Pig Pulmonary Artery Research Articles

Nonadrenergic noncholinergic vasodilation of guinea pig pulmonary arteries is mediated by nitric oxide

Nonadrenergic noncholinergic (NANC) mediated vasodilation may contribute to the maintenance of low pulmonary vascular tone. The NANC neurotransmitters, nitric oxide (NO) and the sensory neuropeptides, substance P and calcitonin gene related peptide (CGRP), were investigated as possible mediators of NANC vasodilation in guinea pig pulmonary arteries. Fresh guinea pig pulmonary artery rings, with and without an intact endothelium, were mounted in organ baths containing Krebs solution and precontracted with the prostaglandin F2alpha analogue U44069. In both endothelium-intact and denuded vessels, electrical field stimulation (1-12 Hz) in the presence of guanethidine and atropine resulted in a frequency-dependent vasodilation. The peptide fragment hCGRP8-37, a competitive antagonist of the CGRP receptors, the peptide fragment NK1 antagonist SP4-11, and the nonpeptide NK1 antagonist RP67580 had no effect on NANC vasodilation. In both endothelium-intact and denuded vessels, N(G)-nitro-L-arginine methyl ester (L-NAME), an inhibitor of NO synthesis, inhibited NANC vasodilation, an effect that was reversible with L-arginine. We conclude that NANC vasodilation in guinea pig pulmonary arteries is mediated predominantly through NO activity.

Characterization of the mechanism involved in the relaxant response of dopexamine in the guinea pig pulmonary artery in vitro.

Dopexamine is a synthetic catecholamine used for the management of low-cardiac-output states. The purpose of this study was to characterize some of the mechanisms underlying dopexamine-mediated relaxation in the guinea pig pulmonary artery (PA) in vitro. Dopexamine (EC50, 1.2 microM; Rmax, 100%), like dobutamine (EC50, 1.4 microM, Rmax, 93.3%), prostacyclin (PGI2; EC50, 37 nM; Rmax, 96.2%), sodium nitroprusside (EC50, 370 pM; Rmax, 96.9%), forskolin (EC50, 47 pM: Rmax, 98.6%), and SKF 38393 (EC50, 120 nM; Rmax, 100%), caused graded relaxation in rings of PA precontracted by phenylephrine. The dopexamine vasorelaxation was antagonized by propranolol (1 microM), SCH 23390 (100 nM, a D1-dopamine antagonist), sulpiride (1 microM), glibenclamide (30 microM), tetraethylammonium (3 mM), apamin (100 nM), charybdotoxin (100 nM), SQ 22536 (10 microM, an adenylyl cyclase inhibitor), KT 5720 (10 microM, a protein kinase A inhibitor) and by calcitonin gene-related peptide (CGRP) or vasoactive intestinal peptide (VIP)-receptor antagonists (both 100 nM), as well as by chymotrypsin (1 U/ml). Neither the prior incubation of N(G)-nitro-L-arginine (100 pM), indomethacin (1 microM), nor removal of the vascular endothelium interfered with dopexamine vasorelaxation response in PA. Thus dopexamine relaxation in PA is mediated by activation of beta-adrenoceptors and dopamine receptors, and by the opening of both low- and high-conductance Ca2+-activated K+ channels, partially through adenosine triphosphate (ATP)-sensitive K+ channels. In addition, dopexamine-induced relaxation in PA seems to involve the release of peptides such as VIP and CGRP, an effect mediated by a cyclic adenosine monophosphate (cAMP)-dependent mechanism.

Nonadrenergic noncholinergic vasodilation of guinea pig pulmonary arteries is mediated by nitric oxide

Nonadrenergic noncholinergic vasodilation of guinea pig pulmonary arteries is mediated by nitric oxide

Bradykinin antagonists with dehydrophenylalanine analogues at position 5

Continuing the studies on structural requirements of bradykinin antagonists, it has been found that analogues with dehydrophenylalanine (deltaPhe) or its ring-substituted analogues (deltaPhe(X)) at position 5 act as antagonists on guinea pig pulmonary artery, and on guinea pig ileum. Because both organs are considered to be bradykinin B2 receptor tissues, the analogues with deltaPhe or deltaPhe(X) at position 5, but without any replacement at position 7, seem to represent a new structural type of B2 receptor antagonist. All the analogues investigated act as partial antagonists; they inhibit the bradykinin-induced contraction at low concentrations and act as agonists at higher concentrations. Ring substitutions by methyl groups or iodine reduce both the agonistic and antagonistic activity. Only substitution by fluorine gives a high potency. Incorporation of deltaPhe into different representative antagonists with key modifications at position 7 does not enhance the antagonist activity of the basic structures, with one exception. Only the combination of deltaPhe at position 5 with DPhe at position 7 increases the antagonistic potency on guinea pig ileum by about one order of magnitude. Radioligand binding studies indicate the importance of position 5 for the discrimination of B2 receptor subtypes. The binding affinity to the low-affinity binding site (KL) was not significantly changed by replacement of Phe by deltaPhe. In contrast, ring-methylation of deltaPhe results in clearly reduced binding to KL. The affinity to the high-affinity binding site (KH) was almost unchanged by the replacement of Phe in position 5 by deltaPhe, whereas the analogue with 2-methyl-dehydrophenylalanine completely failed to detect the KH-site. The peptides were synthesized on the Wang-resin according to the Fmoc/Bu(t) strategy using Mtr protection for the side chain of Arg. The dehydrophenylalanine analogues were prepared by a strategy involving PyBop couplings of the dipeptide unit Fmoc-Gly-deltaPhe(X)-OH to resin-bound fragments.

Bradykinin antagonists with dehydrophenylalanine analogues at position 5.

Continuing the studies on structural requirements of bradykinin antagonists, it has been found that analogues with dehydrophenylalanine (deltaPhe) or its ring-substituted analogues (deltaPhe(X)) at position 5 act as antagonists on guinea pig pulmonary artery, and on guinea pig ileum. Because both organs are considered to be bradykinin B2 receptor tissues, the analogues with deltaPhe or deltaPhe(X) at position 5, but without any replacement at position 7, seem to represent a new structural type of B2 receptor antagonist. All the analogues investigated act as partial antagonists; they inhibit the bradykinin-induced contraction at low concentrations and act as agonists at higher concentrations. Ring substitutions by methyl groups or iodine reduce both the agonistic and antagonistic activity. Only substitution by fluorine gives a high potency. Incorporation of deltaPhe into different representative antagonists with key modifications at position 7 does not enhance the antagonist activity of the basic structures, with one exception. Only the combination of deltaPhe at position 5 with DPhe at position 7 increases the antagonistic potency on guinea pig ileum by about one order of magnitude. Radioligand binding studies indicate the importance of position 5 for the discrimination of B2 receptor subtypes. The binding affinity to the low-affinity binding site (KL) was not significantly changed by replacement of Phe by deltaPhe. In contrast, ring-methylation of deltaPhe results in clearly reduced binding to KL. The affinity to the high-affinity binding site (KH) was almost unchanged by the replacement of Phe in position 5 by deltaPhe, whereas the analogue with 2-methyl-dehydrophenylalanine completely failed to detect the KH-site. The peptides were synthesized on the Wang-resin according to the Fmoc/Bu(t) strategy using Mtr protection for the side chain of Arg. The dehydrophenylalanine analogues were prepared by a strategy involving PyBop couplings of the dipeptide unit Fmoc-Gly-deltaPhe(X)-OH to resin-bound fragments.

Reconstrução da continuidade ventrículo-pulmonar (conexão VD-TP): técnicas e táticas cirúrgicas

Os autores revisam as técnicas cirúrgicas utilizadas para a reconstrução da continuidade ventrículo direito (VD) e o tronco pulmonar (TP), dando ênfase às diferentes técnicas e associação, dependendo das formas anatômicas de cada lesão. Apresentam, para ilustração, 2 casos, operados com o uso de um novo conceito de prótese biológica produzida com a valva pulmonar e o tronco pulmonar suíno, preservados em glutaraldeído mediante dois modelos: retalho bivalvulado e tubo valvulado na experiência atual. No período de maio/91 e junho/95, foram operados 48 pacientes, em 43 foi usado retalho bivalvulado e, em 5 casos, prótese tubular valvulada. Houve 5 (10,4%) óbitos imediatos e 2 (4,6%) tardios. Este novo conceito de reconstrução da valva pulmonar e via de saída do ventrículo direito, usando heteroenxertos valvulados evita a insuficiência pulmonar significativa e apresenta baixa incidência de calcificação. É uma técnica facilmente reprodutível.

Nonadrenergic noncholinergic relaxation of human pulmonary arteries is partially mediated by nitric oxide.

Nonadrenergic noncholinergic (NANC) vasodilator mechanisms may contribute to the maintenance of low vascular resistance characteristic of the pulmonary circulation. Previous studies have demonstrated that nitric oxide (NO) is the principal NANC neurotransmitter in guinea pig pulmonary arteries. We examined whether NANC relaxation could be demonstrated in human pulmonary arteries, and the role of NO in this phenomenon. Fresh human pulmonary artery rings, with and without an intact endothelium, were mounted in organ baths containing Krebs' solution and precontracted with U44069 (0.3 microM). Adrenergic and cholinergic neurotransmitter pathways were blocked with guanethidine and atropine, respectively (10 microM each). In both endothelium-intact and -denuded vessels, electrical field stimulation (EFS, 1 to 10 Hz, 100 V) resulted in a frequency-dependent relaxation (maximal relaxation of 25 +/- 4% and 15 +/- 2% in endothelium-intact and -denuded vessels, respectively). Tetrodotoxin (0.3 microM) abolished the EFS-induced relaxation. L-NG-nitro-arginine methyl ester (L-NAME, 10 microM), was used to block enzymatic synthesis of NO. In both endothelium-intact and -denuded vessels, L-NAME reduced NANC relaxation to approximately 50% of control values. This reduction was reversible with the application of L-arginine (100 microM). We conclude that NANC relaxation exists in human pulmonary arteries and that it is partly mediated through NO.

Pharmacological characterization of histamine H 3 receptors in isolated guinea pig pulmonary artery and ileum

We characterized the histamine H 3 receptors involved in the modulation of electrical field stimulated neurogenic contraction of guinea pig pulmonary artery sympathetic, and guinea pig ileum parasympathetic preparations. Simultaneous measures of electrical field stimulation-evoked 3H overflow and tension in [ 3H]norepinephrine-loaded pulmonary artery were sensitive to tetrodotoxin (300 nM) and insensitive to hexamethonium (100 μM). Only the contractile response was inhibited by prazosin (100 nM). ( R)-α-Methylhistamine's inhibition of the pulmonary artery contraction and 3H overflow were dose-dependently antagonized by thioperamide (30–100 nM). ( R)-α-Methylhistamine also inhibited the neurogenic contractions of the isolated ileum (pD 2 = 8.2). In the pulmonary artery, the relative potency of the histamine H 3 receptor antagonists vs. ( R)-α-methylhistamine innibition of neurogenic contractions (pD 2 = 7.1) was thioperamide (pA 2 = 8.6 ± 0.1) > burimamide (pA 2 = 7.6 ± 0.2) > impromidine (pA 2 = 6.9 ± 0.02). Similarly, the relative potency of histamine H 3 receptor antagonists in the isolated ileum was thioperamide > burimamide ≥ impromidine, with pA 2 estimates of 8.7 ± 0.1, 7.3 ± 0.1 and 7.1 ± 0.1, respectively. Antagonist potencies suggest a predominant histamine H 3A-like receptor population on postganglionic sympathetic neurons innervating the pulmonary artery and parasympathetic neurons innervating the ileum longitudinal muscle.

Regulation of the adenylyl cyclase signaling system in various types of cultured endothelial cells.

We studied the effects of modulators of the adenylyl cyclase pathway on the accumulation of cAMP in endothelial cells isolated from bovine aortas, pig pulmonary arteries, human umbilical veins, and human subcutaneous adipose microvessels. In addition to quantitative differences in the basal levels, cAMP stimulation in different endothelial cell types varied in sensitivity and magnitude in response to both the direct adenylyl cyclase activator forskolin and the beta-adrenergic receptor agonist isoproterenol. Furthermore, the ubiquitous phosphodiesterase inhibitor IBMX differentially enhanced both the basal and the stimulated cAMP levels in the various cell types. Histamine caused an elevation of cAMP only in bovine aortic endothelial cells and in human umbilical vein endothelial cells. Treatment of the cells with cholera and pertussis toxins, which uniquely affect G-protein subunits, resulted in divergent elevation of cAMP in the various cells. Thus, in each cell type, a distinct profile of regulation of the cAMP levels was found. Our results suggest that the adenylyl cyclase signaling system in various types of endothelial cells can be differentially regulated at the levels of receptors, G-proteins, adenylyl cyclase, and phosphodiesterase.

Direct demonstration of no formation in vivo from organic nitrites and nitrates, and correlation to effects on blood pressure and to in vitro effects

Previous studies, utilizing nitric oxide synthase inhibitors and nitric oxide application, indicate that nitric oxide has the capacity to modulate contractile responses in pulmonary vessels. In the present study, in vitro effects of organic nitrates/nitrites were compared with their in vivo ability to generate nitric oxide and their effects on blood pressure. Glyceryl trinitrate, ethyl nitrite, isobutyl nitrate, isobutyl nitrite, isoamyl nitrite and butyl nitrite inhibited contractions in response to nerve stimulation in guinea pig pulmonary artery and vas deferens. Glyceryl trinitrate (also known as nitroglycerin) was the most potent and isobutyl nitrate the least potent substance with this action ( ic 50 4.5 ± 0.2 × 10 −10 and 1.1 ± 0.1 × 10 −5M, respectively). Contractile responses to noradrenaline were inhibited, whereas noradrenaline release was unaffected by organonitrates/nitrites, indicating a post-junctional inhibitory effect. When infused intravenously to anaesthetized rabbits glyceryl trinitrate, ethyl nitrite and isobutyl nitrate generated dose-dependent increments of nitric oxide in exhaled air and dose-dependent decrements in systemic blood pressure. Significant correlations were obtained between in vivo NO generation and effects on blood pressure, as well as between NO generation in vivo and the in vitro activity of the organic nitrites and organic nitrates. In conclusion, organic nitrites and organic nitrates can modulate adrenergic neuroeffector transmission in guinea pig pulmonary artery and vas deferens, and produce detectable concentrations of nitric oxide in exhaled air in vivo, in the rabbit. The observations give direct in vivo evidence that organic nitrites and nitrates generate NO, and strongly support them exerting their action via NO formation.

Thrombin receptor activation peptide induces pulmonary vasoconstriction

We investigated the involvement of the 14-residue thrombin receptor activating peptide SFLLRNPNDKYEPF (TRAP-14) in mediating the pulmonary vasoconstriction in response to alpha-thrombin. Isolated guinea pig lungs were uniformly perfused with Ringer-albumin solution at a constant flow of 28 ml/min. Addition of TRAP-14 or human alpha-thrombin to the perfusate caused dose-dependent increases of pulmonary arterial pressure within 1 min. TRAP-14 at 1 microM increased pulmonary arterial pressure to a similar extent as 10 nM alpha-thrombin (i.e., increase of 7.7 +/- 0.8 and 7.4 +/- 0.9 cmH2(0) from baseline, respectively). The increases in pulmonary venous resistance induced by TRAP-14 and alpha-thrombin were two- to fivefold greater than the increases in pulmonary arterial resistance, indicating that both agonists mediated pulmonary hypertension secondary to pulmonary venoconstriction. Stimulation of cultured guinea pig pulmonary artery smooth muscle cells with 100 microM TRAP-14 or 10 nM alpha-thrombin increased cytosolic Ca2+ concentration about five- to sevenfold over baseline. The increase in cytosolic Ca2+ concentration in smooth muscle cells was not observed with a subsequent challenge with either agonist, indicating desensitization. In the perfused lungs, an initial stimulation with alpha-thrombin or TRAP-14 desensitized the lungs to either agonist. The alpha-thrombin-desensitized lungs remained refractile to alpha-thrombin after 1 h of perfusion with fresh Ringer solution, whereas the TRAP-14-desensitized lungs recovered 79% of the vasoconstrictor response by 10 min and 93% of the response by 30 min.(ABSTRACT TRUNCATED AT 250 WORDS)

Endogenous vasodilators modulate pulmonary vascular anaphylaxis

We examined the role of endothelium-derived nitric oxide during antigen-induced contraction in pulmonary arteries isolated from actively sensitized guinea pigs. Ovalbumin (10(-2) mg/ml)-induced contraction was not sustained, and tension returned to baseline within 15 min. Pretreatment with methylene blue (10(-5) M) increased both the amplitude and the duration of the contractile response in these tissues. At 15 min, tension remained elevated and was > 70% of the peak amplitude. Removal of the endothelium with saponin (200 micrograms/ml) increased the magnitude of the contraction by > 125%; however, the duration of the response was unaffected. After pretreatment with saponin, methylene blue no longer increased the amplitude of antigen-induced contraction but its effect on the duration was unchanged. Pretreatment with nitro-L-arginine methyl ester significantly increased the magnitude of the contraction in each of the tissues. These results suggest that the response of guinea pig pulmonary arteries to antigen is modulated by two types of endogenous vasodilators, endothelium-derived nitric oxide that inhibits the initial phase of the response and an endothelium-independent relaxing factor that is guanosine 3',5'-cyclic monophosphate dependent and attenuates the duration of anaphylactic contraction.

A Comparative Study on the Sites of Nitric Oxide Release on Perivascular Nerve Stimulation in Different Arteries from Guinea Pig

This study investigates the sites of nitric oxide (NO) release on perivascular nerve stimulation (PNS) in organ bath experiments with isolated guinea pig pulmonary (PA) and iliac artery (IA). All the preparations were pretreated with indomethacin (10−5M), to exclude the effects of cyclo-oxygenase-generated eicosanoids. NG-nitro-L-arginine methyl ester (L-NAME, 10 −6 M) significantly augmented the PNS-induced adrenergic contraction and subsequently administered L-arginine (3 × 10 −4 M) reversed this augmentation in both arteries. In PA, endothelial denudation did not abolish but significantly attenuated this L-NAME-induced augmentation, while it abolished the augmentation in IA. In prostaglandin F2α -precontracted PA after blockade of both adrenergic and cholinergic effector responses, PNS induced tetrodotoxin (3 × 10−7M)-sensitive relaxation. This relaxation was attenuated by L-NAME (10−6 M) and reversed by L-arginine (3 × 10−4 M). But it was not abolished by endothelial denudation. On the other hand, PNS...

Bradykinin receptor types and B 2 subtypes

Bradykinin, desArg 9BK, some agonist analogues and several antagonists have been tested in isolated organs in order to identify bradykinin B 2 receptor subtypes. The initial pharmacological characterization was made in the rabbit jugular vein and the guinea pig ileum, two widely used B 2 preparations which have shown marked differences in their sensitivities to both agonists and antagonists. The study has then been extended to peripheral tissues (stomach, colon, urinary bladder) of four species (the rat, guinea pig, rabbit and man) and to isolated vessels (rabbit jugular vein, rabbit vena cava, guinea pig pulmonary artery, rat portal vein) in order to determine if pharmacologic receptor subtypes may be related to species. It has been shown that B 2 receptors in rat and guinea pig tissues belong to a similar pharmacological entity, a receptor which is different from that mediating the responses of rabbit and human tissues. Agonists order of potency ([Hyp 3]BK > BK > [Aib 7]BK) obtained in the rabbit jugular vein is different from that found in the guinea pig ileum (BK ≤ [Aib 7]BK > [Hyp 3]BK). Affinities of competitive antagonists (for instance DArg[Hyp 3,DPhe 7,Leu 8]BK) in rabbit tissues are higher than in guinea pig and rat tissues by at least 2 log units, while the non peptidic compound WIN 64338 is more active (also by two log units) in guinea pig than in human and rabbit tissues. The non competitive long-acting antagonist HOE 140 is very potent and equally active in the four species. Some antagonists (peptides without unnatural residues, peptides with unnatural residues, non peptides) have been shown to be specific for kinin receptors and selective for the B 2. Altogether, the present results a) confirm the existence of two B 2 receptor subtypes, b) suggest that receptor subtypes may be species dependent and c) indicate that the B 2 receptor subtype found in the rabbit is similar to that found in man.

Cyclic Strain and Forskolin Differentially Induce cAMP Production in Phenotypically Diverse Endothelial Cells

The effects of cyclic strain and forskolin on cAMP production were studied in 4 endothelial cell types. Basal cAMP levels varied among the various types. Cyclic strain induced an increase in cAMP levels in endothelial cells isolated from bovine aortae, pig pulmonary arteries and human adipose microvessels, but not in endothelial cells isolated from human umbilical veins. 100μM forskolin induced a differential increase in cAMP which ranged from 5-fold in pig pulmonary artery endothelial cells to 50-fold in human microvascular endothelial cells. Forskolin and cyclic strain, when applied to the cells concomitantly, had no additional effects on cAMP levels. However, when forskolin was applied 5niin after initiation of cyclic strain, a synergistic elevation of cAMP was observed in human umbilical vein endothelial cells. This action of forskolin was not mimicked by isoproterenol (10μM). We suggest that the cAMP signalling system of phenotypically diverse endothelial cells is differentially regulated by cyclic strain and forskolin and that these differences might contribute to endothelial cell phenotypic diversity.

Developmental changes in endothelium-dependent pulmonary vasodilatation in pigs.

1. We compared in vitro endothelium-dependent vasorelaxant responses to acetylcholine (ACh) and the endothelium-independent vasodilator response to sodium nitroprusside (SNP) in prostaglandin F2 alpha (PGF2 alpha)-precontracted muscular pulmonary arteries (PA) from pigs aged 5 min to 2 h (neonatal), 3-10 days, 3-8 weeks and adults. 2. In the pulmonary artery (PA) rings from neonatal animals, the vasodilator response to ACh was negligible. However, responses to ACh were present in all PA rings from older animals, being greatest at 3-10 days and then decreasing with age (P less than 0.001, ANOVA). ACh (30 microM) induced a 1 +/- 1%, 92 +/- 9%, 62 +/- 5% and 51 +/- 6% reduction of the PGF2 alpha-generated tension in neonatal, 3-10 days, 3-8 weeks and adult groups, respectively. 3. The relaxant response to SNP was present in the PA rings from all age groups and increased with age (P less than 0.001, ANOVA). SNP (1 microM)-induced relaxation was 55 +/- 9%, 73 +/- 7%, 97 +/- 5% and 93 +/- 6% in neonatal, 3-10 days, 3-8 week and adult groups, respectively. 4. Removal of the vascular endothelium abolished the relaxant response to ACh but had no effect on the response to SNP in any groups. 5. NG-monomethyl-L-arginine (30 microM), a nitric oxide synthesis inhibitor, inhibited the response to ACh but not to SNP. The lipoxygenase inhibitor, nordihydroguaiaretic acid, had no significant effect on responses to ACh or SNP in any group.6. These findings suggest that the nitric oxide pathway may not play a part in dilating the pig pulmonary arteries at birth, but may be important during the transitional period of establishing a stable post-natal pulmonary circulation. The increase in response to SNP with age parallels the increase in smooth muscle cell myofilaments to which it may be related.

Effects of nanomolar to submillimolar carteolol on noradrenaline release in the absence and presence of uptake1 and uptake2 blockers in guinea pig pulmonary arteries.

Effects of nanomolar to submillimolar carteolol, a non-selective beta-antagonist, on the evoked release at 1 Hz and the spontaneous release in the absence and presence of uptake1 and uptake2 blockers were studied in pulmonary arteries, isolated from guinea pigs, and then preloaded with [3H]noradrenaline. dl-Carteolol at 10(-8), 10(-7) and 10(-6) M applied at the increasing concentrations inhibited the evoked [3H]-release in untreated arteries and in desipramine and corticosterone-treated arteries. The spontaneous [3H]-release slightly but significantly increased or tended to increase in untreated arteries. dl-Carteolol at 10(-5) and 10(-4) M clearly and concentration-dependently increased the spontaneous [3H]-release in untreated and cocaine-treated arteries. This increase was markedly inhibited by further pretreatment with normetanephrine. The evoked [3H]-release was not altered by dl-carteolol at 10(-5) M, but increased at 10(-4) M. This increase was not modified by cocaine and by cocaine and normetanephrine. d-Carteolol at 10(-5) and 10(-4) M produced effects similar to those of dl-carteolol. Nanomolar to micromolar dl-carteolol inhibits the evoked [3H]-release, which supports our previous conclusion that this inhibition is due to blockade of tonically functioning presynaptic beta 2-adrenoceptors. Carteolol at the higher concentrations seems to become a substrate for an uptake2 mechanism and to produce an unknown sympathomimetic activity in guinea pig pulmonary arteries.

Contribution of Na+/H+ Exchange to pH Regulation in Pulmonary Artery Smooth Muscle Cells

In blood vessels in the systemic circulation, the plasmalemmal Na+/H+ exchanger has been implicated in a variety of cellular functions, including the regulation of intracellular pH (pHi) and cell volume, and the response to smooth muscle mitogens. The role of this transport system in pulmonary vascular smooth muscle has not been explored. The present study examined the characteristics of Na+/H+ exchange in cultured guinea pig pulmonary artery smooth muscle cells. These cells were subjected to an acid load, and the recovery from acid loading was monitored using the fluorescent pH-sensitive dye 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein (BCECF). In the absence of HCO3-, pHi recovery from acid loading was dependent on external Na+ and was inhibited by the Na+/H+ exchange inhibitor dimethylamiloride (DMA) (recovery rate was reduced from 54.4 +/- 5.5 to 12.8 +/- 2.0 mmol H+/liter.min). This exchanger was also active in the presence of HCO3-; DMA reduced resting pHi and slowed the rate of recovery from acid loading in HCO3- buffers. However, in the presence of HCO3-, other transport systems, presumably HCO3-/Cl- exchange, also contribute to the regulation of pHi. In HCO3- buffers, the rate of recovery from acid load averaged 40.8 +/- 1.8 mmol H+/liter.min. Addition of 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS), an inhibitor of HCO3-/Cl- exchange, slowed this recovery to 25.5 +/- 1.6 mmol H+/liter.min. A combination of DIDS and DMA further slowed the recovery to 19.7 +/- 1.5 mmol H+/liter.min. These findings indicate that the Na+/H+ exchanger plays a significant role in the regulation of pHi in pulmonary artery smooth muscle cells, even in HCO(3-)-containing buffers.

Modulation of neuroeffector transmission in the guinea pig pulmonary artery by endogenous nitric oxide

The influence of endogenous nitric oxide (NO) on neuroeffector transmission in segments of guinea pig pulmonary artery was analyzed by application of N G - monomethyl- l-argining ( l-NMMA). l-NMMA enhanced contractile responses to nerve stimulation and this enhancement was counteracted by l-arginine. The enhancement remained after removal of the endothelium. l-NMMA enhanced contractions to exogenous noradrenaline. After blockade of adrenergic transmission by phentolamine, l-NMMA enhanced contractions induced by nonadrenergic-noncholinergic (NANC) neurotransmission. Stimulation-induced release of [ 3H]noradrenaline was unchanged by l-NMMA. The results suggest that endogenous NO exerts a postjunctional inhibition on adrenergic neurotransmission in the guinea pig pulmonary artery. A concomitant pre- and/or postjunctional inhibition of NANC transmission is implicated. The neuromodulation by NO does not require an intact endothelium.

A comparative biochemical, pharmacological and immunological study of Clostridium novyi α-toxin, C. difficile toxin B and C. sordellii lethal toxin

The three clostridial cytotoxins, i.e. α-toxin of C. novyi (Toxα-nov), toxin B of C. difficile (ToxB-dif) and lethal toxin of C. sordellii (LT-sor) consist of single peptide chains of about 200,000 (Toxα-nov), 250,000 (LT-sor) and 275,000 (ToxB-dif) mol. wts. ToxB-dif and LT-sor but not Toxα-nov cross-reacted with rabbit polyclonal antibodies. Toxicity upon i.v. injection in mice was similar ( ld 50, 100 hr, 50–200 ng/kg) and was characterized by a slowly developing fluid loss into the interstitial space. When injected into the rat paw the toxins caused a delayed local edema lasting for days. In vitro the three toxins provoked a persistent retraction of endothelial cells cultured from pig pulmonary artery. ToxB-dif and Toxα-nov triggered the accumulation of F-actin in the perinuclear region at the expense of the tight peripheral bands whereas LT-sor led to a random loss of microfilament structure. The toxins inhibited uridine incorporation into endothelial or chicken embryonic cells whereas T 84 cells responded by an about 10-fold increase of uridine incorporation. Neither toxin ADP-ribosylated actin. The similarities between the three cytotoxins warrant their arrangement into a common group which perturbs the microfilament system.