James N. Bates

Diet-induced atherosclerosis increases the release of nitrogen oxides from rabbit aorta.

We examined the hypothesis that impaired endothelium-dependent vasodilation in atherosclerosis is associated with decreased synthesis of nitrogen oxides by the vascular endothelium. The descending thoracic aortae of rabbits fed either normal diet, a high cholesterol diet for 2-5 wk (hypercholesterolemic, HC), or a high cholesterol diet for 6 mo (atherosclerotic, AS) were perfused in a bioassay organ chamber with physiologic buffer containing indomethacin. Despite a dramatic impairment in the vasodilator activity of endothelium-dependent relaxing factor (EDRF) released from both HC and AS aortae (assessed by bioassay), the release of nitrogen oxides (measured by chemiluminescence) from these vessels was not reduced, but markedly increased compared to NL. Thus, impaired endothelium-dependent relaxation in atherosclerosis is neither due to decreased activity of the enzyme responsible for the production of nitrogen oxides from arginine nor to arginine deficiency. Because the production of nitrogen oxides increased in response to acetylcholine in both hypercholesterolemic and atherosclerotic vessels, impairments in signal transduction are not responsible for abnormal endothelium-dependent relaxations. Impaired vasodilator activity of EDRF by cholesterol feeding may result from loss of incorporation of nitric oxide into a more potent parent compound, or accelerated degradation of EDRF.

Nitric oxide generation from nitroprusside by vascular tissue. Evidence that reduction of the nitroprusside anion and cyanide loss are required

Nitric oxide (NO) was produced from sodium nitroprusside in the presence of vascular tissue but was not released spontaneously from the nitroprusside anion. In the absence of tissue in the dark nitroprusside did not release NO. When solutions of nitroprusside alone were irradiated with visible light, nitric oxide was released at rates linearly proportional to nitroprusside concentration and light intensity. Nitric oxide was produced from solutions of nitroprusside in the dark after the addition of vascular tissue, including lengths of rabbit aorta, subcellular fractions of aorta, and human plasma. NO was also released from nitroprusside after reaction with various reducing agents including cysteine and other thiols, ascorbic acid, sodium dithionite, ferrous chloride, hemoglobin, myoglobin, and partially purified cytochrome P450 with an NADPH-regenerating system. HCN was simultaneously produced in these solutions, and addition of KCN blocked NO release. Iodine oxidized intermediate cyanoferrates and blocked nitric oxide release. KCN or iodine also blocked NO production by tissue, but had no effect upon photochemical NO release. These results show that, apart from photolysis which makes no physiological contribution, release of nitric oxide from nitroprusside, in simple solutions and in biological tissue, occurs after nitroprusside has undergone reduction and lost cyanide.

The nitrovasodilators. New ideas about old drugs.

The nitrovasodilators are a diverse group of pharmacological agents that produce vascular relaxation by releasing nitric oxide. The mechanisms by which these compounds release nitric oxide vary, depending on their chemical structure. Compounds with lower oxidation states of nitrogen such as nitroprusside, nitrosamines, and nitrosothiols release nitric oxide nonenzymatically. In the case of nitroprusside, this involves a one-electron reduction that may occur upon exposure to a variety of reducing agents and tissues such as vascular smooth muscle membranes. In the case of the organic nitrates, which have higher oxidation states of nitrogen, the release of nitric oxide in vascular tissue occurs predominantly by a poorly understood enzymatic process. This interesting property of nitroglycerin is important because it “targets” its effect to vascular tissues that are capable of this enzymatic process. In the case of the coronary circulation, nitroglycerin predominantly dilates the larger coronary arteries while having a minimal effect on coronary resistance vessels <100 μm in diameter. This prevents the development of coronary steal, which is often encountered with agents that produce intense vasodilation of the coronary resistance vessels. In this review, the mechanisms by which the nitrovasodilators (particularly nitroglycerin) release nitric oxide will be considered, and recent studies of nitroglycerin bioconversion in various-sized coronary vessels will be discussed in detail.

Continuous infusion epidural analgesia during labor: a randomized, double-blind comparison of 0.0625% bupivacaine/0.0002% fentanyl versus 0.125% bupivacaine.

The analgesic efficacy of the continuous epidural infusion of 0.0625% bupivacaine/0.0002% fentanyl was compared with the infusion of 0.125% bupivacaine alone in a randomized, double-blind study of nulliparous women. Each patient received, in sequence: 1)3 ml of 0.5% bupivacaine with 1:200,000 epinep

Nitric Oxide as an Autocrine Regulator of Sodium Currents in Baroreceptor Neurons

Arterial baroreceptors are mechanosensitive nerve endings in the aortic arch and carotid sinus that play a critical role in acute regulation of arterial blood pressure. A previous study has shown that nitric oxide (NO) or NO-related species suppress action potential discharge of baroreceptors. In the present study, we investigated the effects of NO on Na+ currents of isolated baroreceptor neurons in culture. Exogenous NO donors inhibited both tetrodotoxin (TTX) -sensitive and -insensitive Na+ currents. The inhibition was not mediated by cGMP but by NO interaction with channel thiols. Acute inhibition of NO synthase increased the Na+ currents. NO scavengers (hemoglobin and ferrous diethyldithiocarbamate) increased Na+ currents before but not after inhibition of NO synthase. Furthermore, NO production in the neuronal cultures was detected by chemiluminescence and immunoreactivity to the neuronal isoform of NO synthase was identified in fluorescently identified baroreceptor neurons. These results indicate that NO/NO-related species function as autocrine regulators of Na+ currents in baroreceptor neurons. Modulation of Na+ channels may represent a novel response to NO.

The Influence of Continuous Epidural Bupivacaine Analgesia on the Second Stage of Labor and Method of Delivery in Nulliparous Women

A randomized, double blind, placebo-controlled study was performed to evaluate the analgesic efficacy and influence of continuing an epidural infusion of 0.125% bupivacaine beyond a cervical dilatation of 8 cm in nulliparous women. When the cervix was ≥ 8 cm dilated, coded study solution was substituted for the known 0.125% bupivacaine solution. The study solution for 46 patients was 0.125% bupivacaine; 46 patients received saline. During the first stage of labor, 44 (96%) women in the bupivacaine group, and 45 (98%) in the saline group, had analgesia of excellent or good quality. During the second stage, 36 (82%) women in the bupivacaine group, versus 18 (41%) women in the saline group, had analgesia of excellent or good quality (P < .0001). Six (13%) women in each group underwent cesarean delivery after the start of the study solution. Among the women who delivered vaginally, the mean (±S.D.) duration of the second stage of labor was 124 (±70) min in the bupivacaine group, versus 94 (±54) min in the saline group (P < .05). Twenty-one of 40 (53%) women in the bupivacaine group, versus 11 of 40 (28%) in the saline group, underwent instrumental vaginal delivery (P < .05). Twenty-eight of 40 (70%) women in the bupivacaine group, versus six of 40 (15%) in the saline group, had surgical perineal anesthesia for vaginal delivery (P < .0001). There were no significant differences between groups in Apgar scores or umbilical cord blood acid-base values. We conclude that, under the conditions of the present study, epidural bupivacaine infusion beyond a cervical dilatation of 8 cm provided satisfactory analgesia, but prolonged the second stage of labor and increased the frequency of instrumental delivery in nulliparous women. However, maintenance of epidural bupivacaine analgesia did not result in an increased incidence of abnormal position of the vertex or a more frequent performance of cesarean section.

Continuous Epidural Infusion of 0.0625% Bupivacaine-0.0002% Fentanyl during the Second Stage of Labor

A randomized, double-blind, placebo-controlled study was performed to evaluate the analgesic efficacy and influence of continuing an epidural infusion of 0.0625% bupivacaine-0.0002% fentanyl during the second stage of labor in nulliparous women. When the cervix was fully dilated, coded study solution was substituted for the known bupivacaine-fentanyl solution. The study solution for 29 patients was 0.0625% bupivacaine-0.0002% fentanyl; 34 patients received saline placebo. The two groups had similar pain scores during the first stage of labor. During the second stage, pain scores were significantly higher in the saline-placebo group at each 30-min interval between 60 and 150 min after the diagnosis of full cervical dilation. Similarly, there was a significant difference between the two groups in global assessment of analgesia quality during the second stage, but the difference occurred in those patients with a second-stage duration of greater than or equal to 60 min. Among the women who delivered vaginally, eleven of 28 (39%) women in the bupivacaine-fentanyl group, versus five of 34 (15%) in the saline-placebo group, had surgical perineal anesthesia for vaginal delivery (P less than .05). Six of 28 (21%) women in the bupivacaine-fentanyl group, and five of 34 (15%) in the saline-placebo group, underwent instrumental vaginal delivery (P = NS). The median duration of the second stage of labor was 53 min (range = 5-283) in the bupivacaine-fentanyl group, and 63 min (range = 16-181) in the saline-placebo group (P = NS).(ABSTRACT TRUNCATED AT 250 WORDS)

Nitric oxide inhibits aromatase activity: Mechanisms of action

NO synthase is present in human ovarian granulosa-luteal cells and NO inhibits estradiol secretion by granulosa cells in culture. These findings suggest that NO is an autocrine regulator of ovarian steroidogenesis. The purpose of this investigation was to explore the mechanisms through which NO exerts an inhibitory effect on cytochrome P450 aromatase activity. To examine the effect of NO on aromatase mRNA levels, human granulosa-luteal cells were cultured in the presence or absence of the NO donor SNAP for 16 h. Using a probe for human aromatase, Northern blots revealed a 26% decrease in aromatase mRNA in cells exposed to SNAP. Because this modest decrease in mRNA is unlikely to explain a rapid and profound reduction in estradiol secretion that we have observed, we looked for direct effects of NO on cytochrome P450 aromatase activity. Aromatase activity was assayed in placental microsomes and granulosa-luteal cells by measuring the release of 3H2O from [1 beta-3H] androstenedione. NO (10(-4)-10(-3)M), added as a saturated saline solution, reduced aromatase activity by as much as 90% in a concentration-dependent, non-competitive manner. In contrast, carbon monoxide (CO), a gas known to bind to the heme iron in aromatase, had no effect on aromatase activity when added alone nor could CO reverse the NO-induced inhibition of aromatase. These data suggest that NO binding to the heme is insufficient to inhibit aromatase activity. NO has been reported to alter protein function by reacting with the sulfhydryl group of cysteines, forming a nitrosothiol group. Because a cysteine sulfhydryl group is thought to participate in the catalytic mechanism of all P450 enzymes, experiments were designed to test whether NO might inhibit aromatase via such a mechanism. Addition of increasing amounts of mercaptoethanol, a chemical with free sulfhydryl groups, blocked the NO-induced inhibition of aromatase in microsomes. N-Ethylmaleimide, a chemical which covalently modifies sulfhydryl groups, reduced aromatase activity in a concentration-dependent manner. We conclude that NO inhibits aromatase both by decreasing mRNA for the enzyme and by an acute, direct inhibition of enzyme activity. We hypothesize that the direct inhibition occurs as a result of the formation of a nitrosothiol on the cysteine residue adjacent to the heme in aromatase.

Mechanisms responsible for the heterogeneous coronary microvascular response to nitroglycerin.

Nitroglycerin dilates large (greater than or equal to 100 microns) but not small coronary arterial microvessels, and a putative metabolite of nitroglycerin, S-nitroso-L-cysteine, has been shown in vitro to dilate both large and small coronary microvessels. Based on this evidence, we tested the hypothesis that the lack of response of small coronary microvessels was due to an inability of small coronary microvessels to convert nitroglycerin into its vasoactive metabolite and examined possible explanations for this phenomenon. We studied left ventricular epicardial microvessels in vivo using video microscopy and stroboscopic epi-illumination in anesthetized, open-chest dogs. Diameters were determined while the epicardium was suffused with nitroglycerin, S-nitroso-L-cysteine, or S-nitroso-D-cysteine (all 10 microM) and nitroglycerin in the presence of L- or D-cysteine (100 microM). None of the agents affected systemic hemodynamics. Nitroglycerin dilated large arterioles (20 +/- 2%) but not small arterioles (1 +/- 1%). Both S-nitroso-L-cysteine and S-nitroso-D-cysteine were potent dilators of all size classes of microvessels. Concomitant application of L-cysteine and nitroglycerin evoked dilation in small microvessels (22 +/- 4%, p less than 0.5 versus nitroglycerin alone) and larger microvessels (27 +/- 6%, p = NS versus nitroglycerin alone). D-Cysteine did not alter the microvascular response to nitroglycerin in either small (7 +/- 4%, p = NS versus nitroglycerin alone) or large (18 +/- 3%, p = NS versus nitroglycerin alone) microvessels. Neither L-cysteine nor D-cysteine had a direct effect on microvascular diameter. These findings suggest that 1) sulfhydryl groups are required for the conversion of nitroglycerin to its vasoactive metabolite; 2) the interaction between nitroglycerin and sulfhydryl residues is a stereospecific process, indicating either an intracellular mechanism or a membrane-associated enzymatic reaction; and 3) a lack of available sulfhydryl groups may be responsible for the lack of response of small coronary arterioles to nitroglycerin.

Modulation of Baroreceptor Activity by Nitric Oxide and S-Nitrosocysteine

The goal of this study was to determine whether nitric oxide (NO) and the NO donor, S-nitrosocysteine (cysNO), modulate the activity of carotid sinus baroreceptors. Baroreceptor activity was recorded from the vascularly isolated carotid sinus in anesthetized rabbits. Baroreceptor activity decreased in a dose-dependent manner after injection of either NO or cysNO as constant pressure was maintained, and activity recovered spontaneously over time, within seconds to minutes. The baroreceptor pressure-activity relation was shifted significantly to the right by cysNO, with a profound suppression of activity at high pressure. Baroreceptor activity at 160 mm Hg averaged 76 +/- 8%, 60 +/- 6%, and 36 +/- 5% of the control maximum during exposure to 10(-4), 2 to 3 x 10(-4), and 10(-3) mol/L cysNO, respectively. The inhibition of activity by the L and D isomers of cysNO was equivalent and was blocked by reduced hemoglobin, suggesting that the effect was mediated by NO. The suppression of baroreceptor activity by cysNO was not related to vascular relaxation as measured by videomicrometer. Inhibition of soluble guanylate cyclase with methylene blue or 6-anilinoquinoline-5,8-quinone (LY83583, 10(-5) mol/L) did not attenuate and dibutyryl cGMP (10(-3) mol/L) did not mimic the suppression of baroreceptor activity by cysNO, suggesting a cGMP-independent mechanism. Activation of endogenous NO formation with thimerosal (10(-5) to 10(-4) mol/L) reduced maximum baroreceptor activity in five of eight experiments to 59 +/- 7% of the control maximum.(ABSTRACT TRUNCATED AT 250 WORDS)