The effects of two enkephalinase inhibitors, acetorphan and thiorphan, and the enkephalin analogue [d-Ala2\p=n-\Met5]-enkephalinamide (DAMEA), on spontaneous uterine contractions were studied at day 21 of pregnancy in rats following treatment in vivo or in vitro. Acetorphan (10 mg kg\m=-\1)and thiorphan (1 mg kg\m=-\1),immediately after their i.v. administration, increased the duration of spontaneous contractions 3.4- and 4.6-fold, respectively, but did not modify the maximum amplitude. Similarly, thiorphan (40 \g=m\moll\m=-\1) increased the duration of contractions when administered in vitro. Thiorphan was ineffective during the first 30 min when given into the cerebral ventricles (50 \g=m\gper rat). These results suggest that the enkephalinase inhibitors are acting via a peripheral opioid pathway; and this conclusion is supported by the observation that thiorphan potentiated the stimulatory effect of a submaximal dose of DAMEA administered in vitro. The excitatory effects of DAMEA and the enkephalinase inhibitors were blocked by naloxone. This antagonistic effect of naloxone on uterine motility in the periparturient rat uterus, induced by either acetorphan and thiorphan or DAMEA, seems to be regulated by peripheral opiate receptors. Naloxone (10 mg kg\m=-\1 s.c.) increased both the amplitude and duration of uterine motility in vivo; however, naloxone (26 \g=m\moll\m=-\1 and 52 \g=m\moll\m=-\1) produced a paradoxical dose-dependent biphasic effect in vitro.