Toshiaki Okawa

Roles of potassium channels and nitric oxide in modulation of uterine contractions in rat pregnancy

OBJECTIVE We sought to study the involvement of potassium channels in the inhibition by nitric oxide of spontaneous contractions in isolated uterine rings from midterm and term pregnant rats. STUDY DESIGN Uterine rings from Sprague-Dawley rats at midterm and term gestation were used for isometric tension recording. The inhibition of spontaneous contractile activity by potassium channel openers and nitric oxide was studied in the absence and presence of potassium channel inhibitors. RESULTS The adenosine triphosphate-dependent potassium channel opener levcromakalim inhibited spontaneous contractions in rings from both midterm and term pregnant rats in a concentration-dependent manner, and the effects were significantly attenuated by pretreatment with selective inhibitor of the adenosine triphosphate-dependent potassium channel inhibitor glibenclamide. The opener of calcium-dependent potassium channel NS 1619 inhibited spontaneous contractions in rings from midterm but significantly less so in rings from term pregnant rats in a concentration-dependent manner, and the effect was significantly attenuated by pretreatment with potassium channel inhibitors tetraethylammonium and tetrabutylammonium but not with glibenclamide. Rings from midterm and term pregnant rats were more sensitive to the inhibitory effect of levcromakalim compared with NS 1619. Nitric oxide donor diethylamine-nitric oxide inhibited spontaneous contractions in rings from midterm but significantly less in rings from term pregnant rats in a concentration-dependent manner, and the effect was attenuated by tetraethylammonium and tetrabutylammonium but not by glibenclamide. CONCLUSIONS There is gestational age-dependent refractoriness to calcium-dependent potassium but not adenosine triphosphate-dependent potassium channel opener-induced inhibition of spontaneous contractile activity of isolated rat uterine rings. Nitric oxide inhibits uterine contractions by opening of calcium-dependent potassium channels in pregnant rat myometrium. Refractoriness to nitric oxide toward term may result from decreased probability to open or number of calcium-dependent potassium channels.

The effects of nitric oxide on the contractility of isolated uterine and aortic rings from pregnant rats

OBJECTIVE The object was to compare the effects of nitric oxide on isolated uterus and aorta of pregnant rats. STUDY DESIGN Rings of uterus and thoracic aorta without endothelium from Sprague-Dawley rats at mid and late gestation were used for isometric tension recording. The concentration-response curve for diethylamine/nitric oxide was studied in the presence or absence of oxyhemoglobin (10(-5) mol/L), or oxyhemoglobin was added after the response to diethylamine/nitric oxide. RESULTS Diethylamine/nitric oxide concentration dependently inhibited uterine contractions, and the effect was attenuated by previous treatment with oxyhemoglobin at mid gestation (n = 8). The effects were negligible at late gestation (n = 8). The relaxation of aortic rings by diethylamine/nitric oxide and its attenuation by previous treatment with oxyhemoglobin were similar at mid (n = 6) and late (n = 6) gestation. The sensitivity of aortic rings to diethylamine/nitric oxide is significantly higher than that of uterine rings. Oxyhemoglobin partly restored inhibited diethylamine/nitric oxide phenylephrine tension in aortic rings and had no effect on diethylamine/nitric oxide-inhibited uterine rings. CONCLUSIONS Uterine smooth muscle is less sensitive to nitric oxide than is aortic smooth muscle. Nitric oxide sensitivity of rat uterus but not aorta decreases toward term.

Effect of placental tissue on inhibition of uterine contraction by nitric oxide donors

OBJECTIVE Our purpose was to test the hypothesis that placental tissue modulates the effect of nitric oxide on spontaneous uterine contractility in pregnant rats. STUDY DESIGN Rings (approximately 4 mm) of uterus taken from rats on day 14 (midpregnancy, n = 6), day 18 (late pregnancy, n = 4), and day 22 (term, n = 4) of gestation were placed in organ chambers filled with Krebs-bicarbonate buffer bubbled with 5% carbon dioxide in air (37 degrees C, pH approximately 7.4) for isometric tension recording. In some rings a piece of placenta was left attached to the uterine wall. In the other rings the fetuses, placentas, and membranes were removed completely. Change of spontaneous contractions of the rings (percentage change of basal integral activity for 10 minutes) in response to cumulative concentrations of the nitric oxide donors diethylamine-nitric oxide and nitroglycerin (10(-6) mol/L to 10(-4) mol/L) were compared between rings with and without placenta. RESULTS Diethylamine-nitric oxide and nitroglycerin inhibited spontaneous uterine contractions in rings from midpregnancy, in both the absence and the presence of placenta. In rings from midpregnancy, the maximal inhibition of contractions by diethylamine-nitric oxide but not by nitroglycerin was significantly (P <.05) higher in the presence (26.7% +/- 3.5% of basal activity) than in the absence (39. 6% +/- 3.3%) of placenta. Inhibition of contraction by nitric oxide donors in rings from late and term pregnancy was less than in midpregnancy, and the presence of placental tissue did not influence the responses. CONCLUSIONS The presence of placental tissue enhances inhibition of uterine contractility by agents that spontaneously release nitric oxide, such as diethylamine-nitric oxide, but not by nitroglycerin, which requires metabolic transformation for nitric oxide to be released. Refractoriness to nitric oxide near or at term does not depend on the presence or absence of placental tissue.

Effect of nitric oxide on contractions of uterine and cervical tissues from pregnant rats

Our objective was to evaluate the role of the nitric oxide (NO)–cyclic guanosine monophosphate (cGMP) pathway in rat uterine and cervical contractility at mid- and late gestation. Rings of uterus and cervix from Sprague Dawley rats on day 14 of pregnancy (mid-) and day 21 of pregnancy (late) were equilibrated at 2 g passive tension in organ chambers filled with Krebs-Henseleit solution and bubbled with 5% CO2in air (37°C, pH approximately 7.4). Rings were treated with an inhibitor of outward potassium current, tetraethylammonium, to activate phasic contractions, and the concentration–response relationships to diethylamine/NO and 8-bromo-cGMP (8-br-cGMP) were assessed. Baseline spontaneous activity was significantly higher at term gestation in both uterine and cervical rings compared with mid-gestation. Spontaneous contractile activity was not apparent in cervical rings from rats in mid-gestation, but was persistent after treatment with tetraethylammonium. Oxyhemoglobin did not affect NO-induced inhibition of activation by tetraethylammonium contractile activity in either cervical or uterine tissues in mid- or late gestation. The 8-br-cGMP concentration-dependently inhibited tetraethylammonium-activated contractions that were more pronounced in uterine tissues compared with cervical tissues in both mid- and late gestation. We concluded that activation of the NO- cGMP pathway inhibits both uterine and cervical smooth muscle contractility. Both tissues demonstrated refractoriness to NO at term.

Role of nucleotide cyclases in the inhibition of pregnant rat uterine contractions by the openers of potassium channels

OBJECTIVE Our objective was to study the involvement of adenylate and guanylate cylases in spontaneous uterine contractions and inhibition induced by the opening of potassium channels. STUDY DESIGN Uterine rings from rats at mid and term gestation and from rats at term gestation in labor were suspended in organ chambers for isometric tension recording. Concentration-response relationships to an opener of adenosine triphosphate-dependent potassium channels, levcromakalim, or to an opener of calcium-dependent potassium channels, NS 1619, were studied in the absence and presence of inhibitors of adenylate cyclase (MDL 12330 A, 2 x 10(-5) mol/L; SQ 22536, 10(-4) mol/L) or guanylate cyclase (LY 83583,3 x 10(-6) mol/L). RESULTS MDL 12330 A and SQ 22536 accentuated contractions in rings from rats at mid gestation but not at term gestation or at term gestation in labor. LY 83583 inhibited contractions in the rings from all 3 groups. Levcromakalim was equally effective in inhibiting contractions of rings from all 3 groups. MDL 12330 A, but not SQ 22536, decreased sensitivity and maximal inhibition induced by levcromakalim (term gestation greater than mid gestation greater than term gestation in labor). LY 83583 decreased the sensitivity to and maximal inhibition induced by levcromakalim in rings from pregnant rats at mid gestation. NS 1619 attenuated contraction of rings from rats at mid gestation and, to a lesser extent, at term gestation but accentuated contractions in rings from animals at term gestation in labor. MDL 12330 A, but not SQ 22536 or LY 83583, attenuated the changes induced by NS 1619 in rings from all 3 groups. CONCLUSIONS (1) The influence of nucleotide cyclases on basal uterine contractility depends on gestational age. (2) The inhibition of uterine contractions that results from the opening of calcium-dependent potassium channels depends on adenylate cyclase, whereas that of adenosine triphosphate-dependent potassium channels depends on both adenylate and guanylate cyclases. 3. Activation of adenosine triphosphate-dependent potassium channels is more efficient than activation of calcium-dependent potassium channels. 4. The inhibition induced by calcium-dependent potassium channel openers, but not adenosine triphosphate-dependent potassium channel openers, decreases as pregnancy progresses, and at delivery the activation of spontaneous contractile activity is evident.

Influence of progesterone on myometrial contractility in pregnant mice treated with lipopolysaccharide

Aim:  To evaluate the effect of progesterone on interleukin (IL)‐6, prostaglandin (PG) E2 and nitric oxide (NO) metabolite (NOx) production and contractile activity by NO in pregnant mice treated with lipopolysaccharide (LPS).

The influence of prostaglandins on the contractile response to electrical field stimulation in rabbit myometrial and cervical smooth muscles

Indomethacin (10 microM) significantly reduced the contractile responses to electrical field stimulation (EFS) in the non-pregnant myometrium (5-50 Hz by an average of 25.7%) and the pregnant myometrium (30-50 Hz by an average of 16.5%). On the other hand, it significantly increased the contractile responses of the cervix in non-pregnant (2.5-30 Hz by an average of 21.5%) and pregnant (2.5-10 Hz by an average of 12.5%) muscular strips. Prostaglandin (PG) E2 (0.1-1000 nM) strongly and PGI2 (0.1-1000 nM) slightly and dose-dependently inhibited the contractile response to EFS of the non-pregnant cervical muscle, but not of the myometrium. In the pregnant uterus, the contractile responses of the cervical muscle were similar to those of the non-pregnant cervix, while the response of the myometrium was increased significantly and dose-dependently by the application of PGE2 or PGI2.PGF2 alpha (1000 nM) significantly increased the contractile response of the myometrium but not cervical muscle to EFS in the pregnant rabbits. In non-pregnant muscles, the contractile responses of the myometrium and cervical smooth muscle to EFS were not influenced by the treatment with PGF2 alpha.PGE2 (0.1-1000 nM) also inhibited the contractile response to direct muscle stimulation, but the inhibition was significantly less than that of the response to indirect stimulation. PGI2 slightly inhibited the contractile responses to both indirect and direct stimulation. These findings indicate that PGE2, PGI2 and PGF2 alpha increase the contractile response of pregnant myometrium to EFS.(ABSTRACT TRUNCATED AT 250 WORDS)

Nitric oxide donor-induced inhibition of pregnant rat uterine spontaneous contractile activity and release of nitric oxide from uterus measured by microdialysis.

Our aim was to study whether nitric oxide (NO) donor-induced inhibition of pregnant rat myometrium contractility correlates with the release of NO. Uterine rings from mid-pregnant and late pregnant Sprague-Dawley rats were used for isometric tension recording. Concentration-response relationships to sodium nitroprusside (SNP), nitroglycerine (NTG) and diethylamine (DEA)/NO were assessed. The time course of NO release after addition to the organ chambers of the 3 NO-donors was assessed by the detection of NO products NOx (NO3+NO2) using the microdialysis probe by a HPLC-NO detector system. DEA/NO induced greater inhibition of the spontaneous contractile activity of uterine rings from mid-pregnant rats than SNP or NTG. In uterine rings from late pregnant rats, however, the maximal inhibition of the contractility by all 3 NO-donors were significantly less. The NOx levels measured in the uterine ring walls from either mid-pregnant or late pregnant rats significantly increased after DEA/NO as compared to the basal levels or the levels after NTG or SNP. The decrease of NO-donor-induced inhibition of rat myometrium contractility, with unchanged formation of NOx, at term, suggests that the changes in NO signaling are responsible for gestational age-dependent attenuation of the inhibitory effect.

Oxytocin, PGE2 and PGF2α stimulate the production of inositol phosphates in the rabbit myometrium

SummaryPhosphoinositide breakdown is thought to be important in regulating a variety of transmembrane signal transduction in the action of oxytocic agent during uterine smooth muscle contraction. We investigated the effects of oxytocin and prostaglandins (PGE2 and PGF2α) on phosphoinositide hydrolysis in the myometrium taken from non-pregnant and pregnant rabbits by measuring the accumulation of total inositol phosphates (IP). Oxytocin strongly, and PGE2 and (PGF2α) slightly but significantly, stimulated IP production in both the non-pregnant and pregnant myometria. Oxytocin more markedly accelerated the IP production in pregnant myometrium than in non-pregnant myometrium. However, IP production stimulated by PGE2 and PGF2α was much the same in non-pregnant and pregnant myometria. The amount and time course of the increase in the production of the total IPs by oxytocin are quite different from those by PGs. It seems that the mechanism by which oxytocin stimulates phospholipase C is different from that of the PGs. It is suggested that transmembrane signalling pathways of phosphoinositide hydrolysis play an important role in the each mechanism.

Effects of nitric oxide and prostacyclin on hemodynamic response by big endothelin-1 in near term fetal sheep.

Abstract Aims: To measure plasma concentrations of endothelin (ET)-1, NO metabolites (nitrate/nitrite; NOx) and 6-keto PGF1α (PGF1α) in maternal and fetal sheep blood, and to evaluate the effects of big ET-1 on hemodynamic response, blood gases and NO and 6-keto PGF1α production in near term fetal sheep. Methods: Hemodynamic parameters were measured during infusion of big ET-1 into the carotid vein in chronically catheterized fetal sheep on day 125 of gestation. Fetal arterial blood samples were obtained for ET-1, PGF1α) and nitrate/nitrite (NOx) measurements. Results: ET-1, NOx and PGF1α plasma concentrations were all significantly higher in fetal compared with the maternal plasma. Big ET-1 significantly decreased fetal systolic and diastolic blood pressure and significantly increased fetal heart rate. Big ET-1 stimulated plasma PGF1α), but not NOx, concentration. Conclusions: Circulatory regulating factors in the fetus were up-regulated. The effects of ET-1 on fetal hemodynamic respnse may be mediated via prostacyclin, but not via the NO pathway.