The effects of exogeneous LH and PGF₂α on the progesterone secretion in the Hanwoo were studied using in vitro culture of luteal cells. The corpora lutea(CL) were collected and pooled from the Korean Native Cattle (Hanwoo) ovaries within 2-3 hrs from a local slaughter house. After enzymatic dissociation, combined large and small luteal cells(LLC and SLC)(I × 10^6 live cells/㎖) were incubated in D-MEM(Dulbicco`s-Modified Eagle Medium) in the presence of 10% FCS and antibiotics with or without LH and PGF2α, alone or in various combinations, for 7 days with the medium change every 24 hr. Luteal tissues were apparently formed by two luteal cell populations which were the small and large luteal cells(about 10:1) on day 1 of culture. Treatment with LH(10, 100ng/㎖) markedly stimulated in vitro luteal cell development and proliferation until day 7 of culture. Progesterone secretion in vitro from luteal cell culture was the highest on day 1 and then dramatically decreased to day 7 of culture, and was positively stimulated by LH or PGF2α on day I of culture. However, addition of exogeneous PGF₂α (200ng/㎖) in the presence of LH(10, 100ng/㎖) significantly(P$lt;0.05) inhibited progesterone secretion on day 1 of culture. These observations indicate that luteolysis of luteal cell by PGF₂α may be regulated by inhibition of pituitary LH function and other factors.