Nitric Oxide: A Physiologic Messenger

  1. Charles J. Lowenstein;
  2. Jay L. Dinerman; and
  3. Solomon H. Snyder
  1. From The Johns Hopkins University School of Medicine, Baltimore, Maryland. Requests for Reprints: Solomon H. Snyder, MD, Departments of Neuroscience, Pharmacology, and Molecular Sciences, The Johns Hopkins University School of Medicine, 725 North Wolfe Street, Baltimore, Maryland 21205. Grant Support: By the National Institutes of Health through grants PSA K1102451 (Dr. Lowenstein), MH18501, DA00266, and Research Scientist Award DA-00074; and a grant from the W.M. Keck Foundation (Dr. Snyder).
     
    Next Section

    Abstract

    Purpose: To review the physiologic role of nitric oxide, an unusual messenger molecule that mediates blood vessel relaxation, neurotransmission, and pathogen suppression.

    Data Sources: A MEDLINE search of articles published from 1987 to 1993 that addressed nitric oxide and the enzyme that synthesizes it, nitric oxide synthase.

    Study Selection: Animal and human studies were selected from 3044 articles to analyze the clinical importance of nitric oxide. Descriptions of the structure and function of nitric oxide synthase were selected to show how nitric oxide acts as a biological messenger molecule.

    Data Extraction: Biochemical and physiologic studies were analyzed if the same results were found by three or more independent observers.

    Data Synthesis: Two major classes of nitric oxide synthase enzymes produce nitric oxide. The constitutive isoforms found in endothelial cells and neurons release small amounts of nitric oxide for brief periods to signal adjacent cells, whereas the inducible isoform found in macrophages releases large amounts of nitric oxide continuously to eliminate bacteria and parasites. By diffusing into adjacent cells and binding to enzymes that contain iron, nitric oxide plays many important physiologic roles. It regulates blood pressure, transmits signals between neurons, and suppresses pathogens. Excess amounts, however, can damage host cells, causing neurotoxicity during strokes and causing the hypotension associated with sepsis.

    Conclusions: Nitric oxide is a simple molecule with many physiologic roles in the cardiovascular, neurologic, and immune systems. Although the general principles of nitric oxide synthesis are known, further research is necessary to determine what role it plays in causing disease.

    A noxious, unstable gas is an unlikely candidate to act as a biological messenger. However, in the last 7 years, nitric oxide, a byproduct of automobile exhaust, electric power stations, and lightning, was discovered in the body, where it participates in various functions, including suppression of pathogens, vasodilation, and neurotransmission [1-8]. We describe what is known about nitric oxide, focusing on its clinical relevance.

    Previous SectionNext Section

    Unusual Properties of Nitric Oxide Metabolism

    Nitric oxide is an unusual messenger. It is a small molecule composed of one atom each of nitrogen and oxygen, not to be confused with nitrous oxide, N2O, an inhalational anesthetic commonly referred to as laughing gas. Nitric oxide is an uncharged molecule with an unpaired electron. These characteristics of nitric oxide make it an ideal messenger molecule: Uncharged, nitric oxide can diffuse freely across membranes. With an unpaired electron, it is called a radical molecule, which is highly reactive (having a half-life of 2 to 30 seconds); after transmitting a signal spontaneously, it decays into nitrite.

    Nitric oxide is made by nitric oxide synthase in an unusual reaction that converts arginine and oxygen into citrulline and nitric oxide. The mechanism of nitric oxide synthesis is not completely understood, but it involves the transfer of electrons between various cofactors, including flavin adenine dinucleotide, flavin mononucleotide, nicotinamide adenine dinucleotide phosphate (NADPH), tetrahydrobiopterin, and heme. Finally, one atom of oxygen from oxygen binds with the terminal guanidine nitrogen from arginine to form nitric oxide [9, 10].

    Although several nitric oxide synthase isoforms have been isolated [11-17], all are homologous and divided into two categories with different regulation and activities (Figure 1). The constitutive isoforms in neuronal or endothelial cells are always present [11, 18]. These nitric oxide synthase isoforms are inactive until intracellular calcium levels increase, the calcium-binding protein calmodulin binds to calcium, and the calcium-calmodulin complex binds to and activates nitric oxide synthase [19-22]. The constitutive nitric oxide synthase isoforms then synthesize small amounts of nitric oxide until calcium levels decrease. This intermittent production of small amounts of nitric oxide transmits signals. In contrast, the inducible nitric oxide synthase isoform is normally absent from macrophages and hepatocytes, but when these cells are activated by specific cytokines, an inducible nitric oxide synthase enzyme is produced; once produced, it always synthesizes large amounts of nitric oxide. Induced nitric oxide synthase is transcriptionally regulated [16, 23]. The continuous production of large amounts of nitric oxide kills or inhibits pathogens.

    Figure 1. CAL = binding site for calmodulin; FMN = flavin mononucleotide; FAD = flavin adenine dinucleotide; NADPH = reduced form of nicotinamide adenine dinucleotide phosphate;  = site for phosphorylation by cyclic adenosine monophosphate-dependent protein kinase. The sites for heme, arginine, and tetrahydrobiopterin binding are unknown.
    View larger version:
      Figure 1. CAL = binding site for calmodulin; FMN = flavin mononucleotide; FAD = flavin adenine dinucleotide; NADPH = reduced form of nicotinamide adenine dinucleotide phosphate;  = site for phosphorylation by cyclic adenosine monophosphate-dependent protein kinase. The sites for heme, arginine, and tetrahydrobiopterin binding are unknown. Diagrammatic representation of the structure of cloned forms of nitric oxide synthase with sites for cofactor binding.P

      Other mechanisms of regulating nitric oxide synthase enzymes recently were discovered. Neuronal nitric oxide synthase can be phosphorylated by protein kinases to decrease its activity [24]. The subcellular location of nitric oxide synthase in endothelial cells can also be changed by its phosphorylation [25]. Although the constitutive isoforms are regulated by calcium, the inducible isoforms also appear to bind calmodulin, although calcium has little effect on their activity [26]. The role of calmodulin in inducible nitric oxide synthase function is unknown.

      Most molecules that transmit signals between cells, such as hormones, neurotransmitters, and growth factors, act through specific protein receptors that are often associated with the plasma membrane. In contrast, nitric oxide diffuses out of the cell that generates it and into target cells, where it interacts with specific molecular targets (Table 1). The best-characterized receptor of nitric oxide is iron, contained in certain proteins as a heme group or as an iron-sulfur complex. Nitric oxide exerts some of its effects by binding to iron-containing enzymes and either activating or inactivating the enzymes. When nitric oxide binds to the iron in the heme group of guanylate cyclase, the enzyme is activated. Guanylate cyclase then produces cyclic guanosine monophosphate (cGMP), and the increase in cGMP activates other cellular processes (Figure 2). By changing the activity of guanylate cyclase, nitric oxide dilates arteries, signals neurons, and kills cells.

      View this table:
      Table 1. Molecular Targets of Nitric Oxide*
      Figure 2. A messenger molecule such as acetylcholine binds to the acetylcholine receptor on an endothelial cell, activating inward calcium currents. Calcium binds to calmodulin and activates endothelial cell nitric oxide synthase, which converts arginine plus oxygen into citrulline and nitric oxide. Nitric oxide diffuses out of the endothelial cell into an adjacent smooth muscle cell and activates guanylate cyclase by binding to the iron in its heme group. The increase in cyclic guanosine monophosphate (cGMP) causes smooth muscle relaxation, and thus vasodilation. GTP = guanosine triphosphate.
      View larger version:
        Figure 2. A messenger molecule such as acetylcholine binds to the acetylcholine receptor on an endothelial cell, activating inward calcium currents. Calcium binds to calmodulin and activates endothelial cell nitric oxide synthase, which converts arginine plus oxygen into citrulline and nitric oxide. Nitric oxide diffuses out of the endothelial cell into an adjacent smooth muscle cell and activates guanylate cyclase by binding to the iron in its heme group. The increase in cyclic guanosine monophosphate (cGMP) causes smooth muscle relaxation, and thus vasodilation. GTP = guanosine triphosphate. Nitric oxide arterial smooth muscle.

        Another unusual way in which nitric oxide affects cells is by facilitating transfer of an ADP-ribose group to an accepting molecule (a process called ADP ribosylation). Normally an ADP-ribose group is attached to a protein target by an enzyme, for example, when cholera toxin ADP-ribosylates a guanosine triphosphate-binding protein. However, when nitric oxide diffuses into a cell, it can cause auto-ADP ribosylation, that is, ADP ribosylation of a target without enzyme catalysis. For example, nitric oxide inactivates the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase by attaching an ADP-ribose group to it, thereby blocking the production of adenosine triphosphate from glycolysis [27-32].

        Previous SectionNext Section

        Nitric Oxide in the Cardiovascular System

        Although the effect of exogenously administered nitrates on the vasculature has been studied for decades, the first clue that endothelial cells released a substance that could cause vasodilation was found only recently. In 1980, Furchgott and Zawadski [33] discovered that sections of the aorta would relax in response to agonists only if the inner linings of endothelial cells were intact. However, aortic rings with no endothelial cells could not relax. Endothelial cells thus released an agent that relaxed vascular smooth muscle; within several years, it was discovered that this “endothelial-derived relaxation factor” was nitric oxide [34].

        The Role of Nitric Oxide in Mediating an Active State of Vasodilation

        The vasculature is in a constant state of active dilation mediated by nitric oxide. Endothelial cells continuously release small amounts of nitric oxide, producing a basal level of vascular smooth muscle relaxation. When inhibitors of nitric oxide synthase, such as nitro-arginine methyl ester or n-mono-methyl-arginine, are infused into animals [35-44] or humans [45], nitric oxide production is inhibited, vascular smooth muscle contracts, and blood pressure increases.

        Nitric oxide dilates blood vessels by directly relaxing vascular smooth muscle cells (see Figure 2). An agonist such as acetylcholine binds to its receptor on endothelial cells, causing a transient increase in intracellular calcium. Calcium binds to calmodulin, and the calcium-calmodulin complex activates endothelial nitric oxide synthase, which makes nitric oxide. Nitric oxide then diffuses out of the endothelial cell and into adjacent smooth muscle cells, where it binds to the heme group of guanylate cyclase. Guanylate cyclase is activated to produce cGMP, which, through a cascade of protein kinases, induces smooth muscle relaxation. Because the half-life of nitric oxide in biological fluids is between 2 and 30 seconds, the effect of nitric oxide spontaneously diminishes and the vessel constricts unless more nitric oxide is produced.

        Nitric Oxide: An Autoregulator and Neural Regulator of Blood Flow

        Nitric oxide automatically regulates blood flow in response to local changes in some regions of the vasculature. Ischemia and reperfusion cause vasodilation only in the affected tissue, and this response is mediated by nitric oxide. For example, a study of ischemia in patients' arms and legs showed an increase in nitric oxide production [46]. However, whereas some investigators found that ischemia and reperfusion increase nitric oxide production in animals [47, 48], others observed a decrease in nitric oxide [49-51]. Shear-stress, an increase in blood flow through a vessel, is another physical stimulus to which endothelial cells respond by increasing nitric oxide production [52, 53]. In animal studies, increases in blood flow but not pressure caused an increase in nitric oxide production and dilation in excised hearts and vessels [54-59]. Factors locally released by adjacent tissue, such as bradykinin or acetylcholine, can also induce nitric oxide release in some vessels but not in others. A basal level of nitric oxide regulates blood flow in the brain [60-62], heart [59, 63-67], lung [68], gastrointestinal tract [69], and kidney [38, 42, 43, 70]. Thus, nitric oxide is an endogenous autoregulator of blood flow.

        The release of nitric oxide into the vasculature is also controlled by the autonomic nervous system. Parasympathetic nerves containing nitric oxide synthase terminate in the adventitia of certain large vessels, such as the cerebral and retinal arteries [71]. Nitric oxide is released from the nerves and diffuses into the muscular media from the outside of the vessel, causing vasorelaxation.

        The Role of Nitric Oxide in Hypertension and Vasospasm

        Because nitric oxide plays a central role in regulating blood pressure, defects in the regulation of nitric oxide synthase could lead to vasospasm or hypertension, although this has not been proved. Clearly the endothelium is abnormal in persons with hypertension of unknown cause (“essential hypertension”), because acetylcholine causes less vasodilation in hypertensive than in healthy persons [72-74]. In contrast, the vascular smooth muscle is normal in persons with hypertension because intravenous nitroprusside, which releases nitric oxide, causes equal amounts of vasodilation in both hypertensive and healthy persons [74, 75]. Abnormal endothelial nitric oxide synthase could cause this defective response to acetylcholine in persons with hypertension. In fact, the blood pressure of healthy persons increases when nitric oxide synthase inhibitors are infused, but the blood pressure of persons with hypertension changes much less [74, 75]. These studies imply that persons with hypertension produce less nitric oxide. However, direct measurement in hypertensive persons' arteries shows normal quantities of nitric oxide synthase (Personal communication. Michel T, Brigham and Women's Hospital, Boston, Massachusetts). Perhaps endothelial nitric oxide synthase in persons with hypertension has an abnormal structure or regulation.

        Endothelial dysfunction is characteristic of other diseases, including diabetes and atherosclerosis [76-80]. Some atherosclerotic arteries paradoxically constrict when the vasodilator acetylcholine is infused. Defects in the cascade of events leading to production of nitric oxide might cause less relaxation. Nitric oxide not only regulates the systemic blood pressure but also the local blood flow to specific vascular beds [81], including the brain [60-62], kidney [38, 42, 43, 70], lung [68], heart [59, 63, 64, 66, 67], and gastrointestinal tract [69]. A local defect in nitric oxide production could cause vasospasm in particular organs. Perhaps nitric oxide is involved in hepatorenal syndrome, Prinzmetal angina, Raynaud disease, or preeclampsia.

        However, nitric oxide is not the only vasodilator, and defects in other relaxation pathways could also cause endothelial dysfunction and contractile abnormalities. Agonists such as acetylcholine, bradykinin, and thrombin bind to endothelial cells, inducing the release of vasodilators including nitric oxide, endothelial-derived hyperpolarizing factor, and prostacyclin. These compounds relax smooth muscle cells in different ways. Nitric oxide activates guanylate cyclase. Endothelial-derived hyperpolarizing factor activates a potassium channel, and the potassium flux hyperpolarizes and thus relaxes the smooth muscle cell. Prostacyclin can activate both adenylate cyclase and guanylate cyclase, thereby causing an increase in cyclic adenosine monophosphate and cGMP and ultimately relaxation. Another endothelial cell agonist is lysophosphatidylcholine, a component of oxidized low-density lipoprotein that activates an endothelial pathway that leads to relaxation of smooth muscle. This pathway probably does not involve nitric oxide [82-84] but rather an endothelial cell-surface receptor coupled to a guanosine triphosphate-binding protein. Lysophosphatidylcholine disrupts the interaction of this receptor and its coupled guanosine triphosphate-binding protein [85].

        Although no diseases have been described in which excessive amounts of nitric oxide are produced, rare cases of abnormally low blood pressure, such as idiopathic orthostatic hypotension, may be associated with overproduction of nitric oxide. As discussed below, nitric oxide mediates the hypotension of sepsis. It may be involved in local shunting of blood through the liver in cirrhosis [86, 87], although some studies have not found excessive amounts of nitric oxide in the cirrhotic liver [88, 89].

        Effects of Nitric Oxide on Cardiac Contractility and Platelet Clotting

        Nitric oxide influences areas of the cardiovascular system besides blood vessels. It exerts a negative chronotropic effect on the heart and a negative inotropic effect on cardiac muscle cells similar to its relaxation of smooth muscle cells. Our laboratory has shown that nitric oxide synthase can be induced during viral myocarditis (unpublished data). Reversible left ventricular dysfunction known to occur during myocarditis may be caused by an excess of nitric oxide that relaxes cardiac muscle and resolves as nitric oxide synthase produces less nitric oxide.

        Nitric oxide reduces clotting by acting directly on platelets. The nitric oxide produced by endothelial cells can diffuse not only toward the vessel wall but also into the vessel lumen, where it enters platelets. Nitric oxide synthase has also been found inside platelets [90]. It reduces clotting by inhibiting platelet aggregation [91-95] and adhesion [96-101]. The molecular basis of this action is unknown, but possible mechanisms include nitric oxide activating platelet guanylate cyclase and adenylate cyclase [99, 102-106], inhibiting platelet phospholipase C [107], or attaching ADP-ribose to platelet glyceraldehyde 3-phosphate dehydrogenase [108]. Thus, endothelial generation of nitric oxide not only regulates blood pressure but also blood clotting.

        Previous SectionNext Section

        Nitric Oxide: An Immune Effector Capable of Eliminating Many Pathogens

        In contrast to the neuronal and endothelial nitric oxide synthase, the inducible macrophage nitric oxide synthase produces a large amount of nitric oxide. In large quantities, nitric oxide kills almost any nearby cell. It kills or inhibits the growth of many pathogens, including bacteria [109-113], fungi [114], and parasites [115-122] (Table 2). In particular, nitric oxide eliminates intracellular pathogens such as Mycobacterium tuberculosis, and recent studies from our laboratory (unpublished) and others [123, 124] suggest that nitric oxide blocks viral replication as well. Nitric oxide also kills tumor cells [125]. In addition, nitric oxide might damage normal host cells in autoimmune diseases in which nitric oxide synthase is inappropriately induced. Thus, it is a nonspecific effector capable of killing or inhibiting the growth of many targets, both pathogens and host cells.

        View this table:
        Table 2. Pathogens and Cellular Targets of Nitric Oxide*

        Mechanisms whereby Nitric Oxide Kills Its Targets

        Because nitric oxide has so many molecular targets, it can damage cells in many ways, although precisely how it does so is unclear. Nitric oxide inhibits adenosine triphosphate production at three separate steps, blocking glycolysis by transferring ADP-ribose to glyceraldehyde-3-phosphate dehydrogenase, disrupting the Krebs cycle by binding to the heme group of cis-aconitase, and inhibiting oxidative phosphorylation by binding to the heme group of ubiquinone reductase. Nitric oxide inhibits DNA synthesis by inactivating ribonucleotide reductase [126, 127] and can damage DNA directly by deamination [128]. While continuously secreting large amounts of nitric oxide, activated macrophages avoid the toxic effects of nitric oxide by mechanisms that are also unknown.

        Transcriptional Regulation of Macrophage Nitric Oxide Synthase

        The production of nitric oxide must be controlled precisely, because excessive amounts could damage the host and too little could lead to immunodeficiency. In contrast to the neuronal and endothelial nitric oxide synthase enzymes that are always present but only active when intracellular calcium levels increase, nitric oxide synthase is normally absent in quiescent macrophages; however, once it is synthesized it is always active. Macrophage nitric oxide synthase is thus transcriptionally regulated [16, 23]. The sequence of signals leading to nitric oxide production begins with an infection that induces immune cells to release cytokines, which in turn signal macrophages to make nitric oxide synthase (Figure 3). For example, a viral infection induces CD4+ helper T lymphocytes or natural killer lymphocytes to produce interferon-γ, or lipopolysaccharide, from the cell wall of infecting bacteria cause cells to make tumor necrosis factor-α. Specific combinations of cytokines, such as interferon-γ along with either tumor necrosis factor-α or interleukin-1 β, activate intracellular transcription factors inside macrophages, such as nuclear factor-κ B, to initiate production of nitric oxide synthase [129, 130]. Nuclear factor-κ B itself can be activated by oxygen radicals, suggesting a novel positive feedback loop in which nitric oxide might activate transcription factors, which in turn could increase nitric oxide synthesis by causing additional nitric oxide synthase expression [131]. Once synthesized, macrophage nitric oxide synthase produces nitric oxide continuously. In fact, macrophages activated in culture can make nitric oxide synthase and produce nitric oxide for days, but macrophages activated in vivo must eventually curtail nitric oxide production to avoid damaging the host. This is probably achieved by other inhibitory cytokines, such as transforming growth factor-β [132-136] and mRNA-destabilizing sequences [12, 137].

        Figure 3. Lipopolysaccharide (LPS) is released from the wall of infecting bacteria, causing host cells to release tumor necrosis factor-α (TNF-α). Tumor necrosis factor-α binds to macrophages and activates intracellular proteins such as nuclear factor-κ B (NF- κ B), which induce the transcription of nitric oxide synthase. Once synthesized, macrophage nitric oxide synthase continuously makes nitric oxide, which can kill bacteria. MAC-NOS = macrophage nitric oxide synthase.
        View larger version:
          Figure 3. Lipopolysaccharide (LPS) is released from the wall of infecting bacteria, causing host cells to release tumor necrosis factor-α (TNF-α). Tumor necrosis factor-α binds to macrophages and activates intracellular proteins such as nuclear factor-κ B (NF- κ B), which induce the transcription of nitric oxide synthase. Once synthesized, macrophage nitric oxide synthase continuously makes nitric oxide, which can kill bacteria. MAC-NOS = macrophage nitric oxide synthase. Induction of nitric oxide synthase in macrophages.

          Nitric Oxide in the Human Immune System

          The role of nitric oxide in the immune system is better defined in animals than in humans. Although nitric oxide synthase can be induced easily in murine macrophages (and also in murine neutrophils, hepatocytes, fibroblasts, glial cells, smooth muscle cells, and endothelial cells [138-144]), it cannot be induced reproducibly in human macrophages. However, nitric oxide synthase can be induced in human hepatocytes in culture. The best evidence to show that humans make inducible nitric oxide synthase comes from clinical trials in which patients who were given intravenous interleukin-2 excreted nitric oxide byproducts in their urine [145, 146]. Perhaps inhibitory cytokines, such as transforming growth factor-β, are present more often in humans than in animals, thus preventing the widespread induction of nitric oxide synthase. However, because nitric oxide synthase can be induced in so many different types of animal cells, every human cell might also induce nitric oxide synthase given the correct set of cytokines.

          The Role of Nitric Oxide in Mediating Septic Hypotension

          We would expect excessive amounts of nitric oxide to cause hypotension, and it appears to be responsible for the hypotension associated with sepsis. During bacterial infections, the lipopolysaccharide component released from bacterial walls stimulates production of the inducible isoform of nitric oxide synthase in many cells, including macrophages, hepatocytes, and endothelial cells. Release of massive amounts of nitric oxide into the blood stream overwhelms the arterial smooth muscle and causes excess dilation and hypotension. Treatment of sepsis in animals with inhibitors of nitric oxide synthase reverses hypotension within minutes [147-149]. A few patients with sepsis have been treated with nitric oxide synthase inhibitors. Their mean arterial pressure increased from about 50 to 90 mm Hg within minutes as their systemic vascular resistance doubled [150, 151], which is consistent with the hypothesis that excessive nitric oxide causes hypotension during sepsis. However, the patients died soon thereafter, despite treatment, because the course of sepsis clearly involves much more than nitric oxide-induced hypotension.

          Immunodeficiency syndromes in which patients cannot produce nitric oxide have not been identified. This is in contrast to chronic granulomatous disease, in which patients are susceptible to bacterial infections because they cannot produce another molecule with an unpaired electron, superoxide [152, 153]. One case report identified high levels of an arginine analog in the serum of a patient with renal failure [154]. Perhaps an excess of this compound inhibits nitric oxide synthase, causing the hypertension and immunodeficiency often seen with chronic renal failure. It is tempting to speculate that the immune system of the malnourished patient is compromised in part because of an inability to produce nitric oxide. Arginine, a substrate of nitric oxide synthase, is an essential amino acid, and a diet lacking arginine might reduce nitric oxide synthesis. Nitric oxide suppresses M. tuberculosis and M. leprae, intracellular pathogens that especially affect malnourished patients. Determination of how diet affects intracellular arginine concentrations, coupled with observation of the effect of arginine-deficient diets on mice infected with M. tuberculosis, might lead to the treatment of malnourished patients with tuberculosis with a simple diet rich in arginine.

          The role of nitric oxide in the immune system is unique, because nitric oxide is nonspecific and damages any cell or pathogen. In contrast, antibodies or cytotoxic T lymphocytes act by first recognizing specific pathogens or infected cells and then destroying them. Furthermore, recent work suggests that only one inducible nitric oxide synthase isoform exists [155] and that it can be induced in many, if not all, cells. Thus, nitric oxide functions as a widespread, nonspecific immune system by itself, and it may have developed as a primitive immune system before the complex and specific interactions of lymphocytes evolved.

          Previous SectionNext Section

          Nitric Oxide as a Neurotransmitter

          Abundant evidence indicates that nitric oxide is a neurotransmitter in the peripheral and central nervous systems. Immunohistochemical studies have localized nitric oxide synthase to discrete neuronal populations, such as those in the myenteric plexus throughout the gastrointestinal tract [156-158]. Electric stimulation of gut segments produces relaxation corresponding to the relaxant phase of peristalsis and associated with nitric oxide release. Nitric oxide synthase inhibitors block this relaxation, indicating that nitric oxide is the neurotransmitter of these neurons [159, 160]. Nitric oxide released from neurons in the gastrointestinal tract causes gastric relaxation in response to a food bolus, as well as relaxation of the ileocolonic junction and anal sphincter [161-163]. Pelvic plexus neurons contain nitric oxide synthase, and nitric oxide is a mediator of penile erection [164]. Nitric oxide synthase neurons also innervate the adventitia of blood vessels [158]. Thus, nitric oxide is a neurotransmitter in the peripheral nervous system.

          In the brain, nitric oxide synthase occurs in discrete neuronal groups [158], but the specific pathways in which it is involved are unclear. Because nitric oxide synthase neurons are found in the cerebellum, nitric oxide might be involved in coordination and balance. Nitric oxide synthase neurons are also found in the accessory olfactory bulb, where they may cause olfactory signal processing.

          Nitric Oxide and the Transmission of Signals between Neurons

          Neurons release nitric oxide, which diffuses into adjacent neurons in a series of steps. The presynaptic neuron is triggered by glutamate binding to the n-methyl-d-aspartate subtype receptor. This receptor possesses a calcium channel that opens, and the resulting influx of calcium binds to calmodulin to activate neuronal nitric oxide synthase. Nitric oxide is produced and diffuses out of the presynaptic neuron into the postsynaptic neuron, where it binds to the heme group of guanylate cyclase, activating the enzyme to produce cGMP. Given physiologic conditions, small amounts of nitric oxide allow glutamate to increase cGMP levels in the brain. However, the massive release of glutamate during stroke triggers formation of large amounts of nitric oxide that are neurotoxic to adjacent neurons.

          The neurons in the central nervous system that contain nitric oxide synthase stain with the dye nitroblue tetrazolium in the presence of NADPH and were designated NADPH-diaphorase neurons [156, 165-167]. These cells are selectively resistant to the neurologic damage of stroke and to neurodegenerative conditions such as Huntington and Alzheimer diseases [168]. These nitric oxide synthase neurons appear to mediate neurotoxicity in vascular stroke [169]. Strokes are associated with a massive release of the excitatory transmitter glutamate. Large doses of glutamate and its derivative n-methyl-d-aspartate cause neurologic damage comparable to that occurring in stroke [170]. Further, drugs that block the n-methyl-d-aspartate subtype of the glutamate receptor alleviate stroke damage even when administered after occluding cerebral arteries. Nitric oxide appears to mediate this neurotoxicity, because nitric oxide synthase inhibitors block neurotoxic effects of glutamate in tissue culture and reduce stroke damage in mice, rats, and cats after cerebral artery ligation [171, 172]. Because nitric oxide synthase inhibition also constricts blood vessels, the resulting cerebral ischemia may counteract the beneficial effects of nitric oxide synthase inhibition, perhaps accounting for the failure of very large doses of nitric oxide synthase inhibitors to decrease stroke damage in some animal studies [173].

          Nitric Oxide as a Factor in Penile Erection

          The mechanism of penile erection is a good example of nitric oxide functioning both as a neurotransmitter and as a vasodilator. Penile erection results when cavernosal and pelvic nerves induce vasodilation of arteries that supply the corpora cavernosa, but until recently the neurotransmitter causing erection was unknown. Acetylcholine and norepinephrine were ruled out as neurotransmitters of this nonadrenergic, noncholinergic system. The cavernous nerves contain neuropeptides, such as vasoactive intestinal peptide, but evidence of this peptide's role in penile erection is minimal.

          Several lines of evidence show that nitric oxide transmits the neurons responsible for erection. First, isolated strips of smooth muscle from the corpora relax when electrically stimulated; this relaxation is blocked by nitric oxide synthase inhibitors [174-178]. Next, immunohistochemical studies show that nitric oxide synthase is localized to nerves in the pelvic plexus, the cavernous nerve, and processes extending into the corpus cavernosa, innervating the adventitial layers of penile arteries [164]. Finally, electric stimulation of the pelvic nerves of live rats causes a physiologic erection that can be blocked completely by intravenous nitric oxide synthase inhibitors [164].

          This mechanism of erection involves nitric oxide both as a neurotransmitter and as a vasodilator. After a neural stimulus for erection is introduced, nerves from the pelvic plexus synthesize and release nitric oxide into the corpus cavernosum. The smooth muscle of the corpus cavernosum relaxes in response, and the corpora fill with blood and swell, producing an erection. Whether nitric oxide synthase is involved in erectile diseases is unknown. A lack of nitric oxide might cause impotence, and an excess could lead to priapism.

          Previous SectionNext Section

          Nitric Oxide Therapy

          Although nitrates have been prescribed to treat angina pectoris for more than a century, it was not until recently that their mechanism of action was understood. Nitrates form nitric oxide, which causes direct vasodilation of the coronary arteries. Some nitrates, such as nitroprusside, spontaneously decompose to release nitric oxide; others, such as nitroglycerin and isosorbide dinitrate, are metabolized by cells into nitric oxide.

          Nitrates have been used as vasodilators to treat hypertension, angina, hypertensive crises, and other emergencies requiring immediate afterload reduction. Inhaled nitric oxide relaxes pulmonary arteries, reducing pulmonary vascular resistance, and it is metabolized swiftly so that systemic vascular resistance is not affected. It has been used to treat pulmonary hypertension of neonates by dilating pulmonary arteries [179]. It has also been used to treat the adult respiratory distress syndrome by reducing pulmonary artery pressure and intrapulmonary shunting [180]. Nitric oxide reduces the size of myocardial infarctions in animals [50, 181], although whether it does this by vasodilation, by reducing inflammatory cell infiltration, or by removing other radical molecules is unknown. It is even possible that nitric oxide delivered in a stable preparation directly to the corpus cavernosum could induce erection, thereby treating impotence. Perhaps inhaled nitric oxide could be used as a bronchodilator to treat asthma or chronic obstructive pulmonary disease by directly relaxing constricted bronchial smooth muscle.

          Nitric oxide synthase inhibitors successfully treat the hypotension of sepsis, although they do not reduce mortality. Whether inhibitors can treat other conditions involving an excess of nitric oxide is unresolved. Nitric oxide synthase inhibitors reduce the infarction size of strokes in some animal studies but not in others [171-173]. However, the widespread use of nitric oxide synthase inhibitors is limited because nitric oxide is involved in many different physiologic functions. Blocking nitric oxide synthesis from neuronal nitric oxide synthase to reduce the severity of stroke might also block nitric oxide from endothelial nitric oxide synthase, leading to hypertension and clotting. The commonly used nitric oxide synthase inhibitors are compounds related to the nitric oxide synthase substrate arginine, such as n-mono-methyl-arginine or n-nitro-arginine methyl ester. These compounds nonselectively block all nitric oxide synthase isoforms. Widespread treatment of patients with nitric oxide synthase inhibitors awaits development of compounds that inhibit specific nitric oxide synthase isoforms.

          Previous SectionNext Section

          Conclusion

          In 1987, only 7 articles were written about endogenously produced nitric oxide, but since then more than 3000 have been published. Nitric oxide, once considered a noxious byproduct of combustion, has been shown to be an endogenous messenger that plays various physiologic roles. As this explosion in research continues, the molecular targets of nitric oxide will be more precisely defined, and perhaps novel methods by which it regulates enzymes, such as amino acid nitrosylation, will be discovered. Research into the regulation of nitric oxide synthase should lead to a better understanding of its role in the pathogenesis of various diseases. The development of nitric oxide synthase inhibitors to block specific isoforms of nitric oxide as well as stable compounds that release it is also likely. Finally, the study of nitric oxide may lead to an understanding of how other small molecules, such as carbon monoxide, serve as biological messengers.

          Previous SectionNext Section

          Abbreviations

          ADP: adenosine diphosphate

          cGMP: cyclic guanosine monophosphate

          NADPH: nicotinamide adenine dinucleotide phosphate

          Previous Section
           

          References

          1. 1.↵
            Lowenstein CJ, Snyder SH. Nitric oxide, a novel biologic messenger. Cell. 1992; 70:705-7.
          2. 2.↵
            Moncada S, Palmer RM, Higgs EA. Nitric oxide: physiology, pathophysiology, and pharmacology. Pharmacol Rev. 1991; 43:109-42.
          3. 3.↵
            Nathan C. Nitric oxide as a secretory product of mammalian cells. FASEB J. 1992; 6:3051-64.
          4. 4.↵
            Luscher TF. Endogenous and exogenous nitrates and their role in myocardial ischaemia. Br J Clin Pharmacol. 1992; 34(Suppl 1):29S-35S.
          5. 5.↵
            Marletta MA. Nitric oxide: biosynthesis and biological significance. Trends Biochem Sci. 1989; 14:488-92.
          6. 6.↵
            Ignarro LJ. Biosynthesis and metabolism of endothelium-derived nitric oxide. Annu Rev Pharmacol Toxicol. 1990; 30:535-60.
          7. 7.↵
            Luscher TF. Endothelial control of vascular tone and growth. Clin Exp Hypertens (A). 1990; 12:897-902.
          8. 8.↵
            Stamler JS, Singel DJ, Loscalzo J. Biochemistry of nitric oxide and its redox-activated forms. Science. 1992; 258:1898-902.
          9. 9.↵
            White KA, Marletta MA. Nitric oxide synthase is a cytochrome P-450 type hemoprotein. Biochemistry. 1992; 31:6627-31.
          10. 10.↵
            McMillan K, Bredt DS, Hirsch DJ, Snyder SH, Clark JE, Masters BS. Cloned, expressed rat cerebellar nitric oxide synthase contains stoichiometric amounts of heme, which binds carbon monoxide. Proc Natl Acad Sci U S A. 1992; 89:11141-5.
          11. 11.↵
            Bredt DS, Hwang PM, Glatt CE, Lowenstein C, Reed RR, Snyder SH. Cloned and expressed nitric oxide synthase structurally resembles cytochrome P-450 reductase. Nature. 1991; 351:714-8.
          12. 12.↵
            Lowenstein CJ, Glatt CS, Bredt DS, Snyder SH. Cloned and expressed macrophage nitric oxide synthase contrasts with the brain enzyme. Proc Natl Acad Sci U S A. 1992; 89:6711-5.
          13. 13.↵
            Lamas S, Marsden PA, Li GK, Tempst P, Michel T. Endothelial nitric oxide synthase: molecular cloning and characterization of a distinct constitutive enzyme isoform. Proc Natl Acad Sci U S A. 1992; 89:6348-52.
          14. 14.↵
            Janssens SP, Shimouchi A, Quertermous T, Bloch DB, Bloch KD. Cloning and expression of a cDNA encoding human endothelium-derived relaxing factor/nitric oxide synthase. J Biol Chem. 1992; 267:14519-22.
          15. 15.↵
            Sessa WC, Harrison JK, Barber CM, Zeng D, Durieux ME, D'Angelo DD, et al. Molecular cloning and expression of a cDNA encoding endothelial cell nitric oxide synthase. J Biol Chem. 1992; 267:15274-6.
          16. 16.↵
            Xie QW, Cho HJ, Calaycay J, Mumford RA, Swiderek KM, Lee TD, et al. Cloning and characterization of inducible nitric oxide synthase from mouse macrophages. Science. 1992; 256:225-8.
          17. 17.↵
            Lyons CR, Orloff GJ, Cunningham JM. Molecular cloning and functional expression of an inducible nitric oxide synthase from a murine macrophage cell line. J Biol Chem. 1992; 267:6370-4.
          18. 18.↵
            Marsden PA, Schappert KT, Chen HS, Flowers M, Sundell CL, Wilcox JN, et al. Molecular cloning and characterization of human endothelial nitric oxide synthase. FEBS Lett. 1992; 307:287-93.
          19. 19.↵
            Bredt DS, Snyder SH. Isolation of nitric oxide synthetase, a calmodulin-requiring enzyme. Proc Natl Acad Sci U S A. 1990; 87: 682-5.
          20. 20.↵
            Long CJ, Stone TW. The release of endothelium-derived relaxant factor is calcium dependent. Blood Vessels. 1985; 22:205-8.
          21. 21.↵
            Busse R, Mulsch A. Calcium-dependent nitric oxide synthesis in endothelial cytosol is mediated by calmodulin. FEBS Lett. 1990; 265:133-6.
          22. 22.↵
            Flavahan NA. Atherosclerosis or lipoprotein-induced endothelial dysfunction. Potential mechanisms underlying reduction in EDRF/nitric oxide activity. Circulation. 1992; 85:1927-38.
          23. 23.↵
            Lorsbach RB, Russell SW. A specific sequence of stimulation is required to induce synthesis of the antimicrobial molecule nitric oxide by mouse macrophages. Infect Immun. 1992; 60:2133-5.
          24. 24.↵
            Bredt DS, Ferris CD, Snyder SH. Nitric oxide synthase regulatory sites. Phosphorylation by cyclic AMP-dependent protein kinase, protein kinase C, and calcium/calmodulin protein kinase; identification of flavin and calmodulin binding sites. J Biol Chem. 1992; 267:10976-81.
          25. 25.↵
            Michel T, Li GK, Busconi L. Phosphorylation and subcellular translocation of endothelial nitric oxide synthase. Proc Natl Acad Sci U S A. 1993; 90:6252-6.
          26. 26.↵
            Cho HJ, Xie QW, Calaycay J, Mumford RA, Swiderek KM, Lee TD, et al. Calmodulin is a subunit of nitric oxide synthase from macrophages. J Exp Med. 1992; 176:599-604.
          27. 27.↵
            Brune B, Lapetina EG. Activation of a cytosolic ADP-ribosyltransferase by nitric oxide-generating agents. J Biol Chem. 1989; 264: 8455-8.
          28. 28.↵
            Brune B, Lapetina EG. Properties of a novel nitric oxide-stimulated ADP-ribosyltransferase. Arch Biochem Biophys. 1990; 279: 286-90.
          29. 29.↵
            Zhang J, Snyder SH. Purification of a nitric oxide-stimulated ADP-ribosylated protein using biotinylated β-nicotinamide adenine dinucleotide. Biochemistry. 1993; 32:2228-33.
          30. 30.↵
            Zhang J, Snyder SH. Nitric oxide stimulates auto-ADP-ribosylation of glyceraldehyde-3-phosphate dehydrogenase. Proc Natl Acad Sci U S A. 1992; 89:9382-5.
          31. 31.↵
            Dimmeler S, Brune B. Characterization of a nitric-oxide-catalysed ADP-ribosylation of glyceraldehyde-3-phosphate dehydrogenase. Eur J Biochem. 1992; 210:305-10.
          32. 32.↵
            Kots AY, Skurat AV, Sergienko EA, Bulargina TV, Severin ES. Nitroprusside stimulates the cysteine-specific mono(ADP-ribosylation) of glyceraldehyde-3-phosphate dehydrogenase from human erythrocytes. FEBS Lett. 1992; 300:9-12.
          33. 33.↵
            Furchgott RF, Zawadzki JV. The obligatory role of endothelial cells in the relaxation of arterial smooth muscle by acetylcholine. Nature. 1980; 288:373-6.
          34. 34.↵
            Palmer RM, Ferrige AG, Moncada S. Nitric oxide release accounts for the biological activity of endothelium-derived relaxing factor. Nature. 1987; 327:524-6.
          35. 35.↵
            Ikeda K, Gutierrez OG Jr, Yamori Y. Dietary NG-nitro-L-arginine induces sustained hypertension in normotensive Wistar-Kyoto rats. Clin Exp Pharmacol Physiol. 1992; 19:583-6.
          36. 36.↵
            Baylis C, Mitruka B, Deng A. Chronic blockade of nitric oxide synthesis in the rat produces systemic hypertension and glomerular damage. J Clin Invest. 1992; 90:278-81.
          37. 37.↵
            Kobayashi Y, Ikeda K, Shinozuka K, Nara Y, Yamori Y, Hattori K. L-nitroarginine increases blood pressure in the rat. Clin Exp Pharmacol Physiol. 1991; 18:397-9.
          38. 38.↵
            Ribeiro MO, Antunes E, de Nucci G, Lovisolo SM, Zatz R. Chronic inhibition of nitric oxide synthesis. A new model of arterial hypertension. Hypertension. 1992; 20:298-303.
          39. 39.↵
            Rees DD, Palmer RM, Moncada S. Role of endothelium-derived nitric oxide in the regulation of blood pressure. Proc Natl Acad Sci U S A. 1989; 86:3375-8.
          40. 40.↵
            Elsner D, Muntze A, Kromer EP, Riegger GA. Systemic vasoconstriction induced by inhibition of nitric oxide synthesis is attenuated in conscious dogs with heart failure. Cardiovasc Res. 1991; 25:438-40.
          41. 41.↵
            Klabunde RE, Ritger RC, Helgren MC. Cardiovascular actions of inhibitors of endothelium-derived relaxing factor (nitric oxide) formation/release in anesthetized dogs. Eur J Pharmacol. 1991; 199: 51-9.
          42. 42.↵
            Baumann JE, Persson PB, Ehmke H, Nafz B, Kirchheim HR. Role of endothelium-derived relaxing factor in renal autoregulation in conscious dogs. Am J Physiol. 1992; 263:F208-13.
          43. 43.↵
            Yukimura T, Yamashita Y, Miura K, Okumura M, Yamanaka S, Yamamoto K. Renal effects of the nitric oxide synthase inhibitor, L-NG-nitroarginine, in dogs. Am J Hypertens. 1992; 5:484-7.
          44. 44.↵
            Aisaka K, Gross SS, Griffith OW, Levi R. NG-methylarginine, an inhibitor of endothelium-derived nitric oxide synthesis, is a potent pressor agent in the guinea pig: does nitric oxide regulate blood pressure in vivo? Biochem Biophys Res Commun. 1989; 160:881-6.
          45. 45.↵
            Vallance P, Collier J, Moncada S. Effects of endothelium-derived nitric oxide on peripheral arteriolar tone in man (see comments). Lancet. 1989; 2:997-1000.
          46. 46.↵
            Wennmalm A, Lanne B, Petersson AS. Detection of endothelial-derived relaxing factor in human plasma in the basal state and following ischemia using electron paramagnetic resonance spectrometry. Anal Biochem. 1990; 187:359-63.
          47. 47.↵
            Schror K, Woditsch I, Strobach H, Schroder H. Interactions between nitric oxide and prostacyclin in myocardial ischemia and endothelial cell cultures. Basic Res Cardiol. 1991; 86(Suppl 2):117-25.
          48. 48.↵
            Matheis G, Sherman MP, Buckberg GD, Haybron DM, Young HH, Ignarro LJ. Role of L-arginine-nitric oxide pathway in myocardial reoxygenation injury. Am J Physiol. 1992; 262:H616-20.
          49. 49.↵
            Carey C, Siegfried MR, Ma XL, Weyrich AS, Lefer AM. Antishock and endothelial protective actions of a NO donor in mesenteric ischemia and reperfusion. Circ Shock. 1992; 38:209-16.
          50. 50.↵
            Siegfried MR, Erhardt J, Rider T, Ma XL, Lefer AM. Cardioprotection and attenuation of endothelial dysfunction by organic nitric oxide donors in myocardial ischemia-reperfusion. J Pharmacol Exp Ther. 1992; 260:668-75.
          51. 51.↵
            Ma XL, Weyrich AS, Lefer DJ, Lefer AM. Diminished basal nitric oxide release after myocardial ischemia and reperfusion promotes neutrophil adherence to coronary endothelium. Circ Res. 1993; 72: 403-12.
          52. 52.↵
            Buga GM, Gold ME, Fukuto JM, Ignarro LJ. Shear stress-induced release of nitric oxide from endothelial cells grown on beads. Hypertension. 1991; 17:187-93.
          53. 53.↵
            Kelm M, Feelisch M, Deussen A, Strauer BE, Schrader J. Release of endothelium derived nitric oxide in relation to pressure and flow. Cardiovasc Res. 1991; 25:831-6.
          54. 54.↵
            Ueeda M, Silvia SK, Olsson RA. Nitric oxide modulates coronary autoregulation in the guinea pig. Circ Res. 1992; 70:1296-303.
          55. 55.↵
            Lamontagne D, Pohl U, Busse R. Mechanical deformation of vessel wall and shear stress determine the basal release of endothelium-derived relaxing factor in the intact rabbit coronary vascular bed. Circ Res. 1992; 70:123-30.
          56. 56.↵
            Pohl U, Herlan K, Huang A, Bassenge E. EDRF-mediated shear-induced dilation opposes myogenic vasoconstriction in small rabbit arteries. Am J Physiol. 1991; 261:H2016-23.
          57. 57.↵
            Rubanyi GM, Romero JC, Vanhoutte PM. Flow-induced release of endothelium-derived relaxing factor. Am J Physiol. 1986; 250: H1145-9.
          58. 58.↵
            Pohl U, Holtz J, Busse R, Bassenge E. Crucial role of endothelium in the vasodilator response to increased flow in vivo. Hypertension. 1986; 8:37-44.
          59. 59.↵
            Chu A, Chambers DE, Lin CC, Kuehl WD, Cobb FR. Nitric oxide modulates epicardial coronary basal vasomotor tone in awake dogs. Am J Physiol. 1990; 258:H1250-4.
          60. 60.↵
            Faraci FM, Breese KR. Nitric oxide mediates vasodilatation in response to activation of N-methyl-D-aspartate receptors in brain. Circ Res. 1993; 72:476-80.
          61. 61.↵
            Tanaka K, Gotoh F, Gomi S, Takashima S, Mihara B, Shirai, et al. Inhibition of nitric oxide synthesis induces a significant reduction in local cerebral blood flow in the rat. Neurosci Lett. 1991; 127: 129-32.
          62. 62.↵
            Toda N, Okamura T. Mechanism underlying the response to vasodilator nerve stimulation in isolated dog and monkey cerebral arteries. Am J Physiol. 1990; 259:H1511-7.
          63. 63.↵
            Broten TP, Miyashiro JK, Moncada S, Feigl EO. Role of endothelium-derived relaxing factor in parasympathetic coronary vasodilation. Am J Physiol. 1992; 262:H1579-84.
          64. 64.↵
            Richard V, Berdeaux A, la Rochelle CD, Giudicelli JF. Regional coronary haemodynamic effects of two inhibitors of nitric oxide synthesis in anaesthetized, open-chest dogs. Br J Pharmacol. 1991; 104:59-64.
          65. 65.↵
            Benyo Z, Kiss G, Szabo C, Csaki C, Kovach AG. Importance of basal nitric oxide synthesis in regulation of myocardial blood flow. Cardiovasc Res. 1991; 25:700-3.
          66. 66.↵
            Amezcua JL, Palmer RM, de Souza BM, Moncada S. Nitric oxide synthesized from L-arginine regulates vascular tone in the coronary circulation of the rabbit. Br J Pharmacol. 1989; 97:1119-24.
          67. 67.↵
            Jones LF, Brody MJ. Coronary blood flow in rats is dependent on the release of vascular nitric oxide. J Pharmacol Exp Ther. 1992; 260:627-31.
          68. 68.↵
            Fineman JR, Heymann MA, Soifer SJ. Nω-nitro-L-arginine attenuates endothelium-dependent pulmonary vasodilation in lambs. Am J Physiol. 1991; 260:H1299-306.
          69. 69.↵
            Iwata F, Joh T, Kawai T, Itoh M. Role of EDRF in splanchnic blood flow of normal and chronic portal hypertensive rats. Am J Physiol. 1992; 263:G149-54.
          70. 70.↵
            Deng A, Baylis C. Locally produced EDRF controls preglomerular resistance and ultrafiltration coefficient. Am J Physiol. 1993; 264: F212-5.
          71. 71.↵
            Nozaki K, Moskowitz MA, Maynard KI, Koketsu N, Dawson TM, Bredt DS, et al. Possible origins and distribution of immunoreactive nitric oxide synthase-containing nerve fibers in cerebral arteries. J Cereb Blood Flow Metab. 1993; 13:70-9.
          72. 72.↵
            Panza JA, Quyyumi AA, Brush JE Jr, Epstein SE. Abnormal endothelium-dependent vascular relaxation in patients with essential hypertension. N Engl J Med. 1990; 323:22-7.
          73. 73.↵
            Linder L, Kiowski W, Buhler FR, Luscher TF. Indirect evidence for release of endothelium-derived relaxing factor in human forearm circulation in vivo. Blunted response in essential hypertension. Circulation. 1990; 81:1762-7.
          74. 74.↵
            Panza JA, Casino PR, Kilcoyne CM, Quyyumi AA. Role of endothelium derived nitric oxide in the abnormal endothelium dependent vascular relaxation of patients with essential hypertension. Circulation. 1993; 87:1468-74.
          75. 75.↵
            Calver A, Collier J, Moncada S, Vallance P. Effect of local intra-arterial NG-monomethyl-L-arginine in patients with hypertension: the nitric oxide dilator mechanism appears abnormal. J Hypertens. 1992; 10:1025-31.
          76. 76.↵
            Iwama Y, Kato T, Muramatsu M, Asano H, Shimizu K, Toki Y, et al. Correlation with blood pressure of the acetylcholine-induced endothelium-derived contracting factor in the rat aorta. Hypertension. 1992; 19:326-32.
          77. 77.↵
            Jayakody L, Senaratne M, Thomson A, Kappagoda T. Endothelium-dependent relaxation in experimental atherosclerosis in the rabbit. Circ Res. 1987; 60:251-64.
          78. 78.↵
            Bossaller C, Habib GB, Yamamoto H, Williams C, Wells S, Henry PD. Impaired muscarinic endothelium-dependent relaxation and cyclic guanosine 5′-monophosphate formation in atherosclerotic human coronary artery and rabbit aorta. J Clin Invest. 1987; 79: 170-4.
          79. 79.↵
            Forstermann U, Mugge A, Alheid U, Haverich A, Frolich JC. Selective attenuation of endothelium-mediated vasodilation in atherosclerotic human coronary arteries. Circ Res. 1988; 62:185-90.
          80. 80.↵
            Ludmer PL, Selwyn AP, Shook TL, Wayne RR, Mudge GH, Alexander RW, et al. Paradoxical vasoconstriction induced by acetylcholine in atherosclerotic coronary arteries. N Engl J Med. 1986; 315:1046-51.
          81. 81.↵
            Faraci FM, Heistad DD. Endothelium-derived relaxing factor inhibits constrictor responses of large cerebral arteries to serotonin. J Cereb Blood Flow Metab. 1992; 12:500-6.
          82. 82.↵
            Menon NK, Bing RJ. Nitroarginine does not inhibit lysophosphatidylcholine (LPC)-induced vascular relaxation and accumulation of cyclic GMP. Proc Soc Exp Biol Med. 1991; 196:461-3.
          83. 83.↵
            Menon NK, Wolf A, Zehetgruber M, Bing RJ. An improved chemiluminescence assay suggests non-nitric oxide-mediated action of lysophosphatidylcholine and acetylcholine. Proc Soc Exp Biol Med. 1989; 191:316-9.
          84. 84.↵
            Saito T, Wolf A, Menon NK, Saeed M, Alves C, Bing RJ. Lysolecithins as endothelium-dependent vascular smooth muscle relaxants that differ from endothelium-derived relaxing factor (nitric oxide). Proc Natl Acad Sci U S A. 1988; 85:8246-50.
          85. 85.↵
            Flavahan NA. Lysophosphatidylcholine modifies G protein-dependent signaling in porcine endothelial cells. Am J Physiol. 1993; 264: H722-7.
          86. 86.↵
            Sieber CC, Lopez-Talavera JC, Groszmann RJ. Role of nitric oxide in the in vitro splanchnic vascular hyporeactivity in ascitic cirrhotic rats. Gastroenterology. 1993; 104:1750-4.
          87. 87.↵
            Castro A, Jimenez W, Claria J, Ros J, Martinez JM, Bosch M, et al. Impaired responsiveness to angiotensin II in experimental cirrhosis: role of nitric oxide. Hepatology. 1993; 18:367-72.
          88. 88.↵
            Sogni P, Moreau R, Ohsuga M, Cailmail S, Oberti F, Hadengue A, et al. Evidence for normal nitric oxide-mediated vasodilator tone in conscious rats with cirrhosis. Hepatology. 1992; 16:980-3.
          89. 89.↵
            Pizcueta P, Pique JM, Fernandez M, Bosch J, Rodes J, Whittle BJ, et al. Modulation of the hyperdynamic circulation of cirrhotic rats by nitric oxide inhibition. Gastroenterology. 1992; 103:1909-15.
          90. 90.↵
            Radomski MW, Palmer RM, Moncada S. Characterization of the L-arginine:nitric oxide pathway in human platelets. Br J Pharmacol. 1990; 101:325-8.
          91. 91.↵
            Radomski MW, Palmer RM, Moncada S. An L-arginine/nitric oxide pathway present in human platelets regulates aggregation. Proc Natl Acad Sci U S A. 1990; 87:5193-7.
          92. 92.↵
            Varela AF, Runge A, Ignarro LJ, Chaudhuri G. Nitric oxide and prostacyclin inhibit fetal platelet aggregation: a response similar to that observed in adults. Am J Obstet Gynecol. 1992; 167:1599-604.
          93. 93.↵
            Ivanova K, Schaefer M, Drummer C, Gerzer R. Effects of nitric oxide-containing compounds on increases in cytosolic ionized Ca2+ and on aggregation of human platelets. Eur J Pharmacol. 1993; 244:37-47.
          94. 94.↵
            Vallance P, Benjamin N, Collier J. The effect of endothelium-derived nitric oxide on ex vivo whole blood platelet aggregation in man. Eur J Clin Pharmacol. 1992; 42:37-41.
          95. 95.↵
            Pasqui AL, Capecchi PL, Ceccatelli L, Mazza S, Gistri A, Laghi Pasini F, et al. Nitroprusside in vitro inhibits platelet aggregation and intracellular calcium translocation. Effect of haemoglobin. Thromb Res. 1991; 61:113-22.
          96. 96.↵
            Sneddon JM, Vane JR. Endothelium-derived relaxing factor reduces platelet adhesion to bovine endothelial cells. Proc Natl Acad Sci U S A. 1988; 85:2800-04.
          97. 97.↵
            Radomski MW, Palmer RM, Moncada S. The anti-aggregating properties of vascular endothelium: interactions between prostacyclin and nitric oxide. Br J Pharmacol. 1987; 92:639-46.
          98. 98.↵
            Radomski MW, Palmer RM, Moncada S. Endogenous nitric oxide inhibits human platelet adhesion to vascular endothelium. Lancet. 1987; 2:1057-8.
          99. 99.↵
            Radomski MW, Palmer RM, Moncada S. The role of nitric oxide and cGMP in platelet adhesion to vascular endothelium. Biochem Biophys Res Commun. 1987; 148:1482-9.
          100. 100.↵
            Siney L, Lewis MJ. Endothelium-derived relaxing factor inhibits platelet adhesion to cultured porcine endocardial endothelium. Eur J Pharmacol. 1992; 229:223-6.
          101. 101.↵
            de Graaf JC, Banga JD, Moncada S, Palmer RM, de Groot PG, Sixma JJ. Nitric oxide functions as an inhibitor of platelet adhesion under flow conditions. Circulation. 1992; 85:2284-90.
          102. 102.↵
            Nolte C, Eigenthaler M, Schanzenbacher P, Walter U. Endothelial cell-dependent phosphorylation of a platelet protein mediated by cAMP- and cGMP-elevating factors. J Biol Chem. 1991; 266:14808-12.
          103. 103.↵
            Yamakado T, Nishikawa M, Hidaka H. Stimulation of human platelet guanylate cyclase by nitroso compounds. Thromb Res. 1982; 26:135-40.
          104. 104.↵
            Bowen R, Haslam RJ. Effects of nitrovasodilators on platelet cyclic nucleotide levels in rabbit blood; role for cyclic AMP in synergistic inhibition of platelet function by SIN-1 and prostaglandin E1. J Cardiovasc Pharmacol. 1991; 17:424-33.
          105. 105.↵
            Maurice DH, Haslam RJ. Molecular basis of the synergistic inhibition of platelet function by nitrovasodilators and activators of adenylate cyclase: inhibition of cyclic AMP breakdown by cyclic GMP. Mol Pharmacol. 1990; 37:671-81.
          106. 106.↵
            Gryglewski RJ, Korbut R, Kalecinska A, Zembowicz A. Interaction between stimulators of adenylate and guanylate cyclases in human leukocytes, platelets and arteries. Int J Tissue React. 1989; 11:269-75.
          107. 107.↵
            Durante W, Kroll MH, Vanhoutte PM, Schafer AI. Endothelium-derived relaxing factor inhibits thrombin-induced platelet aggregation by inhibiting platelet phospholipase C. Blood. 1992; 79:110-6.
          108. 108.↵
            Brune B, Molina y Vedia L, Lapetina EG. Agonist-induced ADP-ribosylation of a cytosolic protein in human platelets. Proc Natl Acad Sci U S A. 1990; 87:3304-8.
          109. 109.↵
            Fortier AH, Polsinelli T, Green SJ, Nacy CA. Activation of macrophages for destruction of Francisella tularensis: identification of cytokines, effector cells, and effector molecules. Infect Immun. 1992; 60:817-25.
          110. 110.↵
            Denis M. Interferon-γ-treated murine macrophages inhibit growth of tubercle bacilli via the generation of reactive nitrogen intermediates. Cell Immunol. 1991; 132:150-7.
          111. 111.↵
            Adams LB, Franzblau SG, Vavrin Z, Hibbs JB Jr, Krahenbuhl JL. L-arginine-dependent macrophage effector functions inhibit metabolic activity of Mycobacterium leprae. J Immunol. 1991; 147: 1642-6.
          112. 112.↵
            Beckerman KP, Rogers HW, Corbett JA, Schreiber RD, McDaniel ML, Unanue ER. Release of nitric oxide during the T cell-independent pathway of macrophage activation. Its role in resistance to Listeria monocytogenes. J Immunol. 1993; 150:888-95.
          113. 113.↵
            Mayer J, Woods ML, Vavrin Z, Hibbs JB Jr. Gamma interferon-induced nitric oxide production reduces Chlamydia trachomatis infectivity in McCoy cells. Infect Immun. 1993; 61:491-7.
          114. 114.↵
            Alspaugh JA, Granger DL. Inhibition of Cryptococcus neoformans replication by nitrogen oxides supports the role of these molecules as effectors of macrophage-mediated cytostasis. Infect Immun. 1991; 59:2291-6.
          115. 115.↵
            Liew FY, Li Y, Millott S. Tumour necrosis factor (TNF-α) in leishmaniasis. II. TNF-α-induced macrophage leishmanicidal activity is mediated by nitric oxide from L-arginine. Immunology. 1990; 71:556-9.
          116. 116.↵
            Mauel J, Ransijn A, Buchmuller-Rouiller Y. Killing of Leishmania parasites in activated murine macrophages is based on an L-arginine-dependent process that produces nitrogen derivatives. J Leukoc Biol. 1991; 49:73-82.
          117. 117.↵
            Green SJ, Meltzer MS, Hibbs JB Jr, Nacy CA. Activated macrophages destroy intracellular Leishmania major amastigotes by an L-arginine-dependent killing mechanism. J Immunol. 1990; 144:278-83.
          118. 118.↵
            Munoz-Fernandez MA, Fernandez MA, Fresno M. Synergism between tumor necrosis factor-α and interferon-γ on macrophage activation for the killing of intracellular Trypanosoma cruzi through a nitric oxide-dependent mechanism. Eur J Immunol. 1992; 22:301-7.
          119. 119.↵
            Gazzinelli RT, Oswald IP, James SL, Sher A. IL-10 inhibits parasite killing and nitrogen oxide production by IFN-γ-activated macrophages. J Immunol. 1992; 148:1792-6.
          120. 120.↵
            Vincendeau P, Daulouede S. Macrophage cytostatic effect on Trypanosoma musculi involves an L-arginine-dependent mechanism. J Immunol. 1991; 146:4338-43.
          121. 121.↵
            Lin JY, Chadee K. Macrophage cytotoxicity against Entamoeba histolytica trophozoites is mediated by nitric oxide from L-arginine. J Immunol. 1992; 148:3999-4005.
          122. 122.↵
            Rockett KA, Awburn MM, Aggarwal BB, Cowden WB, Clark IA. In vivo induction of nitrite and nitrate by tumor necrosis factor, lymphotoxin, and interleukin-1: possible roles in malaria. Infect Immun. 1992; 60:3725-30.
          123. 123.↵
            Croen KD. Evidence for antiviral effect of nitric oxide. Inhibition of herpes simplex virus type 1 replication. J Clin Invest. 1993; 91: 2446-52.
          124. 124.↵
            Karupiah G, Xie Q, Buller RML, Nathan C, Duarte C, MacMicking JD. Inhibition of viral replication by interferon-γ induced nitric oxide synthase. Science. 1993; 261:1445-8.
          125. 125.↵
            Hibbs JB Jr, Taintor RR, Vavrin Z, Rachlin EM. Nitric oxide: a cytotoxic activated macrophage effector molecule. Biochem Biophys Res Commun. 1988; 157:87-94.
          126. 126.↵
            Lepoivre M, Fieschi F, Coves J, Thelander L, Fontecave M. Inactivation of ribonucleotide reductase by nitric oxide. Biochem Biophys Res Commun. 1991; 179:442-8.
          127. 127.↵
            Kwon NS, Stuehr DJ, Nathan CF. Inhibition of tumor cell ribonucleotide reductase by macrophage derived nitric oxide. J Exp Med. 1991; 174:761-8.
          128. 128.↵
            Wink DA, Kasprzak KS, Maragos CM, Elespuru RK, Misra M, Dunams TM, et al. DNA deaminating ability and genotoxicity of nitric oxide and its progenitors. Science. 1991; 254:1001-3.
          129. 129.↵
            Lowenstein CJ, Alley E, Raval P, Snyder SH, Russel SW, Murphy W. Nitric oxide synthase gene: two upstream regions mediate its induction by interferon-γ and lipopolysaccharide. Proc Natl Acad Sci U S A. 1993; 90:9730-4.
          130. 130.↵
            Xie Q, Whisnant R, Nathan C. Promoter of the mouse gene encoding calcium-independent nitric oxide synthase confers inducibiltiy by interferon-γ and bacterial lipopolysaccharide. J Exp Med. 1993; 177:1779-84.
          131. 131.↵
            Schreck R, Rieber P, Baeuerle PA. Reactive oxygen intermediates as apparently widely used messengers in the activation of the NF-κ B transcription factor and HIV-1. EMBO J. 1991; 10: 2247-58.
          132. 132.↵
            Schini VB, Durante W, Elizondo E, Scott-Burden T, Junquero DC, Schafer AI, et al. The induction of nitric oxide synthase activity is inhibited by TGF-β 1, PDGFAB and PDGFBB in vascular smooth muscle cells. Eur J Pharmacol. 1992; 216:379-83.
          133. 133.↵
            Gazzinelli RT, Oswald IP, Hieny S, James SL, Sher A. The microbicidal activity of interferon-γ-treated macrophages against Trypanosoma cruzi involves an L-arginine-dependent, nitrogen oxide-mediated mechanism inhibitable by interleukin-10 and transforming growth factor-β. Eur J Immunol. 1992; 22:2501-6.
          134. 134.↵
            Junquero DC, Scott-Burden T, Schini VB, Vanhoutte PM. Inhibition of cytokine-induced nitric oxide production by transforming growth factor-β 1 in human smooth muscle cells. J Physiol (Lond). 1992; 454:451-65.
          135. 135.↵
            Nelson BJ, Ralph P, Green SJ, Nacy CA. Differential susceptibility of activated macrophage cytotoxic effector reactions to the suppressive effects of transforming growth factor-β 1. J Immunol. 1991; 146:1849-57.
          136. 136.↵
            Pfeilschifter J, Vosbeck K. Transforming growth factor β 2 inhibits interleukin 1 β- and tumour necrosis factor α-induction of nitric oxide synthase in rat renal mesangial cells. Biochem Biophys Res Commun. 1991; 175:372-9.
          137. 137.↵
            Geller DA, Lowenstein CJ, Shapiro RA, Nussler AK, Di Silvio M, Wang SC, et al. Molecular cloning and expression of inducible nitric oxide synthase from human hepatocytes. Proc Natl Acad Sci U S A. 1993; 90:3491-5.
          138. 138.↵
            Marletta MA, Yoon PS, Iyengar R, Leaf CD, Wishnok JS. Macrophage oxidation of L-arginine to nitrite and nitrate: nitric oxide is an intermediate. Biochemistry. 1988; 27:8706-11.
          139. 139.↵
            Nussler AK, Di Silvio M, Billiar TR, Hoffman RA, Geller DA, Selby R, et al. Stimulation of the nitric oxide synthase pathway in human hepatocytes by cytokines and endotoxin. J Exp Med. 1992; 176: 261-4.
          140. 140.↵
            Gross SS, Jaffe EA, Levi R, Kilbourn RG. Cytokine-activated endothelial cells express an isotype of nitric oxide synthase which is tetrahydrobiopterin-dependent, calmodulin-independent and inhibited by arginine analogs with a rank-order of potency characteristic of activated macrophages. Biochem Biophys Res Commun. 1991; 178:823-9.
          141. 141.↵
            Galea E, Feinstein DL, Reis DJ. Induction of calcium-independent nitric oxide synthase activity in primary rat glial cultures. Proc Natl Acad Sci U S A. 1992; 89:10945-9.
          142. 142.↵
            Schini VB, Junquero DC, Scott-Burden T, Vanhoutte PM. Interleukin-1 β induces the production of an L-arginine-derived relaxing factor from cultured smooth muscle cells from rat aorta. Biochem Biophys Res Commun. 1991; 176:114-21.
          143. 143.↵
            Busse R, Mulsch A. Induction of nitric oxide synthase by cytokines in vascular smooth muscle cells. FEBS Lett. 1990; 275:87-90.
          144. 144.↵
            Lot TY, Stark G, Wilson VG. Endothelium-dependent contractions to NG-nitro-L-arginine methyl ester in the porcine isolated splenic artery are sensitive to cyclooxygenase and lipoxygenase inhibitors. Naunyn Schmiedebergs Arch Pharmacol. 1993; 347:115-8.
          145. 145.↵
            Ochoa JB, Curti B, Peitzman AB, Simmons RL, Billiar RT, Hoffman R, et al. Increased circulating nitrogen oxides after human tumor immunotherapy: correlation with toxic hemodynamic changes. J Natl Cancer Inst. 1992; 84:864-7.
          146. 146.↵
            Hibbs JB Jr, Westenfelder C, Taintor R, Vavrin Z, Kablitz C, Baranowski RL, et al. Evidence for cytokine-inducible nitric oxide synthesis from L-arginine in patients receiving interleukin-2 therapy. J Clin Invest. 1992; 89:867-77.
          147. 147.↵
            Kilbourn RG, Gross SS, Jubran A, Adams J, Griffith OW, Levi R, et al. NG-methyl-L-arginine inhibits tumor necrosis factor-induced hypotension: implications for the involvement of nitric oxide. Proc Natl Acad Sci U S A. 1990; 87:3629-32.
          148. 148.↵
            Kilbourn RG, Jubran A, Gross SS, Griffith OW, Levi R, Adams J, et al. Reversal of endotoxin-mediated shock by NG-methyl-L-arginine, an inhibitor of nitric oxide synthesis. Biochem Biophys Res Commun. 1990; 172:1132-8.
          149. 149.↵
            Gryglewski RJ, Korbut R, Trabka-Janik E, Zembowicz A, Trybulec M. Interaction between NO donors and iloprost in human vascular smooth muscle, platelets, and leukocytes. J Cardiovasc Pharmacol. 1989; 14(Suppl 11):S124-8.
          150. 150.↵
            Hotchkiss RS, Karl IE, Parker JL, Adams HR. Inhibition of NO synthesis in septic shock. Lancet. 1992; 339:434-5.
          151. 151.↵
            Petros A, Bennett D, Vallance P. Effect of nitric oxide synthase inhibitors on hypotension in patients with septic shock. Lancet. 1991; 338:1557-8.
          152. 152.↵
            Curnutte JT, Babior BM. Chronic granulomatous disease. Adv Hum Genet. 1987; 16:229-97.
          153. 153.↵
            Curnutte JT, Whitten DM, Babior BM. Defective superoxide production by granulocytes from patients with chronic granulomatous disease. N Engl J Med. 1974; 290:593-7.
          154. 154.↵
            Vallance P, Leone A, Calver A, Collier J, Moncada S. Accumulation of an endogenous inhibitor of nitric oxide synthesis in chronic renal failure. Lancet. 1992; 339:572-5.
          155. 155.↵
            Wood ER, Berger H Jr, Sherman PA, Lapetina EG. Hepatocytes and macrophages express an identical cytokine inducible nitric oxide synthase gene. Biochem Biophys Res Commun. 1993; 191:767-74.
          156. 156.↵
            Young HM, Furness JB, Shuttleworth CW, Bredt DS, Snyder SH. Co-localization of nitric oxide synthase immunoreactivity and NADPH diaphorase staining in neurons of the guinea-pig intestine. Histochemistry. 1992; 97:375-8.
          157. 157.↵
            Saffrey MJ, Hassall CJ, Hoyle CH, Belai A, Moss J, Schmidt HH, et al. Colocalization of nitric oxide synthase and NADPH-diaphorase in cultured myenteric neurones. Neuroreport. 1992; 3:333-6.
          158. 158.↵
            Bredt DS, Hwang PM, Snyder SH. Localization of nitric oxide synthase indicating a neural role for nitric oxide. Nature. 1990; 347: 768-70.
          159. 159.↵
            Gustafsson LE, Wiklund CU, Wiklund NP, Persson MG, Moncada S. Modulation of autonomic neuroeffector transmission by nitric oxide in guinea pig ileum. Biochem Biophys Res Commun. 1990; 173:106-10.
          160. 160.↵
            Sanders KM, Ward SM, Thornbury KD, Dalziel HH, Westfall DP, Carl A. Nitric oxide as a non-adrenergic, non-cholinergic neurotransmitter in the gastrointestinal tract. Jpn J Pharmacol. 1992; 58(Suppl 2):220P-5P.
          161. 161.↵
            Desai KM, Sessa WC, Vane JR. Involvement of nitric oxide in the reflex relaxation of the stomach to accommodate food or fluid. Nature. 1991; 351:477-9.
          162. 162.↵
            Bult H, Boeckxstaens GE, Pelckmans PA, Jordaens FH, Van Maercke YM, Herman AG. Nitric oxide as an inhibitory non-adrenergic non-cholinergic neurotransmitter. Nature. 1990; 345:346-7.
          163. 163.↵
            Tottrup A, Glavind EB, Svane D. Involvement of the L-arginine-nitric oxide pathway in internal anal sphincter relaxation. Gastroenterology. 1992; 102:409-15.
          164. 164.↵
            Burnett AL, Lowenstein CJ, Bredt DS, Chang TS, Snyder SH. Nitric oxide: a physiologic mediator of penile erection. Science. 1992; 257:401-3.
          165. 165.↵
            Hope BT, Michael GJ, Knigge KM, Vincent SR. Neuronal NADPH diaphorase is a nitric oxide synthase. Proc Natl Acad Sci U S A. 1991; 88:2811-4.
          166. 166.↵
            Dawson TM, Bredt DS, Fotuhi M, Hwang PM, Snyder SH. Nitric oxide synthase and neuronal NADPH diaphorase are identical in brain and peripheral tissues. Proc Natl Acad Sci U S A. 1991; 88: 7797-801.
          167. 167.↵
            Bredt DS, Glatt CE, Hwang PM, Fotuhi M, Dawson TM, Snyder SH. Nitric oxide synthase protein and mRNA are discretely localized in neuronal populations of the mammalian CNS together with NADPH diaphorase. Neuron. 1991; 7:615-24.
          168. 168.↵
            Ferrante RJ, Kowall NW, Beal MF, Richardson EP Jr, Bird ED, Martin JB. Selective sparing of a class of striatal neurons in Huntington's disease. Science. 1985; 230:561-4.
          169. 169.↵
            Uemura Y, Kowall NW, Beal MF. Selective sparing of NADPH-diaphorase-somatostatin-neuropeptide Y neurons in ischemic gerbil striatum. Ann Neurol. 1990; 27:620-5.
          170. 170.↵
            Choi DW. Cerebral hypoxia: some new approaches and unanswered questions. J Neurosci. 1990; 10:2493-501.
          171. 171.↵
            Dawson TM, Dawson VL, Snyder SH. A novel neuronal messenger molecule in brain: the free radical, nitric oxide. Ann Neurol. 1992; 32:297-311.
          172. 172.↵
            Nowicki JP, Duval D, Poignet H, Scatton B. Nitric oxide mediates neuronal death after focal cerebral ischemia in the mouse. Eur J Pharmacol. 1991; 204:339-40.
          173. 173.↵
            Dawson DA, Kusumoto K, Graham DI, McCulloch J, Macrae IM. Inhibition of nitric oxide synthesis does not reduce infarct volume in a rat model of focal cerebral ischaemia. Neurosci Lett. 1992; 142:151-4.
          174. 174.↵
            Bush PA, Aronson WJ, Buga GM, Rajfer J, Ignarro LJ. Nitric oxide is a potent relaxant of human and rabbit corpus cavernosum. J Urol. 1992; 147:1650-5.
          175. 175.↵
            Knispel HH, Goessl C, Beckmann R. Nitric oxide mediates relaxation in rabbit and human corpus cavernosum smooth muscle. Urol Res. 1992; 20:253-7.
          176. 176.↵
            Adaikan PG, Lau LC, Ng SC, Ratnam SS. Nitric oxide is the likely inhibitory neurotransmitter of erection in the human penis. Jpn J Pharmacol. 1992; 58(Suppl 2):300P.
          177. 177.↵
            Rajfer J, Aronson WJ, Bush PA, Dorey FJ, Ignarro LJ. Nitric oxide as a mediator of relaxation of the corpus cavernosum in response to nonadrenergic, noncholinergic neurotransmission. N Engl J Med. 1992; 326:90-4.
          178. 178.↵
            Trigo-Rocha F, Aronson WJ, Hohenfellner M, Ignarro LJ, Rajfer J, Lue TF. Nitric oxide and cGMP: mediators of pelvic nerve-stimulated erection in dogs. Am J Physiol. 1993; 264:H419-22.
          179. 179.↵
            Roberts JD, Polaner DM, Lang P, Zapol WM. Inhaled nitric oxide in persistent pulmonary hypertension of the newborn. Lancet. 1992; 340:818-9.
          180. 180.↵
            Rossaint R, Falke KJ, Lopez F, Slama K, Pison U, Zapol WM. Inhaled nitric oxide for the adult respiratory distress syndrome. N Engl J Med. 1993; 328:399-405.
          181. 181.↵
            Johnson G 3d, Tsao PS, Lefer AM. Cardioprotective effects of authentic nitric oxide in myocardial ischemia with reperfusion. Crit Care Med. 1991; 19:244-52.
          « Previous | Next Article »Table of Contents

          This Article

          1. Ann Intern Med February 1, 1994 vol. 120 no. 3 227-237
          1. AbstractFREE
          2. Figures
          3. » Full Text
          4. Full Text (PDF)

          Rapid Responses

          1. Submit a response
          2. No responses published

          Navigate This Article

          Current Issue

          1. November 17, 2009, 151 (10)
          1. Alert me to current issues