In smooth muscle cells, the sarcoplasmic reticulum (SR) is an important regulator of cytosolic Ca²⁺, membrane excitability, and a source of Ca²⁺ for contraction. Previous studies on the myometrium have shown that the SR has a role in limiting contraction (1). As the relative quiescence of the uterus clearly changes during gestation, the aim of this study is to investigate the role of the SR throughout pregnancy. Simultaneous measurements of intracellular Ca²⁺ ([Ca²⁺]_i) and force were made on strips of longitudinal myometrium taken from non-pregnant (NP) and pregnant rats at 9-10 days (10), 16-17 days (17) and 20-22 days (22, term day 22-23) gestation, after humane killing. The effect of the SR on spontaneous uterine activity was compared using 20μM cyclopiazonic acid (CPA), a specific inhibitor of the SR Ca²⁺ pump. The myometrial strips were spontaneously active at 32°C and means ± s.e.m. are normalized to the amplitude of control Ca²⁺ and force transients (100%). Significance was tested using the appropriate Student’s t test with significance taken at p<0.05; n is number of animals. Inhibiting SR Ca²⁺ uptake with CPA increased contraction frequency and force in all animals (n=25). The CPA- induced rise in basal force was similar in day 17 and 22 rats and was significantly increased in these late gestational groups compared with day 10 and NP rat uteri (42.7 ± 10.2%; n=10 vs 3.01 ±0.6%; n=15, respectively). To further investigate this gestational difference in response to CPA, the contribution of external Ca²⁺ entry, through voltage-gated (VOCC) or SOCE channels was determined by emptying the SR in zero Ca²⁺ solutions containing CPA (0-CPA) and then returning tissues to physiological solutions containing 2mM Ca²⁺. In NP and day 22 rats, 0-CPA induced a prolonged (>20min) rise in basal [Ca²⁺]_i that was significantly greater in day 22 compared with NP rats (142.8 ± 7.6%, n=5 vs 77.2 ± 14.4%, n=5, respectively). In 22 rats (n=3), nifedipine (1μM; an inhibitor of VOCC) reduced the amplitude of this basal [Ca²⁺]_i rise by 33.0 ± 4.4%, but SKF 96365 (50μM; an inhibitor of SOCE) inhibited almost all CPA-induced rise in basal [Ca²⁺]_i. Preliminary data suggest that the effect of both nifedipine and SKF 96365 are very similar in NP rats. These data suggest an increased contribution of the SR to quiescence towards the end of pregnancy and that SOCE contributes to its re-filling, which could be significant in labour as agonists induce store depletion causing Ca²⁺ entry and enhanced contraction.