OXYTOCIN MOBILIZES CALCIUM FROM A UNIQUE HEPARIN-SENSITIVE AND THAPSIGARGIN-SENSITIVE STORE IN SINGLE MYOMETRIAL CELLS FROM PREGNANT RATS

Intracellular free Ca2+ concentration ([Ca2+](i)) was monitored using the fluorescence from the dye Fura-2-AM in single myometrial cells from pregnant rats. Oxytocin and acetylcholine applied to the cell evoked an initial peak in [Ca2+](i) followed by a smaller sustained rise which was rapidly terminated upon removal of acetylcholine or persisted after oxytocin removal. A Ca2+ channel blocker (oxodipine) and external Ca2+ removal decreased both the transient and sustained rises in [Ca2+](i) suggesting that Ca2+ influx through L-type Ca2+ channels participated in the global Ca2+ response induced by oxytocin. However, the initial peak in [Ca2+](i) produced by oxytocin was mainly due to Ca2+ store release: it was abolished by inclusion of heparin [which blocks inositol 1,4,5-trisphosphate (InsP(3)) receptors] in the pipette (whole-cell recording mode of patch-clamp) and external application of thapsigargin (which blocks sarcoplasmic reticulum Ca2+-ATPases). In contrast, the transient Ca2+ response induced by oxytocin was unaffected by ryanodine. Moreover, caffeine failed to induce a rise in [Ca2+](i) but reduced the oxytocin-induced transient Ca2+ response. The later sustained rise in [Ca2+](i) produced by oxytocin was due to the entry of Ca2+ into the cell as it was suppressed in external Ca2+-free solution. The Ca2+ entry pathway is permeable to Mn2+ ions, in contrast to that described in various vascular and visceral smooth muscle cells. Oxytocin-induced Ca2+ release is blocked by the oxytocin antagonist d(CH2)(5)[Tyr(Me)(2),Thr(4),Tyr-NH29]OVT. The prolonged increase in [Ca2+](i) after oxytocin removal is rapidly terminated by addition of the oxytocin antagonist suggesting that oxytocin dissociation from its receptor is very slow. The oxytocin stimulation of [Ca2+](i) was insensitive to incubation with pertussis toxin, and blocked by a pipette solution containing anti-alpha q/alpha(11) antibody. These data show that myometrial cells possess an unique heparin-sensitive and thapsigargin-sensitive store that can be mobilized by activation of oxytocin receptors which couples with a Gq/G(11)-protein to activate phospholipase C.