GRANULOSA LAYER - PRIMARY SITE OF REGULATION OF PLASMINOGEN-ACTIVATORMESSENGER-RIBONUCLEIC-ACID BY LUTEINIZING-HORMONE IN THE AVIAN OVARY

Plasminogen activator (PA) is hypothesized to be important in the remo deling of the extracellular matrix during follicular growth. The granu losa layer produces high amounts of PA in response to a stimulatory fa ctor, produced by the theca layer, that is inhibited by LH. To determi ne the site and mechanism by which LH inhibits PA production, we asked 1) whether LH acts on the granulosa layer and/or the theca layer to i nhibit PA production by the largest preovulatory follicle (F1), and 2) whether LH affects PA production by acting at the mRNA or protein lev el. Sections (10 mm in diameter) of granulosa layers obtained from the F1 follicle before (14 h before ovulation) or after (2 h before ovula tion) the LH surge were incubated (24 h at 37 degrees C) in theca-cond itioned medium; this medium had been prepared by incubation of 10-mm-d iameter sections of theca layers, obtained before (14 h before ovulati on) or after (2 h before ovulation) the LH surge, in Dulbecco's Modifi ed Eagle's Medium for 24 h at 37 degrees C. PA production in culture m edium was measured with use of the chromogenic substrate S-2251. PA pr oduction was high when granulosa layers obtained before the LH surge w ere incubated in theca-conditioned medium obtained before the LH surge ; it was also high when granulosa layers obtained before the LH surge were incubated in theca-conditioned medium obtained after the LH surge . PA production was low when granulosa layers obtained after the LH su rge were incubated in theca-conditioned medium obtained before the LH surge, and was also low when granulosa layers obtained after the LH su rge were incubated in theca-conditioned medium obtained after the LH s urge. Northern and Western blots and activity assays performed on gran ulosa layer homogenates indicated that PA mRNA, protein, and activity were high before the LH surge and low after the LH surge. Production o f the stimulatory factor by the theca layer is apparently unaffected b y LH. After exposure to LH, the granulosa layer is no longer capable o f producing PA, even in the presence of the theca-derived stimulatory factor. We conclude that the granulosa layer is the site of mRNA and/o r protein regulation of PA production by LH.