Multiparous Holstein cows (n = 18) were fed a total mixed ration containing corn silage, corn grain, whole cottonseed, soybean meal, dried distillers grains, and chopped bermudagrass hay (control) or same diet plus Ca salts of long-chain fatty acids (2.2% of diet DM) for the first 60 d postpartum. Predicted energy balance was calculated from DM intake, milk yield and composition, and BW. On d 25 postpartum, cows were injected with 25 mg of prostaglandin F2-alpha and treated for 15 d with an intravaginal device containing 1.9 g progesterone. Profiles of 15-keto-13,14-dihydro-prostaglandin F2-alpha (d 1 to 21) and plasma triglycerides (d 7 to 60) were similar between groups. Average number of follicles, determined by ultrasonography prior to d 25, tended to differ between groups; controls had more 3- to 5-mm and fewer 6- to 9-mm follicles than the group of fat-fed cows. Basal, smoothed mean concentration, and average luteinizing hormone amplitude, determined by 10-min samples for 8 h on d 10, were not significantly different between groups. Increasing predicted energy balance was associated with increased pulse amplitude and diameter of the largest follicle on d 10. During the progesterone treatment period and the postprogesterone treatment estrous cycle, cows fed fat had greater numbers of 3- to 5-mm and >15-mm follicles. In conclusion, feeding fat did not influence 15-keto-13,14-dihydro-prostaglandin F2-alpha or luteinizing hormone dynamics but did alter the average number of follicles within different size classes and the diameter of largest and second largest follicle after progesterone treatment.