The decrease in the level of estradiol (E-2) in granulosa cells caused by lipopolysaccharide (LPS) is one of the major causes of infertility underlying postpartum uterine infections; the precise molecular mechanism of which remains elusive. This study investigated the role of endoplasmic reticulum (ER) stress in LPS-induced E-2 decrease in mouse granulosa cells. Our results showed that LPS increased the pro-inflammatory cytokines [(interleukin (IL)-1 beta, IL-6, IL-8, and tumor necrosis factor (TNF)-alpha)], activated ER stress marker protein expression [(glucose-regulated protein 78 (GRP78) and CCAAT/enhancer-binding protein homologous protein (CHOP)], and decreased cytochrome P450 family 19 subfamily A member 1 (Cyp19a1) expression and E-2 production. Moreover, inhibition of ER stress by 4-phenylbutyrate (4-PBA) attenuated thapsigargin-(TG, ER stress agonist) or LPS-induced reduction of Cypl 9a1 and E2, pro-inflammatory cytokines expression (IL-1 beta, 1L-6, IL-8, and TNF-alpha), and the expression of CHOP and GRP78. Additionally, inhibition of toll-like receptor 4 (TLR4) by resatorvid (TAK-242) reversed the inhibitory effects of LPS on Cyp19a1 expression and E2 production, activation of GRP78 and CHOP, and expression of IL-1 beta, IL-6, IL-8, and TNF-alpha. In summary, our study suggests that ER stress is involved in LPS-inhibited E-2 production in mouse granulosa cells.