CYTOCHROME-P450 SIDE-CHAIN CLEAVAGE ENZYME IN THE CEREBELLAR PURKINJENEURON AND ITS NEONATAL CHANGE IN RATS

Neurosteroids are de novo synthesized in the nervous system through me chanisms at least partly independent of peripheral steroidogenic gland s. However, the concept of neurosteroidogenesis in neurons is not clea r in mammalian brains. The present study identified the presence of cy tochrome P450scc in the rat Purkinje cell, a typical cerebellar neuron . Immunohistochemical analysis with the antibody against the purified bovine adrenal P450scc showed an immunoreaction restricted to somata a nd dendrites of the Purkinje cells in adult cerebella. Preadsorbing th e antibody with P450scc resulted in a complete absence of the immunore action. The antibody against inositol triphosphate receptor, a marker of the Purkinje cell, recognized P450scc-immunoreactive cerebellar cel ls that showed no immunoreaction with glial fibrillary acidic protein, a specific marker of glial cells. Expression of the P450scc-like prot ein in the cerebellum was verified by Western blot analysis, and cereb ellar P450scc messenger RNA, by RT-PCR analysis in adulthood. On the o ther hand, P450scc-immunoreactive cells were found to scatter througho ut the cerebellum at 0 day of age, before the differentiation of the f irst Purkinje cells, while the site of expression of this protein was localized only in somata of Purkinje cells at 3 days of age. Immunorea ctive dendrites of the Purkinje cell spread into the molecular layer d uring neonatal development concurrently with its maturation. The inten sity of the immunoreaction did not change during neonatal life. Expres sion of the cerebellar P450scc messenger RNA was also detected after b irth, and the level was almost constant during neonatal life. A specif ic RIA indicated that the pregnenolone concentration was unexpectedly high at 0 day and decreased until 7 days. The total amount of pregneno lone in the cerebellum was almost constant from 0-7 days and increased during 7-21 days concurrently with the cerebellar development. In con trast, the pregnenolone sulfate ester level was low and did not signif icantly change among the developmental stages. These results suggest t hat steroidogenic enzyme P450scc appears in the rat Purkinje cell imme diately after its differentiation. The expression of this enzyme may r emain during neonatal development and in adulthood.