Intracellular sodium increase and susceptibility to ischaemia in hearts from type 2 diabetic db/db mice.

AIMS/HYPOTHESIS: An important determinant of sensitivity to ischaemia is altered ion homeostasis, especially disturbances in intracellular Na(+) (Na(i)(+)) handling. As no study has so far investigated this in type 2 diabetes, we examined susceptibility to ischaemia-reperfusion in isolated hearts from diabetic db/db and control db/+ mice and determined whether and to what extent the amount of (Na(i)(+)) increase during a transient period of ischaemia could contribute to functional alterations upon reperfusion. METHODS: Isovolumic hearts were exposed to 30-min global ischaemia and then reperfused. (23)Na nuclear magnetic resonance (NMR) spectroscopy was used to monitor[Formula: see text] and (31)P NMR spectroscopy to monitor intracellular pH (pH(i)). RESULTS: A higher duration of ventricular tachycardia and the degeneration of ventricular tachycardia into ventricular fibrillation were observed upon reperfusion in db/db hearts. The recovery of left ventricular developed pressure was reduced. The increase in[Formula: see text] induced by ischaemia was higher in db/db hearts than in control hearts, and the rate of pH(i) recovery was increased during reperfusion. The inhibition of Na(+)/H(+) exchange by cariporide significantly reduced (Na(i)(+)) gain at the end of ischaemia. This was associated with a lower incidence of ventricular tachycardia in both heart groups, and with an inhibition of the degeneration of ventricular tachycardia into ventricular fibrillation in db/db hearts. CONCLUSIONS/INTERPRETATION: These findings strongly support the hypothesis that increased (Na(i)(+)) plays a causative role in the enhanced sensitivity to ischaemia observed in db/db diabetic hearts.

Modulation of vascular tone and reactivity by nitric oxide in porcine pulmonary arteries and veins.

Isolated porcine pulmonary vessels were studied in order to evaluate the role of nitric oxide in arteries and veins. Leukotriene C4 and noradrenaline contracted porcine pulmonary arteries but induced only negligible contractions of porcine pulmonary veins. After treatment with the nitric oxide synthase inhibitor N(omega)-nitro-L-arginine (L-NOARG), significant contractions to leukotriene C4 and noradrenaline were uncovered in pulmonary veins. In arterial preparations, L-NOARG caused a less marked potentiation of noradrenaline-induced contractions and did not alter leukotriene C4-induced contractions. Endothelium-dependent relaxations to acetylcholine were greater in veins compared with arteries whereas the endothelium-independent relaxations to the nitric oxide donor sodium nitroprusside (SNP) and the cyclic nucleotide analogue 8-bromo-cGMP were similar in the two preparations. Taken together these data suggest that the apparent insensitivity of porcine pulmonary veins to leukotriene C4 and noradrenaline was because of release of nitric oxide. The effect of nitric oxide synthase inhibition was less pronounced in porcine pulmonary arteries, suggesting a preferential functional role of nitric oxide in porcine pulmonary veins, originating in a greater production of nitric oxide by veins as opposed to arteries.

EGTA treatment of human airways in vitro unmasks M1/MUC5AC mucin in submucosal glands.

Mucin staining can be used to evaluate secretory activity of human airways. However, mucin epitopes may be masked by physicochemical properties of the secretions. The aim of this investigation was to examine the effects of the calcium chelator, ethyleneglycol-bis-(beta-aminoethylether)-N,N,N',N'-tetraacetic acid (EGTA) on the detection of M1/MUC5AC mucin in isolated human bronchial preparations. Immunohistochemical investigation and immunoradiometric assays with anti-M1 monoclonal antibodies (Mabs) were used to detect M1/MUC5AC mucin derived from bronchial preparations with an intact surface epithelium, or in tissues where the epithelium had been removed (rubbed preparations). The Mabs labelled both epithelial goblet cells and submucosal glandular cells in EGTA (4 mM)-exposed bronchial preparations, while only goblet cells were stained in EGTA (0.4 mM)-exposed tissues. The quantities of M1/MUC5AC mucin detected in either the bronchial fluids derived from EGTA (4 mM)-exposed intact and rubbed preparations or in bronchial fluids treated with EGTA (4 mM) were significantly increased by two-fold when compared with untreated control values (p<0.001). In addition, lactate dehydrogenase (LDH) activity and protein measurements were unaltered during exposure of human airways to EGTA (4 mM) suggesting that this treatment did not affect tissue viability. These results provide evidence that ethyleneglycol-bis-(beta-aminoethylether)-N,N,N',N'-tetraacetic acid (4 mM) facilitates the detection of M1/MUC5AC mucin by altering the physicochemical properties of respiratory mucin, thereby exposing epitopes with which anti-M1 monoclonal antibodies are reactive. This will allow more accurate measurement of secretory activity in human airways in vitro.

MUC5AC mucin release from human airways in vitro: effects of indomethacin and Bay X1005.

BACKGROUND: Increased secretion of mucus is a hallmark of many respiratory diseases and contributes significantly to the airflow limitation experienced by many patients. While the current pharmacological approach to reducing mucus and sputum production in patients is limited, clinical studies have suggested that drugs which inhibit the cyclooxygenase and/or 5-lipoxygenase enzymatic pathways may reduce secretory activity in patients with airway disease. AIM: This study was performed to investigate the effects of indomethacin (cyclooxygenase inhibitor) and Bay x 1005 (5-lipoxygenase inhibitor) on MUC5AC release from human airways in vitro. METHODS: An immunoradiometric assay was used to determine the quantities of MUC5AC present in the biological fluids derived from human airways in vitro. The measurements were made with a mixture of eight monoclonal antibodies (MAbs; PM8) of which the 21 M1 MAb recognized a recombinant M1 mucin partially encoded by the MUC5AC gene. RESULTS: The quantities of MUC5AC detected in the biological fluids derived from human bronchial preparations were not modified after treatment with indomethacin (cyclooxygenase inhibitor) and/or an inhibitor of the 5-lipoxygenase metabolic pathway (BAY x 1005). CONCLUSION: These results suggest that the cyclooxygenase and 5-lipoxygenase metabolic pathways play little or no role in the release of MUC5AC from human airways.

Development of a functional human bronchial model of mucin secretion.

In an attempt to study the functional aspects of respiratory mucin secretion and the effects of mediators of inflammation on the release of M1/MUC5AC mucins in airways diseases, an ex vivo human bronchial model of mucin secretion was developed. Anti-M1 mucin monoclonal antibodies raised against the peptidic core of ovarian cyst M1 mucins were used. PAS and Alcian blue stainings of sections of bronchial rings revealed the presence of mucins in epithelial goblet cells as well as in glandular mucous cells. Immunohistochemical labelling of these sections with anti-M1 monoclonal antibodies revealed a preferential localization of M1/MUC5AC mucins in epithelial goblet cells. Functional studies were performed on this bronchial model using various secretagogues (methacholine, leukotrienes D4 and anti-human immunoglobulin E antibodies). No statistical difference of M1/MUC5AC mucin secretion was observed after a one-hour stimulation of bronchial rings with these agents. The development of an ex vivo functional human bronchial model of mucin secretion and the use of specific anti-M1 antibodies are essential tools in studying the regulation of the M1/MUC5AC mucin release from human airways.

Evidence for a M(1) muscarinic receptor on the endothelium of human pulmonary veins.

1. To characterize the muscarinic receptors on human pulmonary veins associated with the acetylcholine (ACh)-induced relaxation, isolated venous and arterial preparations were pre-contracted with noradrenaline (10 microM) and were subsequently challenged with ACh in the absence or presence of selective muscarinic antagonists. 2. ACh relaxed venous preparations derived from human lung with a pD(2) value of 5.82+/-0.09 (n=16). In venous preparations where the endothelium had been removed, the ACh relaxations were abolished (n=4). ACh relaxed arterial preparations with a pD(2) value of 7. 06+/-0.14 (n=5). 3. Atropine (1 microM), the non selective antagonist for muscarinic receptors, inhibited ACh-induced relaxations in human pulmonary veins. The affinity value (pK(B) value) for atropine was: 8.64+/-0.10 (n=5). The selective muscarinic antagonists (darifenacin (M(3)), himbacine (M(2),M(4)), methoctramine (M(2)) and pFHHSiD (M(1),M(3))) also inhibited ACh-induced relaxations in venous preparations. The pK(B) values obtained for these antagonists were not those predicted for the involvement of M(2 - 5) receptors in the ACh-induced relaxation in human pulmonary veins. 4. The pK(B) value for darifenacin (1 microM) was significantly greater in human pulmonary arterial (8.63+/-0.14) than in venous (7.41+/-0.20) preparations derived from three lung samples. 5. In human pulmonary veins, the pK(B) values for pirenzepine (0.5 and 1 microM), a selective antagonist for M(1) receptors, were: 7.89+/-0.24 (n=7) and 8.18+/-0.22 (n=5), respectively. In the venous preparations, the pK(B) values derived from the functional studies with all the different muscarinic antagonists used were correlated (r=0.89; P=0.04; slope=0.78) with the affinity values (pK(i) values) previously published for human cloned m1 receptors in CHO cells. 6. These results suggest that the relaxations induced by ACh are due to the activation of M(1) receptors on endothelial cells in isolated human pulmonary veins.

Functional studies of leukotriene receptors in vascular tissues.

The paradoxical effects of cysteinyl-leukotrienes, namely contraction and relaxation, are now well documented in a number of vascular preparations from various species. The vascular smooth muscle contractions are associated with activation of a single receptor subtype and in some vascular smooth muscles with activation of two receptor subtypes. However, the receptors implicated in the contraction of vessels such as pig pulmonary arteries and veins, dog inferior vena cava, and dog splenic and mesenteric veins remain to be established. There are sufficient data concerning some vascular tissues to suggest that relaxations induced by cysteinyl-leukotrienes are via the stimulation of specific receptors present on the endothelium. The endothelium in human pulmonary arteries has one receptor (CysLT2) and activation induced the release of NO. However, in isolated human pulmonary veins two receptors are present, CysLT1 and CysLT2 (Figure 1). Activation of the former induced the release of a contractile factor whereas activation of the CysLT2 receptor released NO. In guinea pig pulmonary artery and guinea pig thoracic aorta, one receptor has been demonstrated since the relaxations are blocked by ICI-198615. These data suggest the presence of a CysLT1 receptor. Activation of this receptor leads to the release of a relaxant factor, namely, nitric oxide. In contrast, in human pulmonary arteries and veins activation of a receptor that is resistant to ICI-198615 is associated with NO release. These results suggest that there may be species differences even when analogous vascular preparations are examined. While the cysteinyl-leukotrienes are known to relax vascular smooth muscle in a variety of preparations from different species, there are presently two pathways known to be involved in this response. One involves the metabolites of arachidonic acid via the cyclooxygenase enzymatic pathway and the other implicates products of the L-arginine enzymatic pathway. Although both pathways may be present and active in the endothelium of the vascular preparations only one of these enzymatic pathways may be dominant and responsible for the relaxations observed. Ortiz and coworkers have demonstrated that in pulmonary veins the dominant pathway for cysteinyl-leukotriene relaxations is the NO pathway. There are some reports from animal studies that support a dominant role for NO in pulmonary veins. In contrast, Allen and co-workers demonstrated that the LTC4-induced relaxations in isolated human saphenous veins were not modified by treatment of tissues with an NO inhibitor but were significantly enhanced after treatment with indomethacin. These authors suggested that a contracting factor derived from the arachidonic acid pathway was released in preparations challenged with LTC4. In addition, these investigators demonstrated that the NO inhibitor had no effect on the LTC4 relaxations. Together, these results suggest that cysteinyl-leukotriene effects in human pulmonary veins are dominated by the NO pathway whereas in human systemic veins these mediator effects are modified by metabolites of the cyclooxygenase pathway. Unfortunately, most studies involving the actions of cysteinyl-leukotrienes on vessels have been performed in the presence of indomethacin, making interpretation of the relative contribution of the cyclooxygenase and NO pathways difficult. In any event, the cysteinyl-leukotrienes may have a prominent role in the activation of these pathways and the receptors involved have not been clearly established.

ATP induced MUC5AC release from human airways in vitro.

BACKGROUND: Chronic airway diseases are often associated with marked mucus production, however, little is known about the regulation of secretory activity by locally released endogenous mediators. AIM: This investigation was performed to determine the release of MUC5AC mucin from human bronchial preparations using the purinergic agonists adenosine 5'-triphosphate (ATP) and uridine 5'-triphosphate (UTP). METHODS: Immunohistochemical and immunoradiometric assays (IRMA) were used to detect the MUC5AC mucin. Immunohistochemical analysis were performed using individual 1-13 M1 and 21 M1 MAbs recognizing a recombinant M1 mucin partially encoded by the MUC5AC gene. IRMA measurments were performed using a mixture of eight anti-M1 mucin MAbs (PM8), which included both 1-13 M1 and 21 M1 MAbs. Lysozyme and protein were also measured in the biological fluids derived from human bronchial preparations obtained from patients who had undergone surgery for lung carcinoma. RESULTS: The anti-M1 monoclonal antibodies labelled epithelial goblet cells. After challenge of human bronchial preparations with ATP, the goblet cells exhibited less staining. In contrast, UTP did not alter the immunolabelling of goblet cells. MUC5AC mucin in the bronchial fluids derived from ATP-challenged preparations was increased while UTP had no effect on release. ATP did not alter either the quantities of lysozyme or protein detected in the biological fluids. CONCLUSION: These results suggest that ATP may regulate epithelial goblet cell secretion of MUC5AC mucin from human airways in vitro.

M1/MUC5AC mucin released by human airways in vitro.

A series of monoclonal antibodies which bind to a mucin known as M1 (anti-M1 MAbs) have also been shown to detect the product of the human gene MUC5AC. The aim of this investigation was to determine the concentration of the M1 mucin in the surface epithelium of human bronchial preparations by means of immunohistochemistry and in the bronchial fluid derived from human airways by means of an immunoradiometric assay. Human bronchial ring preparations from the resection material of 20 patients were challenged with methacholine, leukotriene D4, or anti-immunoglobulin E. Experiments were performed in preparations with an intact epithelium as well as in tissues in which the epithelium had been mechanically removed. The anti-M1 MAbs stained the goblet cells in the epithelium intensely and there was also light and less uniform staining in the submucosa. The M1/MUC5AC mucin in the fluids secreted by the bronchial preparations was not modified during either the experimental protocol or stimulation with the different secretagogues. However, in preparations in which the epithelium had been removed, there was a significant reduction in the amount of M1/MUC5AC mucin detected. These data suggest that the M1/MUC5AC mucin detected in the biological fluids produced by human airways in vitro may be released constantly, and principally from the goblet cells in the epithelial layer.

Cholinergic control of human and animal pulmonary vascular tone.

The regulation of pulmonary vascular tone by acetylcholine (ACh) involves the activation of different subtypes of muscarinic receptors as well as the cholinesterase activities which are responsible for ACh degradation. Most of the studies on the cholinergic control of the pulmonary vascular tone have been performed in vessels derived from animals. The ability of ACh to induce pulmonary vasoconstriction is species dependent. In vessels derived from sheep lung, ACh induced contractions in veins but not in arteries whereas in human pulmonary vessels the reverse was observed. The subtype(s) of the muscarinic receptors involved in the pulmonary vasoconstrictions is also dependent on the species which are studied. M1 receptors are implicated in the rabbit pulmonary vasoconstrictions, M3 in humans, whereas M1 and M2 receptors are involved in the dog. The cholinesterases are implicated in the vasoconstriction produced by ACh in human and rabbit pulmonary arteries. However, in these studies while acetylcholinesterase (EC 3.1.1.7) and butyrylcholinesterase (EC 3.1.1.8) activities were detected in human vessels only acetylcholinesterase activity was found in rabbit vessels. The endothelium-dependent relaxation induced by ACh has been reported in isolated pulmonary vessels from different animals including man. However, the muscarinic receptors involved in the ACh-induced vasodilatation of rat and rabbit pulmonary artery are of the M3 subtype while those characterized in the human pulmonary artery are of the M3 and M1 subtypes. Together these results concerning the cholinergic control of the pulmonary vascular tone indicate that extrapolation of the data obtained in animal models to human vessels requires some caution. In addition, there is considerable evidence to demonstrate that ageing may modify cholinergic responses. However, little information is available concerning the pulmonary vascular bed during ageing.

Prostanoid receptors involved in the relaxation of human bronchial preparations.

1. Iloprost and cicaprost (IP-receptor agonists) induced relaxations in the histamine- (50 microM) contracted human bronchial preparations (pD2 values, 6.63+/-0.12 and 6.86+/-0.08; Emax values, 90+/-04 and 65+/-08% of the papaverine response for iloprost (n=6) and cicaprost (n=3), respectively). 2. Prostaglandin E2 (PGE2) and misoprostol (EP-receptor agonist) relaxed the histamine-contracted human bronchial preparations (pD2 values, 7.13+/-0.07 and 6.33+/-0.28; Emax values, 67+/-04 and 57+/-08% of the papaverine response for PGE2 (n=14) and misoprostol (n=4), respectively). In addition, both relaxations were inhibited by AH6809 (DP/EP1/EP2-receptor antagonist; 3 microM; n=5-6). 3. The PGE2-induced relaxations of human bronchial preparations were not modified by treatment with AH23848B (TP/EP4-receptor antagonist; 30 microM; n=4). 4. The contracted human bronchial preparations were significantly relaxed by prostaglandin D2 (PGD2) or by BW245C a DP-receptor agonist. However, these responses did not exceed 40% of the relaxation induced by papaverine. In addition, the relaxations induced by PGD2 were significantly inhibited by treatment with a DP-receptor antagonist BWA868C (0.1 microM; n=3). 5. These data suggest that the relaxation of human isolated bronchial preparations induced by prostanoids involved IP-, EP2- and to a lesser extent DP-receptors but not EP4-receptor.

Prostanoid receptors involved in the relaxation of human pulmonary vessels.

1. To characterize the prostanoid receptors on human pulmonary smooth muscle involved in vasodilatations, isolated arteries and veins were contracted with norepinephrine (10 microM) and vessels were subsequently challenged with different prostanoid-receptor agonists in the absence or presence of selective antagonists. 2. Prostaglandin D2 (PGD2) and the selective DP-receptor agonist, BW245C, induced relaxations in the contracted human pulmonary venous preparations. The pD2 values were: 6.88+/-0.11 (n=17) and 7.31+/-0.12 (n=5), respectively. The relaxant responses induced by PGD2 were reduced by the selective DP-receptor antagonist, BWA868C, and the estimated pA2 value was 7.84+/-0.16 (n=4). PGD2 and BW245C did not relax contracted human pulmonary arteries. 3. The selective IP-receptor agonists, iloprost and cicaprost, both induced relaxations in the contracted human vascular preparations. The pD2 values for iloprost were: 7.84+/-0.08 (n=6) and 8.25+/-0.06 (n=4) and for cicaprost: 8.06+/-0.12 (n=5) and 8.11+/-0.09 (n=5) in arteries and veins respectively. 4. Prostaglandin E2 (PGE2) and the EP2/EP3-receptor agonist, misoprostol, partially relaxed the contracted venous preparations and the pD2 values were: 8.10+/-0.15 (n=15) and 6.24+/-0.33 (n=3), respectively. These relaxations suggest the presence of an EP receptor in the human pulmonary veins. The contracted human pulmonary arteries did not relax when challenged with PGE2. 5. In human pulmonary venous preparations, the PGE2-induced relaxations were neither modified by treatment with TP/EP4-receptor antagonist, AH23848B (10 and 30 microM, n=6), nor by the DP/EP1/EP2-receptor antagonist, AH6809 (3 microM, n=6). 6. These data suggest that the relaxation induced by prostanoids involved DP-, IP-receptors and to a lesser extent an EP-receptor on human pulmonary venous smooth muscle. In contrast, only the IP-receptor is involved in the prostanoid induced relaxations on human pulmonary arterial smooth muscle.

Airway epithelial damage and release of inflammatory mediators in human lung parenchyma after sulfur mustard exposure.

This study was performed to evaluate the morphological effects of sulfur mustard on human lung parenchyma in vitro and to measure the metabolites of arachidonic acid which are released during acute exposure to the alkylating agent. Histological analysis of the tissue following exposure to sulfur mustard for a period of 45 min at 10 mM revealed the presence of paranuclear vacuoles in the epithelium, specifically, in the ciliated cells. The release of metabolites of arachidonic acid were determined in the bath fluids by an enzymo-immunoassay. The basal release of prostaglandin E2 (PGE2: 1.36 +/- 0.33 ng/g tissue) and 6-keto prostaglandin F1 alpha (6-keto PGF1 alpha: 8.83 +/- 1.17 ng/g tissue) were not modified during tissue exposure to sulfur mustard (45 min, 0.1 mM). In addition, the basal release of cysteinyl-leukotriene E4 (LTE4: 1.55 +/- 0.44 ng/g tissue) was also not altered by challenge of the tissues with sulfur mustard. In contrast, when the human lung parenchyma was stimulated with anti human IgE (anti-IgE) only the basal release of the metabolite of the 5-lipoxygenase pathway was significantly increased (LTE4: 6.84 +/- 1.57 ng/g tissue). These data suggest that sulfur mustard may produce morphological alterations in epithelial cells and at the time point studied (45 min exposure), this effect is not associated with a release of arachidonic acid metabolites. However, the increased release of LTE4 by anti-IgE suggests that the target cells for sulfur mustard and anti-IgE in the human lung may be different.

Cysteinyl-leukotriene receptors in pulmonary vessels.

Two categories of cysteinyl-leukotrienes have been proposed, namely, CysLT1 and CysLT2. These receptors are found not only on the vascular smooth muscle but also on the endothelium. Activation of the receptor(s) on vascular smooth muscle provokes contraction whereas activation of the receptors on the endothelium produces contraction and/or relaxation. These endothelium dependent effects are due to the release of both contractile and relaxant factors derived from the endothelium. While factors derived from either the cyclooxygenase or nitric oxide pathways are involved, in some vascular preparations other mediators such as endothelin may be involved. However, in isolated human pulmonary vascular preparations, this appears not to be the case and presently the nature and origin of the contractile factor remains to be established.

Basal secretion of lysozyme from human airways in vitro.

The aim of this study was to examine the basal release of lysozyme from isolated human lung tissues. Measurements of lysozyme in the fluids derived from lung preparations were performed using a rate-of-lysis assay subsequent to acidification of the biological samples. Lysozyme released from bronchial preparations into fluids was greater than that observed for parenchymal tissues. The lysozyme quantities detected in bronchial fluids were not modified by removal of the surface epithelium. Furthermore, the quantities of lysozyme in bronchial fluids was correlated with the size of the bronchial preparations. These results suggest that the lysozyme was principally secreted by the human bronchi (submucosal layer) rather than by parenchyma tissues and that a greater release was observed in the proximal airways.

Leukotriene D4 contractions in human airways are blocked by SK&F 96365, an inhibitor of receptor-mediated calcium entry.

In human bronchial muscle preparations, nifedipine (3 microM) significantly inhibited the histamine, ACh and KCl contractions. However, the dihydropyridine did not modify the contractile responses induced by either leukotriene D4 (LTD4) or anti-human IgE (a-IgE). In human airways, SK&F 96365 (30 microM and 100 microM) markedly reduced the KCl and, at the higher concentration, LTD4 maximal contractions. In addition, when preparations were treated with nifedipine (3 microM), SK&F 96365 (100 microM) significantly blocked responses to both LTD4 and a-IgE. The calcium chelating agent ethylene glycol-bis (beta-amino-ethyl ether) N,N,N',N'-tetraacetic acid (4 mM) also inhibited the a-IgE-induced contractions. These data demonstrate that the nifedipine-resistant component of the LTD4 and a-IgE contractions was inhibited by SK&F 96365 and suggest that the cysteinyl-leukotriene receptor in human airways may be intimately linked with a receptor-operated calcium-entry mechanism.

Cholinesterase activity in human pulmonary arteries and veins.

1. Human isolated pulmonary vessels were treated with cholinesterase (ChE) inhibitors to determine the role of these enzymes in regulating vascular muscle tone. In addition, kinetic parameters were determined for acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) in human pulmonary vessel homogenates. 2. Carbachol (CCh) and acetylcholine (ACh) were equipotent contractile agonists in human pulmonary arteries (pD2 values, 5.28 +/- 0.05 and 5.65 +/- 0.16; Emax, 0.91 +/- 0.26 and 0.98 +/- 0.30 g wt. for CCh and ACh, respectively; n = 7). In venous preparations, ACh was ineffective and CCh induced small contractions (Emax, 0.08 +/- 0.04 g wt; n = 13). 3. In human pulmonary arteries following pretreatment with tetraisopropylpyrophosphoramide (iso-OMPA, 100 microM), an increased sensitivity to the contractile agonist ACh was observed (pD2 values, 5.80 +/- 0.13 and 6.37 +/- 0.19 for control and treated preparations, respectively; n = 5). This pretreatment had no effect on the CCh concentration response curve. In contrast, human pulmonary veins pretreated with iso-OMPA failed to elicit a contractile response to ACh. 4. Neither Iso-OMPA nor neostigmine elicited concentration-dependent contractions in human isolated pulmonary arteries or veins. These results suggest the absence of sufficient spontaneous release of ACh to modulate human pulmonary vessel basal tone. 5. CCh was less potent than ACh in relaxing precontracted human isolated pulmonary arteries (pD2 value, CCh: 6.55 +/- 0.15 and ACh: 7.16 +/- 0.13, n = 4) and veins (pD2 value, CCh: 4.95 +/- 0.13; n = 5 and ACh: 5.56 +/- 0.17; n = 6). Pretreatment of vessels with either iso-OMPA or neostigmine did not modify ACh relaxant responses in either type of preparation. 6. In human pulmonary veins, the ChE activity was two fold greater than in arteries (n = 6). Vmax for AChE was 1.73 +/- 0.24 and 3.36 +/- 0.26 miu mg-1 protein in arteries and veins, respectively, whereas Vss for BChE was 1.83 +/- 0.22 and 4.71 +/- 0.17 miu mg-1 protein, in these respectively. 7. In human pulmonary arteries, BChE activity may play a role in the smooth muscle contraction but not on the smooth muscle endothelium-dependent relaxation induced by ACh. A role for ChE activity in the control of venous tone is presently difficult to observe, even though this tissue contains a greater amount of enzyme than the artery.

Pulmonary hypertension in infants with congenital heart defects: are leukotrienes involved?

The circulating levels of leukotriene E4 in infants with congenital heart defects, increased pulmonary blood flow and pulmonary arterial hypertension, were determined and compared with infants with decreased pulmonary blood flow (Tetralogy of Fallot). There was no correlation (r=0.38) between the pulmonary arterial pressure (56+/-4 mmHg) and the leukotriene E4 levels (1.37+/-0.67 ng/ml blood) measured in peripheral blood samples from the hypertensive group prior to surgery. There was considerable variation in the detectable leukotriene E4 levels in blood samples from different patients. The levels detected in the blood samples between the two groups of patients was similar. These data suggest that neither the surgical repair during cardiopulmonary bypass nor the pulmonary hypertension appeared to modify the leukotriene E4 blood levels in the small number of patients studied.