AbstractNifedipine, an L‐type voltage‐gated Ca2+ channel (L‐VGCC) blocker, is one of the most used tocolytics to treat preterm labor. In clinical practice, nifedipine efficiently decreases uterine contractions, but its efficacy is limited over time, and repeated or maintained nifedipine‐based tocolysis appears to be ineffective in preventing preterm birth. We aimed to understand why nifedipine has short‐lasting efficiency for the inhibition of uterine contractions. We used ex vivo term pregnant human myometrial strips treated with cumulative doses of nifedipine. We observed that nifedipine inhibited spontaneous myometrial contractions in tissues with high and regular spontaneous contractions. By contrast, nifedipine appeared to increase contractions in tissues with low and/or irregular spontaneous contractions. To investigate the molecular mechanisms activated by nifedipine in myometrial cells, we used the pregnant human myometrial cell line PHM1‐41 that does not express L‐VGCC. The in vitro measurement of intracellular Ca2+ showed that high doses of nifedipine induced an important intracellular Ca2+ entry in myometrial cells. The inhibition or downregulation of the genes encoding for store‐operated Ca2+ entry channels from the Orai and transient receptor potential‐canonical (TRPC) families in PHM1‐41 cells highlighted the implication of TRPC1 in nifedipine‐induced Ca2+ entry. In addition, the use of 2‐APB in combination with nifedipine on human myometrial strips tends to confirm that the pro‐contractile effect induced by nifedipine on myometrial tissues may involve the activation of TRPC channels.