Identification of type 1 5 alpha-reductase in myelin membranes of male and female rat brain

The formation of the 5 alpha-reduced metabolites of testosterone (T) and of progesterone (P) is a very active process in the brain, since the enzyme 5 alpha-reductase (5 alpha-R) is present in almost any central nervous system (CNS) structure. A particularly elevated 5 alpha-R activity has been shown in myelin sheaths. Two isoforms of the enzyme have been cloned, with different localisation as well as different biochemical properties. The present study was performed to determine whether both isoforms of the 5 alpha-R, or only one of them, are/is responsible for the enzymatic activity observed in myelin. Kinetic analyses have been performed on purified myelin membranes prepared from the male or female rat brain, using both T and P as substrates. The 5 alpha-R present appears to possess a pH optimum at basic values. The V-max values obtained in the Lineweaver-Burk analysis were comparable in male and female preparations independently on whether T or P were used as the substrates, suggesting that a single enzymatic form is present in all samples examined; the K-m obtained using [C-14]T (K-m: male 1.14 mu M; female 1.46 mu M) or [C-14]P (K,: male 0.5 mu M; female 0.64 mu M) as substrates, were in good agreement with those obtained for the recombinant type 1 isoform. These data suggest that the type 1 isoform is the most relevant 5 alpha-R present in myelin. To confirm this, a new polyclonal antibody was raised against the type 1 5 alpha-R enzymatic protein, and used in immunohistochemical studies. The experiments were performed on the optic nerve, a myelinated structure very rich in 5 alpha-R activity and the results clearly indicated the presence of a specific type 1 enzyme immunoreactivity in the myelin sheaths of axons. (C) 1997 Elsevier Science Ireland Ltd.