently decreased during the ensuing 4 weeks. This growth curve closely resembles the growth curve obtained with pregnenolone. Androstenediol gave a histochemical reaction with the Leydig cells of all age groups studied, and the sigmoid growth curve resembles that obtained with 3β-hydroxyandrost-5-en-17-one (DHA). These differing growth curves are regarded as further evidence of substrate-specific 3β-hydroxysteroid dehydrogenases.