Our objectives were to investigate the potential presence and nature of pregnancy-derived endocrine signals in the ovaries during maternal recognition of pregnancy (MRP) in heifers. Pubertal heifers (n = 66) had the estrous cycle and ovulation synchronized with 5-d CIDR Cosynch protocol. On the d of the last GnRH (d 0), 3 in each 5 heifers were selected randomly to be inseminated (AI), while the remaining formed a nonbred nonpregnant group (NP). On d 11, all follicles ≥4 mm were ablated using ovum pickup (OPU) equipment to induce recruitment of new follicles. On d 16.5, presence and size of ovarian structures were evaluated. Heifers with a corpus luteum (CL) >18 mm had the dominant follicle (DF) aspirated by OPU. The aspirated DF was classified as ipsilateral (IP) or contralateral (CO) to the ovary bearing the CL. After centrifugation of the recovered follicular fluid (FF), supernatant and granulosa cells (GC) pellet were stored separately at −80°C for analyses of FF composition and GC gene expression. On d 28, pregnant (P) heifers in the AI group were identified by ultrasonography, and those with a negative diagnosis were excluded. The experimental design was a 2 × 2 factorial based on pregnancy status (P or NP) and DF (IP or CO) on d 16.5. Data were analyzed by ANOVA using the GLIMMIX procedure of SAS, and included the effects of the main factors and their interaction. The number of heifers retained for analyses in each factorial combination were PIP = 12, NPIP = 14, PCO = 11, and NPCO = 12. The CL diameter tended (P = 0.06) to be greater in P than NP (22.8 vs. 21.5 mm). The DF diameter and volume were similar between groups and averaged 12.3 mm and 987 μL. The proportion of heifers with successful FF collection and the volume recovered were similar between groups and averaged 84% and 897 μL. Relative gene expression of ISG15 in GC was 3.4-fold greater (P = 0.03) in P than NP. This difference was mainly explained by the higher expression in PIP (NPIP = 1.0; NPCO = 1.6; PCO = 3.2; PIP = 5.5), suggesting a local communication involving IFNT signaling between P uterine horn and ovary during MRP. Proteomics and lipidomics of FF will determine the presence and nature of specific endocrine signals in the ovary.