LPS exacerbates functional and inflammatory responses to ovalbumin and decreases sensitivity to inhaled fluticasone propionate in a guinea pig model of asthma.

BACKGROUND AND PURPOSE: Asthma exacerbations contribute to corticosteroid insensitivity. LPS is ubiquitous in the environment. It causes bronchoconstriction and airway inflammation and may therefore exacerbate allergen responses. This study examined whether LPS and ovalbumin co-administration could exacerbate the airway inflammatory and functional responses to ovalbumin in conscious guinea pigs and whether these exacerbated responses were insensitive to inhaled corticosteroid treatment with fluticasone propionate (FP). EXPERIMENTAL APPROACH: Guinea pigs were sensitized and challenged with ovalbumin and airway function recorded as specific airway conductance by whole body plethysmography. Airway inflammation was measured from lung histology and bronchoalveolar lavage. Airway hyper-reactivity (AHR) to inhaled histamine was examined 24 h after ovalbumin. LPS was inhaled alone or 24 or 48 h before ovalbumin and combined with ovalbumin. FP (0.05-1 mg·mL(-1) ) or vehicle was nebulized for 15 min twice daily for 6 days before ovalbumin or LPS exposure. KEY RESULTS: Ovalbumin inhalation caused early (EAR) and late asthmatic response (LAR), airway hyper-reactivity to histamine and influx of inflammatory cells into the lungs. LPS 48 h before and co-administered with ovalbumin exacerbated the response with increased length of the EAR, prolonged response to histamine and elevated inflammatory cells. FP 0.5 and 1 mg·mL(-1) reduced the LAR, AHR and cell influx with ovalbumin alone, but was ineffective when guinea pigs were exposed to LPS before and with ovalbumin. CONCLUSIONS AND IMPLICATIONS: LPS exposure exacerbates airway inflammatory and functional responses to allergen inhalation and decreases corticosteroid sensitivity. Its widespread presence in the environment could contribute to asthma exacerbations and corticosteroid insensitivity in humans.

Bronchoprotection in conscious guinea pigs by budesonide and the NO-donating analogue, TPI 1020, alone and combined with tiotropium or formoterol.

BACKGROUND AND PURPOSE: Inhaled corticosteroids, anticholinergics and ß2-adrenoceptor agonists are frequently combined for treating chronic respiratory diseases. We examine the corticosteroid, budesonide, and novel NO-donating derivative, TPI 1020, against histamine- and methacholine-induced bronchoconstriction and whether they enhance the ß2-adrenoceptor agonist formoterol or muscarinic antagonist tiotropium in conscious guinea pigs. EXPERIMENTAL APPROACH: Dunkin-Hartley guinea pigs received inhaled histamine (3 mM) or methacholine (1.5 mM) and specific airway conductance (sG(aw)) was measured before and 15 or 75 min after treatment with budesonide, TPI 1020, tiotropium or formoterol alone or in combinations. KEY RESULTS: Formoterol (0.7-10 µM) and budesonide (0.11-0.7 mM) inhibited histamine-induced bronchoconstriction and tiotropium (2-20 µM) inhibited methacholine-induced bronchoconstriction by up to 70.8 ± 16.6%, 34.9 ± 4.4% and 85.1 ± 14.3%, respectively. Formoterol (2.5 µM) or tiotropium (2 µM) alone exerted small non-significant bronchoprotection. However, when co-administered with TPI 1020 0.11 mM, which alone had no significant effect, there was significant inhibition of the bronchoconstriction (45.7 ± 12.2% and 79.7 ± 21.4%, respectively). Co-administering budesonide (0.11 mM) with tiotropium (2 µM), which alone had no effect, also significantly inhibited the methacholine bronchoconstriction (36.5 ± 13.0%), but there was no potentiation of formoterol against histamine. The NO scavenger, CPTIO, prevented the bronchoprotection by SNAPand TPI 1020. CONCLUSIONS AND IMPLICATIONS: TPI 1020 potentiated the bronchoprotection by formoterol and tiotropium. Budesonide also enhanced the effects of tiotropium but not formoterol. Combination of TPI 1020 with a long-acting ß2-adrenoceptor agonist or muscarinic receptor antagonist may therefore be a more potent therapeutic approach for treatment of chronic respiratory diseases.

Vasoconstrictor and vasodilator responses to tryptamine of rat-isolated perfused mesentery: comparison with tyramine and ß-phenylethylamine.

BACKGROUND AND PURPOSE: Tryptamine increases blood pressure by vasoconstriction, but little is known about its actions on the mesentery, in particular the resistance arteries. Tryptamine interacts with trace amine-associated receptors (TAARs) and because of its structural similarity to 5-HT, it may also interact with 5-HT receptors. Our hypothesis is therefore that the rat mesenteric arterial bed will exhibit vasopressor and vasodepressor responses to tryptamine via both 5-HT and TAARs. EXPERIMENTAL APPROACH: Tryptamine-evoked responses were assayed from pressure changes of the rat-isolated mesenteric vasculature perfused at constant flow rate in the absence and presence of adrenoceptor and 5-HT receptor antagonists. KEY RESULTS: Tryptamine caused dose-dependent vasoconstriction of the mesenteric arterial bed as increases in perfusion pressure. These were unaffected by the α(1) -adrenoceptor antagonist, prazosin, but were attenuated by the non-selective α-adrenoceptor antagonist, phentolamine. The 5-HT(2A) receptor antagonists, ketanserin and ritanserin, abolished the tryptamine-induced pressure increases to reveal vasodilator responses in mesenteric beds preconstricted with phenylephrine. These tryptamine-induced vasodilator responses were unaffected by the 5-HT(7) receptor antagonist, SB269970, but were eliminated by the NOS inhibitor, N(ω) -nitro-L-arginine methyl ester (L-NAME). Tyramine and ß-phenylethylamine also caused vasodilatation in pre-constricted vasculature, which was also abolished by L-NAME. CONCLUSIONS AND IMPLICATIONS: Tryptamine causes vasoconstriction of the mesenteric vasculature via 5-HT(2A) receptors, which when inhibited exposed vasorelaxant effects in pre-constricted tissues. The vasodilatation was independent of 5-HT(2A) and 5-HT(7) receptors but like that for tyramine and ß-phenylethylamine was due to NO release. Potency orders suggest TAAR involvement in the vasodilatation by these trace amines.

Mechanisms of anandamide-induced vasorelaxation in rat isolated coronary arteries.

1. The cannabinoid arachidonyl ethanolamide (anandamide) caused concentration-dependent relaxation of 5-HT-precontracted, myograph-mounted, segments of rat left anterior descending coronary artery. 2. This relaxation was endothelium-independent, unaffected by the fatty acid amide hydrolase inhibitor, arachidonyl trifluoromethyl ketone (10 microM), and mimicked by the non-hydrolysable anandamide derivative, methanandamide. 3. Relaxations to anandamide were attenuated by the cannabinoid receptor antagonist, SR 141716A (3 microM), but unaffected by AM 251 (1 microM) and AM 630 (1 microM), more selective antagonists of cannabinoid CB(1) and CB(2) receptors respectively. Palmitoylethanolamide, a selective CB(2) receptor agonist, did not relax precontracted coronary arteries. 4. Anandamide relaxations were not affected by inhibition of sensory nerve transmission with capsaicin (10 microM) or blockade of vanilloid VR1 receptors with capsazepine (5 microM). Nevertheless capsaicin relaxed coronary arteries in a concentration-dependent and capsazepine-sensitive manner, confirming functional sensory nerves were present. In contrast, capsazepine and capsaicin did inhibit anandamide relaxations in methoxamine-precontracted rat small mesenteric arteries. 5. Relaxations to anandamide were inhibited by TEA (1 mM) or iberiotoxin (50 nM), blockers of large conductance, Ca(2+)-activated K(+) channels (BK(Ca)). Gap junction inhibition with 18alpha-glycyrrhetinic acid (100 microM) did not affect anandamide relaxations. 6. This study shows anandamide relaxes the rat coronary artery by a novel mechanism. Anandamide-induced relaxations do not involve the endothelium, degradation into active metabolites, or activation of cannabinoid CB(1) or CB(2) receptors, but may involve activation of BK(Ca). Vanilloid receptor activation also has no role in the effects of anandamide in coronary arteries, even though functional sensory nerves are present.

Angiotensin II reduces infarct size and has no effect on post-ischaemic contractile dysfunction in isolated rat hearts.

1. In order to test the hypothesis that angiotensin II exacerbates myocardial ischaemia-reperfusion (IR) injury, we examined the effects of graded angiotension II concentrations of angiotensin II on IR injury in both working and non-working (Langendorff) isolated rat hearts. 2. Non-working hearts were subjected to 30 min aerobic perfusion (baseline) then 25 min of global, no-flow ischaemia followed by 30 min of reperfusion either in the absence (control, n=7) or presence of 1 (n=6) or 10 nM (n=5) angiotensin II). Recoveries of LV developed pressure and coronary flow after 30 min reperfusion in control hearts (58+/-9 and 40+/-8% of baseline levels, respectively) were no different from hearts treated with 1 or 10 nM angiotensin II. Infarct size (determined at the end of reperfusion by triphenyltetrazolium chloride staining) was reduced by angiotensin II in a concentration-dependent manner (from a control value of 27+/-3 to 18+/-4% and 9+/-3% of the LV, respectively). 3. Working hearts were subjected to 50 min pre-ischaemic (pre-I) aerobic perfusion then 30 min of global, no-flow ischaemia followed by 30 min of reperfusion either in the absence (control, n=14) or presence of 1 (n=8), 10 (n=7) or 100 nM (n=7) angiotensin II). In controls, post-ischaemic (post-I) left ventricular (LV) work and efficiency of oxygen consumption were depressed (43+/-9 and 42+/-10% of pre-I levels, respectively). The presence of angiotensin II throughout IR had no effect on LV work compared with control. 4. Thus, angiotensin II reduces infarct size in a concentration-dependent manner but has no effect on contractile stunning associated with IR in isolated rat hearts.

Influence of beta-adrenoceptor tone on the cardioprotective efficacy of adenosine A(1) receptor activation in isolated working rat hearts.

This study investigated the role of beta-adrenoceptors in the cardioprotective and metabolic actions of adenosine A(1) receptor stimulation. Isolated paced (300 beats min(-1)) working rat hearts were perfused with Krebs-Henseleit solution containing 1.2 mM palmitate. Left ventricular minute work (LV work), O(2) consumption and rates of glycolysis and glucose oxidation were measured during reperfusion (30 min) following global ischaemia (30 min) as well as during aerobic conditions. Relative to untreated hearts, N(6)-cyclohexyladenosine (CHA, 0.5 microM) improved post-ischaemic LV work (158%) and reduced glycolysis and proton production (53 and 42%, respectively). CHA+propranolol (1 microM) had similar beneficial effects, while propranolol alone did not affect post-ischaemic LV work or glucose metabolism. Isoprenaline (10 nM) impaired post-ischaemic function and after 25 min ischaemia recovery was comparable with 30 min ischaemia in untreated hearts (41 and 53%, respectively). Relative to isoprenaline alone, CHA+isoprenaline improved recovery of LV work (181%) and reduced glycolysis and proton production (64 and 60%, respectively). In aerobic hearts, CHA, propranolol or CHA+propranolol had no effect on LV work or glucose oxidation. Glycolysis was inhibited by CHA, propranolol and CHA+propranolol (50, 53 and 52%, respectively). Isoprenaline-induced increases in heart rate, glycolysis and proton production were attenuated by CHA (85, 57 and 53%, respectively). The cardioprotective efficacy of CHA was unaffected by antagonism or activation of beta-adrenoceptors. Thus, the mechanism of protection by adenosine A(1) receptor activation does not involve functional antagonism of beta-adrenoceptors.

Characterization of cardioprotection mediated by AT2 receptor antagonism after ischemia-reperfusion in isolated working rat hearts.

BACKGROUND: Whether cardioprotection induced by the angiotensin II (AngII) type 2 receptor (AT(2)R) antagonist PD123,319 (PD) after ischemia-reperfusion (IR) is influenced by the concentration of PD, presence of AngII, timing of exposure, or inhibition of proton production from glucose metabolism is not known. METHODS AND RESULTS: We examined these factors in isolated working rat hearts subjected to IR injury, no treatment (control), or treatment with N(6)-cyclohexyl adenosine (CHA, 0.5 micromol/L), an adenosine A(1) receptor agonist that induces cardioprotection by decreasing protons ("positive" control). Compared with control, 1 micromol/L PD present throughout IR improved recovery of left ventricular work (73 +/- 5 vs. 40 +/- 8%) to the level with CHA (82 +/- 5%), but 0.1 micromol/L PD did not (58 +/- 6 vs. 40 +/- 8%). AngII (1 nmol/L) did not effect postischemic recovery associated with 1 micromol/L PD (73 +/- 7%) but improved that associated with 0.1 micromol/L PD (86 +/- 3%). PD (1 micromol/L), present solely during reperfusion, enhanced postischemic left ventricular recovery to 72 +/- 5%. Also, PD (1 micromol/L) did not affect glycolytic rates or proton production in nonischemic or IR hearts. CONCLUSION: PD-induced cardioprotection is 1) PD concentration-dependent, 2) AngII-sensitive, 3) mediated during reperfusion, and 4) independent of proton production, suggesting that reduction in IR injury and indirect AT(1)R stimulation might be involved.

Differences between the vasorelaxant activity of adenosine-receptor agonists on guinea-pig isolated aorta precontracted with noradrenaline or phenylephrine.

The relaxant effect of adenosine and 5'-(N-ethylcarboxamido)adenosine (NECA) against alpha-adrenoceptor-mediated contractile tone in guinea-pig isolated aortic rings has been examined to determine if this A2B-receptor-mediated relaxation was dependent upon the contracting agent, and whether the contractions were dependent upon intracellular or extracellular calcium. Relaxation responses were consistently greater for aortic rings pre-contracted with phenylephrine (3x10(-6) M) than for rings pre-contracted with noradrenaline (3x10(-6) M). Maximum inhibition by NECA was significantly greater for phenylephrine-contracted aortae than for noradrenaline-contracted (81.9+/-2.8% compared with 25.0+/-1.5%). These differences persisted in the presence of beta- and alpha2-adrenoceptor blockade and could not, therefore, be attributed to stimulation of these receptors by noradrenaline. The ratio of the contractions obtained before and in the presence of adenosine or NECA was compared with the control ratio obtained before and after vehicle. Experiments were performed both in the presence of normal calcium levels and under calcium-free conditions. In normal-calcium medium, NECA inhibited phenylephrine-induced contractions (test ratio, 76.7+/-3.9%; control ratio, 133.1+/-9.8%) to a greater extent than noradrenaline-induced contractions (108.4+/-4.1 and 123.4+/-4.9%); adenosine similarly inhibited phenylephrine-induced contractions more than those induced by noradrenaline. Under calcium-free conditions, adenosine (36.7+/-11.9 and 110.7+/-26.6%) and NECA (55.2+/-9.1 and 87.1+/-14.9%) were only effective against phenylephrine-induced contractions. This suggests that activation of the A2B-receptor by these agonists inhibited intracellular mobilization of calcium for phenylephrine-induced contractions only. The effects on extracellular calcium influx were examined for phenylephrine- and noradrenaline-induced contractions in normal-calcium medium but in the presence of ryanodine to prevent intracellular calcium mobilization. NECA inhibited phenylephrine-induced contractions (77.3+/-12.4 and 111.4+/-9.3%), presumably by interfering with influx of calcium through receptor-operated calcium channels. In contrast, NECA failed to reduce noradrenaline-induced contractions (121.5+/-10.7 and 122.4+/-11.6%), suggesting that the effect on noradrenaline is predominantly via interaction with intracellular calcium. Adenosine was consistently a more effective relaxant than NECA, possibly because of an additional intracellular component of the response. We conclude that adenosine receptor agonists inhibit phenylephrine-induced contractions of guinea-pig aorta more selectively than noradrenaline-induced contractions. A2B-receptor stimulation might reveal a fundamental difference between the modes of contraction elicited by these two alpha-adrenoceptor agonists.

Effects of adenosine receptor agonists on induction of contractions to phenylephrine of guinea-pig aorta mediated via intra- or extracellular calcium.

The vasorelaxant actions of adenosine and its analogue, 5'-(N-ethylcarboxamido)-adenosine (NECA), were investigated in guinea-pig isolated aortic rings by addition to the tissue prior to induction of a contraction by the alpha1-adrenoceptor agonist phenylephrine (PE, 3x10(-6) M). The effect was calculated from the ratio (C2/C1) of the contraction to PE before (C1) and in the presence of adenosine or NECA (C2). This was compared with a control ratio obtained at the same time in which no vasorelaxant was present during C2. Experiments were performed in either "normal" or "Ca2+ -free" bathing medium. Both adenosine and NECA caused inhibition of contractions in "normal" and "Ca2+ -free" conditions, the latter indicating that the vasorelaxant action was due in part to inhibition of intracellular Ca2+ mobilization. To determine whether inhibition of influx of extracellular Ca2+ is a target for the vasorelaxation, contractions to PE were obtained in "normal" Ca2+ and in the presence of ryanodine (10(-5) M), which prevents the release of intracellular Ca2+. These contractions were inhibited by NECA indicating that stimulation of A2-receptors by NECA interferes with the influx of Ca2+ via the opening of receptor-operated Ca2+ channels (ROCs). This study has demonstrated that cell surface A2-receptor stimulation in the guinea-pig aorta inhibits phenylephrine-induced contractions by interfering with both the release of intracellular Ca2+ and the influx of extracellular Ca2+, presumably via ROCs.

Effects of K(+)-channel blockers on A1-adenosine receptor-mediated negative inotropy and chronotropy of guinea-pig isolated left and right atria.

Adenosine has previously been shown to stimulate K(+)-efflux and to block L-type calcium channels in atrial myocytes. The aim of the present study was to evaluate the contribution of K(+)-channels in the development of the negative inotropic and chronotropic responses to adenosine agonists in guinea-pig left and right atria, respectively. Tetraethylammonium (TEA) potentiated the negative inotropic and chronotropic responses to R-(-)-N6-(2-phenyl-isopropyl)-adenosine (R-PIA), seen as leftward shifts of the concentration-response curves. Glibenclamide had no effect on the negative inotropic response to R-PIA but increased the rate of onset of the negative chronotropic response in right atria. 4-Aminopyridine (4-AP, 10 mM), potentiated the left atrial inotropic responses to R-PIA, seen as a leftward shift of the concentration-response curve, but slowed the speed of onset of the response to a single concentration (10(-6) M) of R-PIA. This reduction in speed of onset of the response can explain the differences in effects of 4-AP on concentration-response curves reported here and previously. In the right atria, 4-AP (10 mM) inhibited the negative chronotropic responses to R-PIA, seen as a rightward shift of the concentration-response curve and reduction of the maximum response. 4-AP also slowed the onset of the right atrial rate response to R-PIA. These results support the theory that K(+)-efflux plays only a minor role in the negative inotropic responses of guinea-pig left atria to R-PIA. This apparently controls the speed of onset of the response. The negative chronotropic response of guinea-pig right atria to R-PIA appears to be much more dependent upon K(+)-efflux than for the negative inotropic response of the left atria.

K(ATP)-channel activation: effects on myocardial recovery from ischaemia and role in the cardioprotective response to adenosine A1-receptor stimulation.

1. Optimization of myocardial energy substrate metabolism improves the recovery of mechanical function of the post-ischaemic heart. This study investigated the role of K(ATP)-channels in the regulation of the metabolic and mechanical function of the aerobic and post-ischaemic heart by measuring the effects of the selective K(ATP)-channel activator, cromakalim, and the effects of the K(ATP)-channel antagonist, glibenclamide, in rat fatty acid perfused, working hearts in vitro. The role of K(ATP) channels in the cardioprotective actions of the adenosine A1-receptor agonist, N6-cyclohexyladenosine (CHA) was also investigated. 2. Myocardial glucose metabolism, mechanical function and efficiency were measured simultaneously in hearts perfused with modified Krebs-Henseleit solution containing 2.5 mM Ca2+, 11 mM glucose, 1.2 mM palmitate and 100 mu l(-1) insulin, and paced at 300 beats min(-1). Rates of glycolysis and glucose oxidation were measured from the quantitative production of 3H20 and 14CO2, respectively, from [5-3H/ U-14C]-glucose. 3. In hearts perfused under aerobic conditions, cromakalim (10 microM), CHA (0.5 microM) or glibenclamide (30 microM) had no effect on mechanical function. Cromakalim did not affect glycolysis or glucose oxidation, whereas glibenclamide significantly increased rates of glycolysis and proton production. CHA significantly reduced rates of glycolysis and proton production but had no effect on glucose oxidation. Glibenclamide did not alter CHA-induced inhibition of glycolysis and proton production. 4. In hearts reperfused for 30 min following 30 min of ischaemia, left ventricular minute work (LV work) recovered to 24% of aerobic baseline values. Cromakalim (10 microM), administered 5 min before ischaemia, had no significant effect on mechanical recovery or glucose metabolism. CHA (0.5 microM) significantly increased the recovery of LV work to 67% of aerobic baseline values and also significantly inhibited rates of glycolysis and proton production. Glibenclamide (30 microM) significantly depressed the recovery of mechanical function to < 1% of aerobic baseline values and stimulated glycolysis and proton production. 5. Despite the deleterious actions of glibenclamide per se in post-ischaemic hearts, the beneficial effects of CHA (0.5 microM) on the recovery of mechanical function and proton production were not affected by glibenclamide. 6. The data indicate that the cardioprotective mechanism of adenosine A1-receptor stimulation does not involve the activation of K(ATP)-channels. Furthermore, in rat fatty acid perfused, working hearts, stimulation of K(ATP)-channels is not cardioprotective and has no significant effects on myocardial glucose metabolism.

Intrinsic ANG II type 1 receptor stimulation contributes to recovery of postischemic mechanical function.

To determine whether intrinsic angiotensin II (ANG II) type 1 receptor (AT1-R) stimulation modulates recovery of postischemic mechanical function, we studied the effects of selective AT1-R blockade with losartan on proton production from glucose metabolism and recovery of function in isolated working rat hearts perfused with Krebs-Henseleit buffer containing palmitate, glucose, and insulin. Aerobic perfusion (50 min) was followed by global, no-flow ischemia (30 min) and reperfusion (30 min) in the presence (n = 10) or absence (n = 14) of losartan (1 mumol/l) or the cardioprotective adenosine A1 receptor agonist N6-cyclohexyladenosine (CHA, 0.5 mumol/l, n = 11). During reperfusion in untreated hearts (controls), left ventricular (LV) minute work partially recovered to 38% of aerobic baseline, whereas proton production increased to 155%. Compared with controls, CHA improved recovery of LV work to 79% and reduced proton production to 44%. Losartan depressed recovery of LV work to 0% without altering proton production. However, exogenous ANG II (1-100 nmol/l) in combination with losartan restored recovery of LV work during reperfusion in a concentration-dependent manner, suggesting that postischemic recovery of function depends on intrinsic AT1-R stimulation.

Effect of the novel angiotensin II type 1 receptor antagonist L-158,809 on acute infarct expansion and acute anterior myocardial infarction in the dog.

OBJECTIVES: To assess the effect of the angiotensin II type 1 receptor (AT1-R) antagonist L-158,809 on acute infarct expansion and left ventricular (LV) function during acute anterior myocardial infarction. METHODS: Dogs were randomized to receive intravenous L-158,809 (0.1 mg/kg bolus and 0.6 microgram/kg/min infusion) or vehicle beginning 1 h after permanent left anterior descending coronary artery ligation and continued for 48 h. In vivo LV remodelling and function (quantitative echocardiography) and hemodynamics over 48 h, and postmortem remodelling after 48 h were measured. RESULTS: L-158,809 produced 90% to 100% inhibition of the angiotensin II pressor response during the infusions. With respect to percentage changes over the 48 h in vivo, compared with vehicle controls, L-158,809 decreased mean arterial pressure (-20 +/- 4 versus -9 +/- 2%, P = 0.03) and left atrial pressure (-38 +/- 5 versus 25 +/- 6%, P < 0.0001) but did not change heart rate. These unloading effects were associated with a smaller percentage increase in infarct expansion index (-5 +/- 7% versus 27 +/- 2%, P = 0.001) and LV diastolic volume (11 +/- 11% versus 52 +/- 6%, P = 0.008), less shape deformation, fewer apical aneurysms (0 versus 100%, P = 0.0003), better global ejection fraction (49 +/- 2% versus 39 +/- 2%, P = 0.005), less ST segment elevation and fewer Q waves. Also compared with vehicle controls, with L-158,809 postmortem infarct size (19.8 +/- 2.4% versus 50.4 +/- 4.7% of risk region, P = 0.0002) and expansion index (2.06 +/- 0.09 versus 2.76 +/- 0.18, P = 0.006) were less and thinning ratio greater (0.92 +/- 0.02 versus 0.60 +/- 0.05, P = 0.0001). CONCLUSIONS: The novel AT1-R antagonist L-158,809 produces significant AT1-R blockade, reduces LV loading, and effectively limits acute infarct expansion and early LV remodelling during canine myocardial infarction.

Changes in myocardial density during postinfarction healing: effect on estimation of in vivo left ventricular mass by echocardiographic imaging.

To determine whether changes in density (rho) of infarct and noninfarct zones during healing and remodeling after myocardial infarction influence estimates of left ventricular mass and detection of temporal changes by imaging, we measured weights (g) and volumes (mL) of infarct, noninfarct, and mixed tissue in hearts removed 1 to 42 days after anterior infarction in three groups of dogs: nonreperfused infarction treated with placebo or captopril, or infarcts reperfused after 2 h. In vivo mass was calculated from in vivo diastolic myocardial volumes (echocardiograms) and an assumed density of 1.05 g/mL or actual values derived from tissue weights and volumes. Over the 42 days, actual density deviated more from the assumed value of 1.05 in infarct than noninfarct zones, and the overall density was higher for reperfused than nonreperfused ventricles (1.09 vs. 1.06 g/mL, p < 0.01). Correction for density improved the correlation between absolute in vivo and postmortem mass slightly but not the detection of relative changes in mass in control, captopril, or reperfusion groups. These findings suggest that (i) densities of infarct and noninfarct zones differ and change during healing, especially after reperfusion, and (ii) correction for density provides more accurate estimates of volume-derived mass in reperfused hearts.

Functional classification of P1-purinoceptors in guinea-pig left and right atria: anomalous characteristics of antagonism by cyclopentyltheophylline.

The P1 purinoceptor subtype mediating the negative inotropic responses of guinea-pig left atria and the negative chronotropic responses of beating right atria were characterized. Guinea-pig isolated paced left atria (2Hz, 5ms, threshold voltage+50%) and spontaneously beating right atria were set up in Krebs-bicarbonate solution and isometric tension and rate of contraction, respectively, were recorded. Concentration-response curves for the reduction of tension and rate, respectively, by adenosine receptor agonists, N6-cyclopentyladenosine (CPA), the R- and S- stereoisomers of N6-(2-phenylisopropyl) adenosine (R-PIA and S-PIA), 5'-(N-carboxamido) adenosine (NECA) and 2-p-((carboxyethyl)-phenethylamino)-5'-(N-carboxamido) adenosine (CGS21680) were obtained. The orders of potency on the left atria (CPA = NECA > R-PIA > S-PIA > CGS21680) and right atria (CPA = R-PIA > S-PIA > CGS21680) were consistent with the responses being mediated via A1 receptors. Antagonism of the responses to CPA or R-PIA by 8-cyclo-1,3-dimethylxanthine (CPT) was examined by a full Schild analysis. Concentration-response curves for CPA or R-PIA were obtained in the absence or presence of five or six concentrations (10(-7)-10(-5) or 3 x 10(-5)M) of CPT. The shift in the concentration-response by CPT was expressed as the concentration-ratio (CR) and plotted as -log(CR-1) against log molar concentration of CPT (Schild plot). pA2 values were calculated from the intercept on the concentration axis and by application of the equation; pA2 = log(antagonist concentration) -log (CR-1). The Schild plots had unity slopes indicating competitive antagonism and the pA2 values derived therefrom indicated that the responses were mediated via A1-receptor. Closer inspection of the Schild plots, however, showed that at the higher concentrations of CPT there was a limit to the displacement of the concentration-response curves of the left and right atria to CPA and of the left atria to R-PIA. There were also significant differences in the apparent pA2 values calculated from the equation, when different concentrations of antagonist were examined. These results indicated that at higher concentrations of agonist there may be a component of the response that is resistant to antagonism by CPT. Whether this is related to the proposal that cardiac responses are mediated via A3 receptors is discussed.

Opposite effects of angiotensin AT1 and AT2 receptor antagonists on recovery of mechanical function after ischemia-reperfusion in isolated working rat hearts.

BACKGROUND: Angiotensin II type 1 (AT1) receptor antagonists, when given over the long term, reduce the deleterious consequences of ischemia-reperfusion injury. Whether short-term administration of AT1 or angiotensin II type 2 (AT2) receptor antagonists is cardioprotective has not been investigated. METHODS AND RESULTS: The effects of short-term administration of selective AT1 and AT2 receptor antagonists on the recovery of mechanical function during reperfusion after 30 minutes of global, no-flow ischemia were studied in left atrium-perfused isolated working rat hearts. Control hearts (n = 8) showed incomplete recovery of left ventricular minute work (LV work) and cardiac efficiency during reperfusion to 51 +/- 15% and 61 +/- 19% of preischemic levels, respectively. Compared with control hearts, the selective AT2 receptor antagonist PD123,319 (0.3 mumol/L) given before ischemia (n = 7) improved the recovery of LV work and efficiency to 82 +/- 4% and 98 +/- 7% of preischemic levels, respectively (P < .01). In contrast, the selective AT1 antagonist losartan (1 mumol/L) blocked the recovery of LV work and depressed efficiency to 0 +/- 0% and 1 +/- 0% (n = 7) of preischemic levels, respectively (P < .01; n = 7). Neither antagonist altered coronary vascular conductance. CONCLUSIONS: This is the first demonstration that short-term treatment with a selective AT1 versus AT2 antagonist exerts different effects on recovery of mechanical function after ischemia-reperfusion: the AT2 antagonist was cardioprotective, whereas the AT1 antagonist was not. These data suggest that AT2 antagonists and AT1 agonists may offer novel approaches for the treatment of mechanical dysfunction after ischemia-reperfusion.

Vasorelaxation of noradrenaline-constricted guinea-pig and rabbit aorta by the adenosine analogue NECA: roles of intra-and extracellular calcium.

The action of the adenosine agonist, 5'-(N-ethylcarboxamido)-adenosine (NECA), at extracellular A2 receptors of guinea-pig and rabbit aortic rings was investigated. A near-maximum relaxant concentration (10(-5) M) of NECA was determined from cumulative concentration-response curves in aortae precontracted with noradrenaline. The effects of this concentration of NECA upon the noradrenaline-induced contractions were measured as the ratio of the contractions obtained before and, in the same tissue, after addition of NECA. This ratio was compared with the control ratio obtained in paired tissues after adding vehicle between the first and second contraction. The roles of intracellular Ca2+ mobilization and influx of extracellular Ca2+ were examined using normal Ca2+ and Ca(2+)-free media. In normal Ca2+ medium, where both sources of Ca2+ are involved in the contraction to noradrenaline, NECA inhibited the contractions. In Ca(2+)-free conditions, the phasic contraction to noradrenaline was mediated via the intracellular Ca2+ pool and was not inhibited by NECA. The contractions of the guinea-pig aorta to angiotensin II (10(-6) M) in both normal and Ca(2+)-free media, which are mediated via release of intracellular Ca2+, were also not inhibited by NECA. These results indicate that the activation of extracellular A2 adenosine receptors by NECA does not cause vasorelaxation by interfering with the release of intracellular Ca2+ by noradrenaline. The effects of NECA on contractions, due to influx of extracellular Ca2+, were examined in guinea-pig aortae in Ca(2+)-free medium and after exposure to angiotensin to deplete intracellular Ca2+ stores. Contractions were then induced by restoring the Ca2+ to the medium. These contractions were not inhibited by NECA, but when noradrenaline was present during the restoration of Ca2+, NECA was inhibitory. This and the evidence in normal Ca2+ medium, suggests that NECA causes vasorelaxation in the aorta by interfering with the Ca2+ influx via receptor-operated channels induced by noradrenaline.

Potassium channel blockade of atrial negative inotropic responses to P1-purinoceptor and muscarinic receptor agonists and to cromakalim.

The negative inotropic responses of guinea pig isolated left atria to the P1-receptor agonists adenosine and L-N6-phenylisopropyladenosine (L-PIA), the muscarinic receptor agonist carbachol, and the potassium channel activator cromakalim were examined. The potassium channel blocker 4-aminopyridine (4-AP, 10 mM) decreased the concentration-response curve for L-N-6-phenylisopropyladenosine (L-PIA) so that the maximum reduction in atrial tension was significantly reduced from 77.8 to 61.9%. The concentration-response curve for the negative inotropic response to carbachol was displaced to the right by a substantially greater amount (521-fold), partly due to the muscarinic receptor blocking activity of 4-AP. The response to a single submaximum dose of L-PIA (520 nM) was slow in onset and monophasic, whereas adenosine induced a rapid reduction in tension followed by a gradual return toward the resting level. In the presence of 4-AP (10 mM), the peak response to L-PIA was significantly reduced from a 59.7 to 31.0% inhibition of tension. In addition, the rate of development of the response was significantly slowed. The peak inhibition of tension by adenosine (112 microM) (60.0%) was also significantly reduced to 37.4% in the presence of 4-AP (10 mM). Furthermore, there was no rapid decrease in tension but a gradual decrease to a peak effect which was no different from that in the absence of 4-AP. These results suggest that the P1-receptor agonists exert negative inotropy through a K+ channel-dependent component which is blocked by 4-AP and a slower-onset K+ channel-independent component. The potassium channel blocker, bromobenzoylmethyladamantylamine (BMA 100 microM) also shifted (11-fold) the concentration-response curve for the negative inotropic response to adenosine, and to a lesser extent that to cromakalim (2.1-fold) and carbachol. Glibenclamide (0.3, 3, and 30 microM), a selective antagonist of ATP-regulated potassium channels, was a potent antagonist of the negative inotropic responses to cromakalim. However, 30 microM failed to antagonize the responses to L-PIA, adenosine, or carbachol, indicating that they do not mediate their responses through ATP-regulated potassium channels. 4-AP (10 mM) caused a slight shift (2.3-fold) of the concentration-response curve for cromakalim, indicating weak activity against ATP-regulated potassium channels in the left atrium. This study showed that P1-receptor agonists exert negative inotropic responses in guinea pig left atria only in part through activation of potassium channels. These channels differ from the ATP-regulated potassium channels activated by cromakalim.

Converting an acute care unit into a skilled nursing facility: a methodology.

To combat the problem of excess capacity and to meet the needs of its elder community. Highland Park Hospital (HPH) in Highland Park, Illinois decided to convert an unused acute care unit into a skilled nursing facility (SNF). Detailed here is the methodology used to determine the number of SNF beds needed by HPH, the options for development available, and the factors considered when choosing the type of facility to be designed.