Better tocolytics are required to help prevent preterm labour. The gaseotransmitter Hydrogen sulphide (H 2S) has been shown to reduce myometrial contractility and thus is of potential interest. However previous studies used NaHS, which is toxic and releases H 2S as a non-physiological bolus and thus alternative H 2S donors are sought. GYY4137 has been developed to slowly release H 2S and hence better reflect endogenous physiological release. We have examined its effects on spontaneous and oxytocin-stimulated contractility and compared them to NaHS, in human and rat myometrium, throughout gestation. The effects on contractility in response to GYY4137 (1 nM–1 mM) and NaHS (1 mM) were examined on myometrial strips from, biopsies of women undergoing elective caesarean section or hysterectomy, and from non-pregnant, 14, 18, 22 day (term) gestation or labouring rats. In pregnant rat and human myometrium dose-dependent and significant decreases in spontaneous contractions were seen with increasing concentrations of GYY4137, which also reduced underlying Ca transients. GYY4137 and NaHS significantly reduced oxytocin-stimulated and high-K depolarised contractions as well as spontaneous activity. Their inhibitory effects increased as gestation advanced, but were abruptly reversed in labour. Glibenclamide, an inhibitor of ATP-sensitive potassium (K ATP) channels, abolished the inhibitory effect of GYY4137. These data suggest (i) H 2S contributes to uterine quiescence from mid-gestation until labor, (ii) that H 2S affects L-type calcium channels and K ATP channels reducing Ca entry and thereby myometrial contractions, (iii) add to the evidence that H 2S plays a physiological role in relaxing myometrium, and thus (iv) H 2S is an attractive target for therapeutic manipulation of human myometrial contractility.