Prostaglandin E2 (PGE2) plays a key role in the maintenance of human pregnancy and labour onset. PGE2 can elicit diverse actions within the uterus depending on the PGE2 receptors (EP1, EP2, EP3 and EP4) expressed. By signalling through different intracellular pathways the EP receptors may inhibit or promote smooth muscle contractility. Nine different EP3 receptor splice variants have been identified with divergent signalling pathways. RT-PCR and immunohistochemistry were utilized to identify and localize EP receptor isoforms within the upper segment (US) and lower segment (LS) myometrium. EP1 was significantly increased in the LS myometrium with term labour. EP3 (and EP3 splice variants EP3I(1b), EP3II, EP3III and EP3IV) was down-regulated in pregnancy (US and/or LS) with a further decrease at term labour in the LS. Overall, expression of EP2 was significantly higher in the LS while EP3 was significantly higher in the US. No significant EP4 changes were observed. Consistent with the RT-PCR results, immunohistochemistry confirmed the presence and, interestingly, showed nuclear localization of EP receptors in the myometrium with higher EP1 expression and lower expression of EP3. The differential regulation of EP receptors within the myometrium indicates that they may play a role in controlling the onset and maintenance of human labour.