Protein oxidation in aging brain.
Explore the source record for details and available documents.
Aging↗
Biomedical subjects
J M Carney
Publications and source records attributed to J M Carney.
Free radical damage to protein and DNA: mechanisms involved and relevant observations on brain undergoing oxidative stress.
Iron mediates damage to proteins and DNA. The mechanisms of damage not only involve iron but also oxygen free radical intermediates. Oxidative damage to DNA causes not only strand breaks, but also formation of specific base adducts, such as 8-hydroxy-2'-deoxyguanosine. Oxidative damage also inactivates certain enzymes such as glutamine synthetase. Novel methods of assessing oxidative damage to tissue, including quantitation of salicylate hydroxylation as an index of hydroxyl free radical flux as well as specific lesions to proteins and DNA, have yielded results that clearly show that ischemia/reperfusion injury to mongolian gerbil brain involves oxidatively damaging events. Aging in gerbil as well as human brain is also associated with increased oxidative damage. Recent novel observations have shown that the spin-trapping agent phenyl alpha-tert-butylnitrone (PBN) offers protection in gerbil brain during ischemia/reperfusion injury. We also show that oxidative damage to brain during aging is decreased by chronic administration of PBN. The mechanism of action of PBN may be related to its trapping of specific free radicals, which triggers a cascade of oxidative events that eventually lead to tissue injury.
Animals↗
Protein modification in aging.
During aging a number of enzymes accumulate as catalytically inactive or less active forms. The age-related changes in catalytic activity are due in part to reactions of the protein with "active" oxygen species such as ozone, singlet oxygen, or with oxygen free radicals as are produced during exposure to ionizing radiation or to metal ion catalyzed oxidation (MCO) systems. The levels of oxidized proteins in cultured human fibroblasts from individuals of various ages and in liver and brain extracts of rats of different ages increase progressively with age, and in old rats can represent 30-50% of the total cellular protein. The age-related increase in oxidized protein in rat liver and brain tissue is accompanied by a loss of glutamine synthetase (GS) and glucose-6-P dehydrogenase (G-6-PDH) activities, and to a decrease in the level of cytosolic neutral protease activity which is responsible for the degradation of oxidized (denatured) protein. Of particular significance are the results of experiments showing that similar age-related changes occur in the gerbil brain and that these changes are accompanied by a loss of short-term memory as measured by the radial arm maze technique. Chronic treatment (intraperitoneal injections) of old animals with the free radical spin-trap reagent, N-tert-butyl-alpha-phenylnitrone (PBN) resulted in normalization of the several biochemical parameters to those characteristic of the young animals; coincidentally, the short-term memory index was restored to the young animal values. These results provide the first evidence that there is likely a linkage between the age-dependent accumulation of oxidized enzymes and the loss of physiological function.
Aging↗
Phencyclidine-induced desensitization of striatal dopamine release.
Increased release of dopamine (DA) and dihydroxyphenylacetic acid (DOPAC) from slices of striatum of DBA/2J mouse in response to administration of phencyclidine (PCP) in vitro has been observed to be transient, despite continued exposure to PCP. To determine whether this transient response was a result of depletion of releasable pools, toxicity or an adaptive response (desensitization), the recovery of the response to PCP was evaluated. Slices in the control condition were exposed to PCP (300 microM) during test-exposure only. Slices in the PCP pre-exposure condition, were exposed first to PCP (30 microM; pre-exposure) and subsequently to PCP (300 microM; test-exposure). During a washout period (0, 30, 60 or 120 min) between exposures to PCP, the slices were superfused in the absence of PCP. Pre-exposure to PCP diminished the subsequent response to test-exposure to PCP (49 and 37% of control for DA and DOPAC, respectively) after 0 min washout. Overflow of DA evoked by PCP returned towards control values but remained decreased (66% of control) after up to 120 min washout. However, overflow of DOPAC did not return to control values after 60 min washout. Thus, the diminished dopaminergic response, resulting from continued exposure to PCP was not due to depletion of the releasable pool or cytotoxicity but rather to a dynamic adaptive response of the dopaminergic neuron to PCP.
3,4-Dihydroxyphenylacetic Acid↗
Excess brain protein oxidation and enzyme dysfunction in normal aging and in Alzheimer disease.
The relationship between Alzheimer disease (AD) and aging is not currently known. In this study, postmortem frontal- and occipital-pole brain samples were obtained from 16 subjects with AD, 8 age-matched controls, and 5 young controls. These samples were analyzed both for protein oxidation products (carbonyl) and the activities of two enzymes vulnerable to mixed-function oxidation, glutamine synthetase and creatine kinase. Glutamine synthetase is more sensitive to mixed-function oxidation than creatine kinase. Carbonyl content rises exponentially with age, at double the rate in the frontal pole compared with the occipital pole. Compared with young controls, both aged groups (AD and age-matched controls) have increased carbonyl content and decreased glutamine synthetase and creatine kinase activities, which are more marked in the frontal than occipital pole in all instances. We conclude that protein oxidation products accumulate in the brain and that oxidation-vulnerable enzyme activities decrease with aging in the same regional pattern (frontal more affected than occipital). However, only glutamine synthetase activity distinguishes AD from age-matched controls: Because glutamine synthetase activity is differentially reduced in the frontal pole in AD, we suggest that AD may represent a specific brain vulnerability to age-related oxidation.
Adult↗
Reversal of age-related increase in brain protein oxidation, decrease in enzyme activity, and loss in temporal and spatial memory by chronic administration of the spin-trapping compound N-tert-butyl-alpha-phenylnitrone.
Oxygen free radicals and oxidative events have been implicated as playing a role in bringing about the changes in cellular function that occur during aging. Brain readily undergoes oxidative damage, so it is important to determine if aging-induced changes in brain may be associated with oxidative events. Previously we demonstrated that brain damage caused by an ischemia/reperfusion insult involved oxidative events. In addition, pretreatment with the spin-trapping compound N-tert-butyl-alpha-phenylnitrone (PBN) diminished the increase in oxidized protein and the loss of glutamine synthetase (GS) activity that accompanied ischemia/reperfusion injury in brain. We report here that aged gerbils had a significantly higher level of oxidized protein as assessed by carbonyl residues and decreased GS and neutral protease activities as compared to young adult gerbils. We also found that chronic treatment with the spin-trapping compound PBN caused a decrease in the level of oxidized protein and an increase in both GS and neutral protease activity in aged Mongolian gerbil brain. In contrast to aged gerbils, PBN treatment of young adult gerbils had no significant effect on brain oxidized protein content or GS activity. Male gerbils, young adults (3 months of age) and retired breeders (15-18 months of age), were treated with PBN for 14 days with twice daily dosages of 32 mg/kg. If PBN administration was ceased after 2 weeks, the significantly decreased level of oxidized protein and increased GS and neutral protease activities in old gerbils changed in a monotonic fashion back to the levels observed in aged gerbils prior to PBN administration. We also report that old gerbils make more errors than young animals and that older gerbils treated with PBN made fewer errors in a radial arm maze test for temporal and spatial memory than the untreated aged controls. These data can be interpreted to indicate that oxidation of cellular proteins may be a critical determinant of brain function. Moreover, it also implies that there is an age-related increase in vulnerability of tissue to oxidation that can be modified by free radical trapping compounds.
Aging↗
Protection against oxidative damage to CNS by alpha-phenyl-tert-butyl nitrone (PBN) and other spin-trapping agents: a novel series of nonlipid free radical scavengers.
Brain is extremely susceptible to oxidative damage. Utilizing a series of novel approaches, we have demonstrated that oxidative damage occurs during an ischemia/reperfusion insult (IRI) to brain. Thus, we have demonstrated that an IRI to Mongolian gerbil brain results in: (1) an enhanced rate of salicylate hydroxylation, implicating an increased flux of hydroxyl free radicals; (2) an enhanced flux of free radicals as determined by spin-trapping; (3) an enhanced level of endogenous protein oxidation; (4) a decrease in glutamine synthetase (GS) activity, an enzyme very sensitive to oxidative damage; and (5) demonstration of protection from an IRI by administering the spin-trapping agent alpha-phenyl-tert-butyl nitrone (PBN). The novel observation that PBN offers protection from the lethality brought on by a brain IRI appears to be clearly linked to the ability of the administered spin-trap to inhibit oxidative damage as evidenced by the decreased amount of brain protein oxidation and the prevention of an IRI-mediated loss of GS activity in treated animals. Aged gerbils are more sensitive to the lethal action of a brain IRI than younger animals, but they are protected by PBN administration as are the younger animals. Older gerbils have a significantly higher level of oxidized protein in the brain. Older gerbils have decreased activities of GS and neutral protease, the enzyme that removes oxidized protein, than younger animals. Chronic twice daily administration of PBN (32 mg/kg) for 14 days to older animals significantly lowered brain oxidized protein levels and raised GS and neutral protease activity to those observed in younger animals. Cessation of PBN administration resulted in a time-dependent restoration of protein oxidation levels and enzyme activities back to those observed prior to spin-trap administration. Older gerbils exhibit significantly higher errors in a radial arm maze than younger animals, but older gerbils that had received chronic daily treatments of PBN (32 mg/kg) for 14 days committed significantly less errors than untreated controls. The errors committed in PBN-treated animals was decreased down to the level of those observed in younger animals. Clearly the spin-trapping agent, PBN, appears to have promise in: (1) elucidation of the role of oxidative damage in normal brain function during aging, (2) understanding the development of pathological conditions, and (3) development of treatment regimens for prevention of damage that occurs during the development of pathological conditions and in aging.
Animals↗
Selective effects of behaviorally active doses of methamphetamine on mRNA expression in the gerbil brain.
Administration of methamphetamine results in neuronal damage that may be mediated through the production of oxygen-free radicals and modulations in levels of calcium and glutamate in the brain. These changes have been associated with alterations in gene expression, which may play a role in cell damage. To assess the differences in gene expression related to treatment with methamphetamine, levels of mRNA were evaluated for the proto-oncogene c-fos, heat-shock protein (HSP 70) and actin. It was found that c-fos mRNA expression increased in a dose-dependent manner after administration of methamphetamine. Whereas, levels of HSP mRNA dropped at small doses of methamphetamine and increased dramatically at large doses. In addition, both c-fos and HSP mRNA showed increased levels throughout the brain. Actin mRNA expression was unaffected by any dose of the drug. At the doses that altered gene expression, methamphetamine produced dose-related behavioral changes. Spontaneous locomotor activity was increased at 1.0 mg/kg of methamphetamine, while, larger doses did not alter activity. The data demonstrated that effects of methamphetamine on gene expression occurred within the behaviorally-active dose range and might correlate with the degree of neuronal damage. Selective modulation of gene expression may have a role in determining the acute quantitative and qualitative effects of methamphetamine and the long-term changes that occur after administration of methamphetamine.
Animals↗
Modulation of ornithine decarboxylase mRNA following transient ischemia in the gerbil brain.
Ornithine decarboxylase (ODC) is the rate-limiting enzyme that catalyzes the synthesis of polyamines from ornithine and is thought to be involved in the cellular response to growth, differentiation, and stress. Previous studies have demonstrated that transient cerebral ischemia results in an increase in ODC activity and polyamine synthesis. We have used the Mongolian gerbil as a model system to test the hypothesis that the cellular response to ischemia induces a distinct pattern of ODC gene expression. Our results indicate that transient ischemia, induced by bilateral carotid occlusion, elevates ODC mRNA within 1-4 h after reperfusion, which correlates with increased ODC activity and polyamine synthesis. Increased ODC mRNA can be detected in the forebrain, striatum, hippocampus, and midbrain but not the cerebellum, which is not subject to ischemic injury. In contrast, c-fos mRNA increased by 15 min after reperfusion and actin mRNA did not demonstrate alterations in level after ischemia. Pentobarbital prevented the increase in ODC mRNA, whereas the glutamate antagonist MK-801 had no effect on the elevation of ODC gene expression after ischemia. We conclude that the ischemia-induced increase in ODC enzyme activity may be attributed in part to transcriptional activation of the ODC gene.
Animals↗
Establishment of chronic intravenous drug self-administration in the C57BL/6J mouse.
A wide variety of drugs that have significant human abuse potential have been demonstrated to function as positive reinforcers in animals. The present study was designed to characterize a new mouse model of chronic intravenous drug self-administration. Adult male C57BL/6J mice, implanted with external jugular infusion catheters, were given access to response-contingent injections. They did not initiate responding for saline delivery, whereas the C57BL/6J mice initiated morphine, cocaine, methamphetamine and pentobarbital self-administration. Drug-maintained responding was consistently and significantly higher for each compound than for saline responding. In contrast to C57BL/6J mice, DBA/2J mice failed to initiate cocaine self-administration. Thus, chronic intravenous drug self-administration procedures can be adapted to the inbred mouse.
Animals↗
Genetic determinants of susceptibility to the rewarding and other behavioral actions of cocaine.
The role of genotype as a determinant of biologically based inter-individual differences in vulnerability to substance abuse has received little systematic investigation except in the case of alcohol. This report describes the use of an animal model, the inbred mouse, to identify and to characterize variants with inherently altered susceptibilities to the rewarding and other behavioral actions of cocaine. Among a battery of nine inbred strains chosen solely for their genetic diversity, genetic polymorphisms commonly occurred which altered the potency and/or efficacy of cocaine to induce conditioned place preference, oral self-administration, motor activity activation, seizures and lethality. These changes in cocaine sensitivity generally were of a behavior-specific and pharmacodynamic nature. One strain, DBA/2J, found to be markedly hyporesponsive to the rewarding action of cocaine, also was hyporesponsive to the rewarding effects of amphetamine, etonitazene, phencyclidine, caffeine and procaine. We speculate that this strain has an inherent generalized appetitive defect. The frequent occurrence and large magnitude of inherent phenotypic changes in cocaine responsiveness which we have identified among inbred mouse strains now permits an analytical genetic study of processes underlying cocaine-mediated reinforcement.
Animals↗
Issues surrounding the assessment of the genetic determinants of drugs as reinforcing stimuli.
A wide variety of drugs used in medicine and on a non-prescription basis have abuse liability. However, not all psychoactive drugs have demonstrable reinforcing properties either in animal models or in humans. In order to predict the abuse liability of a compound or to understand determinants of the abuse liability, several animal testing procedures have been developed. These procedures include tests for physical dependence, tolerance, disruption of ongoing behaviors, discriminative stimulus properties, and the direct or indirect assessment of the reinforcement. While tests for the production of tolerance and physical dependence may not predict abuse liability, they would significantly modify the abuse risk. This paper will focus on the methods available for the assessment of the reinforcing properties and the advantages of using genetically defined mice to better understand the determinants of the reinforcing properties and to isolate phenotypes that are hyper-responsive and hypo-responsive to the reinforcing properties of drugs under study. Examples of genetically determined differences are provided and areas of inadequate information are discussed.
Animals↗
Induction of the protooncogene c-fos and recovery of cytosolic adenosine triphosphate in reperfused liver after transient warm ischemia: effect of nitrone free-radical spin-trap agents.
Ischemia and reperfusion stimulate several adenosine triphosphate (ATP)-dependent processes involving release of substances including free radicals. This cellular response is mediated through receptors responsive to transcriptional products of gene expression; c-fos acts as a transcriptional factor involved in the regulation of genes associated with cellular proliferation and differentiation. We hypothesized that nitrone free-radical spin traps promote restoration of cytosolic ATP during reperfusion and prevent c-fos induction. Four control rats had no ischemia. Global hepatic ischemia was induced in 19 rats in four groups: saline solution, phenyl-N-tert-butyl nitrone (PBN), alpha 1-pyridyl-N-oxide N-tert-butyl nitrone (POBN), and 5,5-dimethyl-1-pyrroline-N-oxide (DMPO). ATP and intracellular pH were measured at intervals before, during, and after ischemia. At 90 minutes of reperfusion, liver c-fos mRNA was measured. A fourfold elevation of c-fos occurred in the saline-treated group (p less than 0.001). PBN and POBN groups did not differ from the saline group. DMPO resulted in significantly less induction of c-fos than did NS. ATP depletion and recovery in all treatment groups was similar to that of the saline group. We conclude that (1) nitrone spin traps do not prevent c-fos induction or alter the pattern of ATP recovery after hepatic ischemia and reperfusion and (2) c-fos induction is not necessary for restoration of ATP, but the rate of ATP restoration is inversely related to c-fos induction.
Adenosine Triphosphate↗
Ischemic induction of protooncogene expression in gerbil brain.
Cerebral ischemia and reperfusion results in an active series of metabolic events, eventually leading to cell death. The expression of specific genes during cerebral ischemia and reperfusion may play an important, determinant role in the mechanisms controlling cellular processes. Ten minutes of bilateral carotid occlusion in the Mongolian gerbil was found to increase the messenger RNA for both the c-fos and c-jun protooncogenes. The changes in gene expression were detected in the regions of ischemia, specifically the cortex and striatum, and no increases were seen in either the brain stem or the cerebellum, which possess a separate circulation. Induction of protooncogene mRNA is correlated to the duration of ischemia, i.e., the longer the time of ischemia, the greater the increase in c-fos expression. Pretreatment of animals with pentobarbital reduced the effect of the ischemic insult and prevented the increase in c-fos mRNA. Analysis of the c-fos and c-jun proteins after ischemia demonstrated an increase in the formation of a functional transcriptional complex and association with the AP-1 binding region. These findings suggest that ischemic cell death and recovery in neurodegenerative disorders such as stroke may involve the regulated expression of these protooncogenes early in the pathway of ischemia.
Animals↗
Inducible ornithine decarboxylase expression in brain subject to vasogenic oedema after transient ischaemia: relationship to C-fos gene expression.
Ornithine Decarboxylase (ODC) is the rat controlling enzyme of polyamine biosynthesis and has been shown to be produced in a delayed fashion in response to cerebral ischaemia. Its appearance has been linked to the development of vasogenic brain oedema. To understand the genetic control of this protein, Mongolian gerbils were studied for the possible expression of the ODC gene as compared to that of the inducible proto oncogenes c-fos and c-jun after transient bilateral carotid artery occlusion. Total cellular RNA was isolated from gerbil brains by guanidine-thiocyanate extraction and characterized by northern blot analysis for c-fos, c-jun, and ODC mRNA over reperfusion times. c-fos and c-jun expression rose rapidly with peak level reached at 60 min of reperfusion (70 x control, p less than or equal to 0.01). Peak levels of ODC mRNA induction were seen at 4 hrs reperfusion (2.83 x control, p less than or equal to 0.01) consistent with the period of maximum of brain oedema as measured by specific gravity (1.0386 +/- 0.0009, p less than or equal to 0.05). These data indicate the differential timing of genetic expression during the reperfusion period after transient ischaemia. Such studies suggest that potential therapies may be possible by addressing the delayed ODC component of ischaemic oedema formation and allow a greater understanding of the role of gene induction in the multifaceted cerebral response to ischaemia.
Animals↗
Oxidative damage to brain proteins, loss of glutamine synthetase activity, and production of free radicals during ischemia/reperfusion-induced injury to gerbil brain.
Free radical-mediated oxidative damage has been implicated in tissue injury resulting from ischemia/reperfusion events. Global cortical ischemia/reperfusion injury to Mongolian gerbil brains was produced by transient occlusion of both common carotid arteries. Protein oxidation, as measured by protein carbonyl content, increased significantly during the reperfusion phase that followed 10 min of ischemia. The activity of glutamine synthetase, an enzyme known to be inactivated by metal-catalyzed oxidation reactions, decreased to 65% of control levels after 2 hr of reperfusion that followed 10 min of ischemia. We also report that the free radical spin trap N-tert-butyl-alpha-phenylnitrone [300 mg/kg (body weight)] administered 60 min before ischemia/reperfusion is initiated, partially prevents protein oxidation and protects from loss of glutamine synthetase activity. In addition, we report a N-tert-butyl-alpha-phenylnitrone-dependent nitroxide radical obtained in the lipid fraction of the ischemia/reperfusion-lesioned brains, but there was very little radical present in the comparable sham-operated control brains. These data strengthen the previous observation utilizing in vivo-trapping methods, that free radical flux is increased during the reperfusion phase of the ischemia-lesioned gerbil brain. The loss of glutamine synthetase would be expected to increase the levels of brain L-glutamate. Thus, the oxidative inactivation of glutamine synthetase may be a critical factor in the neurotoxicity produced after cerebral ischemia/reperfusion injury.
Animals↗
Behavioral interaction between cocaine and caffeine: a drug discrimination analysis in rats.
The effects of caffeine upon the discriminative and rate-altering effects of cocaine were examined in rats. Using a food-reinforced two-lever operant procedure, 12 Sprague-Dawley male rats were trained to discriminate between 10 mg/kg cocaine and saline. Stimulus generalization tests with both cocaine and amphetamine resulted in a dose-related increase in cocaine-appropriate responding. A variable response rate topography was produced by cocaine. Caffeine also engendered a dose-related increase in cocaine-appropriate responding and resulted in a potency ratio of 15:1 when compared to cocaine. In contrast, increasing doses of caffeine produced a biphasic response rate function (first increases and then decreases). Response choice data suggested a potency relationship of amphetamine greater than cocaine greater than caffeine. Caffeine potentiated the discriminative stimulus properties of cocaine. Isobolographic analysis characterized this interaction as simple additivity. However, caffeine's effects upon the rate-altering effects of cocaine resulted in a biphasic interaction pattern. With low doses of cocaine in combination with various doses of caffeine, the interaction for rate reduction is best categorized as "supra-additive," in contrast, increasing either the cocaine dose or caffeine dose could change the interaction to simple additivity and/or infra-additivity.
Animals↗
Effects of caffeine, cocaine and their combination on fixed-interval behavior in rats.
The effects of the central nervous system stimulants, caffeine and cocaine, on schedule-controlled behavior were determined in rats trained to perform a fixed-interval (FI) 5-minute task. When given alone caffeine produced a doubling of FI response rate at a dose of 10 mg/kg and reduced responding at a dose of 32 mg/kg. Cocaine, which was also expected to increase FI responding, did not increase response rate at doses of 3.2 or 10 mg/kg and decreased the rate of responding at a dose of 32 mg/kg. Caffeine had minimal effects on quarter life and appeared to increase local rates of responding across the interval. Cocaine decreased quarter life dramatically at a dose that had no effect on overall response rate. Local rates of responding were increased early in the interval and decreased in the later segments. The effects of both drugs were found to be rate-dependent. When these compounds were given in combination the results obtained appeared to be related to the rate of responding that caffeine alone would produce.
Animals↗