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Nalbandov (1972) suggested that the ovum produces a "luteostatic" substance which prevents luteinization of granulosa cells. This suggestion was derived from three lines of observation: (1) surgical ovectomy in vivo induced luteinization and increased the output of progesterone into the ovarian venous blood of rabbits (El-Fouly, Cook, Nekola & Nalbandov, 1970) and pigs (Nalbandov, 1972); (2) rat granulosa cell monolayers cultured with only a few oocytes luteinized, whereas those cultured in the presence of many ova retained their granulosa cell morphology (Nekola & Nalbandov, 1971); and (3) oocytic degeneration on the 3rd day of culture of rat follicles coincided with morphological luteinization of the mural granulosa (Stoklosowa & Nalbandov, 1972). To test this hypothesis porcine granulosa cells were cultured with and without the addition of cumulus-enclosed porcine oocytes. Granulosa cells and oocytes were collected from medium and large follicles (3-12 mm) of pig ovaries removed at a local slaughterhouse. Within 20-30 min after death the ovaries were excised and placed in 0-9% (w/v) NaCl containing 100 i.u. penicillin/ml, and 100 pg streptomycin/ml and kept at room temperature until culture 3-5 h later. Details of granulosa cell and oocyte collection have been described elsewhere (Channing & Ledwitz-Rigby, 1975 ; Tsafriri & Channing, 1975a, b). About 1 IO6 granulosa cells were placed in compartments of Multiwell Tissue Culture plates (Falcon, Oxnard, California) containing glass coverslips and grown in 0-5 ml medium 199 with Earle's salts and 25 mM-Hepes buffer (Grand Island Biological Co., New York) supplemented with 15 % pig serum (GIBCO), 1 mM-L-glutamine (GIBCO), 003 mM-sodium pyruvate (Eastman Organic Chemicals, Rochester, New York), 2-5 mM-sodium lactate (Sigma Chemical Co., St. Louis, Missouri), 5 pg crystalline bovine zinc insulin (Lilly Laboratories, Indianapolis, Indiana)/ ml, and 50 pg Gentamicin (Schering Diagnostics, Port Reading, New Jersey)/ml. The pig serum used had an LH activity (in terms ofNIH-oLH-Sl units) of < 2 ng/ml as assayed by a radioligand displace¬ ment assay (Channing, 1974). The serum contained 66 pg total oestrogens/ml as measured by radio¬ immunoassay (Channing & Coudert, 1976). The progesterone content of the serum was 3-8-4-1 ng/ ml, using a modification (Channing & Coudert, 1976) of the radioimmunoassay developed by Thorneycroft & Stone (1970). Morphological luteinization and the number of cells in the cultures

Lack of an inhibitory influence of oocytes upon luteinization of porcine granulosa cells in culture