Oxytocin induced a biphasic increase in the intracellular Ca2+ concentration of porcine myometrial cells: Participation of a pertussis toxin—insensitive G‐protein, inositol 1,4,5‐trisphosphate—sensitive Ca2+ pool, and Ca2+ channels

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Abstract
This study investigated the underlying mechanisms of oxytocin (OT)‐induced increases in intracellular Ca2+ concentrations ([Ca2+]i) in acutely dispersed myometrial cells from prepartum sows. A dosedependent increase in [Ca2+]i was induced by OT (0.1 nM to 1 μM) in the presence and absence of extracellular Ca2+ ([Ca2+]e). [Ca2+]i was elevated by OT in a biphasic pattern, with a spike followed by a sustained plateau in the presence of [Ca2+]e. However, in the absence of [Ca2+]e, the [Ca2+]i response to OT became monophasic with a lower amplitude and no plateau, and this monophasic increase was abolished by pretreatment with ionomycin, a Ca2+ ionophore. Administration of OT (1 μM) for 15 sec increased inositol 1,4,5‐trisphosphate (IP3) formation by 61%. Pretreatment with pertussis toxin (PTX, 1 μg/ml) for 2 hr failed to alter the OT‐induced increase in [Ca2+]i and IP3 formation. U‐73122 (30 nM to 3 μM), a phospholipase C (PLC) inhibitor, depressed the rise in [Ca2+]i by OT dose dependently. U‐73122 (3 μM) also abolished the OT‐induced IP3 formation. Thapsigargin (2 μM), an inhibitor of Ca2+‐ATPase in the endoplasmic reticulum, did not increase [Ca2+]i. However, it did time‐dependently inhibit the OT‐induced increase in [Ca2+]i. Nimodipine (1 μM), a Voltage‐dependent Ca2+ channel (VDCC) blocker, inhibited the OT‐induced plateau by 26%. La3+ (1 μM), a nonspecific Ca2+ channel blocker, abrogated the OT‐induced plateau. In whole‐cell patch‐clamp studies used to evaluate VDCC activities, OT (0.1 μM) increased Ca2+ Current (Ica) by 40% with no apparent changes in the current‐voltage relationship. The OT‐induced increase in Ica reached the maximum in 5 min, and the increase was abolished by nimodipine (1 μM). These results suggested that (1) activation of OT receptors in porcine myometrium evokes a cascade in the PTX‐insensitive G‐protein–PLC‐IP3 signal transduction, resulting in an increase in [Ca2+]i; (2) the OT‐induced increase in [Ca2+]i is characterized by a biphasic pattern, in which the spike is predominately contributed by the intracellular Ca2+ release from the IP3‐sensitive pool, and to a lesser extent by Ca2+ influx, whereas the plateau is from increased Ca2+ influx; and (3) the influx is via VDCC and receptor‐operated Ca2+ channels.