Supplemental treatment to atropine improves the efficacy to reverse nerve agent induced bronchoconstriction.

Exposure to highly toxic organophosphorus compounds causes inhibition of the enzyme acetylcholinesterase resulting in a cholinergic toxidrome and innervation of receptors in the neuromuscular junction may cause life-threatening respiratory effects. The involvement of several receptor systems was therefore examined for their impact on bronchoconstriction using an ex vivo rat precision-cut lung slice (PCLS) model. The ability to recover airways with therapeutics following nerve agent exposure was determined by quantitative analyses of muscle contraction. PCLS exposed to nicotine resulted in a dose-dependent bronchoconstriction. The neuromuscular nicotinic antagonist tubocurarine counteracted the nicotine-induced bronchoconstriction but not the ganglion blocker mecamylamine or the common muscarinic antagonist atropine. Correspondingly, atropine demonstrated a significant airway relaxation following ACh-exposure while tubocurarine did not. Atropine, the M3 muscarinic receptor antagonist 4-DAMP, tubocurarine, the ß2-adrenergic receptor agonist formoterol, the Na+-channel blocker tetrodotoxin and the K+ATP-channel opener cromakalim all significantly decreased airway contractions induced by electric field stimulation. Following VX-exposure, treatment with atropine and the Ca2+-channel blocker magnesium sulfate resulted in significant airway relaxation. Formoterol, cromakalim and magnesium sulfate administered in combinations with atropine demonstrated an additive effect. In conclusion, the present study demonstrated improved airway function following nerve agent exposure by adjunct treatment to the standard therapy of atropine.

The effect of progesterone on myometrial contractility, potassium channels, and tocolytic efficacy.

OBJECTIVES: Recent clinical trials have demonstrated a beneficial effect of supplementation with progesterone to prevent preterm labor. We aimed to determine the effects of progesterone treatment in vitro and in vivo and 17alpha-hydroxyprogesterone caproate (17OHPC) in vitro on myometrial contractions. METHODS: Myometrial strips were taken from women undergoing cesarean delivery at term. We also obtained myometrial biopsies from women participating in a clinical trial of progesterone to prevent preterm labor in twins (STOPPIT). After establishment of spontaneous contractions, strips were exposed to progesterone or 17OHPC. Separate strips were exposed to oxytocin and tocolytics alone and in combination with progesterone. Potassium channel blockers were added in conjunction with progesterone. STOPPIT samples were used to compare the effects of in vivo progesterone and placebo. We measured amplitude, frequency and activity integral of contractions. RESULTS: Maximum inhibition of contraction amplitude was 93 +/- 2% and 67 +/- 14% for progesterone at 30 microM and vehicle (70% ethanol), respectively, P < 0.05. 17OHPC did not exert an inhibitory effect. Water soluble progesterone exerted a maximal inhibitory effect on amplitude of contractions of 82 +/- 10% at 100 microM, P < 0.05. The inhibitory effect of progesterone was unaffected by potassium channel blockers. There was no difference between in vivo placebo and progesterone-treated groups in either amplitude or frequency of contractions, nor was there any difference in the response to oxytocin or the tocolytic drugs. CONCLUSIONS: Progesterone exerts rapid inhibition of the amplitude of myometrial contractions in vitro but 17OHPC does not. The action of progesterone does not appear to operate via potassium channels nor does it enhance the activity of certain tocolytic drugs.

New R/S-3,4-dihydro-2,2-dimethyl-6-halo-4-(phenylaminothiocarbonylamino)-2H-1-benzopyrans structurally related to (+/-)-cromakalim as tissue-selective pancreatic beta-cell K(ATP) channel openers.

The present work was aimed at exploring a series of R/S-3,4-dihydro-2,2-dimethyl-6-halo-4-(phenylaminothiocarbonylamino)-2H-1-benzopyrans structurally related to (+/-)-cromakalim and differently substituted at the 4- and 6-positions. The biological effects of these putative activators of ATP-sensitive potassium channels (K(ATP)) were characterized in vitro on the pancreatic endocrine tissue (inhibition of insulin release) and on the vascular smooth muscle tissue (relaxation of aorta rings). The biological activity of these new dimethylchroman derivatives was further compared to that of (+/-)-cromakalim, (+/-)-pinacidil, diazoxide and BPDZ 73. Structure-activity relationships indicated that an improved potency for the pancreatic tissue was obtained by introducing a meta- or a para-electron-withdrawing group such as a chlorine atom on the C-4 phenyl ring, independently of the nature of the halogen atom at the 6-position of the benzopyran nucleus. Most original dimethylchroman thioureas were more potent than their 'urea' homologues and even more potent than diazoxide at inhibiting insulin release. Moreover, and unlike (+/-)-cromakalim or (+/-)-pinacidil, such compounds appeared to be highly selective towards the pancreatic tissue. Radioisotopic and fluorimetric investigations indicated that the new drugs activated pancreatic K(ATP) channels. Lastly, conformational studies suggested that the urea/thiourea dimethylchromans can be regarded as hybrid compounds between cromakalim and pinacidil.

Activation of the iberiotoxin-sensitive BKCa channels by salvianolic acid B of the porcine coronary artery smooth muscle cells.

In this study, we examined the effects of Salvia miltiorrhiza (Danshen) crude extract, some of its lipid-soluble components (tanshinone I, tanshinone II(A), cryptotanshinone, dihydroisotanshinone I) and the water-soluble compounds (danshensu and salvianolic acid B) on the K(+) channels such as the iberiotoxin-sensitive Ca(2+)-activated K(+) (BK(Ca)) channels and the glibenclamide-sensitive ATP-dependent K(+) (IK(ATP)) channels of the porcine left anterior descending coronary artery smooth muscle cells. Cumulative application of salvianolic acid B (30-300 microM) caused a l-NNA (100 microM)-insensitive, potentiation of the outward BK(Ca) current amplitude with no apparent effect on the IK(ATP) channels opening. Salvianolic acid B (300 microM) caused an ODQ (10 microM, a guanylate cyclase inhibitor)-sensitive enhancement of the outward BK(Ca) current amplitude. In contrast, none of the other isolated chemical constituents of S. miltiorrhiza modified the openings of the two types of K(+) channels studied. In conclusion, our results suggest that salvianolic acid B, a major hydrophilic constituent found in Radix S. miltiorrhiza, activated the opening of the BK(Ca) channels of the porcine coronary artery smooth muscle cells through the activation of guanylate cyclase without the involvement of the nitric oxide synthase activation.

Synthesis and pharmacological evaluation of some N-arylsulfonyl-N-methyl-N'-(2,2-dimethyl-2H-1-benzopyran-4-yl)ureas structurally related to cromakalim.

Some N-arylsulfonyl-N-methyl-N'-(2,2-dimethyl-2H-1-benzopyran-4-yl)ureas were prepared and evaluated as putative potassium channel openers on the vascular and uterine smooth muscle tissue (myorelaxant effect), as well as on insulin-secreting pancreatic islets (inhibition of insulin release). The pharmacological results indicated that these compounds exhibited a marked biological activity on these three tissues.

Antispasmodic and blood pressure lowering effects of Valeriana wallichii are mediated through K+ channel activation.

Crude extract of Valeriana wallichii rhizome (Vw.Cr) and its fractions were studied for possible antispasmodic and blood pressure lowering activities to rationalize some of the folkloric uses. In rabbit jejunum preparations, Vw.Cr (0.1-3.0 mg/mL) caused relaxation of spontaneous contractions. When tested against high K(+) (80 mM)-induced contractions it produced weak inhibitory effect, while caused complete relaxation of the contractions induced by low K(+) (20 mM). In the presence of glibenclamide (3 microM), the inhibitory effect of low K(+) was shifted to the right, similar to that produced by cromakalim while, verapamil caused no differentiation in its inhibitory effect against low and high K(+)-induced contractions. In guinea pig ileum, the plant extract produced similar results as in rabbit jejunum. Intravenous administration of Vw.Cr, produced fall in arterial blood pressure in normotensive anaesthetized rats and this effect was partially blocked by glibenclamide. In rabbit aortic preparations, plant extract also caused a selective and glibenclamide-sensitive relaxation of low K(+) (20 mM)-induced contractions. Activity-directed fractionation studies revealed that the observed activity was distributed both in the chloroform and aqueous fractions. These results indicate that the antispasmodic and hypotensive effects of Valeriana wallichii are mediated possibly through K(ATP) channel activation, which justify its use in gastrointestinal and cardiovascular disorders.

In vivo evaluation of the potency and bladder-vascular selectivity of the ATP-sensitive potassium channel openers (-)-cromakalim, ZD6169 and WAY-133537 in rats.

OBJECTIVE: To compare in vivo the potency and bladder-vascular selectivity of ATP-sensitive potassium channel openers (KCOs) (-)-cromakalim, WAY-133537 and ZD6169 and a muscarinic antagonist, tolterodine in rats. MATERIALS AND METHODS: Bladder and arterial pressures were monitored simultaneously, before and after increasing intravenous doses of compounds, in each of two urethane-anaesthetized rat bladder hyperactivity models: spontaneous non-voiding myogenic contractions secondary to partial outlet obstruction and volume-induced neurogenic contractions. RESULTS: (-)-Cromakalim, WAY-133537 and ZD6169 caused a dose-dependent suppression of spontaneous contractions in the obstructed model, with a 50% inhibition of the contraction area under the curve at doses of 0.06, 0.14 and 2.4 micro mol/kg (intravenous), respectively. Corresponding decreases in mean arterial pressure at these effective doses were 24%, 15% and 15%, respectively. The KCO potency rank order was the same and their relative potency highly comparable in the neurogenic model. There was complete inhibition of spontaneous contractions in obstructed rats at doses corresponding to approximately 50% inhibition of the neurogenic contractions. While tolterodine caused a dose-dependent inhibition of contractions in the neurogenic model, it was ineffective at inhibiting non-voiding contractions in obstructed rats. CONCLUSIONS: All KCOs tested caused significant decreases in arterial pressure at doses effective on the bladder in the model of obstructive instability, suggesting a lack of bladder-vascular selectivity. Similar KCO potency in both assays suggests no appreciable changes in KATP channel function as a result of partial outlet obstruction.

The JT-area indicates dispersion of repolarization in dogs with atrioventricular block.

UNLABELLED: Heterogeneity in cardiac repolarization (Delta APD) is known to be arrhythmic. In the dog model of chronic complete AV-block and acquired long QT syndrome, an increase in Delta MAPD (defined as left ventricular monophasic action potential duration (MAPD) minus right ventricular MAPD) is often associated with changes in T-wave morphology. The purpose of this study was to correlate known changes in Delta MAPD with the planimetric total area of the T-wave on the surface ECG (integral of J-T, mVx ms). METHODS: The relationship between Delta MAPD and total area of the T-wave (i.e., JT-area) was assessed in four different protocols with different types of dispersion: (1) class III drugs followed by levcromakalim (n= 7), (2) LAD coronary artery occlusion and reperfusion (n = 6), (3) dronedarone i.v., an amiodarone like agent (n = 5) and (4) steady state pacing at cycle lengths of 1000 ms and 500 ms (n = 5). RESULTS: Class III drugs increased Delta MAPD (55 +/- 40 ms to 120 +/- 50 ms(#), P<0.05), which was correlated (r = 0.74, P < 0.001) with JT-area (50 +/- 40 mV. ms to 95 +/- 35 mV x ms(#)). Ischemia increased both Delta MAPD (30 +/- 25 ms to 90 +/- 40 ms(#)) and JT-area (60 +/- 55 mV x ms to 75 +/- 50 mV x ms(#)). Both levcromakalim and reperfusion reversed these conditions. Dronedarone had no effect on Delta MAPD or on JT-area while a faster frequency reduced both Delta MAPD and JT-area. CONCLUSION: Changes in dispersion of ventricular repolarization are reflected by alterations in JT-area. This non-invasive parameter may therefore be used to indicate changes in heterogeneity in ventricular repolarization.

Interaction of isoflurane and cromakalim, a KATP channel opener, on coronary and systemic haemodynamics in chronically instrumented dogs.

BACKGROUND: Although isoflurane has been shown to cause coronary and systemic vasodilation through KATP channel activation, the interaction of KATP channel openers and isoflurane has not been fully investigated. The present study was carried out to determine the haemodynamic actions of cromakalim, a KATP channel opener, under the conscious state and during isoflurane anaesthesia in chronically instrumented dogs. METHODS: Fourteen dogs were chronically instrumented to measure systemic and coronary haemodynamics. Each dog was randomly assigned to receive doses of either cromakalim, 4 and 10 microg x kg(-1) i.v., or isoflurane, 2.1% end-tidal (1.5 MAC), plus cromakalim, 4 and 10 microg x kg(-1) i.v. RESULTS: Cromakalim dose-relatedly decreased mean arterial pressure and systemic vascular resistance and increased coronary blood flow in both conscious and anaesthetized states. With isoflurane, the duration of effects of cromakalim were prolonged. Isoflurane exerted an additive effect on the increase in coronary blood flow induced by a low-dose cromakalim, whereas it did not influence the effect of a high-dose cromakalim. The maximum rate of increase in left ventricular pressure and segment shortening were increased by cromakalim in the conscious state but unchanged during isoflurane anaesthesia. CONCLUSION: The results suggest that the coronary vasodilating effects of isoflurane and cromakalim are basically additive until cromakalim exerts the maximal effect, and that the action of cromakalim on the coronary vasculature is prolonged by isoflurane.

Analysis of responses of garlic derivatives in the pulmonary vascular bed of the rat.

Allicin, an extract from garlic, has been shown to be a systemic and pulmonary arterial vasodilator that acts by an unknown mechanism. In the present experiments, pulmonary vascular responses to allicin (10-100 microg), allyl mercaptan (0.3-1 mg), and diallyl disulfide (0.3-1 mg) were studied in the isolated lung of the rat under constant-flow conditions. When baseline tone in the pulmonary vascular bed of the rat was raised to a high-steady level with the thromboxane A(2) mimic U-46619, dose-related decreases in pulmonary arterial pressure were observed. In terms of the mechanism of action of allicin vasodilator activity in the rat, responses to allicin were not significantly different after administration of the nitric oxide synthase inhibitor N(omega)-nitro-L-arginine methyl ester, the K(ATP)(+) channel antagonist U-37883A, or the cyclooxygenase inhibitor sodium meclofenamate, or when lung ventilation was interrupted. These data show that allicin has significant vasodilator activity in the pulmonary vascular bed of the rat, whereas allyl mercaptan and diallyl disulfide produced no significant changes in pulmonary arterial perfusion pressure. The present data suggest that pulmonary vasodilator responses to allicin are independent of the synthesis of nitric oxide, ATP-sensitive K(+) channels, activation of cyclooxygenase enzyme, or changes in bronchomotor tone in the pulmonary vascular bed of the rat.

Preventing ventricular fibrillation by flattening cardiac restitution.

Ventricular fibrillation is the leading cause of sudden cardiac death. In fibrillation, fragmented electrical waves meander erratically through the heart muscle, creating disordered and ineffective contraction. Theoretical and computer studies, as well as recent experimental evidence, have suggested that fibrillation is created and sustained by the property of restitution of the cardiac action potential duration (that is, its dependence on the previous diastolic interval). The restitution hypothesis states that steeply sloped restitution curves create unstable wave propagation that results in wave break, the event that is necessary for fibrillation. Here we present experimental evidence supporting this idea. In particular, we identify the action of the drug bretylium as a prototype for the future development of effective restitution-based antifibrillatory agents. We show that bretylium acts in accord with the restitution hypothesis: by flattening restitution curves, it prevents wave break and thus prevents fibrillation. It even converts existing fibrillation, either to a periodic state (ventricular tachycardia, which is much more easily controlled) or to quiescent healthy tissue.

Reconstituted human cardiac KATP channels: functional identity with the native channels from the sarcolemma of human ventricular cells.

ATP-sensitive potassium (KATP) channels in striated myocytes are heteromultimers of KIR6.2, a weak potassium inward rectifier, plus SUR2A, a low-affinity sulfonylurea receptor. We have cloned human KIR6.2 (huKIR6.2) and a huSUR2A that corresponds to the major, full-length splice variant identified by polymerase chain reaction analysis of human cardiac poly A+ mRNA. ATP- and glibenclamide-sensitive K+ channels were produced when both subunits were coexpressed in COSm6 and Chinese hamster ovary cells lacking endogenous KATP channels, but not when huSUR2A or huKIR6.2 were transfected alone. Recombinant channels activated by metabolic inhibition in cell-attached configuration or in inside-out patches with ATP-free internal solution were compared with sarcolemmal KATP channels in human ventricular cells. The single-channel conductance of approximately 80 pS measured at -40 mV in quasi-symmetrical approximately 150 mmol/L K+ solutions, the intraburst kinetics that were dependent on K+ driving force, and the weak inward rectification were indistinguishable for both channels. Similar to the native channels, huSUR2A/huKIR6.2 recombinant channels were inhibited by ATP at quasi-physiological free Mg2+ ( approximately 0. 7 mmol/L) or in the absence of Mg2+, with an apparent IC50 of approximately 20 micromol/L and a pseudo-Hill coefficient of approximately 1. They were "refreshed" by MgATP and stimulated by ADP in the presence of Mg2+ when inhibited by ATP. The huSUR2A/huKIR6.2 channels were stimulated by cromakalim and pinacidil in the presence of ATP and Mg2+ but were insensitive to diazoxide. The results suggest that reconstituted huSUR2A/huKIR6.2 channels represent KATP channels in sarcolemma of human cardiomyocytes and are an adequate experimental model with which to examine structure-function relationships, molecular physiology, and pharmacology of these channels from human heart.

Effect of JTV-506, a novel vasodilator, on experimental angina model in rats.

The antianginal effects of JTV-506, a newly synthesized 2,2-bis-methoxymethyl benzopyran-derivative potassium channel opener, were evaluated in experimental angina models in rats and compared with those of levcromakalim, nicorandil, and nifedipine. JTV-506 (0.01-0.1 mg/kg, i.d.) dose-dependently inhibited S-wave elevation induced by injection of methacholine but caused almost no changes in blood pressure or heart rate. Other vasodilators including levcromakalim, nicorandil, and nifedipine, when administered intraduodenally, also inhibited the S-wave elevation and elicited a decrease in blood pressure. Oral administration of JTV-506 (1 and 3 mg/kg), levcromakalim (1 and 3 mg/kg), and nicorandil (30 mg/kg) significantly inhibited the S-wave depression induced by intravenous administration of arginine vasopressin (AVP). Thus JTV-506 exerts a potent protective effect on angina pectoris models to an extent similar to those of levcromakalim, nicorandil, and nifedipine, whereas its action on systemic blood pressure is different from that of other vasodilators, including reference potassium channel openers. These results suggest that JTV-506 may clinically be useful as an antianginal agent.