IMMUNOFLUORESCENT LOCALIZATION OF CONSTITUTIVE AND INDUCIBLE PROSTAGLANDIN-H SYNTHASE IN OVINE ASTROGLIA

We have used immunofluorescent techniques to examine the distribution of prostaglandin H synthase (PGHS) in ovine astrocyte-enriched seconda ry cultures and in mixed cortical cells in primary culture. A battery of monoclonal and polyclonal antibodies specific for the constitutive (PGHS-1) or inducible (PGHS-2) forms of the enzyme were used to examin e the cells in culture. Varying levels of PGHS-1- and PGHS-2-specific immunofluorescence were seen in astrocytes as well as in other cells. The fluorescent pattern and localization seen with antisera to both PG HS-1 and PGHS-2 were similar but were not identical. Both immunoreacti ve species were confined to nuclear and perinuclear regions of the cel l, with no immunoreactivity evident in plasmalemma. In addition, PGHS- 2-specific fluorescence was concentrated often as a homogeneous ring a round the nucleus in heavily stained astrocytes. Mixed cortical glia/f ibroblasts in primary culture were double labeled with antibodies to g lial fibrillary acidic protein (GFAP) and to PGHS-2. GFAP and PGHS-2 w ere colocalized in clusters of astrocytes, but PGHS-2 was evident in G FAP(-) cells as well. Cells treated with the mitogenic agent phorbol d ibutyrate displayed more PGHS-2(+) immunofluorescence compared to eith er vehicle control or cells pretreated with dexamethasone. We conclude that astrocytes cultured in serum express both constitutive and induc ible forms of PGHS and that PGHS-2 is induced by mitogens in this cell type. (C) 1996 Wiley-Liss, Inc.