Atherosclerosis and nitric oxide production.
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Animals↗
Biomedical subjects
R W Caldwell
Publications and source records attributed to R W Caldwell.
Role of L-arginine in the vascular actions and development of tolerance to nitroglycerin.
The goal of this work was to test the role of nitric oxide synthase (NOS) and its substrate L-arginine in development of tolerance to nitroglycerin's (GTN) vasodilator actions. GTN's effects on NOS activity and NO formation were tested in cultured bovine aortic endothelial cells (BAECs). The arginine to citrulline conversion assay showed that GTN stimulated NOS basal activity in BAECs by approximately 40%, comparable with acetylcholine (ACh)-treated controls. Both effects were blocked by L-NMMA. Photometric assays showed that both GTN and ACh-stimulated NO formation. Both effects were potentiated by L-arginine and inhibited by L-NAME. L-NAME inhibited ACh responses approximately 80% compared with approximately 40% for GTN responses. The aortic ring assay showed that 2 h pretreatment with GTN caused substantial tolerance to GTN's vasodilating effects as evidenced by a 38 fold rightward shift of the concentration-relaxation curve. In contrast to D-arginine, addition of L-arginine substantially inhibited this effect, reducing the rightward shift to 4.4 fold of control values. GTN tolerance was associated with a 40% reduction in L-arginine tissue levels. GTN had a biphasic effect on BAEC uptake of L-arginine, stimulating uptake at 5 and 15 min, and suppressing uptake after 1 and 4 h In summary, acute GTN treatment stimulates endothelial NOS activity in producing NO and increases cellular uptake of L-arginine. Prolonged GTN exposure reduces GTN's vasodilator actions, decreases L-arginine tissue levels and depresses BAECs uptake of L-arginine. Supplementation of L-arginine reduces development of GTN tolerance. These data indicate that GTN tolerance depends in part on activation of the NOS pathway.
Animals↗
Vascular and endothelial actions of inhibitors of substance P amidation.
Formation of mature active neuropeptides such as substance P (SP) from their glycine extended precursors entails alpha-amidation of peptide precursors by the sequential enzymatic action of peptidylglycine alpha-monooxygenase (PAM) and peptidylamidoglycolate lyase (PGL). We reported that these two enzymes that can produce mature active neuropeptides are present in cultured bovine aortic endothelial cells (BAECs). We hypothesize that alpha-amidation of peptides occurs in endothelial cells and that these peptides are critically involved in the overall regulation of cardiovascular function. In this study, this hypothesis was tested using specific amidation inhibitors to determine their effects on the actions of SP and its glycine-extended precursor (SP-Gly). We have found that SP and SP-Gly are equipotent in stimulating nitric oxide (NO) release by BAECs. At 10(-5) M, the specific inhibitors of PAM (4-phenyl-3-butenoic acid; PBA) and PGL (5-acetamido-2,4-diketo-6-phenyl-hexanoic acid and its methyl ester) reduced NO basal release by 40, 34, and 45%, respectively. They also reduced the production of NO induced by SP-Gly by 63, 68, and 69%, respectively, but had no effect on NO production in response to either SP or acetylcholine. SP and SP-Gly also were equipotent in relaxing rat aortic segments. The vasorelaxation to SP-Gly was endothelium dependent and inhibited by the NOS antagonist L-nitroarginine methyl ester (L-NAME), but it was not affected by inhibition of prostaglandin synthesis. Inhibitors of both PAM and PGL significantly reduced the vasorelaxing actions of SP-Gly, whereas responses to SP were not affected. A cumulative infusion of PBA into the femoral artery of rabbits, at final concentrations of 2.4, 24, and 240 microM for 20 min each, increased the vascular resistance (VR), indicating the tonic production of vasodilating amidated peptide(s). This effect was maximum at 60 min after infusion (20.5 +/- 4.7 vs. 8.2 +/- 0.7 mm Hg/ml/min; p < 0.05). These results suggest that endothelial cells can produce mature SP from its SP-Gly precursor and that a product of peptide alpha-amidation tonically stimulates endothelial cell NO release to control vascular tone.
Amidine-Lyases↗
Effects of NO donors and synthase agonists on endothelial cell uptake of L-Arg and superoxide production.
It is commonly believed that the activity of NO synthase (NOS) solely controls NO production from its substrates, L-Arg and O(2). The Michaelis-Menten constant (K(m)) of NOS for L-Arg is in the micromolar range; cellular levels of L-Arg are much higher. However, evidence strongly suggests that cellular supply of L-Arg may become limiting and lead to reduced NO and increased superoxide anion (O(-)(2)*) formation, promoting cardiovascular dysfunction. Uptake of L-Arg into cells occurs primarily (approximately 85%) through the actions of a Na(+)-independent, carrier-mediated transporter (system y(+)). We have examined the effects of NOS agonists (substance P, bradykinin, and ACh) and NO donors (S-nitroso-N-acetyl-penicillamine and dipropylenetriamine NONOate) on transport of L-Arg into bovine aortic endothelial cells (BAEC). Our results demonstrate that NOS agonists increase y(+) transporter activity. A rapidly acting NO donor initially increases L-Arg uptake; however, after longer exposure, L-Arg uptake is suppressed. Exposure of BAEC without L-Arg to substance P and a Ca(2+) ionophore (A-23187) increased O(-)(2)* formation, which was blocked with concurrent presence of L-Arg or the NOS antagonist N(omega)-nitro-L-arginine methyl ester. We conclude that factors including NO itself control y(+) transport function and the production of NO and O(-)(2)*.
Animals↗
In situ localization of cholesterol in skeletal muscle by use of a monoclonal antibody.
A common perception is that cholesterol, the major structural lipid found in mammalian membranes, is localized nearly exclusively to the plasma membrane of living cells and that it is found in much smaller quantities in internal membranes. This perception is based almost exclusively on cell fractionation studies, in which density gradient centrifugation is used for purification of discrete subcellular membrane fractions. Here we describe a monoclonal antibody, MAb 2C5-6, previously reported to detect purified cholesterol in synthetic membranes (Swartz GM Jr, Gentry MK, Amende LM, Blanchette-Mackie EJ, and Alving CR. Proc Natl Acad Sci USA 85: 1902-1906, 1988), that is capable of detecting cholesterol in situ in the membranes of skeletal muscle sections. Localization of cholesterol, the dihydropyridine receptor of the T tubule, and the Ca(2+)-ATPase of the sarcoplasmic reticulum (SERCA2) by means of double and triple immunostaining protocols clearly demonstrates that cholesterol is primarily localized to the sarcoplasmic reticulum membranes of skeletal muscle rather than the sarcolemmal or T tubule membranes. The availability of this reagent and its ability to spatially localize cholesterol in situ may provide a greater understanding of the relationship between membrane cholesterol content and transmembrane signaling in skeletal muscle.
Animals↗
Pravastatin sodium activates endothelial nitric oxide synthase independent of its cholesterol-lowering actions.
OBJECTIVES: We tested the hypothesis that pravastatin (PRA) activates endothelial nitric oxide synthase (eNOS). BACKGROUND: Pravastatin has been found to have clinical benefits beyond those predicted by its actions in reducing plasma low density lipoprotein cholesterol (LDL). Both PRA and simvastatin (SIM) are equally effective in reducing LDL, but only PRA reduces platelet aggregation and is an effective vasodilator. Nitric oxide (NO) also inhibits platelet aggregation and vasodilates. METHODS: We determined PRA and SIM effects on vasorelaxation in aortic rings and NO production by cultured bovine aortic endothelial cells. Nitric oxide was measured by using a NO electrode and by an assay for conversion of hemoglobin to methemoglobin. Specificity of NOS activation was tested by using the NOS inhibitor nitro-L-arginine methyl ester (L-NAME, 1 mmol/liter) in the presence or absence of excess L-arginine (L-ARG, 1 mmol/liter). RESULTS: Endothelium-dependent vasorelaxation was maximal with acetylocholine (ACH, 100%), followed by PRA (62.8%) and then SIM (37.1%). Direct measurement of NO confirmed that vasorelaxation is due to NO release and showed that PRA and ACH had similar dose-dependent effects on NO production, while SIM was only 25% to 30% as effective. Methemoglobin assay confirmed these results and demonstrated their specificity for NOS activity. The L-NAME blunted the responses to 45% of initial values. Excess L-ARG reversed this effect and potentiated NO production to 133% of initial levels. CONCLUSIONS: Both PRA and SIM activate eNOS, but SIM is much less effective. Clinical benefits with PRA not explained by LDL reductions may be the result of an independent action of PRA on eNOS activation.
Animals↗
Certain norditerpenoid alkaloids and their cardiovascular action.
Thirteen new derivatives of norditerpenoid alkaloids, namely, 8-deacetyl-8-p-aminobenzoyldelphinine (1), 8-deacetyl-8-anthranoyldelphinine (2), 8-deacetyl-8-(4-hydroxy-3-methoxycinnamoyl)delphinine (3), 16-demethoxy-15,16-didehydro-8-p-anisoyl-14-benzoyldelpho nine (4), 6-acetylheteratisine N-oxide (6), 3,8-diacetylfalconerine (7), 8-stearoylfalconerine (8), 8-linolenylfalconerine (9), 13-acetylpyrodelphinine (11), 13-acetyldelphinine N-oxide (13), N-deacetyl-8,9-diacetyllappaconitine (14), 8, 9-(methylenedioxy)lappaconine (15), and 16-epipyroaconitine N-oxide (17), were prepared, and their structures were established by analysis of spectroscopic data (1D and 2D NMR, HRFABMS). The preliminary in vivo cardiovascular action (hypotensive, bradycardic, and ventricular arrhythmias) of these new compounds was tested in male Sprague-Dawley rats. The results are reported herein.
Alkaloids↗
Zn2+ inhibits nitric oxide formation in response to lipopolysaccharides: implication in its anti-inflammatory activity.
There is compelling evidence to indicate an anti-inflammatory action of Zn2+. Most inflammatory diseases are associated with an increase of the inducible form of nitric oxide (NO) synthase. Additionally, inflammatory mediators such as histamine or bradykinin stimulate the constitutive NO synthase. Thus, the present study was undertaken to investigate whether Zn2+ inhibits production of inducible NO synthase and/or constitutive NO synthase activity to produce NO. Lipopolysaccharide, 5 mg/kg i.v., administered to Zn2+-deficient (ZD) rats, rats supplemented with Zn2+ sulfate (ZG), 10 mg/kg s.c., or controls resulted in a significant reduction of their serum Zn2+. The levels of N(G)-nitro-L-arginine methylester (L-NAME)-sensitive cyclic GMP (cGMP) in aortas isolated from ZD or ZG were significantly lower than those obtained from control animals. Zinc (100-150 microM) produced a dose-dependent inhibition of lipopolysaccharide or interleukin-1beta-induced NO formation in isolated rat aortic smooth muscle cells. Compared to cyclohexamide or actinomycin-D, the time course of inhibition of NO formation by 150 microM Zn2+ did not suggest an effect of Zn2+ on inducible NO synthase protein synthesis. Moreover, Zn2+ (150 microM) significantly reduced the rate of conversion of [3H]arginine to [3H]citrulline in lung homogenates from lipopolysaccharide-treated rats. Incubation of rat aortic smooth muscle cells and bovine pulmonary artery endothelial cell co-cultures with Zn2+ (150 microM) caused a significant reduction in basal and bradykinin- or A-23187-induced formation of cGMP. Thus, our results indicate that Zn2+ is capable of inhibiting lipopolysaccharide- or interleukin-1beta-induced NO formation as well as NO formation by constitutive NO synthase basally or in response to bradykinin or A-23187, and may explain the reported anti-inflammatory activity of Zn2+.
Animals↗
Amidative peptide processing and vascular function.
Substance P (SP), an amidated peptide present in many sensory nerves, is known to affect cardiovascular function, and exogenously supplied SP has been shown to activate nitric oxide synthase (NOS) in endothelial cells. We now report that SP-Gly, the glycine-extended biosynthetic precursor of SP (which is enzymatically processed to the mature amidated SP), causes relaxation of rat aortic strips with an efficacy and potency comparable to that of SP itself. Pretreatment of the aortic strips with 4-phenyl-3-butenoic acid (PBA), an irreversible amidating enzyme inactivator, results in marked inhibition of the vasodilation activity induced by SP-Gly but not of that induced by SP itself. Isolated endothelial cell basal NOS activity is also decreased by pretreatment with PBA, with no evidence of cell death or direct action of PBA on NOS activity. Both bifunctional and monofunctional forms of amidating enzymes are present in endothelial cells, as evidenced by affinity chromatography and Western blot analysis. These results provide evidence for a link between amidative peptide processing, NOS activation in endothelial cells, and vasodilation and suggest that a product of amidative processing provides intrinsic basal activation of NOS in endothelial cells.
Amino Acid Sequence↗
A novel catecholamine, arbutamine, for a pharmacological cardiac stress agent.
Arbutamine, developed for use as a cardiac stress agent, was compared with isoproterenol and dobutamine in anesthetized dogs for cardiovascular actions prior to and after beta-adrenergic blockade with propranolol. The efficacy and safety of arbutamine were also evaluated in a canine model of myocardial ischemia obtained by partially occluding the left anterior descending coronary artery. Comparison of hemodynamic variables in normal dogs showed that arbutamine was approximately equipotent to isoproterenol in increasing heart rate and cardiac contractility, and in decreasing total peripheral vascular resistance and mean arterial blood pressure. Arbutamine was 210 times more potent than dobutamine in increasing cardiac contractility by 70%; however, at this dose dobutamine exhibited a negative chronotropic response. Beta-adrenergic blockade with propranolol shifted the agonist's dose-response curves for heart rate and contractility to the right; however, low doses of dobutamine exhibited a negative chronotropic effect and increased the total peripheral vascular resistance. In dogs subjected to partial left anterior descending coronary artery occlusion, arbutamine produced significant ST-segment deflections, beginning at a dose of 0.1 nmol/kg/min. Impairment of segment shortening, reflecting cardiac wall motion abnormality, was evident at a dose of 0.3 nmol/kg/min. Isoproterenol did not cause significant changes in these parameters. These results show that arbutamine is capable of producing graded increments in cardiac contractility and rate before and after beta-adrenergic blockage in normal dogs. In dogs subjected to coronary artery occlusion, it is capable of provoking myocardial ischemia at dose levels devoid of toxicity.
Animals↗
Characterization of the adrenergic activity of arbutamine, a novel agent for pharmacological stress testing.
In this study, we characterized the interactions of arbutamine, a novel catecholamine developed for use as a cardiac stress testing agent, with different adrenergic receptor subtypes in vitro. These effects were compared with those of isoproterenol. In the electrically stimulated left atria of rats, arbutamine increased contractile force. The pD2 values (- log of the dose that produces 50% of the maximal responses) for arbutamine and isoproterenol were 8.45 +/- 0.15 and 8.55 +/- 0.02, respectively. Metoprolol shifted the concentration-effect curves for both isoproterenol and arbutamine to the right with a pA2 value (- log of the dose of the antagonist that reduces the maximal responses of an agonist to 50%) of 7.22-7.5. Both arbutamine and isoproterenol increased the rate of spontaneously beating rat right atria with pD2 values of 9.0 +/- 0.19 and 8.82 +/- 0.18, respectively. The affinity constants (KA) of arbutamine and isoproterenol for cardiac beta1-adrenergic receptors, as determined by competition binding assays, were found to be 7.32 and 6.04, respectively. In guinea pig trachea, arbutamine and isoproterenol produced a concentration-dependent relaxation that was blocked by propranolol. Their pD2 values were 7.9 +/- 0.1 and 8.2 +/- 0.1, respectively. Arbutamine contracted isolated rat aortic rings with a maximal increase of 38.1 +/- 6.7% that of 10 microM of norepinephrine. In rat white adipocytes, arbutamine, isoproterenol, and BRL-37344 stimulated glycerol release, with the order of potency being BRL-37344 > arbutamine > isoproterenol. In hamster brown adipocytes, the order was arbutamine > isoproterenol > BRL-37344. Moreover, arbutamine stimulated beta3-adrenergic receptors in guinea pig ileum. In conclusion, arbutamine is a novel catecholamine with similar potency and efficacy to that of isoproterenol. It stimulates cardiac beta1-, tracheal beta2-, and adiopocyte beta3-adrenergic receptors. Arbutamine does not stimulate alpha-adrenergic receptors at concentrations that were high enough to maximally activate the beta-adrenergic receptors.
Adipocytes↗
The role of substance P in myocardial dysfunction during ischemia and reperfusion.
Impairment of myocardial contraction ("myocardial stunning") occurs during reperfusion after short ischemic periods. Substance P (SP) is widely distributed in heart and can be released by various stimuli including myocardial hypoxia. Our previous study shows SP has a negative inotropic effect in guinea pig heart. The objective of this study was to investigate whether SP contributes to the myocardial stunning after brief global ischemia. Guinea pig hearts in a Langendorff preparation were subjected to 15 min of global ischemia followed by 60 min reperfusion. Experiments were performed without and with pretreatment with neurokinin-1 (NK1) receptor antagonists, spantide (10(-6)M) or CP-99,994-01 (10(-6)M) in order to study the role of SP. Experiments were also performed in hearts which were perfused with atropine, phentolamine, and nadolol (10(-6)M each) to examine the role of neurotransmitters and autonomic receptors. A group of hearts obtained from capsaicin-pretreated guinea pigs was also investigated. Left ventricular developed pressure (LVDP), left ventricular end-diastolic pressure (LVEDP), heart rate, and perfusion pressure were monitored. At the end of reperfusion, the LVDP of control hearts recovered to only 55 +/- 6% (+/- SEM) of preischemic baseline and the LVEDP increased significantly (P > 0.05). With pretreatment with spantide or CP-99,994-01, LVDP recovered to 88 +/- 2% or 78 +/- 2% of the preischemic baseline, respectively. The LVEDP of these hearts was not different from preischemic baseline and much smaller than in control hearts. There were no differences in heart rate and perfusion pressure compared to baseline among all groups. Similar results were obtained in hearts perfused with autonomic blockers. However, recoveries of LVDP and LVEDP were faster in hearts perfused with autonomic blockers during the first 10 min of reperfusion. Pretreatment with capsaicin also significantly improved recovery of LVDP and LVEDP. In conclusion, substance P is involved in postischemic myocardial dysfunction and neurokinin-1 receptors mediate this action. The NK1 receptor antagonists may be useful in prevention of "myocardial stunning".
Analgesics↗
Downregulation of nitrovasodilator-induced cyclic GMP accumulation in cells exposed to endotoxin or interleukin-1 beta.
1. Induction of nitric oxide synthase (iNOS) results in overproduction of nitric oxide (NO), which may be a principal cause of the massive vasodilatation and hypotension observed in septic shock. Since NO-induced vasorelaxation is mediated via the soluble isoform of guanylate cyclase (sGC), the regulation of sGC activity during shock is of obvious importance, but yet poorly understood. The aim of the present study was to investigate the activation of sGC by sodium nitroprusside (SNP) before and after exposure of rat aortic smooth muscle cells to endotoxin (LPS) or interleukin-1 beta (IL-1 beta). 2. Exposure of rat aortic smooth muscle cells to SNP (10 microM) elicited up to 200 fold increases in cyclic GMP. This effect was attenuated by 30-70% in IL-1 beta- or LPS-pretreated cells, in a pretreatment time-and IL-1 beta- or LPS-concentration-dependent manner. When, however, cells were exposed to IL-1 beta or LPS and then stimulated with the particulate guanylate cyclase activator, atriopeptin II, no reduction in cyclic GMP accumulation was observed. 3. Pretreatment of rats with LPS (5 mg kg-1, i.v.) for 6 h led to a decrease in aortic ring SNP-induced cyclic GMP accumulation. 4. The IL-1 beta-induced reduction in SNP-stimulated cyclic GMP accumulation in cultured cells was dependent on NO production, as arginine depletion abolished the downregulation of cyclic GMP accumulation in response to SNP. 5. Reverse-transcriptase-polymerase chain reaction analysis revealed that the ratio of steady state mRNA for the alpha, subunit of sGC to glyceraldehyde phosphate dehydrogenase was decreased in LPS- or IL-1 beta-treated cells, as compared to vehicle-treated cells. 6. Protein levels of the alpha 1 sGC subunit remained unaltered upon exposure to LPS or IL-1 beta, suggesting that the early decreased cyclic GMP accumulation in IL-1 beta- or LPS-pretreated cells was probably due to reduced sGC activation. Thus, the observed decreased responsiveness of sGC to NO stimulation following cytokine or LPS challenge may represent an important homeostatic mechanism to offset the extensive vasodilatation seen in sepsis.
Animals↗
Effect of diterpenoid alkaloids on cardiac sympathetic efferent and vagal afferent nerve activity.
The diterpenoid alkaloid, lappaconitine, at a dose of 150 micrograms/kg (i.v.) increased cardiac vagal afferent nerve activity (16.2%) and reduced cardiac sympathetic efferent nerve activity (12.5%). A polar analog, N-deacetyllappaconitine, at this same dose, increased cardiac vagal afferent nerve activity (40%) and reduced cardiac sympathetic efferent nerve activity (23.5%). Both of these agents also reduced arterial blood pressure and heart rate. A larger dose of lappaconitine (300 micrograms/kg i.v.) produced the same changes in nerve activities and cardiac function as the lower dose. Two other structurally related agents, lycoctonine and aconine, failed to alter these variables in doses up to 300 micrograms/kg. These data suggest that certain diterpenoid alkaloids activate autonomic reflex receptors, including cardiac reflex receptors. The polar agent, N-deacetyllappaconitine, appears to be more effective on cardiac reflex receptors than the non-polar agent, lappaconitine. Such agents may be useful in the treatment of hypertension.
Aconitine↗
Local cardiac effects of substance P: roles of acetylcholine and noradrenaline.
1. The local cardiac actions of substance P were examined in isolated perfused hearts and atria of the guinea-pig. 2. In both hearts and right atria, substance P caused negative inotropic and chronotropic effects. 3. Atropine (10(-6) M) or depletion of acetylcholine, by electrical stimulation and hemicholinium-3 perfusion, significantly attenuated the negative inotropic and chronotropic effects of substance P. alpha- and beta-adrenoceptor blockade by nadolol and phentolamine (10(-6) M each) did not prevent the negative inotropic and chronotropic effects of substance P. This indicates that cholinergic neurones, but not adrenergic neurones, partially mediate the effects of substance P. 4. There was no significant difference in the effects of substance P observed between groups with acetylcholine depletion and with cholinoceptor blockade. This suggests that substance P elicits its effects mainly through release of acetylcholine. 5. These results indicate that substance P has negative inotropic and chronotropic effects in guinea-pig hearts and right atria mediated partly by release of acetylcholine. Substance P also appears to have direct effects on cardiac tissue.
Acetylcholine↗
Contraction-induced cell wounding and release of fibroblast growth factor in heart.
The heart hypertrophies in response to certain forms of increased mechanical load, but it is not understood how, at the molecular level, the mechanical stimulus of increased load is transduced into a cell growth response. One possibility is that mechanical stress provokes the release of myocyte-derived autocrine growth factors. Two such candidate growth factors, acidic and basic fibroblast growth factor (aFGF and bFGF, respectively), are released via mechanically induced disruptions of the cell plasma membrane. In the present study, we demonstrate that transient, survivable disruption (wounding) of the cardiac myocyte plasma membrane is a constitutive event in vivo. Frozen sections of normal rat heart were immunostained to reveal the distribution of the wound event marker, serum albumin. Quantitative image analysis of these sections indicated that an average of 25% of the myocytes contained cytosolic serum albumin; ie, this proportion had suffered a plasma membrane wound. Wounding frequency increased approximately threefold after beta-adrenergic stimulation of heart rate and force of contraction. Heparin-Sepharose chromatography, enzyme-linked immunosorbent assay, growth assay coupled with antibody neutralization, and two-dimensional SDS-PAGE followed by immunoblotting were used to demonstrate that both aFGF and bFGF were released from an ex vivo beating rat heart. Importantly, beta-adrenergic stimulation of heart rate and force of contraction increased FGF release. Cell wounding is a fundamental but previously unrecognized aspect of the biology of the cardiac myocyte. We propose that contraction-induced cardiac myocyte wounding releases aFGF and bFGF, which then may act as autocrine growth-promoting stimuli.
Animals↗
HP-228, a novel synthetic peptide, inhibits the induction of nitric oxide synthase in vivo but not in vitro.
alpha-Melanocyte stimulating hormone has been shown to prevent endotoxin shock. A heptapeptide analog (HP-228) has recently been synthesized and shown to be an even more potent protective agent. Because the hypotensive and toxic actions of lipopolysaccharide (LPS) appear to involve the induction of type II nitric oxide synthase (iNOS), we have examined the actions of HP-228 on nitric oxide production using an endotoxemia model in conscious rats given E. coli LPS (5 mg/kg i.v.) and monitored for 6 h. A group of rats received HP-228 (30 micrograms/kg) 30 min before LPS. Using nitro L-arginine methyl ester-sensitive cGMP production as an estimate of nitric oxide synthase activity in aortic segments, ex vivo, we determined that LPS increases iNOS activity and that HP-228 pretreatment markedly reduces this response. Additionally, the rate of conversion of 3[H]-arginine to 3[H]-citrulline was significantly reduced in lung homogenates from HP-228-treated rats. HP-228 did not alter the activity of the constitutive nitric oxide synthase in aortic rings or in cerebella. In isolated rat aortic smooth muscle cells, LPS or interleukin-1 beta caused prominent rises in nitric oxide generated by iNOS. HP-228 did not antagonize the effect of these inducing agents. However, in these cells, plasma obtained from rats 1 h after administration of HP-228 prevented the induction of iNOS by both LPS and interleukin-1 beta. In conclusion, HP-228 prevents the in vivo induction of nitric oxide synthase by LPS.(ABSTRACT TRUNCATED AT 250 WORDS)
Animals↗
Effect of halothane on in vivo and in vitro cardiotoxicity of an aminocardenolide.
Halothane opposes cardiotoxicity of neutral-sugar digitalis compounds in intact animals, presumably by depressing a sympathetic component of arrhythmogenesis. However, halothane also produces a dose-related reduction in arrhythmogenicity of ouabain in isolated canine Purkinje fibers, suggesting that the anesthetic may oppose direct mechanisms of cardiotoxicity as well. The present study examined in vivo and in vitro the effect of halothane on the arrhythmogenicity of ASI-222 (3-beta-O[4-amino-4-6-dideoxy-beta-D-galactopyranosyl] digitoxigen in HCl), a highly polar aminocardenolide with no sympathetic component to cardiotoxicity. For in vivo studies, ASI-222 was infused at a rate of 1 microgram/kg/min until appearance of third-degree atrioventricular (AV) block or sustained ventricular arrhythmias in 5 conscious (control) and 6 halothane-anesthetized (1.4% end-tidal) dogs. For in vitro studies, standard microelectrode techniques were used to measure action potentials (AP) in seven excised canine Purkinje fibers superfused with oxygenated Krebs-Henseleit buffer. AP were recorded during control superfusion, after induction of toxicity with 10(-7) M ASI-222, and during exposure to 0.5, 1.0, and 2.0% halothane. Purkinje fibers were paced at 500-ms cycle lengths (CL) for 20 beats, and the amplitude of delayed afterdepolarizations (DAD) were recorded. Pacing at 250 ms CL was used to trigger ectopy. In vivo studies showed no difference in the cardiotoxic dose of ASI-222 between control dogs and those anesthetized with 1.4% halothane. However, in 4 of 6 anesthetized dogs, acutely increasing the inspired halothane concentration suppressed arrhythmias once end-tidal concentration were >2.2%.(ABSTRACT TRUNCATED AT 250 WORDS)
Anesthesia↗