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Biomedical subjects

L J Ignarro

Publications and source records attributed to L J Ignarro.

At least 19 recordsLinked to original sources
PubMed1992

Basal release of nitric oxide from aortic rings is greater in female rabbits than in male rabbits: implications for atherosclerosis.

Estradiol is known to exert a protective effect against the development of atherosclerosis, but the mechanism of this hormonal action is unknown. One of the early events in the development of atherosclerosis is the adhesion of macrophages to endothelial cells, and nitric oxide (NO) inhibits this process. We show that basal release of NO is greater with endothelium-intact aortic rings from female rabbits than those from males. Oophorectomy diminishes both circulating estradiol concentration and basal release of NO to levels seen in male rabbits. These data establish that basal NO release from endothelium-intact aortic rings depends on circulating estradiol concentration and offer an explanation for the protective effect of estradiol against the development of atherosclerosis.

Acetylcholine↗
PubMed1992

Constitutive nitric oxide synthase from cerebellum is reversibly inhibited by nitric oxide formed from L-arginine.

The objective of this study was to determine whether constitutive nitric oxide (NO) synthase from rat cerebellum could be regulated by the two products of the reaction, NO and L-citrulline, utilizing L-arginine as substrate. NO synthase activity was determined by monitoring the formation of 3H-citrulline from 3H-L-arginine in the presence of added cofactors. The rate of citrulline formation in enzyme reaction mixtures was non-linear. Addition of superoxide dismutase (SOD; 100 units) inhibited NO synthase activity and made the rate of product formation more non-linear, whereas addition of oxyhemoglobin (HbO2; 30 microM) increased NO synthase activity, made the rate of product formation linear and also abolished the effect of SOD. Added NO (10 microM) inhibited NO synthase activity and this inhibition was potentiated by SOD and abolished by HbO2. Added L-citrulline (1 mM) did not alter NO synthase activity. The two NO donors, S-nitroso-N-acetylpenicillamine (200 microM) and N-methyl-N'-nitro-N-nitrosoguanidine (200 microM) mimicked the inhibitory effect of NO and inhibition of NO synthase activity by NO was reversible. These observations indicate clearly that NO formed during the NO synthase reaction or added to the enzyme reaction mixture causes a reversible inhibition of NO synthase activity. Thus, NO may function as a negative feedback modulator of its own synthesis.

Amino Acid Oxidoreductases↗
PubMed1992

Biosynthesis of nitric oxide and citrulline from L-arginine by constitutive nitric oxide synthase present in rabbit corpus cavernosum.

The objective of this study was to determine whether a constitutive isoform of nitric oxide (NO) synthase is present in rabbit corpus cavernosum that could account for the involvement of the L-arginine-NO pathway in neurogenically-elicited relaxation of the corpus cavernosum and, therefore, penile erection. Citrulline was determined by monitoring the formation of 3H-citrulline from 3H-L-arginine. NO was determined by monitoring the formation of total NO(x) (NO+nitrite [NO2-]+nitrate [NO3-]) by chemiluminescence after reduction of NO(x) to NO by acidic vanadium (III). Equimolar quantities of NO plus citrulline were generated from L-arginine and the formation of both products was time-dependent at 37 degrees C. NO synthase activity was distributed almost entirely to the cytosolic fraction. Enzymatic activity was completely dependent on NADPH, calmodulin, and calcium. Addition of tetrahydrobiopterin increased NO synthase activity by about 30 percent. The NO synthase inhibitor NG-nitro-L-arginine, abolished enzymatic activity. The Km for L-arginine was 17 microM and the Vmax of the reaction was 18 pmol/min/mg protein. These observations indicate that a cytosolic, constitutive isoform of NO synthase, like that found in brain neuronal tissue, is present in rabbit corpus cavernosum.

Amino Acid Oxidoreductases↗
PubMed1992

Nitric oxide synthase from cerebellum catalyzes the formation of equimolar quantities of nitric oxide and citrulline from L-arginine.

This study examined whether constitutive nitric oxide (NO) synthase from rat cerebellum catalyzes the formation of equimolar amounts of NO plus citrulline from L-arginine under various conditions. Citrulline was determined by monitoring the formation of 3H-citrulline from 3H-L-arginine. NO was determined by monitoring the formation of total NOx (NO+nitrite [NO2-] + nitrate [NO3-]) by chemiluminescence after reduction of NOx to NO by acidic vanadium (III). Equal quantities of NO plus citrulline were generated from L-arginine and the formation of both products was linear for about 20 min at 37 degrees C provided L-arginine was present in excess to maintain a zero order reaction rate. Deletion of NADPH, addition of the calmodulin antagonist calmidazolium, or addition of NO synthase inhibitors (NG-methyl-L-arginine, NG-amino-L-arginine) abolished or markedly inhibited the formation of both NO and citrulline. The Km for L-arginine (14 microM; 18 microM) and the Vmax of the reaction (0.74 nmol/min/mg protein; 0.67 nmol/min/mg protein) were the same whether NO or citrulline formation, respectively, was monitored. These observations indicate clearly that NO and citrulline are formed in equimolar quantities from L-arginine by the constitutive isoform of NO synthase from rat cerebellum.

Amino Acid Oxidoreductases↗
PubMed1992

Nitric oxide as a mediator of relaxation of the corpus cavernosum in response to nonadrenergic, noncholinergic neurotransmission.

BACKGROUND: Nitric oxide has been identified as an endothelium-derived relaxing factor in blood vessels. We tried to determine whether it is involved in the relaxation of the corpus cavernosum that allows penile erection. The relaxation of this smooth muscle is known to occur in response to stimulation by nonadrenergic, noncholinergic neurons. METHODS: We studied strips of corpus cavernosum tissue obtained from 21 men in whom penile prostheses were inserted because of impotence. The mounted smooth-muscle specimens were pretreated with guanethidine and atropine and submaximally contracted with phenylephrine. We then studied the smooth-muscle relaxant responses to stimulation by an electrical field and to nitric oxide. RESULTS: Electrical-field stimulation caused a marked, transient, frequency-dependent relaxation of the corpus cavernosum that was inhibited in the presence of N-nitro-L-arginine and N-amino-L-arginine, which selectively inhibit the biosynthesis of nitric oxide from L-arginine. The addition of excess L-arginine, but not D-arginine, largely reversed these inhibitory effects. The specific liberation of nitric oxide (by S-nitroso-N-acetylpenicillamine) caused rapid, complete, and concentration-dependent relaxation of the corpus cavernosum. The relaxation caused by either electrical stimulation or nitric oxide was enhanced by a selective inhibitor of cyclic guanosine monophosphate (GMP) phosphodiesterase (M&B 22,948). Relaxation was inhibited by methylene blue, which inhibits cyclic GMP synthesis. CONCLUSIONS: Our findings support the hypothesis that nitric oxide is involved in the nonadrenergic, noncholinergic neurotransmission that leads to the smooth-muscle relaxation in the corpus cavernosum that permits penile erection. Defects in this pathway may cause some forms of impotence.

Adult↗
PubMed1992

Nitric oxide and prostacyclin inhibit fetal platelet aggregation: a response similar to that observed in adults.

OBJECTIVE: We evaluated the relative importance of two endothelium-derived substances, prostacyclin and nitric oxide, in their ability to inhibit aggregation of fetal and maternal platelets. STUDY DESIGN: The effects of various concentrations of prostacyclin and S-nitroso-N-acetylpenicillamine (which releases nitric oxide) on platelet aggregation were studied by means of platelet-rich plasma from at least five to six subjects per group. Fetal blood was collected from umbilical vein at delivery. Maternal venous blood was collected within 4 hours of delivery. Platelet aggregation was monitored with a platelet aggregation profiler. Adenosine diphosphate was used as the aggregating agent. Statistical differences between means were evaluated with two-way analysis of variance or Student t test. RESULTS: Prostacyclin and S-nitroso-N-acetylpenicillamine inhibited aggregation of fetal and maternal platelets, but prostacyclin was more potent. Fetal platelets were more sensitive than maternal platelets to prostacyclin and S-nitroso-N-acetylpenicillamine. CONCLUSION: Prostacyclin appears to be more important in preventing aggregation of platelets in the feto placental circulation.

Adenosine Diphosphate↗
PubMed1992

Nitric oxide-mediated neuronal injury in multiple sclerosis.

Although several explanations have been proposed for destruction of myelin and oligodendrocytes in multiple sclerosis, there is no proven mechanism of injury. We postulate that the autoimmune response seen in multiple sclerosis results in a cytokine-mediated increase in nitric oxide production by macrophages/microglia, smooth muscle cells and/or endothelium of the central nervous system. 3 mechanisms of cellular damage due to nitric oxide are proposed: 1. direct nitric oxide cytotoxicity; 2. injury due to peroxynitrite formation from superoxide anion and nitric oxide; and 3. nitric oxide-mediated elevations of cellular cGMP that enhance tumor necrosis factor-alpha toxicity. In support of these hypotheses, the anti-inflammatory effectors, dexamethasone and transforming growth factor-beta, ameliorate symptoms seen in clinical multiple sclerosis and experimental allergic encephalitis, respectively. These 2 immunomodulators also inhibit induction of cytokine-mediated nitric oxide production by macrophages. An experimental design and therapeutic interventions which will evaluate the role of nitric oxide in the pathophysiology of experimental allergic encephalitis are presented.

Animals↗
PubMed1992

Nitric oxide is a potent relaxant of human and rabbit corpus cavernosum.

Nitric oxide (NO) caused a potent, marked, and transient relaxation of precontracted strips of corpus cavernosum isolated from humans and rabbits. The relaxation response elicited by NO was very similar to the relaxation evoked by electrical field stimulation via the nonadrenergic-noncholinergic pathway. Sodium nitroprusside, nitroglycerin, and S-nitroso-N-acetylpenicillamine, which are nitrovasodilators known to generate NO, also caused marked concentration-dependent relaxation of corpus cavernosum. Relaxant responses to NO were enhanced by the cyclic GMP phosphodiesterase inhibitor M&B 22,948 and inhibited by oxyhemoglobin. Similarly, relaxation of corpus cavernosum in response to electrical field stimulation or acetylcholine was enhanced by M&B 22,948 and inhibited by oxyhemoglobin. NO stimulated cyclic GMP formation in corpus cavernosum and a close positive correlation was found between the magnitudes of relaxation and cyclic GMP formation. The data suggest that NO-elicited activation of guanylate cyclase and cyclic GMP formation represents the signal transduction mechanism responsible for relaxation and nonadrenergic-noncholinergic-mediated penile erection. These observations indicate that NO is a potent relaxant of human and rabbit corpus cavernosum and support our hypothesis that endogenous NO is the principal mediator of penile erection caused by nonadrenergic-noncholinergic stimulation.

3',5'-Cyclic-GMP Phosphodiesterases↗
PubMed1992

Role of L-arginine-nitric oxide pathway in myocardial reoxygenation injury.

In view of the recent findings that NO reacts with superoxide anion to generate hydroxyl radical, the present study was conducted to ascertain the role of endogenous NO in mediating myocardial reoxygenation injury in the hypoxic piglet on cardiopulmonary bypass. Anesthetized piglets were made hypoxic (PaO2 = 20-30 mmHg) for up to 120 min, followed by reoxygenation on cardiopulmonary bypass for 30 min. Reoxygenation caused rapidly developing myocardial injury characterized by decreased contractility (expressed as end-systolic elastance) and increased lipid peroxidation (measured as conjugated dienes). Systemic venous and coronary sinus blood content of NO decreased significantly during hypoxia and increased substantially above prehypoxic levels during reoxygenation on cardiopulmonary bypass. Administration of either the antioxidants mercaptopropionyl glycine and catalase or the NO synthase inhibitor, NG-nitro-L-arginine methyl ester, to the extracorporeal circuit afforded similar and nearly complete protection against myocardial reoxygenation injury. The protective effects of NG-nitro-L-arginine methyl ester were nullified by adding an excess of L-arginine to the pump circuit, suggesting that the L-arginine-NO pathway is involved in myocardial reoxygenation injury.

Animals↗
PubMed1992

Electrical field stimulation causes endothelium-dependent and nitric oxide-mediated relaxation of pulmonary artery.

The objective of this study was to ascertain the mechanism by which electrical field stimulation (EFS) of bovine pulmonary arterial rings causes endothelium-dependent smooth muscle relaxation. Like acetylcholine-elicited relaxation, EFS-elicited relaxation was endothelium-dependent and accompanied by accumulation of guanosine 3',5'-cyclic monophosphate (cGMP) in the vascular smooth muscle. Relaxation in response to EFS was unaltered by tetrodotoxin, guanethidine, atropine, propranolol, chlorpheniramine, cimetidine, indomethacin, aminophylline, alpha, beta-methylene ATP, nifedipine, capsaicin, and certain antioxidants and free radical scavengers. Thus the relaxation was not neurogenically mediated and was not attributed to free radical formation during EFS. Like nitric oxide-elicited relaxation, EFS-elicited relaxation was antagonized by oxyhemoglobin and methylene blue. Relaxation was also antagonized by the three NG-substituted L-arginine analogues: NG-methyl-L-arginine, NG-nitro-L-arginine, and NG-amino-L-arginine. NG-amino-L-arginine also inhibited the tissue cGMP accumulation in response to EFS. The inhibitory effect of the NG-substituted L-arginine analogues was reversed by addition of excess L-arginine but not D-arginine. Relaxation in response to EFS was dependent on the presence of extracellular calcium and intracellular calmodulin, as removal of extracellular calcium or addition of trifluoperazine nearly abolished relaxation. EFS-elicited relaxation was inhibited also by tetraethylammonium chloride and elevated extracellular potassium concentration. These observations indicate that EFS-elicited relaxation of bovine pulmonary artery is mediated by neuronally independent, but endothelium- and calcium-dependent, stimulation of nitric oxide and cGMP formation.

Animals↗
PubMed1992

Oxygen-mediated regulation of neonatal and adult rabbit aortic tension.

To determine the influence of oxygen on aortic tension in young and mature animals, we evaluated the response of neonatal and adult rabbit aortic rings to changes in oxygen tension. In 95% nitrogen/5% carbon dioxide, endothelium-derived nitric oxide was not a factor in the development or maintenance of low resting aortic ring tensions. However, the production of endothelium-derived nitric oxide increased in 21% oxygen/74% nitrogen/5% carbon dioxide and 95% oxygen/5% carbon dioxide, and attenuated a concomitant endothelium-independent mechanism of increasing vascular tension. A protein kinase inhibitor, 0.2 mM 1-(5-isoquinolinyl-sulfonyl)-2-methyl piperazine (H-7) inhibited oxygen-mediated constriction of neonatal vessels, but also significantly reduced the contractile response to 60 mM KCl. In conclusion, resting aortic tension is in part determined by oxygen-mediated control of the production of endothelium-derived nitric oxide and one or more opposing endothelium-independent mechanisms of constriction.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1991

Heme-dependent activation of guanylate cyclase by nitric oxide: a novel signal transduction mechanism.

The interaction between nitric oxide (NO) synthesized in one cell and cytosolic guanylate-cyclase-bound heme located in adjacent target cells to generate the NO-heme adduct of guanylate cyclase represents a novel and widespread signal transduction mechanism that links extracellular stimuli to the biosynthesis of cyclic GMP in target cells. A variety of chemical factors interact with selective extracellular receptors and trigger the biosynthesis of NO from L-arginine. The unique chemistry of NO endows this molecule with the capacity to diffuse rapidly into nearby cells and stimulate cyclic GMP formation. Cyclic GMP acts as a messenger in each cell type to trigger different but complementary cellular responses within a localized environment. This transcellular signaling is a form of rapid intercellular communication allowing the simultaneous local initiation of increased blood flow, inhibition of platelet-induced thrombosis and other cellular functions.

Animals↗
PubMed1991

Comparison of the inhibitory potencies of N(G)-methyl-, N(G)-nitro- and N(G)-amino-L-arginine on EDRF function in the rat: evidence for continuous basal EDRF release.

The relative potencies of the argininolytic agents NG-methyl-L-arginine (L-NMA), NG-nitro-L-arginine (L-NNA) and NG-amino-L-arginine (L-NAA) were assayed by their inhibitory effect on both basal and stimulated release of endothelium-derived NO in vitro and in vivo. Basal NO release was indirectly assessed by the ability of the analogs to contract phenylephrine-preconstricted rat aortic rings and their ability to produce a hypertensive response in awake, unanesthetized normotensive rats. In aortic rings, the three analogs induced vasocontraction and inhibited the vasorelaxation mediated by ACh-stimulated endothelial NO release. In this latter assay, L-NNA was 30 times more potent than either L-NMA or L-NAA. In free-moving rats, the agents caused dose-dependent increases in arterial pressure due to the blockade of endogenous NO formation. Dose-response analysis indicated that L-NNA was 87 and 230 times more potent than L-NMA and L-NAA, respectively. Pretreatment with L-NNA was also found to selectively inhibit, but not abolish, the depressor effects of acetylcholine in unanesthetized and phenylephrine- or vasopressin-infused normotensive-pithed rats. These studies indicate that L-NNA is a potent antagonist of endothelium-derived relaxing factor formation in vitro and in vivo. The contractile and hypertensive effects of the argininolytic agents clearly demonstrates that a continuous basal release of endothelium-derived relaxing factor/NO occurs in both isolated vascular rings and whole animals.

Acetylcholine↗
PubMed1979

Arachidonic acid activation of guinea pig lung guanylate cyclase by two independent mechanisms.

The purpose of this study was to elucidate the mechanisms by which arachidonic acid activates guanylate cyclase from guinea pig lung. Guanylate cyclase activities in both homogenate and soluble fractions of lung were examined. Guanylate cyclase activity was determined by measuring formtion of [32-P] cyclic GMP from alpha-[32-P] GTP in the presence of Mn2+, a phosphodiesterase inhibitor and a suitable GTP regenerating system. Arachidonic acid, and to a slight extent dihomo-gamma-linolenic acid, activated guanylate cyclase in homogenate but not soluble fractions. Similarly, phospholipase A2 activated homogenate but not soluble guanylate cyclase. Methyl arachidonate, linolenic, linoleic and oleic acids did not activate guanylate cyclase in either fraction. High concentrations of indomethacin, meclofenamate and aspirin inhibited activation of homogenate guanylate cyclase by arachidonic acid and phospholipase A2, without altering basal enzyme activity. These data suggested that a product of cyclooxygenase activity, present in the microsomal fraction, may have accounted for the capacity of arachidonic acid to activate homogenate guanylate cyclase. This view was supported by the findings that addition of the microsomal fraction to be soluble fraction enabled arachidonic acid to activate soluble guanylate cyclase, an effect which was reduced with cycloooxygenase inhibitors. Lipoxygenase activated guanylate cyclase in homogenate and soluble fractions. Arachidonic acid potentiated the activation of soluble guanylate cyclase by lipoxygenase, and this effect was inhibited with nordihydroguairetic acid, 1-phenyl-3-pyrazolidone and hydroquinone, but not with high concentrations of indomethacin, meclofenamate or aspirin. These data suggest that arachidonic acid activates guinea pig lung guanylate cyclase indirectly, via two independent mechanisms, one involving the microsomal fraction and the other involving lipoxygenase.

Animals↗