The causes of the inherently high variation in number of follicles and oocytes in ovaries of mammals are unknown. Evidence suggests that the ovarian reserve (total number of healthy follicles and oocytes in ovaries) can be programmed by events occurring during fetal life. For instance, maternal nutritional restriction during the first trimester of pregnancy negatively affects the size of the ovarian reserve in calves. The aim of the present study was to establish whether exposure of pregnant dairy cows to high environmental temperatures from conception to the end of the first trimester of pregnancy impairs establishment of the ovarian reserve in their offspring. This work was conducted in four commercial dairy farms with similar nutrition and farming systems located in Sardinia, Italy, on a total of 310 Holstein-Friesian dairy heifers (16 months old) that were conceived and born at different times of year coincident with different environmental temperatures. We tested whether exposure of the heifer's dams to a mean temperature-humidity index (THI) >68 from conception to the end of the first trimester of pregnancy resulted in a diminished ovarian reserve in their offspring. To estimate the size of the ovarian reserve, a single blood sample was collected to measure serum anti-Müllerian hormone (AMH; n=310), and the number of follicles >3mm (antral follicle count, AFC) was assessed using transrectal ovarian ultrasonography (n=258) on a random day of the oestrous cycle (16.09±0.07 months of age). Relations among variables were analysed with Pearson correlation with SAS (SAS Institute Inc.). Anti-Müllerian hormone and AFC were analysed with a mixed model (PROC MIXED of SAS) considering the main effects of season during the first trimester of fetal life and age at sampling; the effect of farm was included as a random effect. Tukey's test was used for comparisons. Circulating AMH concentrations and AFC were highly positively correlated (P<0.0001), as previously reported. The results also showed that both AMH concentrations and AFC were lower (419.27±22.81 pgmL−1, 9.32±0.42 follicles; P<0.0001) in young adult heifers of the dams exposed to a THI >68 compared with dams exposed to an average THI of 55 (634.91±47.60 pgmL−1, 11.84±0.46 follicles). Neither AMH nor AFC were influenced by farm and age at sampling of the daughters. In conclusion, maternal exposure to THI >68 (typical high temperatures during summers in Sardinia) during the first trimester of pregnancy has a negative effect on the development of the ovarian reserve in female fetuses, which may subsequently impair their reproductive performance as adults. Research was funded by Regione Autonoma della Sardegna, Legge Regionale 7, Bando 2015.