Brain NOx Research Articles

Benfloreks: Dost mu, Düşman mı? Deneysel Diyabette, Benfloreksin Beyindeki Oksidatif Olaylara Etkileri

Benfluorex is a pharmacological agent with antidiabetic and antihyperlipidemic properties. In this study, the brain's oxidative and non-enzymatic antioxidant status in diabetic and benfluorex administrated diabetic rats have been investigated. For the experimental procedures, three groups of 18 Wistar albino rats were used as the control, diabetes (induced by streptozotocin), and benfluorex treated diabetic rats (benfluorex administration intragastric 50 mg/kg daily for 21 days). Brain NOx, TBARS, GSH, AA levels, and MPO activity were determined spectrophotometrically. Benfluorex administration was caused that decreased lipid peroxidation and MPO activity while increased non-enzymatic antioxidant and NOx levels. These results showed that benfluorex treatment positively affects lipid peroxidation and non-enzymatic antioxidant status of the brain during diabetes.

Gender Based Differences in Stress-induced Gastric Ulcer Formation and its Regulation by Nitric Oxide (NO): An Experimental Study

A variety of physiological and pharmacological factors are known to influence stress responses. Cold restraint stress (CRS) induced gastric ulcerogenesis in both the sexes but such ulceration was found to be markedly higher in male than in female rats. In males, CRS induced significant increases in both ulcer number and ulcer severity; while the females though showed a trend towards increase in both the parameters, the extent of changes was far less than in males. Pre-administration of the NO mimetic, L-Arginine (500 and 1000 mg/kg), prior to CRS, dose dependently decreased ulcer number and severity in male rats. In female rats, L-Arginine also induced a gastric cytoprotective effect during CRS but to a much lesser extent. On the other hand, inhibition of NO synthesis by LNAME (25 and 50 mg/kg) further aggravated such stress ulcerogensis in both males and females, with aggravations being more extensive in males. CRS induced ulcerogenesis was associated with reductions in levels of brain and plasma NOx and GSH levels while MDA levels were elevated in both male and female rats- the magnitude of these changes being higher in males than in females. In female rats, pretreatment with formestane (aromatase inhibitor) but not tamoxifen (estrogen receptor blocker) aggravated stress ulcer formation as compared to vehicle treated CRS exposed rats. Formestane pretreatment also induced greater suppressions in brain NOx and GSH and elevations in brain MDA, as compared to vehicle treated CRS rats. These results indicate that estrogen and its interactions with oxidative stress markers and NO plays a key role in the gender based differences in stress induced gastric ulcerogenesis. It may be speculated that, in males, CRS induces greater reductions in brain NO and enhancement in oxidative injury resulting in greater severity of gastric ulceration. On the other hand, greater resistance of females to ulcerogenic effects of CRS may be due to the protection conferred by estrogen and this effect seems to be related to interactions with brain NO.

Effects of morphine on stress induced anxiety in rats: Role of nitric oxide and Hsp70

The present study evaluated the effects of morphine on acute and chronic restraint stress (RS) induced anxiety modulation and the possible involvement of nitric oxide (NO) and heat shock proteins (Hsp70) during such effects. Acute RS (×1) induced anxiogenesis in the elevated plus maze (EPM) test which was associated with lowered brain NO metabolites (NOx) and elevated Hsp70 levels. Pretreatment with morphine (1 and 5mg/kg) and l-arginine (500mg/kg) attenuated the RS effects on EPM activity and brain NOx, whereas, Hsp70 levels were further augmented. Co-administration of both agents showed synergistic effects. By contrast, repeated RS (×15) did not induce any significant changes in EPM activity or brain NOx, but brain Hsp70 levels stayed elevated. Administration of morphine or l-arginine prior to chronic RS did not influence such chronic stress induced changes in behavioral and biochemical markers, but appreciably attenuated chronic RS induced elevation in Hsp70 levels. These results suggest that acute and chronic RS induced anxiety modulations were differentially influenced by morphine and l-arginine and that complex interactions involving brain NO and unregulated Hsp70 could regulate such effects.

Pharmacological evidence for the role of nitric oxide in the modulation of stress-induced anxiety by morphine in rats

The present study evaluated the effects of the opioid agonist, morphine on stress induced anxiogenesis and the possible involvement of nitric oxide (NO) in such effects in rats. Acute restraint stress consistently induced an anxiety-like response in the elevated plus maze test, i.e. reduced number of open arm entries and time spent in the open arms as compared to controls. Pretreatment with morphine (1 and 5 mg/kg), attenuated the restraint stress induced anxiogenic response in a dose related manner. Restraint stress induced neurobehavioral suppression was associated with reductions in brain NO oxidation products (NOx) levels, which were also reversed with morphine. Interaction studies showed that sub-effective doses of morphine and l-arginine (a NO precursor) had synergistic effects on stress induced elevated plus maze activity and brain NOx, whereas, l-NAME (a NO synthase inhibitor) neutralized these effects of morphine. Repeated restraint stress (× 5) induced adaptative changes as evidenced by normalization of behavioral suppression and elevations in brain NOx, as compared to acute stress. Pretreatment with morphine in combination with repeated stress (× 5) showed potentiating effects in the induction of behavioral adaptation in the elevatedplus maze and elevations in brain NOx, as compared to repeated stress alone. Further, l-NAME, when administered prior to morphine, blocked this effect of morphine on stress adaptation. These results suggest differential morphine–NO interactions during acute and repeated restraint stress.

Involvement of nitric oxide in the protective effects of dehydroepiandrosterone sulphate on stress induced neurobehavioral suppression and brain oxidative injury in rats

The involvement of nitric oxide (NO) in the effects of dehydroepiandrosterone sulphate (DHEAS) on restraint stress induced neurobehavioral and brain oxidative/nitrosative stress markers was investigated in rats. Exposure of rats to restraint stress suppressed behavioral activity in the elevated plus maze and this was associated with increases in malondialdehyde (MDA) and decrease in reduced glutathione (GSH) and brain NO metabolite (NOx) levels in brain homogenates. Pretreatment with DHEAS (5–40 mg/s.c.) reversed the stress induced changes in behavioral and oxidative stress markers and also brain NOx levels. The beneficial effect of DHEAS (40 mg/kg s.c.) was blocked by pretreatment with nitric oxide synthase inhibitor, L-NAME (50 mg/kg i.p.) while pretreatment of rats with NO-precursor l-Arginine (100 mg/kg i.p.) produced potentiation of action of sub effective dose of DHEAS (5 mg/kg s.c.). The DHEAS effects were stress specific as these behavioral and biochemical parameters were not much influenced in non-stressed rats. These observations suggest that pretreatment with DHEAS has a protective effect on restraint stress induced alteration of neurobehavioral changes and brain oxidative injury in rats and NO-dependent mechanisms may be involved in this effect.

Leptin inhibits food intake without affecting brain NOx levels in food-deprived mice.

Effects of leptin on food intake and nitric oxide (NO) metabolites (nitrite and nitrate, NOx) levels of brain were investigated in mice. Leptin dose-dependently decreased milk intake in food-deprived mice. Administration of leptin at a dose of 1 mg/kg, which induces an apparent hypophagia, did not affect NOx levels in the hypothalamus and frontal cortex. These results suggest that leptin reduces food intake in food-deprived mice without altering NO production in the hypothalamus, which plays an important role in regulation of feeding.

Influences of caffeine to nitric oxide production and zonisamide concentration in the brain of seizure-susceptible EL mice.

To investigate changes of nitric oxide (NO) productions and zonisamide (ZNS) concentrations in the brain of seizure-susceptible EL mice given caffeine orally, mice were given caffeine (600 microg/mL) solution ad libitum as a drinking fluid for 1-3 weeks. Nitric oxide production in the brain was determined by measuring levels of nitrite plus nitrate (NOx). The brain NOx levels of mice treated with caffeine for 3 weeks were significantly higher than the control. Seizures in mice treated with caffeine for 2 and 3 weeks were not suppressed by ZNS at a dose of 75 mg/kg. Serum ZNS concentrations of mice with caffeine intake for 1-3 weeks were higher than in untreated mice. Conversely, brain ZNS concentrations of mice with caffeine intake for the same periods were significantly lower than in untreated mice. These results suggested that caffeine influenced brain NO production and ZNS concentrations in the seizure susceptibility of EL mice.

Alterations of nitric oxide and monoamines in the brain of the EL mouse treated with phenobarbital and zonisamide.

The effects of phenobarbital (PB; doses, 5, 10, and 25 mg/kg, intraperitoneally (i.p.)) and zonisamide (ZNS; doses, 30, 75, and 150 mg/kg, i.p.) on nitric oxide (NO) production, and those of coadministration of PB (5 mg/kg, i.p.) and ZNS (75 mg/kg, i.p.) on monoamines in the brain of the seizure-susceptible EL mouse were investigated. Nitric oxide production was obtained by measuring the combined level of nitrite plus nitrate (NOx). Zonisamide and PB dose-dependently suppressed the seizure of the EL mouse, and coadministration of PB (5 mg/kg) and ZNS (75 mg/kg) induced a greater degree of seizure suppression than treatment with ZNS or PB alone. Although PB (5 mg/kg) had no effect on brain NOx levels, ZNS (150 mg/kg) and coadministration of ZNS (75 mg/kg) and PB (5 mg/kg) decreased NOx levels significantly. Phenobarbital (5 mg/kg) did not influence monoamines, while coadministration of PB (5 mg/kg) and ZNS (75 mg/kg) decreased dihydroxyphenylacetic acid and increased 5-HT concentrations. The effect of the coadministration of two drugs on monoamines were similar to that of ZNS alone. These results suggest that one of the anticonvulsant effects of coadministration of PB and ZNS may be caused by changes in NOx levels.

Long‐term maternal ethanol administration decreases nitric oxide metabolites in brains of postnatal mice

AbstractWe examined the effect of long‐term maternal ethanol consumption on nitric oxide metabolites (NOx) in brains of postnatal mice. Three female adult ddY mice were given 10% ethanol solution ad libitum as drinking water for a 4 weeks before mating and continued on their corresponding ethanol solution during gestation and lactation. Other 2 mice receiving tap water ad libitum during gestation and lactation served as controls. The offspring in both groups were weaned at 22 days of age, and they were maintained on the chow and water ad libitum. In brain NOx levels in the offspring at 42 days of age, offspring with maternal alcohol consumption were significantly lower than those in controls, but serum NOx levels did not differ between groups. No significant differences in monoamines in the brain were evident between offspring with and without maternal alcohol administration. Present results indicated that long‐term maternal ethanol administration reduced brain NO metabolite levels in offspring, but not serum NO metabolites or brain monoamines.

Measurement of Nitric Oxide Metabolites in Brain Microdialysates by a Sensitive Fluorometric High-Performance Liquid Chromatography Assay

Nitric oxide (NO), formed from arginine by a specific neuronal NO synthase, is an important neurotransmitter in various regions of the central nervous system. While intracerebral microdialysis is an elegant technique to study local extracellular neurotransmitter concentrations in vivo, NO metabolites (nitrate, nitrite (NOx)) are difficult to study at high temporal resolution because of low tissue concentrations and small sample volumes. We developed a sensitive fluorometric high-performance liquid chromatography (HPLC)-coupled NOx assay adapted for the use in brain microdialysate samples. The assay includes an initial enzymatic step in which nitrate is reduced to nitrite. Nitrite is acidified to N2O3, which reacts with 2,3-diaminonaphthalene to form 1-(H)-naphthotriazole. This reaction product can be readily isolated and quantitated by HPLC with fluorometric detection. The theoretical assay sensitivity is less than 1 nM, but numerous sources of contamination must be eliminated in the sampling and assaying process to reliably monitor brain NOx outflow by microdialysis.

Hyperbaric Oxygen Reduces Cerebral Blood Flow by Inactivating Nitric Oxide

Based on recent evidence that nitric oxide (NO•) is involved in hyperoxic vasoconstriction, we tested the hypothesis that decreases in NO• availability in brain tissue during hyperbaric oxygen (HBO2) exposure contribute to decreases in regional cerebral blood flow (rCBF). rCBF was measured in rats exposed to HBO2 at 5 atmospheres (ATA) and correlated with interstitial brain levels of NO• metabolites (NOX) and production of hydroxyl radical (•OH). Changes in rCBF were also correlated with the effects of NO• synthase inhibitor (l-NAME), NO• donor PAPANONOate, and intravascular superoxide dismutase (MnSOD) during HBO2. After 30 min of O2 exposure at 5 ATA, rCBF had decreased in the substantia nigra, caudate putamen, hippocampus, and parietal cortex by 23 to 37%. These reductions in rCBF were not augmented by exposure to HBO2 in animals pre-treated with l-NAME. After 30 min at 5 ATA, brain NOX levels had decreased by 31 ± 9% and correlated with the decrease in rCBF, while estimated •OH production increased by 56 ± 8%. The decrease in rCBF at 5 ATA was completely abolished by MnSOD administration into the circulation before HBO2 exposure. Doses of NO• donor that significantly increased rCBF in animals breathing air had no effect at 5 ATA of HBO2. These results indicate that decreases in rCBF with HBO2 are associated with a decrease in effective NO• concentration and an increase in ROS production in the brain. The data support the hypothesis that inactivation of NO• antagonizes basal relaxation of cerebral vessels during HBO2 exposure, although an effect of HBO2 on NO• synthesis has not been excluded.

Calcium-independent NO-synthase activity and nitrites/nitrates production in transient focal cerebral ischaemia in mice.

1. The temporal changes in constitutive NO-synthase (cNOS) and in calcium-independent NO-synthase activities were studied in mice subjected to 2 h of transient focal cerebral ischaemia. The changes in brain nitrites/nitrates (NOx) content were also studied. 2. NOS activities were measured by the conversion of L-[14C]-arginine to L-[14C]-citrulline. Brain NOx contents were investigated by the Griess colourimetric method. 3. cNOS activity in the infarcted cortical area was significantly reduced after 6 h of reperfusion and this activity remained attenuated for up to 10 days after ischaemia. A calcium-independent NOS activity began to increase 48 h after reperfusion, reached a maximum at 7 days and returned to baseline at 10 days. 4. There was a significant increase of brain NOx content beginning after 3 days of reperfusion. This increase was maximal at 7 days and returned to baseline at 10 days. 5. Thus, ischaemia followed by recirculation leads to a rapid, prolonged drop in cNOS activity in the infarcted cortex. There is also a substantial appearance of calcium-independent NOS activity in the later phase of transient ischaemia, leading to an important increase of NOx production.

Ex Vivo Measurement of Brain Tissue Nitrite and Nitrate Accurately Reflects Nitric Oxide Synthase Activity In Vivo

The ex vivo tissue concentration of nitrite and nitrate (NOx) was found to correlate closely with the activity of nitric oxide synthase (NOS; EC 1.14.13.39) in various brain regions. Systemic administration of the nonselective NOS inhibitor N omega-nitro-L-arginine (L-NA) at doses that completely inhibited both central and peripheral NOS, depleted whole-brain and CSF NOx by up to 75% but had no effect on plasma NOx. Selective inhibition of central NOS by intracerebroventricular administration of L-NA methyl ester produced similar decreases in levels of whole-brain NOx. A residual concentration of NOx of 10-15 microM remained in all brain regions even after complete inhibition of brain NOS. Brain NOx content decreased rapidly and in parallel with the inhibition of brain NOS. The ex vivo measurement of levels of brain NOx was found to reflect the in vivo efficacy of several different types of NOS inhibitor: L-NA, N omega-monomethyl-l-arginine, and 7-nitroindazole. Intraperitoneal administration of the NOS substrate L-arginine increased brain NOx concentrations by up to 150% of control values. These results demonstrate that the ex vivo measurement of levels of brain tissue NOx is a rapid, reliable, and straightforward technique to determine NOS activity in vivo. This method can be used to assess both the regional distribution and the degree of inhibition of NOS activity in vivo.