This study examined seasonal differences in progesterone (P4) production by granulosa cells (GC) and thecal cells (TC) that were luteinized in vitro during the winter or the summer; it also compared plasma P4 concentrations of lactating dairy cows in the two seasons. First-wave dominant follicles obtained from Holstein cows were dissected on day 6 of the cycle, GC and TC were separated, enzymatically dispersed, and cultured for 9 days in media containing 1% fetal calf serum, forskolin (10 micromol/mL) and insulin (2 microg/mL), to induce cell luteinization. All experimental procedures were identical and characteristics of the follicles were similar in the two seasons. During 9 days of culture, P4 production by luteinized GC was higher in winter than in summer, but the difference only tended to be significant. In contrast, luteinized TC produced three times as much P4 in winter as in summer (324 versus 100 ng/10(5)cells). In the in vivo experiment, P4 concentrations in plasma collected during entire estrous cycles in winter and summer were compared. The cows were, on average, at 70 days postpartum and yielded similar amounts of milk. Concentrations of progesterone in plasma were significantly higher in winter than in summer; during the mid-luteal phase the difference between the two seasons was 1.5 ng/mL. These results indicate that chronic effects of heat-stress are possibly carried over from an impaired follicle to an impaired corpus luteum (CL), and that luteinized TC are more susceptible to heat-stress than luteinized GC.