Tatsuhiko Kawarabayashi

Effect of oxytocin on spontaneous electrical and mechanical activities in pregnant human myometrium

Spontaneous intracellular electrical activity and contraction of pregnant human myometrium were recorded by the single sucrose-gap method, and the effects of oxytocin on the muscle were studied. In pregnant human myometrium at term, both plateau and spike types of action potentials were observed. All contractions were well synchronized with each action potential. Oxytocin, 10(-2) U/ml, potentiated spontaneous contractions by enhancing the plateau part of action potentials; spike-type configuration became plateau. When extracellular ionized calcium was removed, spontaneous activities disappeared, while 10(-2) U/ml of oxytocin could evoke action potentials and contractions but these were smaller than those of the controls. Spontaneous activities also disappeared when ionized calcium was increased to 5 mmol/L, but oxytocin evoked plateau potentials and contractions remarkably. Diltiazem (ionized calcium antagonist), 10(-6) gm/ml, suppressed the spontaneous activity, but oxytocin evoked action potentials and contractions in high frequency, the duration of the action potential being short and the contraction being small. In the presence of 10(-4) gm/ml of diltiazem, 10(-2) U/ml of oxytocin could not evoke any action potentials but did evoke small and long contractures, while in a high ionized potassium contracture experiment, oxytocin potentiated the tonic phase. These results suggest that oxytocin can increase spontaneous contractions by enhancing plateau potentials and that this effect requires sufficient extracellular ionized calcium. In this potentiation, the effects on frequency and amplitude of contractions might vary. It is also suggested that oxytocin may evoke a contracture in the absence of an action potential by releasing calcium from intracellular storage sites.

Oxytocin inhibits nonphasically firing supraoptic and paraventricular neurons in the virgin female rat.

Nonphasically firing activities were recorded from neurons in the supraoptic (s.o.) and paraventricular (p.v.) nucleus of the virgin female, male, and ovariectomized rat hypothalamic slice preparations. Following bath application of oxytocin (OXT), less than 1.0 x 10(-8) M, 24 (75%) of 32 virgin female s.o. neurons showed inhibitory responses and only one neuron showed excitatory response. In contrast to virgin female, five (63%) of eight male s.o. neurons were excited by OXT application. The same tendency was shown in the p.v. neurons. Among six s.o. neurons obtained from ovariectomized virgin rats, five neurons increased their firing rate to the application of OXT. After blocking synaptic transmission, the inhibitory responses of virgin s.o. neurons and the excitatory responses of ovariectomized virgin s.o. neurons to application of OXT were still observed with equal configurations. These findings suggest that the responses of nonphasic neurons to OXT may not due to synaptic drives.

Oxytocin modulates oxytocin neurons in the paraventricular nuclei of female rats throughout pregnancy and parturition

OBJECTIVE The physiologic roles of nonphasic firing oxytocin neurons in the paraventricular nucleus of female rats throughout pregnancy and parturition was studied from the viewpoint of changes in the membrane activities. STUDY DESIGN Thin colonal hypothalamic slices containing the paraventricular nucleus were prepared. Immunohistochemical studies of oxytocin in the paraventricular nucleus were performed to determine the localization of oxytocin-containing neurons. Extracellular single neural activities were recorded with a glass micropipette from the oxytocin-dense area, and physiologic concentrations of oxytocin were applied. RESULTS Nonphasic firing activity was recorded from 61 neurons in virgin female, pregnant, delivering and lactating rats. After bath application of < 5.0 x 10(-11) mol/L of oxytocin, firing rates of 12 of 17 paraventricular neurons in virgin rats decreased and only one increased, whereas the remaining four showed no response to oxytocin. In pregnancy at day 15 all four neurons decreased in activity, whereas at day 21 rats 20 of 26 neurons decreased, two increased, and four showed no response. On the contrary, in delivering rats seven of eight neurons exhibited excitatory responses, and only one showed no response. This excitation reversed to inhibition again after the lactating period ended. CONCLUSION Negative feedback by oxytocin in virgin and pregnant rats might reverse to positive feedback in delivering and lactating animals as a result of changes in hormonal conditions. This reverse might be closely related to the initiation of delivery.

Characteristics of Action Potentials and Contractions Evoked by Electrical-Field Stimulation of Pregnant Human Myometrium

Evoked electrical and mechanical activities of pregnant human myometrium were recorded by the single sucrose-gap method. Each muscle strip has an individual rhythm of membrane excitability and the evoked activity was affected by spontaneous membrane fluctuation. However, high-frequency stimulation caused changes in the rhythm of fluctuation and the pattern of action potential converted from plateau to spike type. Twitch contractions evoked by spike potentials were summated and became fused tetanus in accordance with shortening of the pulse interval. This method is suitable for clarifying the excitation-contraction relationship in human myometrium.

Clinical features of small contraction wave recorded by an external tocodynamometer

Clinical features of the small contraction wave recorded by a guardring tocodynamometer were examined retrospectively. This study included 578 patients and 6363 cardiotocographs ranging from 20 to 42 weeks of gestation. The small wave was observed in 7.5% of the cardiotocographs examined, and the rate of small-wave appearance in each gestational week tended to decrease gradually as the pregnancy progressed. This was not observed after 41 weeks of gestation. The small wave-positive group had relatively poor obstetric parameters and fetal outcome, and the small wave was frequently observed in cases of effective beta 2-stimulant intravenous infusion for the treatment of preterm labor. These results suggest that the small wave represents some degree of contractility and is ominous in general; however, the appearance of the wave does not lead to a poor prognosis in preterm labor if large phasic contractions can be abolished by beta 2-stimulant treatment.

Changes in beta-adrenergic receptors under long-term application of ritodrine in pregnant-rat myometrium.

Changes in beta-adrenergic receptor density under long-term application of ritodrine were investigated using longitudinal muscle strips of day-21-pregnant-rat myometrium. Membrane (-)[3H]dihydroalprenolol binding sites were increased by 3.1 x 10(-8) M ritodrine, with a maximum increase at 3.1 x 10(-7) M, followed by a gradual decrease at higher concentrations. Under long-term application of a lower dose (1.5 x 10(-7) M) of ritodrine, the number of the binding sites increased up to 20 min, then decreased to the control level and fluctuated after 90 min. These results suggest that at lower doses, ritodrine might activate adenylate cyclase, resulting in conformational changes in the receptors, and that receptor density might vary in parallel with cellular cAMP production under long-term application of ritodrine.

Relationship between Changes in Contraction and Cyclic AMP Contents under Long-Term Application of Ritodrine in Pregnant Rat Myometrium

The relationship between contraction changes and cAMP production under long-term application of ritodrine was investigated using longitudinal muscle strips of pregnant (Day 21) rat uterus. Cellular cAMP gradually increased up to 20 min, then decreased to the control level and fluctuated after 90 min in the presence of 1.5 x 10(-7) M ritodrine. When the dose was raised to 3.1 x 10(-4) M, cAMP contents markedly increased. However, it then decreased after 30 min and reached the control level after 180 min. To clarify the actual relationship between the changes in spontaneous contraction and cAMP contents of the same muscle strips, the strips were obtained during control contraction, suppression by 1.5 x 10(-7) M ritodrine and reactivation after suppression in the presence of the drug. Consequently, the cAMP content of reactivated muscle was significantly lower than that of suppressed muscle (p less than 0.01), however it was still higher than the control (p less than 0.05). It is speculated that the reactivation of spontaneous contractions might not depend solely on the cellular cAMP level, and that another activating mechanism, such as changes in ion permeability, might be involved in this type of desensitization phenomenon.

Effect of Methylergometrine Maleate (Methergin) on Electrical and Mechanical Activities of Pregnant Human Myometrium

The effect of methylergometrine maleate (methergin) on electrical and mechanical activities of pregnant human isthmic myometrium was examined. Methergin (10(-8)-10(-6) g/ml) enhanced plateau potential and contraction, however, the effects were not prominent. Plateau potential and contraction were also potentiated after pretreatment with 10(-7) g/ml phentolamine (alpha-blocker), however 10(-7) g/ml methergin further increased these effects. It is suggested that methergin might potentiate contractions by changing the pattern of action potentials and not by alpha-adrenoceptor.

Intracellular calcium of longitudinal muscles isolated from pregnant rat myometrium

Longitudinal muscle cells were successfully isolated from pregnant rat myometrium (21 days of gestation) with more than a 95% survival rate. The approximate size of relaxed cells was 232.2 +/- 74 microns in length and 16.2 +/- 7.0 microns in width. Using the fluorescent indicator Fura-2, the concentration of intracellular free calcium ([Ca2+]i) in resting state cells was calculated to be 116 +/- 18.5 nM. The isolated cells responded well to K+, acetylcholine and oxytocin in terms of contraction as well as the increase in [Ca2+]i. The increase in [Ca2+]i induced by acetylcholine and K+ appeared to be mainly due to an influx of extracellular Ca2+. On the other hand, the oxytocin-induced increase in [Ca2+]i was mainly due to a release of Ca2+ from intracellular storage sites in the isolated cells. Isolated longitudinal muscle cells can serve as a useful tool in establishing the relationship between [Ca2+]i and regulation of the uterine contraction at the final stage of pregnancy.

Changes in serum calcium, magnesium, cyclic AMP and monoamine oxidase levels during pregnancy and under prolonged ritodrine treatment for preterm labor

Changes in serum levels of calcium, magnesium, cAMP and monoamine oxidase (MAO) activity throughout pregnancy and during prolonged treatment for preterm labor with ritodrine were studied. cAMP levels in the third trimester of pregnancy were found to be significantly high and postpartum MAO activity was significantly low. In cases of preterm labor, serum levels of calcium, magnesium and MAO activity were significantly low during the initial stage of ritodrine administration, then recovered after more than 1 week of treatment despite the fact that given doses and serum levels of ritodrine remained constant and that the serum level of cAMP was still high. These results suggested that a control mechanism for MAO activity during pregnancy might exist due to an increase in catecholamine and that the beta 2-effect of ritodrine induced a lowering in serum calcium and magnesium levels. However, some homeostatic mechanism to regulate the ions might come into play later on the mechanism of down-regulation of ritodrine.