Local Release Of Nitric Oxide Research Articles

Evidence for a peripheral mechanism of action for the potentiation of the antinociceptive effect of morphine by dipyrone

The potentiation of the antinociceptive effect of morphine by dipyrone (metamizol) and the possible participation of a peripheral mechanism on such synergism were studied with the use of the formalin test in the rat. Nociception was induced by the intraplantar injection of diluted formalin (1%) in the right hind paw. Local administration of either dipyrone or morphine in the site of injury produced a dose-dependent antinociceptive effect. When combined, noneffective doses of morphine (1.25 μg/paw) and dipyrone (100 μg/paw) produced a significantly greater antinociceptive effect compared with either drug alone or saline. The opioid antagonist naloxone partly reversed the effect of the dipyrone–morphine combination. On the other hand, the inhibitor of nitric oxide (NO) synthesis, N G-L-nitro-arginine methylester (L-NAME), but not its inactive isomer, D-NAME, completely antagonized the effect of the dipyrone–morphine combination. These results suggest that the potentiation of morphine-induced antinociception by dipyrone in the formalin test requires an important participation of local release of NO, activating the NO–cyclic GMP pathway at the peripheral level.

Nitric oxide and thromboxane A2-mediated pulmonary microvascular dysfunction.

To examine whether the lung releases nitric oxide (NO) in response to thromboxane A2 and to examine the local release of NO as a protective compensatory mechanism by which the lung responds to the proinflammatory and vasoactive effects of thromboxane A2. The lungs of anesthetized Sprague-Dawley rats were perfused in vitro with Krebs-Henseleit buffer that contained an inhibitor of NO synthase (nitroglycerinenitro-L-arginine methyl ester [L-NAME]) (10(-4) mol/L), an NO donor (sodium nitroprusside) (10(-8) mol/L), or perfusate alone. Following equilibration, the thromboxane A2 receptor agonist 9,11-dideoxy-11alpha, 9alpha-epoxymethanoprostaglandin F2alpha(U-46619) (7.1 X 10(-8) mol/L) was added to the perfusate. Fifteen minutes later, the capillary filtration coefficient, pulmonary arterial pressure, and vascular resistance were measured. Pulmonary NO release was assessed by quantitating the release of cyclic guanosine monophosphate into the perfusate. The capillary filtration coefficient of lungs exposed to U-46619 was 3.5 times greater than that of lungs perfused with buffer alone (P<.05). The addition of sodium nitroprusside reduced the increase in capillary filtration coefficient associated with U-46619 by 50% (P<.05) whereas L-NAME had no effect. The addition of U-46619 to the perfused lung caused a 3.0+/-0.4 mm Hg increase in pulmonary artery pressure (P<.01) with a corresponding rise in total vascular resistance (P<.05). This effect was exacerbated by L-NAME (P<.05) and inhibited by sodium nitroprusside (P<.05). Exposure of the isolated lungs to U-46619 caused a 4-fold increase in cyclic guanosine monophosphate levels within the perfusate. These data are consistent with the hypothesis that NO release may be an important protective mechanism by which the lung responds to thromboxane A2.

Safety and efficacy of sildenafil citrate in the treatment of male erectile dysfunction

Sildenafil citrate, an oral therapy for erectile dysfunction, is a selective inhibitor of cyclic guanosine monophosphate (cGMP)—specific phosphodiesterase type 5 (PDE5), the predominant isozyme metabolizing cGMP in the corpus cavernosum. Chemically, it is a compound of the pyrazolo-pyrimidinyl-methylpiperazine class. Sildenafil has no direct relaxant effect on human corpus cavernosum but enhances the relaxant effect of nitric oxide (NO) on the corpus cavernosum by inhibiting PDE5, which is responsible for degradation of cGMP in this tissue. When sexual stimulation causes local release of NO, inhibition of PDE5 by sildenafil increases concentrations of cGMP in the corpus cavernosum, causing smooth muscle relaxation and blood flow into the penis, resulting in an erection. Sildenafil at recommended doses has no effect in the absence of sexual stimulation. The drug is rapidly absorbed after oral administration, with absolute bioavailability of 40%. Its pharmacokinetics are dose proportional over the recommended dosage range. Maximum plasma concentrations are reached within 30 to 120 minutes after oral dosing in the fasting state. Sildenafil is cleared predominantly by the hepatic microsomal isoenzymes CYP3A4 (major route) and CYP2C9 (minor route). Clinical studies assessed the effect of sildenafil on the ability of men with erectile dysfunction to engage in sexual activity and, specifically, to achieve and maintain an erection sufficient for satisfactory sexual intercourse. Sildenafil was evaluated at doses of 25, 50, and 100 mg in randomized, double-masked, placebo-controlled clinical trials of up to 6 months' duration. The drug was administered to hundreds of patients aged 19 to 87 years having erectile dysfunction of various etiologies for a mean duration of 5 years. Sildenafil was associated with statistically significant improvement in erectile function compared with placebo. Adverse effects reported at a rate of > 2% were headache, flushing, dyspepsia, nasal congestion, urinary tract infection, abnormal vision, diarrhea, dizziness, and rash. No cases of priapism were reported. The use of sildenafil is contraindicated in men who are taking organic nitrates, because of the potential for a precipitous decrease in blood pressure. Postmarketing reports and surveillance have revealed at least 39 deaths with sildenafil use in men having a history of heart disease, men taking nitrate medications, and men in poor physical health due to lack of exercise. Many of the men who experienced serious adverse effects or death had a variety of concomitant diseases and were taking multiple medications.

Immunohistochemically detected protein nitration indicates sites of renal nitric oxide release in Goldblatt hypertension.

In the kidney, nitric oxide (NO) from the macula densa (MD) is considered an integral modulator of the tubulovascular message system, whereas endothelium-derived NO is a major vasorelaxing factor. The goal of the present study was to determine extracellular pathways of NO in rats with renovascular two-kidney, one clip Goldblatt hypertension (2K1C). To localize NO in the tissue, immunohistochemical detection of NO-dependent tyrosine nitration was performed using a monoclonal antibody against nitrotyrosine. Nitration of phenolic compounds such as tyrosine results from the reaction with peroxynitrite (ONOO ) formed by NO and molecular oxygen or superoxide and may therefore be used as a footprint for local release of NO. Significant nitrotyrosine immunoreactivity was detected in the extraglomerular mesangium (EGM) of the stenotic kidney in 2K1C rats, whereas in the nonclipped contralateral kidney and in control animals no signal was detected at this site. Positive staining of the EGM was paralleled by enhanced NADPH diaphorase (NADPH-d) staining of the adjacent MD, signifying increased type I nitric oxide synthase (NOS) activity in the stenotic kidney. In contrast, in the cortical vasculature selectively enhanced nitrotyrosine immunoreactivity was detected in the arteriolar wall of the nonclipped contralateral kidney, and endothelial NADPH-d signal, indicating NOS Type III activity, was enhanced in parallel. Our results suggest that in MD, stimulation of NOS in the stenotic Goldblatt kidney induces the release of NO into the EGM. From there an NO-dependent intermediate stimulus may reach the glomerular vasculature. Footprints of NO-dependent effects in the vascular smooth muscle layer of the non-clipped contralateral kidney indicate a marked vasodilatory response that may have been caused by enhanced shear stress and/or angiotensin II levels.

Role of nitric oxide in the vasodilator response to mental stress in normal subjects

Vascular production of nitric oxide (NO) plays an important role in a variety of physiologic processes. This study examines the contribution of NO to the vasodilator response to mental stress. The effects of mental arithmetic testing on forearm vascular dynamics were analyzed in 15 normal subjects (9 men; age 45 ± 12 years) during intraarterial infusion of either saline or N G-monomethyl- l-arginine ( l-NMMA; 4 μmol/min for 15 minutes), an inhibitor of NO synthesis. The effect of l-NMMA on endothelium-independent vasodilation induced by intraarterial infusion of sodium nitroprusside was also studied in 11 of the 15 subjects. Forearm blood flow was measured by plethysmography. Mental stress increased forearm blood flow from 2.35 ± 0.84 to 5.06 ± 2.66 ml/min/dl (115%) during saline and from 1.72 ± 0.59 to 2.81 ± 0.99 ml/min/dl (63%) during l-NMMA infusion. The vasodilator effect of mental stress was significantly lower during l-NMMA infusion than during saline (1.1 ± 0.65 vs 2.71 ± 2.15 ml/min/dl; p = 0.01). l-NMMA administration did not significantly change mean arterial pressure and heart rate responses to mental stress. In contrast, the vasodilator effect of sodium nitroprusside (1.6 μg/min) was similar during infusion of l-NMMA and during saline (3.75 ± 1.55 vs 2.85 ± 1.38 ml/min/dl; p = 0.16). These findings indicate that local release of NO is involved in the forearm vasodilator response to mental stress.

Minimal effects of nitric oxide on spatial blood flow heterogeneity of the dog heart.

Eleven Beagle dogs were studied to elucidate the possible role of L-arginine-derived nitric oxide on local blood flow distribution in left and right ventricular myocardium. Local blood flow was determined in 256 samples from the left and 64 samples from the right ventricle per heart using the tracer microsphere technique (mean sample mass 319 +/- 131 mg). Nitric oxide production was effectively inhibited by intravenous infusion of 20 mg/kg nitro-L-arginine methylester (L-NAME) as evidenced by a shift of the dose/response curve for the effect of intracoronary administration of bradykinin (0.004-4.0 nmol/min) on coronary blood flow. L-NAME enhanced left and right ventricular systolic pressures from 132 +/- 18 to 155 +/- 15 mm Hg and from 26 +/- 3 to 29 +/- 3 mm Hg respectively (both P = 0.043). Mean left ventricular blood flow was 1.14 +/- 0.38 before and 0.99 +/- 0.28 ml min-1 g-1 after L-NAME (P = 0.068), while right ventricular blood flow fell from 0.72 +/- 0.28 to 0.53 +/- 0.20 ml min-1 g-1 (P = 0.043). Coronary conductance of left and right ventricular myocardium fell by 31 and 43% respectively (both P = 0.043). The coefficient of variation of left ventricular blood flow was 0.26 +/- 0.07 before and 0.29 +/- 0.07 after L-NAME (P = 0.068), that of right ventricular blood flow was 0.27 before and after L-NAME. Skewness (0.51) and kurtosis (4.23) of left ventricular blood flow distribution were unchanged after L-NAME, while in the right ventricle skewness decreased from 0.54 to 0.09 (P = 0.043) and kurtosis (3.68) tended to decrease after L-NAME (P = 0.080). The fractal dimension (D = 1.20-1.27) and the corresponding nearest-neighbor correlation coefficient (rn = 0.37-0.53) of left and right ventricular myocardium remained unchanged after infusion of L-NAME. From these results it is concluded that firstly, local nitric oxide release does not explain the higher perfusion of physiological high flow samples and secondly, that spatial myocardial blood flow coordination is not dependent on nitric oxide.

The Role of Endothelins in the Paracrine Control of the Secretion and Growth of the Adrenal Cortex

Endothelins (ETs) are a family of vasoactive peptides (ET-1, ET-2, and ET-3) mainly secreted by vascular endothelium and widely distributed in the various body systems, where they play major autocrine/paracrine regulatory functions, acting via two subtypes of receptors (ETA and ETB): Adrenal cortex synthesizes and releases ETS and expresses both ETA and ETB. Zona glomerulosa possesses both ETA and ETB, whereas zona fasciculata/reticularis is almost exclusively provided with ETB. ETS exert a strong mineralocorticoid and a less intense glucocorticoid secretagogue action, mainly via ETB receptors. ETS also appear to enhance the growth and steroidogenic capacity of zona glomerulosa and to stimulate its proliferative activity. This trophic action of ETS is likely to be mediated mainly by ETA receptors. The intraadrenal release of ETS undergoes a multiple regulation, with the rise in blood flow rate and the local release of nitric oxide being the main stimulatory factors. Data are also available that indicate that ETS may also have a role in the pathophysiology of primary aldosteronism caused by adrenal adenomas and carcinomas.

Possible role for endothelins in penile erection

The effects of endothelins (endothelin-1 and endothelin-3) on penile erection were studied in vivo in the pithed rat. Penile erectile responses were evaluated by measurements of intracorporal pressure via a needle inserted into one corpus cavernosum. Injection of endothelins (0.1–10 μg/kg i.v.) was followed by a rapid rise in corporal pressure. The responses to endothelin-1 and endothelin-3 were equivalent at the lower doses, but at 10 μg/kg the response to endothelin-3 was greater than that to endothelin-1. N G -Nitro- l-argininemethyl ester (20 mg/kg) but not indomethacin (10 mg/kg) antagonized the rise in corporal pressure caused by submaximal doses of endothelins. Electrical nerve stimulation-induced increase in corporal pressure was reduced after 5 μg/kg endothelin-1 but not after the same dose of endothelin-3. The results indicate that in corporal vasculature endothelin-3 has a predominantly vasodilator effect, while endothelin-1 has a vasodilator effect at lower doses and a vasoconstrictor effect at higher doses. The vasodilator action of endothelins is probably mediated by activation of endothelin ET B receptors and local release of nitric oxide. Thus, endothelins may participate in initiation and/or maintenance of penile erection.

Blood flow regulation in the transplanted fetal endocrine pancreas. Acquisition of a nitric oxide-dependent glucose-induced increase in blood flow.

Islet-like cell clusters (ICCs) were prepared from the fetal porcine pancreas by a culture technique. The ICCs (approximately 500) were implanted under the left renal capsule of nude (nu/nu) C57BL/6J mice. Six weeks, months, 12 months, or 16-24 months later, the animals were anesthetized and the blood flows to the xenogeneic islet graft and the adjacent kidney parenchyma were measured with laser-Doppler flowmetry. After the blood flow measurements, the graft-bearing kidneys were prepared for enzyme and immunohistochemistry. The blood perfusion of the graft was higher than that of the kidney at all times investigated. Intraperitoneal administration of glucose caused only slight and parallel changes in renal and graft blood flows 6 weeks, 6 months, or 12 months after transplantation. However, in all but 1 animal (n=16) transplanted >16 months before the blood flow measurements, glucose caused a marked increase in graft blood flow but did not affect renal blood flow. Injection of 2-deoxy-glucose also increased graft blood perfusion in animals transplanted > 16 months earlier (n=5). Treatment with NG-monomethyl-L-arginine (n=6), an inhibitor of nitric oxide synthase, prevented this glucose-induced flow increase. Nicotinamide adenine dinucleotide phosphate diaphorase histochemistry revealed nitric oxide synthase only in the endothelium and media of graft arterioles in animals in the oldest age group. Thus, with the passage of time after implantation, the grafted xenogeneic ICCs seem to achieve an autonomous blood flow regulation, different from that of the implantation organ. The reactivity to an increment in blood glucose concentration in the graft is similar to that seen in native islets in the pancreas but is not present until >16 months after implantation. The mechanisms for the glucose-induced blood flow increase are obscure but probably depend on local release of nitric oxide within graft arterioles.

Modulation of coronary autoregulatory responses by endothelium-derived nitric oxide

There is increasing evidence that endothelium-derived nitric oxide production is an important mechanism contributing to the regulation of myocardial perfusion during ischemia distal to a coronary stenosis. Studies in conscious chronically instrumented animals have extended observations in isolated arterioles to demonstrate that inhibiting nitric oxide synthase with l-arginine analogs increases the vulnerability of the myocardium to ischemia. The variable extent to which endothelium-dependent function is impaired in human atherosclerosis raises the possibility that abnormalities in resistance vessel control contribute to the functional significance of a fixed epicardial coronary stenosis. This may explain the wide variability between the physiological effects of a given coronary stenosis and its angiographic severity. Aggressive intervention to normalize endothelium-dependent vasodilation and local nitric oxide release may have beneficial effects on the functional significance of a coronary stenosis.

In the hippocampus in vivo, nitric oxide does not appear to function as an endogenous antiepileptic agent.

Using a reverberatory epilepiform discharge of hippocampal-parahippocampal circuits termed "maximal dentate activation", this study investigated whether the local release of nitric oxide within these circuits functions as an antiepileptic agent. Two nitric oxide synthase inhibitors (L-nitro-arginine methyl ester and 7-nitro-indazole) and a guanylate cyclase inhibitor (methylene blue) were tested, and none had a significant effect on the time to onset or duration of maximal dentate activation. A membrane-permeable analogue of cyclic guanosine monophosphate (cGMP), 8-bromo-cGMP, caused an increase in the time to onset and a decrease in the duration of maximal dentate activation. The number of neurons expressing NADPH diaphorase activity (a marker for nitric oxide synthase) was also examined after repeated elicitation of maximal dentate activation. After 18 seizures there was a significant, but transient, decrease in the number of hilar/subgranular neurons that were NADPH diaphorase-positive. The decrease was only seen at 1 h after the last seizure. There was no induction of NADPH diaphorase activity. These results are not consistent with the hypothesis that, in hippocampal-parahippocampal circuits in vivo, nitric oxide is released in response to neuronal activity and then acts to terminate the neuronal activity.