Non-specific Nitric Oxide Synthase Inhibitor Research Articles

This month in J Lab Clin Med: Issue highlights for November 2002

This month in J Lab Clin Med: Issue highlights for November 2002

Nitric oxide synthase inhibition during development: effect on apoptotic death of dopamine neurons

Naturally occurring cell death via apoptosis occurs in the substantia nigra pars compacta (SNc) during rat development, culminating during the perinatal period. We previously showed that lipid peroxidation-mediated oxidative stress is not involved in this cell death process. Nitric oxide (NO) has been proposed to be critical for many developmental processes in brain and has been shown to mediate cell death in neurotoxin models of neurodegenerative disorders. Here, we reported that in vivo pre- and postnatal treatment with the non-specific NO synthase (NOS) inhibitor, l-NAME (60 mg/kg), or with the neuronal NOS inhibitor, 7-NI (30 mg/kg), dramatically decreased the NOS activity as well as the NADPH-diaphorase staining in brain. However, those treatments did not rescue dopamine neurons from developmental death, suggesting that NO is not involved in vivo in developmental death of these neurons or in the overall development of the SNc.

Adenosine Enhances Functional Activation of Blood Flow in Cat Optic Nerve Head during Photic Stimulation Independently from Nitric Oxide

Blood flow studies in the brain, heart, and other organs suggest that there could be interaction between nitric oxide (NO) and adenosine. This possibility was investigated in the optic nerve head (ONH) during photic stimulation of the dark-adapted cat eye. Functional activation of ONH blood flow was measured by laser Doppler flowmetry, simultaneously with NO and PO 2 using double-barrel recessed electrochemical sensors. Photic stimulation (diffuse luminance flickering light at 30 Hz) increased ONH blood flow to 127.4 ± 4.7% (mean ± SEM) of baseline with a transient increase in NO by 79.8 ± 12.8 nM, while PO 2 decreased from 24.5 ± 2.7 to 22.7 ± 2.4 Torr (control responses, 15 trials, 10 cats). Adenosine (3 mg/kg iv) increased baseline ONH blood flow to 113.8 ± 8.4% of control within 5 min. Functional activation of ONH blood flow was enhanced during photic stimulation, reaching a maximum of 155.8 ± 8.1% within 5 min, and remained enhanced for 30 to 45 min. NO responses during photic stimulation were not different from control responses. Treatment with a nonspecific NO synthase inhibitor ( N G-nitro- l-arginine methyl ester, 40 mg/kg iv, 5 cats) did not alter the increase in resting ONH blood flow or the enhanced functional activation after adenosine. We conclude that there is no interaction between NO and adenosine during functional activation of cat ONH blood flow by photic stimulation.

The role of nitric oxide in the proconvulsant effect of δ-opioid agonist SNC80 in mice

The involvement of nitric oxide (NO) in modulation of seizure susceptibility by δ-opioid agonist (+)-4-((alpha R)-alpha-((2 S, 5 R)-4-allyl-2, 5-dimethyl-1-piperazinyl)-3-methoxybenzyl)- N, N-diethyl-benzamide (SNC80) was examined in mice. Systemic administration of SNC80 (0.1–5 mg/kg, intraperitoneally (i.p.)) decreased the threshold for clonic seizures induced by pentylenetetrazole. The non-specific NO synthase (NOS) inhibitor, N G-nitro- l-arginine methyl ester (3–20 mg/kg, i.p.), but not the specific inducible NOS inhibitor, aminoguanidine (50 and 100 mg/kg, i.p.) inhibited the proconvulsant effect of SNC80. On the other hand, NO substrate, l-arginine (30 and 60 mg/kg, i.p.) potentiated the proconvulsant effect of a lower dose of SNC80 (0.5 mg/kg). These results support the involvement of NO, produced by constitutive NOS, in the proconvulsant effect of the δ-opioid agonist.

Inhibition of the neuronal isoform of nitric oxide synthase significantly attenuates 1-methyl-4-phenylpyridinium (MPP(+)) toxicity in vitro.

The possible protection against the toxicity of 1-methyl-4-phenylpyridinium (MPP(+)) afforded by inhibitors of nitric oxide synthase (NOS) and the antagonist of N-methyl-D-aspartate receptor function, MK-801, was studied in a brain-slice superfusion system. Significant decreases in levels of dopamine and its metabolites 3,4-dihyroxyphenylacetic acid (DOPAC) and homovanillic acid were observed following incubation of slices with 25 microM MPP(+). The activity of intracellular lactate dehydrogenase (LDH), a marker of cell viability, was also significantly decreased. These effects were attenuated by preincubation with I mM 7-nitroindazole (7NI), a selective inhibitor of the neuronal isoform of nitric oxide synthase (NOS). In contrast, the nonspecific NOS inhibitor N(omega)-nitro-L-arginine, also at 1 mM, had no effect on levels of dopamine metabolites but did show a small attenuation of the levels of dopamine. 7NI alone caused some increase in levels of dopamine and a decrease in the metabolite DOPAC, which is consistent with it also acting as an inhibitor of monoamine oxidase-B. MK-801 afforded no significant protection of aminergic cells, although changes in LDH activity suggested that there may have been some protection of non-aminergic neurons affected by this, relatively high concentration of MPP(+).

The involvement of endogenous opioids and nitricoxidergic pathway in the anticonvulsant effects of foot-shock stress in mice

The involvement of endogenous opioids and nitric oxide (NO) in the anticonvulsant effects of stress against pentylenetetrazole (PTZ)- or electroconvulsive shock-induced seizures was assessed in mice. The prolonged and intermittent foot-shock stress, which induced opioid-mediated analgesia, had significant protective effects against both seizure types which was reversible by naloxone (0.3, 1 or 2 mg/kg), while brief and continuous foot-shock did not alter the seizure susceptibility. Pre-treatment with non-specific nitric oxide synthase (NOS) inhibitor, N G-nitro- l-arginine methyl ester ( l-NAME, 1, 2, 5, 10 or 30 mg/kg), but not with specific inducible NOS (iNOS) inhibitor, aminoguanidine (50 or 100 mg/kg), blocked the stress-induced anticonvulsant effects. The lower doses of naloxone (0.3 mg/kg) and l-NAME (2 mg/kg) showed additive effects in blocking the stress-induced anticonvulsant properties. l-arginine at a per se non-effective dose of 20 mg/kg potentiated the stress-induced anticonvulsant properties, an effect which was inhibited by l-NAME but not by aminoguanidine. Furthermore, a low dose of morphine (0.5 mg/kg) showed potentiation with stress in increasing PTZ seizure threshold. This potentiation was reversed by either naloxone or l-NAME at low doses but not by aminoguanidine. Taken together, these results show that NO synthesis, through constitutive but not iNOS, is involved in opioid-dependent stress-induced anticonvulsant effects against electrical and PTZ-induced convulsions.

Modulation of renal disease in MRL/lpr mice by pharmacologic inhibition of inducible nitric oxide synthase

MRL-MPJFaslpr (MRL/lpr) mice spontaneously develop lupus-like disease characterized by immune complex glomerulonephritis and overproduction of nitric oxide (NO). Blocking NO production pharmacologically by a non-specific nitric oxide synthase (NOS) inhibitor ameliorated renal disease in MRL/lpr mice while genetically deficient inducible NOS (iNOS) mice developed proliferative glomerulonephritis similar to wild-type controls. To clarify the role of iNOS in the pathogenesis of nephritis in MRL/lpr mice, we treated mice with two different NOS inhibitors. Either NG-monomethyl-l-arginine (L-NMMA), a nonspecific NOS inhibitor, or l-N6-(1-iminoethyl)lysine (L-NIL), an iNOS specific inhibitor, was administered in the drinking water from 10 through 22 weeks of age with disease progression monitored over time. Control mice received water alone. Both L-NMMA and L-NIL blocked NO production effectively in MRL/lpr mice. As expected, neither L-NNMA nor L-NIL had an effect on antibody production, immune complex deposition or complement activation. Although both NOS inhibitors decreased protein excretion, L-NMMA was more effective than L-NIL. Pathologic renal disease was significantly decreased at 19 weeks in both treatment groups. At 22 weeks the L-NIL treated mice, but not the L-NMMA mice, had significantly reduced renal disease scores compared to controls. These results indicate that specific inhibition of iNOS blocks the development of pathologic renal disease in MRL/lpr mice.

In situ localization of nitric oxide synthase and direct evidence of NO production in rat retinal ganglion cells

The expression of isoforms of nitric oxide synthase (NOS), enzymes responsible for NO production, and the synthesis of nitric oxide (NO) in rat retinal ganglion cells (RGCs) during synaptogenesis for various phases of the pre- and postnatal developmental periods were investigated. The retinas from prenatal, lactating, young, and adult rats were fixed in paraformaldehyde. The cryosections or paraformaldehyde-fixed ganglion cells purified from rat pups were immunostained for constitutive isoforms of NOS (n and eNOS) and observed with a confocal laser scanning microscope. Synthesis of NO in the RGCs was achieved by in vitro stimulation with glutamate. The intracellular NO levels were measured in real time using diaminofluorescein-2 diacetate, a fluorescence indicator of NO. Immunohistochemical analysis revealed nNOS and eNOS expressed in retinal ganglion cells during the first 2 postnatal weeks. Cultured RGCs also expressed nNOS and eNOS in vitro. Intracellular NO levels in cultured RGCs showed spontaneous fluctuation during a 20-min observation. The presence of both a non-specific NOS inhibitor, l-NAME, and a specific nNOS inhibitor, 7-NI, significantly inhibited ( P<0.001) the increase of intracellular NO 6 and 8 min after the introduction of l-arginine and glutamate to the medium. This study revealed that all constitutive NOS isoforms are expressed in RGCs and demonstrated that NO is produced by nNOS mainly through stimulation by glutamate in cultured RGCs.

Evidence that nitric oxide production increases γ-amino butyric acid permeability of blood-brain barrier

Blood-brain barrier permeability (BBB) to the inhibitory neurotransmitter γ-amino butyric acid (GABA) was studied in rats following intraperitoneal (i.p) injections of GABA alone and in combination with L-Arginine (L-Arg). Administration of GABA (600 mg/kg body weight [b. wt.]) alone increased brain GABA concentration (33%, p < 0.01), when compared to untreated rats and administration of L-Arg (2000 mg/kg b. wt.) alone also increased GABA concentration (65%, p < 0.01) in the brain. Moreover, GABA + L-Arg treated brains showed a fourfold increase in GABA level (383.3%, p < 0.01) when compared to controls. Dose-dependent increase in nitric oxide production was observed 10 min after i.p injections of L-Arg (400, 800, 1000, and 2000 mg/kg b. wt.) and a peak nitric oxide (NO) production was observed at the dose level of 2000 mg/kg b. wt. On the other hand, administration of GABA failed to increase NO production in the brain. Rats pretreated (10 min) with a nonspecific nitric oxide synthase (NOS) inhibitor N ω-nitro-L-Arginine methyl ester (L-NAME, 50 mg/kg b. wt.) completely blocked the production of NO induced by L-Arg. In addition, L-NAME attenuated GABA entry into the brain after the administration of GABA alone or in combination with L-Arg. We conclude that high NO concentrations in the brain following L-Arg administration may increase the permeability of BBB to peripheral GABA.

Nitric oxide modulates sympathoexcitatory cardiac-cardiovascular reflexes elicited by bradykinin.

A number of studies have demonstrated an important role for nitric oxide (NO) in central and peripheral neural modulation of sympathetic activity. To assess the interaction and integrative effects of NO release and sympathetic reflex actions, we investigated the influence of inhibition of NO on cardiac-cardiovascular reflexes. In anesthetized, sinoaortic-denervated and vagotomized cats, transient reflex increases in arterial blood pressure (BP) were induced by application of bradykinin (BK, 0.1-10 microg/ml) to the epicardial surface of the heart. The nonspecific NO synthase (NOS) inhibitor NG-monomethyl-L-arginine (L-NMMA, 10 mg/kg iv) was then administered and stimulation was repeated. L-NMMA increased baseline mean arterial pressure (MAP) from 129 +/- 8 to 152 +/- 9 mmHg and enhanced the change in MAP in response to BK from 32 +/- 3 to 39 +/- 5 mmHg (n = 9, P < 0.05). Pulse pressure was significantly enhanced during the reflex response from 6 +/- 4 to 27 +/- 6 mmHg after L-NMMA injection due to relatively greater potentiation of the rise in systolic BP. Both the increase in baseline BP and the enhanced pressor reflex were reversed by L-arginine (30 mg/kg iv). Because L-NMMA can inhibit both brain and endothelial NOS, the effects of 7-nitroindazole (7-NI, 25 mg/kg ip), a selective brain NOS inhibitor, on the BK-induced cardiac-cardiovascular pressor reflex also were examined. In contrast to L-NMMA, we observed significant reduction of the pressor response to BK from 37 +/- 5 to 18 +/- 3 mmHg 30 min after the administration of 7-NI (n = 9, P < 0.05), an effect that was reversed by L-arginine (300 mg/kg iv, n = 7). In a vehicle control group for 7-NI (10 ml of peanut oil ip), the pressor response to BK remained unchanged (n = 6, P > 0.05). In conclusion, neuronal NOS facilitates, whereas endothelial NOS modulates, the excitatory cardiovascular reflex elicited by chemical stimulation of sympathetic cardiac afferents.

The pathophysiology of migraine.

Migraine results from episodic changes in central nervous system physiologic function in hyperexcitable brain manifested by abnormal energy metabolism, lowered threshold for phosphene generation, and increased contingent negative variation. Human functional magnetic resonance imaging and magnetoencepholography data strongly suggest that aura is caused by cortical spreading depression. Brain hyperexcitability may be caused by low magnesium levels, mitochondrial abnormalities with abnormal phosphorylation of adenosine 5'-diphosphate, a dysfunction related to nitric oxide, or calcium channelopathy. Low magnesium can result in opening of calcium channels, increased intracellular calcium, glutamate release, and increased extracellular potassium, which may in turn trigger cortical spreading depression. Mitochondrial dysfunction has been suggested by a low phosphocreatine:Pi ratio and a possible response by migraine patients to riboflavin prophylaxis. Nitroglycerine administration results in a delayed migraine-like headache in migraine patients but not in control patients, and a nonspecific nitric oxide synthase inhibitor aborted migraine at 2 hours in the majority of tested migraine patients compared to controls. Many patients with familial hemiplegic migraine have a missense mutation in the P/Q calcium channel, so that this form of migraine, at least, is associated with a demonstrable calcium channelopathy. The generation of migraine occurs centrally in the brain stem, sometimes preceded by cortical spreading depression and aura. Activation of the trigeminovascular system stimulates perivascular trigeminal sensory afferent nerves with release of vasoactive neuropeptides, resulting in vasodilation and transduction of central nociceptive information. There is then a relay of pain impulses to central second- and third-order neurons and activation of brain stem autonomic nuclei to induce associated symptoms.

Modulatory role of nitric oxide and cyclooxygenase enzyme pathway in LPS-mediated hyperalgesia

Nitric oxide (NO) is a free radical, is known to play an important role in many physiological and pathological processes. Evidence suggests that NO participates in the pathogenesis of inflammatory reactions. It has been reported that exposure of tissues to endotoxin (LPS) leads to induction of both inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). In the present study we have investigated the role of iNOS in LPS-mediated hyperalgesia in mice and the regulatory role between prostanoids in vivo using L-NAME, a non-specific nitric oxide synthase inhibitor, and spirulina purified protein (SPP) containing C-phycocyanin, NS-398 and rofecoxib, COX-2 selective inhibitor in various nociceptive assays. Acute administration of LPS (50 μg/mouse, i.p. or 10 μg/paw, i.pl.) significantly demonstrated hyperalgesia in chemical, thermal and mechanical nociception in mice, respectively. Treatments of mice with L-NAME (5-20 mg/kg, i.p. or 10-40 μg/paw, i.pl.) significantly reversed the LPS-mediated hyperalgesia in peripheral or central nociception in mice. Treatment with SPP (50 and 100 mg/kg, p.o.), NS398 (10 mg/kg, p.o.) and rofecoxib (10 mg/kg, p.o.), significantly exhibited antihyperalgesic effect in chemical hyperalgesia. These COX-2 inhibitors significantly potentiated the L-NAME reversal of LPS-mediated hyperalgesia. It is concluded that iNOS plays a significant role in LPS-mediated hyperalgesia and simultaneous inhibition of COX-2 enzyme leads to modulatory effect between NO and prostanoids.

Pravastatin reduces myocardial lesions induced by acute inhibition of nitric oxide biosynthesis in normocholesterolemic rats

Pravastatin is useful in restoring endothelium-dependent relaxation in hypercholesterolemic animals. A single intravenous bolus injection of N ω-nitro- l-arginine methyl ester ( l-NAME), a non-specific inhibitor of NO synthase, causes myocardial necrosis and reduces coronary flow in rats. Since rats do not develop hypercholesterolemia and atherosclerosis, we have tested the hypothesis that pravastatin protects the heart from myocardial lesions induced by l-NAME in the absence of alterations in cholesterol levels and plaque formation. Male Wistar rats fed standard chow were divided into four groups: CONTROL ( n=14) – rats that received tap water alone for 18 days; l-NAME ( n=14) – rats that received l-NAME (15 mg/kg, i.v.) on the 14th day of the study ; PRAVASTATIN ( n=11) – rats that received pravastatin (6 mg/kg/day) in their drinking water for 18 days; PRAVASTATIN+ l-NAME ( n=12) – rats that received pravastatin (6 mg/kg/day) and l-NAME (15 mg/kg, i.v.) as indicated in the preceding groups. At the end of 18 days, the rats were sacrificed and the hearts removed for stereological analysis by light microscopy. Plasma nitrate/nitrite and thromboxane B 2 concentrations were determined immediately before and after l-NAME administration. Pravastatin prevented the ischemic lesions induced by the acute inhibition of NO biosynthesis (the area of myocardial lesions in the l-NAME group was greater than in the Pravastatin+ l-NAME group: 101.6 μm 2 vs. 1.2 μm 2, respectively; P<0.0001) and markedly increased the plasma nitrate/nitrate concentrations, even before l-NAME administration. There were no significant changes in the plasma thromboxane B 2 concentrations.

ORGAN SPECIFIC FORMATION OF NITROSYL COMPLEXES UNDER INTESTINAL ISCHEMIA/REPERFUSION IN RATS INVOLVES NOS-INDEPENDENT MECHANISM(S)

Intestinal ischemia/reperfusion may lead to local and distant organ damage involving nitric oxide (NO). NO rapidly reacts with heme/non-heme-iron-yielding nitrosyl complexes, which can be determined directly by electron paramagnetic resonance spectroscopy. The aim of the present study was to characterize nitrosylation reactions induced by transient intestinal ischemia in blood and tissues. We used electron paramagnetic resonance spectroscopy and reverse transcription polymerase chain reaction analyses to estimate nitrosyl complex levels and inducible NO synthase mRNA expression in rats subjected to superior mesenteric artery occlusion for 60 min followed by the reperfusion. Nitrosyl hemoglobin concentrations in circulating blood were significantly increased during ischemia and reperfusion. Nitrosyl hemoglobin complexes were detected in ischemic intestine, but not in normoxic lung and liver or reperfused intestine. Administration of N-G-monomethyl-L-arginine, a non-specific NO synthase inhibitor, did not affect the formation of circulating nitrosyl complexes. Moreover, inducible NO synthase mRNA was not found in intestinal tissues at 30 min of reperfusion. Our data suggest an organ-specific NO formation indicated by the increased nitrosylation reaction in ischemic intestinal tissue, but not in the distant normoxic organs, in spite of high circulating nitrosyl hemoglobin levels. NO involved in nitrosylation under intestinal ischemia/reperfusion is probably formed by NO synthase-independent mechanism(s).

Increase in rat plasma antioxidant activity after E. coli lipopolysaccharide administration.

It is well recognized that the sensitivity of animals to lipopolysaccharide (LPS) endotoxin varies tremendously. And, it has been recently observed that Sprague-Dawley rats dramatically increase the activity of hepatic endogenous antioxidative enzyme systems after LPS administration. This finding suggests that the relative resistance of rats to LPS may be related to a concomitant increase in the activities of the hepatic antioxidant systems. This study was designed to examine if the above reported hepatic change in rats given LPS could be observed at the systemic level. Male Sprague-Dawley or Wistar rats, weighing 250 - 350 g, were given increasing doses (10 - 100 mg/kg) of LPS i.p. under 1.0% isoflurane anesthesia. Antioxidant capacity (AOC), blood gas analysis, and the cardiovascular parameters of the arterial blood of animals were determined over a 4 hour period following LPS administration. In addition, we studied the effect of pretreatment with the non-specific nitric oxide synthase inhibitor, L-N(G)-Nitroarginine methyl ester hydrochloride (L-NAME), given 50 mg/kg s.c. one and 24 hours before the administration of 20 mg/kg LPS i.p. in Sprague-Dawley rats. Rats given sufficiently high doses of E. coli LPS to produce behavioral effects also showed increased plasma AOCs in the early period after the administration of LPS. Similar changes were noted in Sprague-Dawley and Wistar rat strains, but at different doses that reflect their differential sensitivities to the LPS induced inflammatory response. Also, the resistance of the Sprague-Dawley strain of rats to LPS was not altered by the prior administration of L-NAME, nor was the plasma AOC altered. In conclusion, our study suggests that the rat strains are relatively resistant to develop the toxic signs of LPS in the early period after the administration of LPS, especially in Sprague-Dawley rats. Moreover, endotoxin-induced increases in plasma AOC may contribute to the rats' resistance to LPS intoxication.

Pre-synaptic NO-cGMP Pathway Modulates Vagal Control of Heart Rate in Isolated Adult Guinea Pig Atria

The role of nitric oxide (NO) in the vagal modulation of heart rate (HR) is controversial. We tested the hypothesis that NO acts via a pre-synaptic, guanylyl cyclase (GC) dependent pathway. The effects of inhibiting NO synthase (NOS) and GC were evaluated in isolated atrial/right vagal nerve preparations from adult (550–750 g) and young (150–250 g) female guinea pigs. Levels of NOS protein were quantified in right atria using Western blotting and densitometry. The non-specific NOS inhibitor N- ω -nitro- L -arginine (L -NA, 100 μM,n =5) significantly reduced the negative chronotropic response to vagal nerve stimulation (VNS) at 3 and 5 Hz in the adult guinea pig. This effect was reversed with 1 m ML -arginine. Similar results were observed with the specific neuronal NOS inhibitor vinyl-N5-(1-imino-3-butenyl)- L -ornithine (L -VNIO, 100 μM,n =7). Inhibition of GC with 1H-(1,2,4)-oxadiazolo-(4,3-a)-quinoxalin-1-one (ODQ, 10 μM,n =7) also significantly reduced the negative chronotropic response to VNS at 3 and 5 Hz in adult guinea pigs. Neither L -NA (n=6), L -VNIO (n=5) nor ODQ (n=6) changed the HR response to cumulative doses of carbamylcholine in adult guinea pig atria suggesting that the action of NO is pre-synaptic. The HR response to VNS was unaffected by L -NA (n=7) or ODQ (n=7) in young guinea pigs and Western blot analysis showed significantly lower levels of nNOS protein in right atria from young animals. These results suggest a pre-synaptic NO-cGMP pathway modulates cardiac cholinergic transmission, although this may depend on the developmental stage of the guinea pig.

Experimental Study of the Effect of Nitric Oxide Inhibition on Mesenteric Blood Flow and Interleukin‐10 Levels with a Lipopolysaccharide Challenge

The septic shock-induced decrease in mesenteric blood flow and release of proinflammatory cytokines are among the major pathophysiologic changes presumed to lead to multiple organ dysfunction syndrome (MODS). Increased nitric oxide (NO) levels are associated with both decreased mesenteric blood flow and positive modulation of proinflammatory cytokine release. In this study we aimed to determine the effect of the timing of the inhibition of nitric oxide synthase (NOS) on mesenteric blood flow and serum interleukin-10 (IL-10) concentrations during endotoxin shock. A nonspecific NOS inhibitor NG-nitro-L-arginine methyl ester (L-NAME), a specific NOS inhibitor aminoguanidine (AG), or placebo were injected 20 minutes before or 20 minutes after a lipopolysaccharide (LPS) or placebo challenge to Swiss-albino mice, as pretreatment or posttreatment, respectively. At 120 minutes after LPS or placebo injection the mesenteric blood flow was measured, and blood samples from the heart were obtained for IL-10 levels in both groups. Pretreatment and posttreatment with both NOS inhibitors prevented the LPS-induced decrease in mesenteric blood flow. Pretreatment was more effective for this purpose. Pretreatment accentuated the LPS-induced increase in serum IL-10 concentrations, whereas posttreatment had no significant effect. We conclude that the timing of NOS inhibition is important for attenuating some deleterious effects of endotoxin.

The nitric oxide pathway is an important modulator of stress-induced fever in rats

Psychological stress evokes a number of physiological responses, including a rise in body temperature ( T b), which has been suggested to be the result of an elevation in the thermoregulatory set point, i.e., a fever. This response seems to share similar mechanisms with infectious fever. A growing number of studies have provided evidence that nitric oxide (NO) has a modulatory role in infectious fever, but no report exists about the participation of NO in stress fever. Thus, the present study aimed to verify the hypothesis that NO modulates stress fever by using restraint stress as a model. To this end, we tested the effects of the non-specific NO synthase (NOS) inhibitor N G-nitro- l-arginine methyl ester ( l-NAME) or its inactive enantiomer N G-nitro- d-arginine methyl ester ( d-NAME) on colonic T b of restrained or unrestrained rats. A rapid increase in T b was observed when animals were submitted to restraint. Intravenous (i.v.) injection of l-NAME at a dose (10 mg/kg) that caused no change in T b when administered alone significantly attenuated the elevation in T b elicited by stress, indicating that the NO pathway may mediate stress fever. Moreover, intracerebroventricular (i.c.v.) l-NAME (250 μg/μl) caused a rise in T b of euthermic animals and enhanced stress fever, supporting that NO in the central nervous system (CNS) leads to a reduction in T b and, therefore, this is unlikely to be the site where NO may mediate stress fever. Taken together, these data indicate that the NO pathway plays an important role in modulating restraint stress-induced fever in rats.

Associative learning is enhanced by selective neuronal nitric oxide synthase inhibitors and retarded by a nitric oxide donor in the rabbit.

Previous studies had reported that the nitric oxide (NO) donor, sodium nitroprusside (SNP), retarded and the non-specific NO synthase (NOS) inhibitor, Nomega-nitro-L-arginine methyl ester (L-NAME), enhanced acquisition of classically conditioned responses (CRs). These effects of IV SNP and IP L-NAME on CR acquisition occurred in the absence of any effect on non-associative processes or performance variables and at a time when there were no alterations in blood pressure or heart rate. In this study, we examined whether the changes in associative learning produced by L-NAME and SNP were due to their central effects on NO content of brain. To this end, we examined the effects of the selective neuronal NOS inhibitors 7-nitroindazole (7-NI) and AR-R 17477 and the effects of central (ICV) administration of the NO donor SNP on learning. Effects of drugs on CR acquisition were determined during classical conditioning of the rabbit's nictitating membrane (NM) response. Explicitly unpaired presentations of conditioned stimuli (CSs) and unconditioned stimuli (USs) were employed to measure non-associative levels of responding and unconditioned response (UR) topography. The SC injection of 7-NI and AR-R 17477 significantly enhanced associative learning while ICV administration of SNP significantly retarded learning. Production of NO within the brain by neuronal NOS normally acts to retard associative learning presumably by decreasing excitability within neuronal circuits involved in the acquisition of the classically conditioned NM reflex.

Clinical trials in sepsis: an update

In this article, we place clinical sepsis trials from the past year in the context of similar sepsis trials run over the past three decades. These recent clinical sepsis trials include studies of agents administered to limit the effects of specific host proinflammatory mediators (tumor necrosis factor, platelet-activating factor and prostaglandins), studies of use of corticosteroids at low doses late in sepsis, and studies of administration of high doses of a nonspecific nitric oxide synthase inhibitor to decrease nitric oxide production in septic shock. The three trials of agents designed to limit host proinflammatory mediators showed minimal beneficial effects, results that are similar to those of the approximately 20 previous trials of similar agents. Low-dose corticosteroid therapy reversed shock and showed nonsignificant trends towards improvements in survival rates. In contrast, high doses of corticosteroids given early in sepsis have shown harmful effects in clinical sepsis trials. Finally, inhibition of nitric oxide production was lethal, indicating that high doses of nonspecific inhibitors of nitric oxide production are contraindicated in septic shock.