In vitro contractile effect of platelet-activating factor on guinea-pig myometrium

Platelet-activating factor (PAF) evoked myometrial contractions in two different patterns, depending on whether spontaneous activity was present. In spontaneously active myometrial strips (58%), both PAF and oxytocin enhanced the amplitude of myometrial contractions. In quiescent myometrial strips, PAF induced contractions characterized by a prompt development of tension, a plateau, and a final, rapid relaxation. In 54% of these strips, PAF-induced contraction was followed by rhythmic activity. PAF contractile response was dependent upon the concentration (0.1-100 nM); the minimal effective concentration of PAF was 0.1 nM and the EC50 was 1 nM. The response to oxytocin (0.01-10 mU/ml), assumed as reference stimulus, was characterized by a prompt development of tension, which was followed by a sustained, slow contraction and relaxation. PAF response was almost completely dependent on cyclooxygenase and partially on lipoxygenase pathways, as inferred from studies with indomethacin and FPL 55712, respectively. A receptor mediated mechanism of PAF action was suggested by specific desensitization of the myometrium to a second challenge with an equimolar concentration of PAF (but not with oxytocin) and the blocking effect of CV 3988, a specific PAF receptor antagonist.     

In vitro contractile effect of platelet-activating factor on guinea-pig myometrium

Platelet-activating factor (PAF) evoked myometrial contractions in two different patterns, depending on whether spontaneous activity was present. In spontaneously active myometrial strips (58%), both PAF and oxytocin enhanced the amplitude of myometrial contractions. In quiescent myometrial strips, PAF induced contractions characterized by a prompt development of tension, a plateau, and a final, rapid relaxation. In 54% of these strips, PAF-induced contraction was followed by rhythmic activity. PAF contractile response was dependent upon the concentration (0.1–100 nM); the minimal effective concentration of PAF was 0.1 nM and the EC₅₀ was 1 nM. The response to oxytocin (0.01–10 mU/ml), assumed as reference stimulus, was characterized by a prompt development of tension, which was followed by a sustained, slow contraction and relaxation. PAF response was almost completely dependent on cyclooxygenase and partially on lipoxygenase pathways, as inferred from studies with indomethacin and FPL 55712, respectively. A receptor mediated mechanism of PAF action was suggested by specific desentization of the myometrium to a second challenge with an equimolar concentration of PAF (but not with oxytocin) and the blocking effect of CV 3988, a specific PAF receptor antagonist.     

Beta-adrenergic relaxation of dog trachealis: contractile agonist-specific interaction

The phenomenon of contractile agonist-dependent relaxation by isoproterenol (ISO) of active tension elicited by acetylcholine (ACh), histamine (HIS), serotonin (5-HT), and potassium chloride-substituted Krebs-Henseleit solution (KCl) was studied in 210 tracheal smooth muscle (TSM) strips from 28 mongrel dogs in vitro. All TSM strips were contracted to similar active tensions [target tension (TT) = 50% of the maximal active tension elicited by 127 mM KCl] with ACh, HIS, 5-HT, or KCl and relaxed with either ISO, forskolin (FSK), N6,2'-O-dibutyryladenosine 3',5'-cyclic monophosphate (db-cAMP), or 3-isobutyl-1-methylxanthine (IMX). The concentrations of ISO causing 50% relaxation from TT (RC50) were ACh (2.9 +/- 1.1 x 10(-6) M) greater than 5-HT (8.4 +/- 1.5 x 10(-8) M) approximately KCl (8.1 +/- 2.1 x 10(-8) M) greater than HIS (1.6 +/- 0.2 x 10(-8) M). FSK and IMX relaxed TSM in the same rank order of potency as ISO. In contrast to the contractile agonist-dependent relaxation elicited by ISO, FSK, and IMX, db-cAMP was nearly equipotent in relaxing similarly contracted strips. These results are consistent with contractile agonist-specific interaction with cAMP production by ISO and FSK. These data demonstrate that the phenomenon of contractile agonist-dependent relaxation by ISO is not related specifically to the beta-adrenoceptor.     

Antagonism of relaxation to isoproterenol caused by agonist interactions

The interaction of contractile agonists on the relaxation elicited with isoproterenol (ISO) was studied in 112 tracheal smooth muscle (TSM) strips from 20 dogs in vitro. Strips were contracted to the same active target tension (TT) with acetylcholine (ACh), histamine (HIS), serotonin (5-hydroxytryptamine, 5-HT), potassium chloride (KCl), or the combinations of ACh + HIS, ACh + 5-HT, HIS + KCl, HIS + 5-HT (50% TT from each agonist). Although a less potent agonist, adding HIS to cause 50% of the TT reduced the concentration of ACh to elicit the remaining 50% TT and substantially altered relaxation by ISO compared with HIS alone [concentration required to achieve 50% relaxation (RC50) = 9.2 +/- 2.4 X 10(-8) vs. 9.0 +/- 4.4 X 10(-9) M to HIS alone; P less than 0.003]. Relaxation for TSM strips contracted with ACh + HIS was comparable to that elicited from the same TT with ACh alone, although concentrations required in combination were lower than for either agonist alone. Trachealis strips contracted equivalently with KCl + HIS also had augmented contraction and attenuated relaxation (RC50 = 3.7 +/- 0.8 X 10(-8) M; P less than 0.015 vs. HIS alone). However, combinations of 5-HT + ACh and 5-HT + HIS did not alter relaxation to ISO from that elicited by the weaker agonist alone. We demonstrate that TSM relaxation depends on the combination of agonists eliciting contraction and may be inhibited substantially by interactions among contractile agonists.     

Long-term hypoxia changes myometrial responsiveness and oxytocin receptors in the pregnant ewe: differential effects on longitudinal versus circular smooth muscle

Previous studies from our laboratory demonstrated that long-term hypoxia (LTH) altered in vitro contractile responses to oxytocin in full-thickness myometrial strips from pregnant sheep. The present study was designed to determine, first, if the reduced contractile response to oxytocin following LTH is the result of combined effects on longitudinal and circular smooth muscle or if the effect is specific to a single muscle layer and, second, if the reduced contractile response to oxytocin following LTH is caused by changes in oxytocin-receptor protein. Pregnant ewes were maintained at high altitude (3820 m) from Day 30 to Days 137-142 of gestation, when the ewes were killed for collection of myometrial tissue. Tissue was also collected from age-matched, normoxic controls. Longitudinal and circular layers were separated, length-tension curves generated to determine optimal resting tension, and all strips exposed to increasing half-log doses of oxytocin ranging from 10-12 to 10-6.5 M. The expression of oxytocin-receptor protein was measured using Western blot analysis. We found that LTH did not affect KCl-induced contraction of either smooth muscle layer, whereas the sensitivity of both myometrial layers to oxytocin was altered. A decreased maximum contractile response of the circular layer to oxytocin was also observed. Additionally, LTH decreased expression of oxytocin-receptor protein in the circular layer and increased levels in the longitudinal layer. Results from the present study indicate that LTH alters contractile responses and oxytocin-receptor protein expression in a layer-specific manner in the pregnant sheep myometrium.     

Morinda lucida reduces contractility of isolated uterine smooth muscle of pregnant and non-pregnant mice.

The present work investigated the effect of Morinda lucida (M. lucida) extract on isolated uterine smooth muscle of pregnant and non-pregnant mice. Pregnant and non-pregnant mice were pretreated with oral stilboesterol (0.1 mg/kg body weight) and killed by cervical dislocation. Thin strips of the uterus were cut and mounted in a 20ml organ bath containing De Jalon solution bubbled with 95%O2-5% CO2 gas mixture. The strips were connected to a force transducer coupled to a Grass 7D Polygraph for the recording of isometric tension. Effects of graded concentrations of oxytocin (OXY; 10-5-10-2 mol/L), acetylcholine (ACh; 10-9-10-5 mol/L) and M. lucida extract (0.015-1.5 mg/ml) were recorded. Fresh uterine strips were then incubated with M. lucida extract for 5mins and cumulative response to OXY was repeated. Another set of fresh strips was incubated in L-NAME for 15mins and the cumulative responses to M.lucida extract were repeated. OXY resulted in increased contractile responses in both pregnant and non-pregnant uterine muscles. M. lucida resulted in relaxation of the uterine smooth muscle in both pregnant and non-pregnant mice at all doses. However, at 1.5mg/ml, M. lucida completely blocked spontaneous uterine contractions. Following incubation with L-NAME, M. lucida extract led to a slightly greater relaxation of the uterine strips. In conclusion, M. lucida reduced contractility of uterine smooth muscle in both pregnant and non-pregnant mice as well as blocking contractile responses to OXY and Ach in uterine smooth muscle of pregnant and non-pregnant mice. There was no significant alteration of M. lucida activity by L-NAME suggesting that the action of the compound on uterine muscle is not associated with impaired nitric oxide synthase.     

Effects of calcium channel blocker, mibefradil, and potassium channel opener, pinacidil, on the contractile response of mid-pregnant goat myometrium

The present study was undertaken to investigate the in vitro influence of mibefradil, a calcium channel blocker, and pinacidil, a potassium channel opener, on pregnant goat myometrial spontaneous rhythmic contractility and contractions induced with the agonist, oxytocin. Longitudinal strips from the distal region of uterus, collected from goats at midgestation, were mounted in an organ bath for recording isometric contractions. Mibefradil (10(-8)-10(-4) M) or pinacidil (10(-10)-10(-4) M), added cumulatively to the bath at an increment of 1 log unit, caused concentration-dependent inhibition of the spontaneous rhythmic contractions of isolated uterine strips. The rhythmic contraction was, respectively, abolished at 100 and 10 microM concentrations of mibefradil and pinacidil. In a concentration-dependent manner, mibefradil (1 and 10 microM) antagonized the contractions elicited with oxytocin (10(-5)-10(-2) IU). Pretreatment of uterine strips with glibenclamide (10 microM), a selective KATP channel blocker, caused a rightward shift of the concentration-response curve of pinacidil with a concomitant decrease in its pD2 value. Pinacidil (0.3, 1 and 3 microM), in a concentration-related manner, antagonized the oxytocin (10(-5)-10(-2) IU)-induced contractile response. The inhibition of spontaneous rhythmic contractions and antagonism of oxytocin-induced contraction by mibefradil in the pregnant goat myometrium may be related to the antagonism of voltage-dependent Ca2+ channels, while by pinacidil suggests that KATP channel could be a therapeutic target for tocolysis.     

Oxytocin enhances the basal release of uterine prostaglandin F2 alpha, but not that of PGE1, or of PGE2, and changes the metabolism of exogenous arachidonate, favouring the formation of prostaglandin F2 alpha and 5-HETE. Relationships with its uterotonic action and modulation by estradiol

We attempted to explore possible mechanism(s) subserving the influence of oxytocin on uterine motility by studying the action of the hormone on: 1) the contractile activity of isolated rat uteri in the presence or absence of indomethacin; 2) the synthesis and release of prostaglandins (PGs) into the solution incubating the uterine tissue as well as the metabolism of labelled arachidonic acid; 3) the uptake of 45Ca2+ by uterine strips. The experiments were bone with uterine preparations isolated from spayed rats treated or not with 17-beta-estradiol. The values of isometric developed tension (IDT) and of frequency of contractions (FC) induced by oxytocin in uterine strips isolated from spayed and spayed-estrogenized rats, were not modified by indomethacin at 10(-6) M. On the other hand, uterine strips from untreated spayed rats, release into the incubating medium approximately equal amounts of PGE1, PGE2 and PGF2 alpha. The in vitro presence of oxytocin (50 mU/ml) increased significantly (p 0.05) the output of PGF 2 alpha without changing the release of PGE1 or PGE2. Uteri from spayed rats injected prior to sacrifice with 17-beta-estradiol released significantly less PGE1 and PGE2 (p less than 0.005) than preparations from non-injected animals, whereas the output of PGF2 alpha in the suspending solution remained unchanged. Following estrogenization the addition of oxytocin to preparations obtained from spayed-estrogenized rats also increased the output of uterine PGF2 alpha (p less than 0.001) without changing that of PGs E1 or E2.(ABSTRACT TRUNCATED AT 250 WORDS)     

Parasympathetic involvement in PAF-induced contraction in canine trachealis in vivo

We studied the contractile response elicited by platelet-activating factor (PAF) administered intra-arterially into the tracheal circulation of 34 dogs in vivo. A method that avoided tachyphylaxis encountered in prior investigations was developed for isometric measurement of multiple dose-response effects. PAF was a very potent contractile agent; active tension was elicited with 10(-11) mol ia PAF. To determine the mechanism by which contraction was induced, dose-response curves were generated in groups of five animals each treated with either 0.5 mg/kg (approximately 1.5 X 10(-5) mol) iv + 10(-3) mg/kg (3 X 10(-8) mol) ia atropine, 5 mg/kg iv indomethacin (INDO), or 7.5 mg/kg iv hexamethonium (HEX). After pretreatment with atropine, contraction still was elicited with 10(-11) mol ia PAF. However, maximal contraction was only 16.2 +/- 2.74 g/cm (vs. 35.7 +/- 5.74 g/cm for untreated controls; P less than 0.02). The dose at which maximal contraction was elicited after atropine was 10(-7) mol ia (vs. 1.9 X 10(-9) mol for controls; P less than 0.001). Pretreatment with INDO caused minimal attenuation, and HEX had no effect on the response elicited by ia PAF. We demonstrate a method for assessing the effects of PAF in central airways that avoids tachyphylaxis and permits dose-response studies in the same animal. We also demonstrate that PAF is an extremely potent mediator that elicits tracheal smooth muscle contraction at least in part by postganglionic activation of parasympathetic nerves. A direct contractile effect of PAF which is not related to secretion of products of the cyclooxygenase pathway is also suggested.     

Effects of stimulation of nervous fibres upon contractile activity of the tracheobronchial lymphatic node capsules' smooth muscles

The contractile activity of smooth muscle of tracheobronchial lymph nodes' capsules was recorded in vitro. The field electric stimulation (0.5 ms pulses, 55 V nominal, 4 min trains at frequencies 0.5, 1, 2, 4, 8, 16 and 32 Hz) of strips from lymph node produced a frequency-dependent increase in baseline tension and frequency of phase contractions. Evoked contractions were significantly (about 80%) reduced by tetrodotoxin (1 x 10(-6) M). The blockage of M-cholinoreceptors with atropine (1 x 10(-6) M) did not affect the field-evoked responses. Contractile field-evoked effects were significantly reduced by the phentolamine (1 x 10(-7)-1 x 10(-6) M) but not completely. Field-evoked contractions were slightly affected by the propranolol (1 x 10(-7)-1 x 10(-6) M). We conclude that the contractile activity of bovine tracheobronchial lymph node capsular smooth muscle is modulated by excitatory adrenergic nerves.     

A novel anti-asthmatic quinone derivative, AA-2414 with a potent antagonistic activity against a variety of spasmogenic prostanoids

The anti-asthmatic activity of AA-2414 [(+/-)-7-(3,5,6-trimethyl-1,4-benzoquinon-2-yl)-7-phenylheptano ic acid] has been studied in vivo and in vitro. Experimental allergic asthma was inhibited by orally administered AA-2414 in a dose-dependent manner. AA-2414, 0.08-1.25 mg/kg (p.o.), inhibited the bronchconstriction in guinea pigs induced by a prostaglandin endoperoxide analogue (U-46619), leukotriene D4 (LTD4), and platelet activating factor (PAF) with a long duration of action. The compound did not inhibit histamine-induced bronchoconstriction. AA-2414 reduced the induction of pulmonary inflation caused by LTD4 aerosol inhalation. AA-2414 competitively inhibited the contractile response to U-46619 in guinea pig tracheal and parenchymal strips and dog saphenous vein strips with pA2 values of 7.69, 8.29 and 6.79, respectively. Furthermore, the contractile responses of guinea pig tracheal strip to PGD2, 9 alpha, 11 beta-PGF2 and PGF2 alpha were inhibited with pA2 values of 7.20, 7.79 and 5.71, respectively. These results suggest that AA-2414, a quinone derivative, is a novel, potent and orally active antagonist of a variety of spasmogenic prostanoids.     

Pharmacological analysis of reactivity changes in airways due to acute inflammation in cats

Our recent in vitro studies on airways smooth muscles of the cat with turpentine oil inflammation showed the occurrence of a contractile response of tracheal preparations and a significant increase in the isometric tension of lung strips to histamine application. This study was aimed to establish whether histamine H2-receptors participated in the changed in vitro reactivity of the airways smooth muscles of cats suffering from experimentally induced airway inflammation. Pretreatment of control tracheal preparations, control and experimental groups of the lung strips by cimetidine did not change the character of the histamine response. Similarly, the amplitude of histamine relaxation, of the tracheal preparations partially contracted by carbachol was unchanged by experimental inflammation. Clemastine significantly shifted the histamine dose-response curves to the right in both groups of lung strips. However, significant differences in lung strip reactivity between control and experimental groups of cats were not eliminated. Our results do not support the role of histamine H2-receptors in the pathologically increased airway reactivity to histamine in vitro.     

Role of extracellular Ca2+ in diaphragmatic contraction: effects of ouabain, monensin, and ryanodine.

The effects of zero extracellular Ca2+ on the contractility of rat diaphragmatic strips in vitro were studied in conjunction with various pharmacological agents known to influence the intracellular Ca2+ concentration: the Na+ ionophore, monensin, and the Na(+)-K+ pump inhibitor, ouabain, which enhance [Ca2+]i, caffeine, which induces Ca2+ release from the sarcoplasmic reticulum (SR), and ryanodine, which prevents Ca2+ retention by the SR. The effect of increasing [Ca2+]i on diaphragmatic contraction was assessed by comparing contractions induced by 120 mM K+ in the small muscle strips before and after the addition of ouabain or monensin. Monensin (20 microM) and ouabain (1-100 microM) augmented contractions up to threefold. Treatment of diaphragm strips with 3 nM ryanodine increased baseline tension 360% above the original resting tension but only if the diaphragm was electrically stimulated concurrently; 100 microM ryanodine induced contracture in quiescent tissue. High K+ contractures were of greater magnitude in the presence of ryanodine compared with control, and relaxation time was prolonged by greater than 200%. Ca(2+)-free conditions ameliorated these actions of ryanodine. Ryanodine reduced contractions induced by 10 mM caffeine and nearly abolished them in Ca(2+)-free solution. The data demonstrate that extracellular Ca2+ is important in certain types of contractile responses of the diaphragm and suggest that the processes necessary to utilize extracellular Ca2+ are present in the diaphragm.     

Cardiodepressive effect of platelet activating factor

The cardiodepressive effect of PAF has been studied on the electrical and mechanical activities of isolated auricles of guinea pig. Intracellular resting potential, action potential (AP) and isometric contractions elicited by electrical stimulation (0.5 Hz) were measured. PAF (10(-7) M) induced negative inotropic effect, which reached its peak after 5 min with 23.5 +/- 6.6% in respect to prechallenge values (n = 8). After 20 min negative inotropic effect relaxed to 39.6 +/- 8.8%. 1 min after the beginning of washing in Tyrode solution, positive inotropic effect of PAF was evident, that reached its peak (217 +/- 49.5%) after 2 min, decayed after 5-10 min to normal values. PAF did not modify the resting membrane potential, produced a decrease in the amplitude and Vmax of the upstroke AP, shortened the AP duration. Ca-AP and contractions, elicited in partially depolarized myocardium were decreased by PAF (10(-7) M). PAF-produce the change of the AP and the negative effect on auricle contractile force was inhibited in muscles pretreated with 3mM 4 aminopyridine. Histamine (10(-4) M) was also capable of neutralizing the depressant effect of PAF. The obtained results suggested that PAF effects on the membrane of cardiac cells could be related to a change in Ca and K conductance.     

Effect of a diabetic state on myometrial ultrastructure and isolated uterine contractions in the rat

Alterations in rat myometrial ultrastructure and in vivo uterine contractile responses to oxytocin were examined in estradiol-treated (40 micrograms/kg) euglycemic and streptozotocin-induced (85 mg/kg) diabetic rats. Myometrial morphology was examined 18, 24, and 36 hr after estradiol administration. At the time points examined, nuclei of myometrial cells from euglycemic and diabetic rats were pleomorphic and contained large areas of heterochromatin dispersed throughout the nuclei. Mitochondria were round to oval in shape and contained a dense matrix with cristae that extended across the mitochondria. Myofilaments were found in both euglycemic and diabetic cells but the relative number of myofilaments in diabetic cells appeared to be less than the number found in myometrial cells removed from euglycemic animals. The number of free cytoplasmic ribosomes in diabetic cells also appeared to be less than those found in euglycemic cells. In order to determine if apparent differences in the number of myofilament found in diabetic myometrial cells could be correlated with changes in uterine contractile responses to hormones, oxytocin dose-response curves (10(-8) to 10(-5) M) were examined in isolated uteri removed from saline-injected and estradiol-injected (24-hr pretreatments) euglycemic and diabetic rats. The maximal contractile responses (milligrams tension developed per milligrams tissue) in saline-injected euglycemic and diabetic rats were 49 +/- 5 and 36 +/- 4, respectively, while maximal contractile responses in estradiol-injected euglycemic and diabetic rats were 68 +/- 7 and 45 +/- 5, respectively. Maximal contractile responses induced by oxytocin in estradiol-treated diabetic uteri were significantly smaller than the contractile responses measured in euglycemic estradiol-treated uteri. This study demonstrates that estradiol-induced changes in both myometrial cell morphology and in vitro uterine contractile responses to oxytocin are altered in diabetic rats.     

Impaired load dependence of diaphragm relaxation during congestive heart failure in the rabbit.

The load dependence (LD) of relaxation was studied in the diaphragm of rabbits with congestive heart failure (CHF). CHF (n = 15) was induced by combined chronic volume and pressure overload. Aortic insufficiency was induced by forcing a catheter through the aortic sigmoid valves, followed 3 wk later by abdominal aortic stenosis. Six weeks after the first intervention, animals developed CHF. Sham-operated animals served as controls (C; n = 12). Diaphragm mechanics were studied in vitro on isolated strips, at 22 degrees C, in isotonic and isometric loading conditions. Contractility was lower in the CHF group, as reflected by lower total tension: 1.11 +/- 0.10 in CHF vs. 2.38 +/- 0.15 N/cm(2) in C in twitch (P < 0.001) and 2.46 +/- 0.22 in CHF vs. 4.90 +/- 0.25 N. cm(-2) in C in tetanus (P < 0.001). The index LD was used to quantify the load dependence of relaxation: LD is <1 in load-dependent muscles and tends toward 1 in load-independent muscles. LD was significantly higher in CHF than in C rabbits, in both twitch (0.99 +/- 0.01 vs. 0.75 +/- 0.03; P < 0. 001) and tetanus (0.95 +/- 0.02 vs. 0.84 +/- 0.02; P < 0.001). In the CHF rabbits' diaphragm, the fall in total tension was linearly related to the fall in load dependence of relaxation. The decrease in load dependence of relaxation in CHF animals suggests sarcoplasmic reticulum abnormalities. Impairment of the sarcoplasmic reticulum may also partly account for the decrease in contractile performance of diaphragm in CHF animals.     

Effect of oxytocin receptor blockade on rat myometrial responsiveness to prostaglandin f(2)(alpha)

In the present study we have shown that the genetic expression of prostaglandin (PG)F(2alpha) receptor (R) and cyclooxygenase (COX)-2 increases in laboring rat myometrium. This finding was associated with a relatively weak contractile in vitro response (E:(max)) of isolated uterine strips when challenged with PGF(2alpha). Five days postpartum PGF(2alpha)-R mRNA values exceeded those during labor while COX-2 mRNA was reduced to preparturient values. Maximal contractility of isolated strips stimulated with PGF(2alpha) at this time was enhanced and E:C(50) decreased. Oxytocin treatment of estrogen-primed nonpregnant rats down-regulated uterine contractile responsiveness to PGF(2alpha), leaving mRNA values for this receptor unchanged, whereas oxytocin receptor blockade with atosiban (an oxytocin receptor antagonist) left E:(max) unaltered. In contrast, atosiban treatment of pregnant rats resulted in a 2.5-fold increase in E:(max) and a considerably reduced EC(50) during labor when compared to untreated delivering rats. The increased contractile ability was associated with a threefold increase in PGF(2alpha)-R mRNA production, indicating that the regulation by atosiban of the PGF(2alpha)-induced response is exerted at the genetic level. Based on the present data we suggest that 1) PGF(2alpha)-R stimulation may not primarily exert a contracting role in the normally delivering myometrium, and 2) the presence of the PGF(2alpha)-R system in rat myometrium may explain the apparent functional redundancy of the oxytocinergic system during the process of birth in animals lacking oxytocin or where the oxytocin receptor is blocked. In this context PGF(2alpha) receptor stimulation may, in the absence of oxytocin receptor stimulation, exert the contractile forces needed for proper propulsion of the fetus.     

Role of platelets in contraction of canine tracheal muscle elicited by PAF in vitro

To elucidate mechanisms of platelet-activating factor (PAF)-induced contraction, we studied the effect of PAF on 203 canine tracheal smooth muscle (TSM) strips from 45 dogs in vitro in the presence and absence of platelets. PAF (10(-11) to 10(-7) M) alone caused no contraction of TSM even in the presence of airway epithelium. In the presence of 2 x 10(5) platelets/microliter, PAF was an extremely potent contractile agonist (threshold 10(-11) M). This response was inhibited by the PAF antagonist, CV-3988 (10(-6) M), and reversed by the serotonin antagonist, methysergide (EC50 = 3.7 +/- 0.79 x 10(-9) M). Neither atropine nor chlorpheniramine (10(-9) to 10(-6) M) attenuated the response to PAF + platelets. In the presence of platelets, 10(-7) M PAF caused an increase in perfusate concentration of serotonin from 0.93 +/- 0.037 x 10(-8) to 1.7 +/- 0.046 x 10(-8) M (P less than 0.001). Tachyphylaxis, previously demonstrated to be irreversible, was shown to be a platelet-dependent phenomenon; contraction could be repeated in the same TSM after addition of fresh platelets. We demonstrate that PAF-induced contraction of canine TSM is caused by the release of cellular intermediates such as serotonin from platelets. We also demonstrate the site of PAF-induced tachyphylaxis in airway smooth muscle contraction.     

The effect of atrial natriuretic peptide on small intestinal contractility and transit.

Isometric tension in response to ANF (10(-10) to 10(-4) M) was recorded from longitudinally and circularly oriented rat jejunal smooth muscle strips. Conscious, fasted rats received an IV infusion of 1.25 nmol ANF/100 g body weight in 0.5 ml normal saline and controls received saline alone. Five minutes later 10 muCi Na2 51CrO4 in 0.5 ml saline was instilled through a jejunostomy. Fifteen minutes later animals were sacrificed, and the gut divided into 8 equal segments of small intestine, cecum and remaining colon. The radioactivity of each segment was measured and a geometric center of transit determined for each group. ANF induced relaxation of longitudinally oriented strips (Tm = -72.3 +/- 10.7 mN/g, ED50 7.3 +/- 3.6 x 10(-8) M), and contraction of circularly oriented strips (Tm = 35.0 +/- 5.0 mN/g, ED50 1.3 +/- 1.0 x 10(-8) M). This response was unaffected by 10(-6) M tetrodotoxin. The geometric mean center of transit was significantly (p less than 0.001) further aboral in ANF-treated compared to control animals (intestinal segment 4.2 +/- 0.2 vs. 3.2 +/- 0.2).     

The contractile action of platelet-activating factor on gallbladder smooth muscle

Platelet-activating factor (PAF) may be a mediator of some sequelae of cholecystitis, a disorder with gallbladder motor dysfunction. The aims of this study were to determine the effect and mechanism of PAF on gallbladder muscle. Exogenous administration of PAF-16 or PAF-18 caused dose-dependent contractions of gallbladder muscle strips in vitro with threshold doses of 1 ng/ml and 10 ng/ml, respectively. The PAF-induced contractions were not significantly reduced by TTX, atropine, or hexamethonium but were significantly inhibited with the PAF receptor antagonists ginkolide B and CV-3988. The PAF-induced contraction was reduced by indomethacin. Preventing influx of extracellular calcium with a calcium-free solution nearly abolished the PAF contractile response. Nifedipine inhibited the PAF contractile response, whereas ryanodine had no effect. Pertussis toxin reduced the PAF contractile response. In conclusion, PAF causes gallbladder contraction through specific PAF receptors on gallbladder muscle. These PAF receptors appear to be linked to a prostaglandin-mediated mechanism and to pertussis toxin-sensitive G proteins. The contractile response is largely mediated through the utilization of extracellular calcium influx through voltage-dependent calcium channels.