Young Rat Brain Research Articles

Oxidative stress and prevention of the adaptive response to chronic iron overload in the brain of young adult rats exposed to a 150 kilohertz electromagnetic field

Iron surcharge may induce an oxidative stress-based decline in several neurological functions. In addition, electromagnetic fields (EMF) of frequencies up to about 100 kHz, emitted by electric/electronic devices, have been suggested to enhance free radical production through an iron dependent pathway. The purpose of this study was therefore to determine a possible relationship between iron status, exposure to EMF, and brain oxidative stress in young adult rats. Samples were micro-dissected from prefrontal cortex, hippocampus, striatum, and cerebellum after chronic saline or iron overload (IO) as well as after chronic sham exposure or exposure to a 150 kHz EMF or after combining EMF exposure with IO. The brain samples were used to monitor oxidative stress-induced lipid peroxidation and activity of the antioxidant enzymes superoxide dismutase and catalase. While IO did not induce any oxidative stress in young adult rats, it stimulated antioxidant defenses in the cerebellum and prefrontal cortex in particular. On the contrary, EMF exposure stimulated lipid peroxidation mainly in the cerebellum, without affecting antioxidant defenses. When EMF was coapplied with IO, lipid peroxidation was further increased as compared to EMF alone while the increase in antioxidant defenses triggered by the sole IO was abolished. These data suggest that EMF exposure may be harmful in young adults by impairing the antioxidant defenses directed at preventing iron-induced oxidative stress.

Oxidative stress induced by the Fe/ascorbic acid system or model ischemia in vitro: effect of carvedilol and pyridoindole antioxidant SMe1EC2 in young and adult rat brain tissue.

New effective strategies and new highly effective neuroprotective agents are being searched for the therapy of human stroke and cerebral ischemia. The compound SMe1EC2 is a new derivative of stobadine, with enhanced antioxidant properties compared to the maternal drug. Carvedilol, a non-selective beta-blocker, possesses besides its cardioprotective and vasculoprotective properties also an antioxidant effect. We compared the effect of carvedilol and SMe1EC2, antioxidants with a similar chemical structure, in two experimental models of oxidative stress in young and adult rat brain tissue. SMe1EC2 was found to improve the resistance of hippocampal neurons to ischemia in vitro in young and even in 18-month-old rats and inhibited formation of protein carbonyl groups induced by the Fe2+/ascorbic acid pro-oxidative system in brain cortex homogenates of young rats. Carvedilol exerted a protective effect only in the hippocampus of 2-month-old rats and that at the concentration 10-times higher than did SMe1EC2. The inhibitory effect of carvedilol on protein carbonyl formation induced by the pro-oxidative system was not proved in the cortex of either young or adult rats. An increased baseline level of the content of protein carbonyl groups in the adult versus young rat brain cortex confirmed age-related changes in neuronal tissue and may be due to increased production of reactive oxygen species and low antioxidant defense mechanisms in the adult rat brain. The results revealed the new pyridoindole SMe1EC2 to be more effective than carvedilol in neuroprotection of rat brain tissue in both experimental models involving oxidative stress.

Age and sex differences in the effects of the immunosuppressants cyclosporine, sirolimus and everolimus on rat brain metabolism

Application of the widely used immunosuppressant (ISS) cyclosporine (CsA) is severely limited by a number of serious side-effects such as kidney and neurotoxicity. As we have shown before, CsA exhibits metabolic toxicity in brain-models. The macrolide ISSs sirolimus (SRL) and everolimus (RAD) are capable of modulating these CsA-induced effects. It was our aim to study the age-dependent metabolic changes in the rat brain after ISS-treatment and the possible role of the blood–brain-barrier in modulation of CsA metabolic toxicity.Young and adult rats were treated orally with one ISS alone or in combination with CsA for six days. Metabolic changes were assessed by nuclear magnetic resonance (NMR) spectroscopy of brain extracts as toxicodynamic endpoints. Brain P-glycoprotein (P-gp) and ISS concentrations were determined as pharmacokinetic endpoints. Young rats were more susceptible to CsA-induced inhibition of the Krebs cycle (glutamate: 78% of controls, glutamine: 82%, GABA: 71% in young vs. 85%, 89%, 92% in adult rats). Increased glycolysis after CsA-treatment was sufficient to maintain the energy state at control levels in adult brains, but not in the young rat brains (phosphocreatine: 35%). Tissue concentrations of CsA and SRL within the brain of young rats were three-fold higher, while concentrations of P-gp were three-fold higher in adult rat brains. Our results suggest that age-dependent differences in the blood–brain barrier led to increased ISS brain concentrations and hence inhibition of brain energy metabolism.

Delayed transplantation of human neural precursor cells improves outcome from focal cerebral ischemia in aged rats

Neural precursor cell (NPC) transplantation may have a role in restoring brain function after stroke, but how aging might affect the brain's receptivity to such transplants is unknown. We reported previously that transplantation of human embryonic stem cell (hESC)-derived NPCs together with biomaterial (Matrigel) scaffolding into the brains of young adult Sprague-Dawley rats 3 weeks after distal middle cerebral artery occlusion (MCAO) reduced infarct volume and improved neurobehavioral performance. In this study, we compared the effect of NPC and Matrigel transplants in young adult (3-month-old) and aged (24-month-old) Fisher 344 rats from the National Institute on Aging's aged rodent colony. Distal MCAO was induced by electrocoagulation, and hESC-derived NPCs were transplanted into the infarct cavity 3 weeks later. Aged rats developed larger infarcts, but infarct volume and performance on the cylinder and elevated body swing tests, measured 6-8 weeks post-transplant, were improved by transplantation. We conclude that advanced age does not preclude a beneficial response to NPC transplantation following experimental stroke.

In situ detection of gliosis and apoptosis in the brains of young rats exposed in utero to a Wi-Fi signal

Pregnant rats were daily whole-body exposed or sham-exposed to a Wi-Fi signal in a free-running reverberation chamber at 0, 0.08, 0.4, and 4 W/kg for 2 h during the last 2 weeks of gestation (5 days/week). Following this in utero exposure, the pups were divided into two groups and 1 group continued exposure for 5 weeks after birth. Several brain areas were examined for gliosis and apoptotic cells. Comparison among sham and exposed groups revealed no significant differences, suggesting that in utero and post-natal exposure to Wi-Fi did not damage the brains of the young rats.

Effect of oseltamivir on catecholamines and select oxidative stress markers in the presence of oligoelements in the rat brain

The effect that osteltamivir has on the metabolism of catecholamines and oxidative damage in the brains of young patients remains unclear. The purpose of this study was to measure the effects of oseltamivir, in the presence of oligoelements, on biogenic amines and select oxidative biomarkers in the brains of uninfected, young rats under normal conditions. The study was conducted using male Wistar rats intraperitoneally treated for three days with either a control dose of 0.9 % NaCl, oseltamivir (50 mg/kg), oligoelements (50 μL/rat), or oseltamivir (50 mg/kg) and oligoelements (50 μL/rat). The brain tissue extracted from the treated rats was used to determine the concentrations of adrenaline, noradrenaline, and dopamine, as well as the levels of GSH, lipid peroxidation, and ATPase activity. An increase in the concentration of adrenaline and noradrenaline and in the level of GSH in the group treated with oligoelements (p < 0.001) was observed, while the group treated with oseltamivir and oligoelements, the levels of dopamine increased (p < 0.001), and in the groups treated with oligoelements alone or combination with oseltamivir a decrease in lipid peroxidation was observed (p < 0.001). The results of this study suggest that the consumption of oseltamivir and oligoelements induce biphasic changes in the metabolism of catecholamines; thereby, inducing a protective mechanism against oxidative damage in the brains of young rats.

Mannosylated liposomal cytidine 5′ diphosphocholine prevent age related global moderate cerebral ischemia reperfusion induced mitochondrial cytochrome c release in aged rat brain

Mitochondrial dysfunctions generating from cerebral ischemia-reperfusion exert a potential threat on neuronal cell survival and hence, accelerate the aging process and age dependent neuropathology. Thirty min moderate cerebral ischemia induced by bilateral common carotid artery occlusion (BCCAO) followed by 30 min reperfusion caused an increased diene production, depleted glutathione (GSH) content, reduced superoxide dismutase (SOD) and catalase activities and pyramidal neuronal loss in young (2 months old) and aged (20 months old) rat brain compared to sham operated controls. Cytidine 5′ diphosphocholine (CDP-Choline) is a known neuroprotective drug. CDP-Choline after metabolism in the liver suffers hydrolysis and splits into cytidine and choline before entering systemic circulation and hardly circumvents blood brain barrier (BBB) as such. Previous reports show CDP-Choline liposomes significantly increased in vivo uptake compared to “free drug” administration in cerebral ischemia. To enhance the therapeutic concentration build up in brain we sought to formulate mannosylated liposomal CDP-Choline (MLCDP) utilizing the mannose receptors. We tested the therapeutic supremacy of MLCDP over liposomal CDP-Choline (LCDP) in global moderate cerebral ischemia reperfusion induced neuronal damage. CDP-Choline in MLCDP delivery system was found potent to exert substantial protection against global moderate cerebral ischemia reperfusion induced mitochondrial damage in aged rat brain. Membrane lipid peroxidation, GSSG/GSH ratio and reactive oxygen species (ROS) generation in cerebral tissue were found to be higher in aged, compared to young rat. Further decline of those parameters was observed in aged rat brain by the induction of global moderate cerebral ischemia and reperfusion. MLCDP treatment when compared to free or LCDP treatment prevented global moderate cerebral ischemia-reperfusion induced mitochondrial damage as evident ultra structurally and release of cytochrome c (cyt c) from mitochondria into cytosol and protected mitochondria to restore its normal structure and functions.

Electroacupuncture enhances cell proliferation and neuronal differentiation in young rat brains

To investigate the effect of electroacupuncture on cell proliferation and differentiation in young rat hippocampus, postnatal day-14 rats were assigned randomly to control, ketamine anesthesia, nonacupoint, or acupoint electroacupuncture groups. Electroacupuncture was applied at bilateral acupoints (Quchi, Waiguan, Huantiao, and Zusanli) 30 min daily for 7 successive days. The proliferation and neuronal differentiation of proliferated surviving cells in the dentate gyrus were evaluated at 4 weeks after last stimulation. The stimulatory effect of electroacupuncture on cell proliferation had a long-lasting effect, as indicated by the increased phosphor-histone H3-positive cells. The number of proliferated and survival cells, indicated by BrdU labeling, was highest in the electroacupuncture group. The number of newly differentiated neurons, as indicated by BrdU/NeuN double labeling, was significantly higher in the electroacupuncture group than in any of the other groups. This finding provides a theoretical basis for the clinical application of acupuncture to cerebral injury rehabilitation in children.

Aging and antioxidants modulate rat brain levels of homocysteine and dehydroepiandrosterone sulphate (DHEA-S): Implications in the pathogenesis of Alzheimer's disease

The study has shown that in aged (22–24 months) rat brains an elevation of homocysteine level (42%) and a decrease in dehydroepiandrosterone sulphate (DHEA-S) content (32%) occur compared to those in the brains of young rats (4–6 months). Such changes in the brain levels of homocysteine and DHEA-S in aged rats are prevented, when the diet daily of the rats is supplemented with a combination of antioxidants (N-acetyl cysteine 50 mg, α-lipoic acid 3 mg and α-tocopherol 1.5 mg – each per 100 g of body weight) starting from 18 months until these are sacrificed between 22 and 24 months. The brain content of reduced glutathione is also decreased in aged rats as compared to that in young ones and the phenomenon can again be prevented completely by the same regimen of antioxidant supplementation. The changes in the levels of homocysteine and DHEA-S in aged rat brain have been related to associated glutathione depletion and oxidative stress and the implications of the results highlighted in the pathogenesis of Alzheimer's disease.

Effect of Acute and Chronic Administration of Methylphenidate on Mitochondrial Respiratory Chain in the Brain of Young Rats

Methylphenidate is commonly used for the treatment of attention deficit/hyperactivity disorder. There are still few works regarding the effects of methylphenidate on brain energy metabolism. Thus, in the present study we evaluated the effect of chronic administration of methylphenidate on the activities of mitochondrial respiratory chain complexes I and III in the brain of young rats. The effect of acute administration of methylphenidate on mitochondrial respiratory chain complexes I, II, III and IV in the brain of young rats was also investigated. For acute administration, a single injection of methylphenidate was given to rats on postnatal day 25. For chronic administration, methylphenidate injections were given starting at postnatal day 25 once daily for 28 days. Our results showed that complexes I and III were not affected by chronic administration of methylphenidate. Moreover, the acute administration of methylphenidate decreased complex I activity in cerebellum and prefrontal cortex, whereas complexes II, III and IV were not altered.

Mitochondrial DNA sequences are present inside nuclear DNA in rat tissues and increase with age

Mitochondrial DNA (mtDNA) mutations increase with age. However, the number of cells with predominantly mutated mtDNA is small in old animals. Here a new hypothesis is proposed: mtDNA fragments may insert into nuclear DNA contributing to aging and related diseases by alterations in the nucleus. Real-time PCR quantification shows that sequences of cytochrome oxidase III and 16S rRNA from mtDNA are present in highly purified nuclei from liver and brain in young and old rats. The sequences of these insertions revealed that they contain single nucleotide polymorphisms identical to those present in mtDNA of the same animal. Interestingly, the amount of mitochondrial sequences in nuclear DNA increases with age in both tissues. In situ hybridization of mtDNA to nuclear DNA confirms the presence of mtDNA sequences inside nuclear DNA in rat hepatocytes. Bone marrow metaphase cells from both young and old rats show mtDNA at centromeric regions in 20 out of the 2 n = 40 chromosomes. Consequently, mitochondria can be a major trigger of aging but the final target could also be the nucleus.

Proline impairs energy metabolism in cerebral cortex of young rats

In the present study we investigated the effect of acute hyperprolinemia on some parameters of energy metabolism, including the activities of succinate dehydrogenase and cytocrome c oxidase and (14)CO(2) production from glucose and acetate in cerebral cortex of young rats. Lipid peroxidation determined by the levels of thiobarbituric acid-reactive substances, as well as the influence of the antioxidants alpha-tocopherol plus ascorbic acid on the effects elicited by Pro on enzyme activities and on the lipid peroxidation were also evaluated. Wistar rats of 12 and 29 days of life received one subcutaneous injection of saline or proline (12.8 or 18.2 micromol/g body weight, respectively) and were sacrificed 1 h later. In another set of experiments, 5- and 22-day-old rats were pretreated for a week with daily intraperitoneal administration of alpha-tocopherol (40 mg/kg) plus ascorbic acid (100 mg/kg) or saline. Twelve hours after the last injection, rats received one injection of proline or saline and were sacrificed 1 h later. Results showed that acute administration of proline significantly reduced cytochrome c oxidase activity and increased succinate dehydrogenase activity and (14)CO(2) production in cerebral cortex, suggesting that Pro might disrupt energy metabolism in brain of young rats. In addition, proline administration increased the thiobarbituric acid-reactive substances levels, which were prevented by antioxidants. These findings suggest that mitochondrial dysfunction and oxidative stress may be important contributors to the neurological dysfunction observed in some hyperprolinemic patients and that treatment with antioxidants may be beneficial in this pathology.

Disturbance of mitochondrial energy homeostasis caused by the metabolites accumulating in LCHAD and MTP deficiencies in rat brain

Aims We investigated the in vitro effects of 3-hydroxydodecanoic (3HDA), 3-hydroxytetradecanoic (3HTA) and 3-hydroxypalmitic (3HPA) acids, which accumulate in tissues of patients affected by mitochondrial trifunctional protein (MTP) and isolated long-chain 3-hydroxyacyl-CoA dehydrogenase (LCHAD) deficiencies, on various parameters of energy homeostasis in mitochondrial preparations from brain of young rats. Main methods We measured the respiratory parameters state 4, state 3, respiratory control ratio (RCR) and ADP/O ratio by the rate of oxygen consumption, as well as the mitochondrial membrane potential and the matrix NAD(P)H levels in the presence of the fatty acids. Key findings We found that 3HDA, 3HTA and 3HPA markedly increased state 4 respiration and diminished the RCR using glutamate plus malate or succinate as substrates. 3HTA and 3HPA also diminished the mitochondrial membrane potential and the matrix NAD(P)H levels. In addition, 3HTA decreased state 3 respiration using glutamate/malate, but not pyruvate/malate or succinate as substrates. Our data indicate that the long-chain 3-hydroxy fatty acids that accumulate in LCHAD/MTP deficiencies act as uncouplers of oxidative phosphorylation, while 3HTA also behaves as a metabolic inhibitor. Significance It is presumed that impairment of brain energy homeostasis caused by these endogenous accumulating compounds may contribute at least in part to the neuropathology of LCHAD/MTP deficiencies.

Neurochemical evidence that glycine induces bioenergetical dysfunction

Glycine tissue concentrations are increased particularly in nonketotic and ketotic hyperglycinemia, inherited metabolic disorders characterized by severe neurologic damage and brain abnormalities. The present work investigated the in vitro effects of glycine on important parameters of energy metabolism in the brain of young rats. The parameters analyzed were CO2 generated from glucose, acetate and citrate and the activities of the respiratory chain complexes I–IV, of the citric acid cycle enzymes citrate synthase, aconitase, isocitrate dehydrogenase, α-ketoglutarate dehydrogenase, succinate dehydrogense, fumarase and malate dehydrogenase, of creatine kinase and Na+,K+-ATPase. Our results show that glycine significantly reduced CO2 production from acetate, but not from glucose and citrate, reflecting an impairment of the citric acid cycle function. We also observed that the activity of the mitochondrial enzyme citrate synthase was markedly inhibited by glycine, whereas the other activities of the citric acid cycle were not altered. Furthermore, the activity of the respiratory chain was reduced at complexes I–III, II–III and II, as well as of the mitochondrial isoform of creatine kinase and Na+,K+-ATPase. The data indicate that glycine severely impairs brain bioenergetics at the level of energy formation, transfer and utilization. Considering the importance of energy metabolism for brain development and functioning, it is presumed that glycine-induced impairment of brain energy homeostasis may be involved at least in part in the neurological damage found in patients affected by disorders in which brain glycine concentrations are increased.

γ-Secretase Dependent Production of Intracellular Domains Is Reduced in Adult Compared to Embryonic Rat Brain Membranes

Backgroundγ-Secretase is an intramembrane aspartyl protease whose cleavage of the amyloid precursor protein (APP) generates the amyloid β-peptide (Aβ) and the APP intracellular domain. Aβ is widely believed to have a causative role in Alzheimer's disease pathogenesis, and therefore modulation of γ-secretase activity has become a therapeutic goal. Besides APP, more than 50 substrates of γ-secretase with different cellular functions during embryogenesis as well as adulthood have been revealed. Prior to γ-secretase cleavage, substrates are ectodomain shedded, producing membrane bound C-terminal fragments (CTFs).Principal FindingsHere, we investigated γ-secretase cleavage of five substrates; APP, Notch1, N-cadherin, ephrinB and p75 neurotrophin receptor (p75-NTR) in membranes isolated from embryonic, young or old adult rat brain by analyzing the release of the corresponding intracellular domains (ICDs) or Aβ40 by western blot analysis and ELISA respectively. The highest levels of all ICDs and Aβ were produced by embryonic membranes. In adult rat brain only cleavage of APP and Notch1 could be detected and the Aβ40 and ICD production from these substrates was similar in young and old adult rat brain. The CTF levels of Notch1, N-cadherin, ephrinB and p75-NTR were also clearly decreased in the adult brain compared to embryonic brain, whereas the APP CTF levels were only slightly decreased.ConclusionsIn summary our data suggests that γ-secretase dependent ICD production is down-regulated in the adult brain compared to embryonic brain. In addition, the present approach may be useful for evaluating the specificity of γ-secretase inhibitors.

Specific monoamine exchange in the brains of young and aged male rats with hypothyroidism

We analyzed monoamine exchange in different brain structures of young (2-month-old) and aged (22–24 month-old) male rats under conditions of thyroid hormone (TH) deficit or 14-day-long intraperitoneal (T3) administration of triiodothyronine at a dose of 70 μg/kg. The contents of monoamines and their metabolites were assayed using HPLC with an electrochemical detector. In the brain structures directly related to animal behavior a TH deficit resulted in changes in monoamines, which depended on the ages of the rats. During experimental hypothyroidism, the most substantial changes in the contents and turnover of norepinephrine, dopamine, and serotonin were observed in the brains of young male rats. In aged animals, a TH deficit increased impairments in neurotransmitter exchange. Substitutive T3 administration did not improve monoamine exchange in the brain structures of young male rats with hypothyroidism. In the aged rats with hypothyroidism, T3 administration restored monoamine exchange only in the hippocampus but not in other brain structures.

Diphenyl diselenide and diphenyl ditelluride: neurotoxic effect in brain of young rats, in vitro

This study examined whether maturity of rat brain may be relevant for the sensitivity to diphenyl diselenide (PhSe)(2) and diphenyl ditelluride (PhTe)(2) on [(3)H]glutamate uptake and release, in vitro. Brain synaptosomes were isolated from young (14- and 30-day-old) and adult rats and incubated at different concentrations of (PhSe)(2) or (PhTe)(2). The results demonstrated that the highest concentration (100 microM) of (PhSe)(2) and (PhTe)(2) inhibited the [(3)H]glutamate uptake by synaptosomes of brain at all ages. In the adult brain, (PhSe)(2) did not inhibit the [(3)H]glutamate uptake at the lowest concentration (10 microM). The highest concentration of (PhTe)(2) inhibited the [(3)H]glutamate uptake more in the 14-day-old than in the 30-day-old rats or adult rats. In the 30-day-old animals, the highest concentration of (PhSe)(2), and the lowest concentration of (PhTe)(2), increased the basal [(3)H]glutamate release. At the highest concentration, (PhTe)(2) increased the basal and K(+)-stimulated glutamate release on all ages evaluated. The results suggest that (PhSe)(2) and (PhTe)(2) caused alterations on the homeostasis of the glutamatergic system at the pre-synaptic level. These alterations were age-, concentration-, and compound-dependent. The maturity of rat brain is relevant for the glutamatergic system sensitivity to (PhSe)(2) and (PhTe)(2) .

Cortical and Putamen Age-Related Changes in the Microvessel Density and Astrocyte Deficiency in Spontaneously Hypertensive and Stroke-Prone Spontaneously Hypertensive Rats

Cerebral small vessel disease (SVD) is a major contributor to dementia in the elderly, and hypertension represents a major cause for developing the disease. However, little is known about its development and progression. Modifications of large cerebral arteries due hypertension are thought to participate to the development of small ischemic infarcts, but the status of the small vessels before the establishment of hypertension is not well defined. Using spontaneously hypertensive rats (SHR) and stroke-prone SHR (SP-SHR) as a models for SVD, we analysed the effect of hypertension on the microvasculature in the cortex and putamen, and on its relationship with astrocytes in animals aged 2 to 9 months. Compared with the normotensive Wistar-Kyoto rats (WKY), the densities of the collagen type IV-positive capillaries were significantly higher in both brain areas of young SHR and SP-SHR. In contrast, the expression of the astrocytic marker GFAP was significantly lower in these animals, whereas astrogliosis was observed after 6 months in their cortex only. To investigate if chronic hypoxia occurs due to the lower number of astrocytes in young SHR and SP-SHR, we evaluated the levels of HIF-1alpha in both brain regions. The accumulation of HIF-1alpha was not observed at the youngest ages, but was apparent in neurons of 9-month-old SHR and SP-SHR. Our results indicate that the brains of young SHR and SP-SHR rats show evidence of cellular imbalance between microvessels and astrocytes at the neurovascular unit that may lead to their higher vulnerability to hypoxic events at older ages.

Evidence that the major metabolites accumulating in medium-chain acyl-CoA dehydrogenase deficiency disturb mitochondrial energy homeostasis in rat brain

Medium-chain acyl-CoA dehydrogenase deficiency (MCADD) is an inherited metabolic disorder of fatty acid oxidation in which the affected patients predominantly present high levels of octanoic (OA) and decanoic (DA) acids and their glycine and carnitine by-products in tissues and body fluids. It is clinically characterized by episodic encephalopathic crises with coma and seizures, as well as by progressive neurological involvement, whose pathophysiology is poorly known. In the present work, we investigated the in vitro effects of OA and DA on various parameters of energy homeostasis in mitochondrial preparations from brain of young rats. We found that OA and DA markedly increased state 4 respiration and diminished state 3 respiration as well as the respiratory control ratio, the mitochondrial membrane potential and the matrix NAD(P)H levels. In addition, DA-elicited increase in oxygen consumption in state 4 respiration was partially prevented by atractyloside, indicating the involvement of the adenine nucleotide translocator. OA and DA also reduced ADP/O ratio, CCCP-stimulated respiration and the activities of respiratory chain complexes. The data indicate that the major accumulating fatty acids in MCADD act as uncouplers of oxidative phosphorylation and as metabolic inhibitors. Furthermore, DA, but not OA, provoked a marked mitochondrial swelling and cytochrome c release from mitochondria, reflecting a permeabilization of the inner mitochondrial membrane. Taken together, these data suggest that OA and DA impair brain mitochondrial energy homeostasis that could underlie at least in part the neuropathology of MCADD.

Evidence that the major metabolites accumulating in hyperornithinemia–hyperammonemia–homocitrullinuria syndrome induce oxidative stress in brain of young rats

Ornithine and homocitrulline are the major metabolites accumulating in hyperornithinemia–hyperammonemia–homocitrullinuria syndrome, a genetic disorder characterized by neurological regression whose pathogenesis is still not understood. The present work investigated the in vitro effects of ornithine and homocitrulline on important parameters of oxidative stress in cerebral cortex from young rats. Ornithine significantly increased chemiluminescence and thiobarbituric acid-reactive substances levels, indicators of lipid peroxidation, while homocitrulline only augmented chemiluminescence values. Furthermore, ornithine-induced increase of thiobarbituric acid-reactive substances levels was attenuated (melatonin and reduced glutathione) or totally prevented (α-tocopherol) by free radical scavengers, suggesting that reactive species were involved in the lipid oxidative damage. We also observed that ornithine and homocitrulline significantly decreased the tissue antioxidant defenses, determined by reduced glutathione concentrations, the major non-enzymatic antioxidant defense found in the brain. Homocitrulline reduction of glutathione levels was completely prevented by melatonin and α-tocopherol, whereas ornithine-induced decrease of glutathione levels was only attenuated by these free radical scavengers. Ornithine and homocitrulline also induced protein oxidative damage, increasing carbonyl formation and sulfhydryl oxidation. In contrast, these amino acids did not affect nitric oxide production, indicating that nitrogen reactive species were not implicated in the lipid and oxidative damage provoked by ornithine and homocitrulline. Therefore, it is presumed that the major metabolites accumulating in hyperornithinemia–hyperammonemia–homocitrullinuria syndrome elicit oxidative stress and that this pathomechanism may possibly be involved in the brain damage found in patients affected by this disorder.