Effect of a 3-hydroxypyridine-class antioxidant on the functional activity of the central nervous system in mice of different age groups

Characteristics of emotional reactivity, orienting-investigating behaviour and of preservation of conditioned reflex of passive avoidance were used for estimation of the functional activity of animals CNS and also of psychotropic effect of antioxidant 2-ethyl-6-methyl-3-oxypyridine (3-OP) on mice of different age. Studies were carried out on white outbred young (3 months) and elderly (15 months) male mice. Different character was observed of the emotional reactions of different age mice to aversive stimuli and to the open field conditions. Differences were noted of psychotropic effects of 3-OP in 3 and 15-months animals.     

The effect of a 3-hydroxypyridine-class antioxidant on disorders of the microcirculatory system in experimental dyslipoproteinemia and its alimentary correction

The effect of antioxidant (AO) of the 3-hydroxypyridin class on microcirculatory system (MC) disorders of rabbits with dyslipoproteinemia (DLP) and its alimentary correction (the standard ration--9 months) was studied. Aorta, the intestinal mesentery microcirculatory bed, the microvessels adrenergic innervation, erythrocyte morphology were examined. The various lipoprotein fractions and lipid peroxidation (LP) products were studied in venous blood. After AO including into atherogenic diet (ATD) the lipid homeostasis disorders, the LP activation, MC disturbances, anomalous erythrocyte form appearance and atherosclerosis changes in aorta were less pronounced. Under the AO influence when DLP was corrected the initial level of lipid metabolism and LP restored, the MC disorders and the atherosclerosis changes in aorta regressed in a shorter period of time. The interrelation was observed between the DLP level, the structural and functional MC disorders and the extent of aorta atherosclerosis. Thus, DLP and LP correction at the early stages of pathological process can lead to the disappearance of MC disorders, what seems important for the prevention of atherosclerosis. The role of the LP activation in MC disorders in DLP and mexidol effect on them, especially in complex with DLP correction were evaluated.     

Age-dependent accumulation of lipofuscin in perivascular and subretinal microglia in experimental mice

Fundus autofluorescence (AF) imaging by confocal scanning laser ophthalmoscopy has been widely used by ophthalmologists in the diagnosis/monitoring of various retinal disorders. It is believed that fundus AF is derived from lipofuscin in retinal pigment epithelial (RPE) cells; however, direct clinicopathological correlation has not been possible in humans. We examined fundus AF by confocal scanning laser ophthalmoscopy and confocal microscopy in normal C57BL/6 mice of different ages. Increasingly strong AF signals were observed with age in the neuroretina and subretinal/RPE layer by confocal scanning laser ophthalmoscopy. Unlike fundus AF detected in normal human subjects, mouse fundus AF appeared as discrete foci distributed throughout the retina. Most of the AF signals in the neuroretina were distributed around retinal vessels. Confocal microscopy of retinal and choroid/RPE flat mounts demonstrated that most of the AF signals were derived from Iba-1+ perivascular and subretinal microglia. An age-dependent accumulation of Iba-1+ microglia at the subretinal space was observed. Lipofuscin granules were detected in large numbers in subretinal microglia by electron microscopy. The number of AF+ microglia and the amount of AF granules/cell increased with age. AF granules/lipofuscin were also observed in RPE cells in mice older than 12 months, but the number of AF+ RPE cells was very low (1.48 mm(-2) and 5.02 mm(-2) for 12 and 24 months, respectively) compared to the number of AF+ microglial cells (20.63 mm(-2) and 76.36 mm(-2) for 6 and 24 months, respectively). The fluorescence emission fingerprints of AF granules in subretinal microglia were the same as those in RPE cells. Our observation suggests that perivascular and subretinal microglia are the main cells producing lipofuscin in normal aged mouse retina and are responsible for in vivo fundus AF. Microglia may play an important role in retinal aging and age-related retinal diseases.     

Effect of ethanol on lipofuscin accumulation in cultured rat cardiac myocytes

The objective of this study was to determine the effect of ethanol on in vitro life span, rate of contraction and lipofuscin content of neonatal rat cardiac myocytes. Lipofuscin was quantified by microspectrofluorometry. The effects of 0, 3.1, 6.5, and 12.5 mM ethanol on myocytes, kept under an ambient oxygen concentration of 20% and 40%, were studied. Exposure to low concentrations of ethanol resulted in a decrease in the amount of lipofuscin whereas exposure to high concentration of ethanol caused an increase in the level of lipofuscin. The length of cell survival in controls and 3.1 mM ethanol exposed myocytes was similar under 20% oxygen, but was longer in the latter group under 40% oxygen, as compared to controls. The total number of contractions in 3.1 mM ethanol-exposed myocytes were, respectively, 4% and 8% higher under 20% and 40% oxygen atmosphere than in control cells.     

Effect of chronically administered methylxanthines on ethanol-induced motor incoordination in mice

The effect of chronic (10 days) administration of methylxanthines, caffeine, IBMX and theophylline on acute ethanol-induced motor incoordination has been investigated in the mice. In animals that received caffeine, 45 and 90 mg/kg/24 h, ethanol, 1.5 g/kg, produced motor incoordination significantly greater compared to that in the control groups. Significantly greater ethanol-induced motor incoordination was seen in animals fed IBMX, 30 and 60 mg/kg/24 h, compared to controls. Ethanol-induced increased motor incoordination in caffeine and IBMX-fed animals was also associated with significantly greater 3H-R-PIA binding in whole brains compared to tap water controls indicating an increase in brain adenosine binding sites. However neither motor incoordination nor 3H-R-PIA binding was altered in theophylline 75 and 150 mg/kg/24 h, fed animals. The increased motor incoordination associated with increased adenosine binding sites in the brains of caffeine and IBMX-fed animals suggests an involvement of central adenosine mechanisms in the motor incoordinating effect of ethanol and further supports our earlier suggestion for the role of adenosine in some of the central effects of ethanol.     

Effect of Maharishi Amrit Kalash an ayurvedic herbal mixture on lipid peroxidation and neuronal lipofuscin accumulation in ageing guinea pig brain

The effects of ayurvedic herbal mixture Maharishi Amrit Kalash(MAK) were studied on brain lipid peroxidation, oxygen consumption, and lipofuscin accumulation in 10 months and 32 months old guinea pigs. Brain regions studied were cerebral cortex, hypothalamus, cerebellum and spinal cord. Parameters assessed were lipid peroxidation, oxygen consumption, and lipofuscin accumulation. The endogenous lipid peroxide was found to be increased significantly (P < 0.05) in the 32-month-old animals. Neuronal lipofuscin accumulation in the neurons of cerebral motor cortex, cerebellum and cervical spinal cord was increased (P < 0.05) in the older animals. Oxygen consumption was found to be decreased significantly(P < 0.05) in the 32-month old guinea pigs. Treatment with MAK at a dose of 500 mg/kg body weight daily for two months reduced the lipid peroxidation and lipofuscin pigment accumulation significantly in brain regions and it also helped in restoring the normal oxygen consumption in the older animals. This indicates antioxidant properties of MAK.     

Effect of alpha-tocopherol and some metal chelators on lipofuscin accumulation in cultured neonatal rat cardiac myocytes

The objective of this study was to test further the hypothesis that oxidative stress is a major causal factor in lipofuscin formation. We have previously shown that cultured cardiac myocytes constitute a suitable model system for the study of factors influencing lipofuscinogenesis. The specific aim of the present study was to elucidate the effects of the chain-breaking free radical scavenger alpha-tocopherol, and the chelators desferrioxamine, EDTA and DTPA on the accumulation of lipofuscin. The effects were examined at different degrees of oxidative stress, obtained by varying the ambient oxygen concentration from 5 to 40%. Lipofuscin was quantified by microspectrofluorometry. Lipofuscin-specific, yellow autofluorescence increased with time in culture, and with enhanced oxidative stress. Increasing concentration of alpha-tocopherol, up to 40 microM, had an inhibitory effect on lipofuscin accumulation that was most pronounced at high oxidative stress. Desferrioxamine and DTPA, both caused a pronounced reduction in lipofuscin formation, while EDTA had no significant effect. The findings are interpreted to support the concept that oxidative stress is a causal factor in lipofuscinogenesis, and that lipofuscin is a product of autophagocytosed, membrane-rich material subjected to free radical-induced, metal-catalyzed peroxidation, fragmentation, and polymerization within the lysosomal vacuome.     

The effect of dietary antioxidants on lipofuscin accumulation in the crustacean brain

Lipofuscin accumulation is associated with ageing at the subcellular level. A strong correlation between lipofuscin and age has been found in crustaceans using histological techniques. This association has been proposed as the basis for a methodology to age crustaceans and in some cases lipofuscin levels were found to be better correlated with age than size. The experiment presented here was designed to test the potential effect of diet, in particular dietary antioxidants, on lipofuscin accumulation and age estimation.The shrimp, Penaeus japonicus, was reared in an aquaculture facility and fed commercial pellets with modified vitamins C and E contents. One group was fed with levels of vitamins C and E of 1000 and 150 mg/kg, respectively, and another group with 2500 and 5000 mg/kg, respectively. The experiment started when the shrimp were 19 weeks old. Samples were obtained at this point and at ages 33 and 43 weeks. Lipofuscin was measured in the nerve cords (antennal neuropils and oesophageal connectives) in an area adjacent to the brain.Dietary antioxidants significantly affected lipofuscin levels. High vitamin content in the diet resulted in lower percentage of the observed area covered with lipofuscin, lower lipofuscin granule density and lower average granule size. Gender had no effect on any of these variables and granule size did not significantly change within each treatment. Lipofuscin area and granule density increased with age in both vitamin treatments.These results suggest that age estimation using lipofuscin indices may be biased when: (1) wild populations are dispersed over diverse environments; (2) the age estimation of wild individuals is based on the results obtained using laboratory-reared individuals.     

Use it and lose it: lipofuscin accumulation in the midbrain of a coral reef fish

Lipofuscin, an autofluorescent biomarker of physiological wear-and-tear, was concentrated in those areas of a fish's midbrain responsible for visual performance, suggesting a potentially strong link between physiological specialization, ecological adaptation and senescence.     

C20-D3-vitamin A slows lipofuscin accumulation and electrophysiological retinal degeneration in a mouse model of Stargardt disease

Stargardt disease, also known as juvenile macular degeneration, occurs in approximately one in 10,000 people and results from genetic defects in the ABCA4 gene. The disease is characterized by premature accumulation of lipofuscin in the retinal pigment epithelium (RPE) of the eye and by vision loss. No cure or treatment is available. Although lipofuscin is considered a hallmark of Stargardt disease, its mechanism of formation and its role in disease pathogenesis are poorly understood. In this work we investigated the effects of long-term administration of deuterium-enriched vitamin A, C20-D(3)-vitamin A, on RPE lipofuscin deposition and eye function in a mouse model of Stargardt's disease. Results support the notion that lipofuscin forms partly as a result of the aberrant reactivity of vitamin A through the formation of vitamin A dimers, provide evidence that preventing vitamin A dimerization may slow disease related, retinal physiological changes and perhaps vision loss and suggest that administration of C20-D(3)-vitamin A may be a potential clinical strategy to ameliorate clinical symptoms resulting from ABCA4 genetic defects.     

Oxidative stress and antioxidant defenses in ethanol-induced cell injury

Although in the past several mechanisms and factors have been proposed to be responsible for alcoholic liver disease (ALD), at present the involvement of oxygen free radicals and consequently of oxidative stress has acquired remarkable credit. In numerous experimental studies it has been shown the occurrence of alcohol-induced generation of oxygen- and ethanol-derived free radicals through different pathways and from different sources. Mitochondria appear to be both an important source of reactive oxygen species (ROS) and also a primary target of ethanol-induced damage. The consistent induction of the mitochondrial antioxidant enzyme manganese superoxide dismutase (Mn-SOD) observed in experimental animals after acute and chronic ethanol administration has all the characteristics of a "stress response" to an oxidative insult.     

On the mechanism of ethanol-induced accumulation of triglycerides in the liver

The possibility that ethanol or acetaldehyde has a direct effect on the activity of acyl-CoA-ligases or $\text{sn}$-glycerophosphate acyltransferases or on the biosynthesis of phosphatidic acid and triglycerides from free fatty acids was studied with subcellular preparations from rat liver. No stimulatory effect of ethanol or acetaldehyde could be observed in any case. It was further shown that the microsomal fraction of homogenate of livers of rats treated with ethanol (single peroral dose of 4.5 g of ethanol per kg body weight) did not have an increased capacity to biosynthesize phosphatidic acid. The possibility was excluded that excess cofactors necessary for formation of phosphatidic acid are responsible for the higher accumulation of triglycerides in livers of rats treated with ethanol.The results indicate that the increased formation of triglycerides in liver of rats treated with ethanol is not due to increased activity of acyl-CoA-ligase or $\text{sn}$-glycerophosphate acyltransferase or due to increased availability of $\text{sn}$-glycerophosphate, ATP or CoA-SH. It is suggested that increased availability of fatty acids is the major explanation for the increased accumulation of triglycerides in the liver after ethanol administration.     

Effect of ambient oxygen concentration on lipofuscin accumulation in cultured rat heart myocytes--a novel in vitro model of lipofuscinogenesis

The objective of this study was to elucidate the factors involved in the accumulation of lipofuscin in post-mitotic cells. The hypothesis that oxidative stress accelerates the rate of lipofuscin accumulation was tested by examining the effects of 5%, 20%, and 40% ambient oxygen concentration on lipofuscin content in cultured rat cardiac myocytes. Lipofuscin was quantified by microspectrofluorometry at 7 and 12 days of in vitro age. Lipofuscin-emitted yellow autofluorescence increased in direct relationship to ambient oxygen concentration with age. Transmission electron microscopic examination of the cells after 3, 8, and 12 days in culture indicated a progressive time and oxygen dependent increase in the frequency and size of lipofuscin organelles. The results are interpreted to suggest that oxidative stress is one of the causal factors in the accumulation of lipofuscin.     

Effects of prolonged ACTH-stimulation on adrenocortical accumulation of lipofuscin granules in aged rats

Subcellular deposition of lipofuscin granules is a marker of aging. Human and rodent adrenal cortices accumulate lipofuscin granules with age, but the mechanism that leads to the accumulation is not known. The ultrastructural appearance of lipofuscin granules resembles that of secondary lysosomes. Since adrenocortical subcellular events are predominantly influenced by ACTH action, we therefore studied the effect of prolonged ACTH-stimulation on adrenocortical accumulation of secondary lysosome-like granules, designated herein as lipofuscin granules. Using aged Fischer 344 male rats as a model, we found that a 7 day ACTH stimulation exerts a reducing effect on adrenocortical lipofuscin accumulation. Thus, adrenocortical accumulation of lipofuscin granules with age in vivo may not be an irreversible process.     

Effects of anabolic steroids and antioxidant vitamins on ethanol-induced tissue injury

Various mechanisms are involved in the process of ethanol-induced tissue impairment. Oxidative stress and its effects are among the most important. We compared the effects of antioxidant vitamins (vitamin C and E in combination) and steroids (testosterone and nandrolone separately) on the toxicity of ethanol in rats. Animals (male Wistar rats, n = 48) were randomised into following groups-Control, Ethanol, Testosterone, Ethanol + Testosterone, Ethanol + Nandrolone, Ethanol + Vitamins. Alcohol was given daily by gavage in a dose of 5 g/kg of body weight. On the 27th day of the study the animals were sacrificed by decapitation and tissue samples were taken. Metabolic status, parameters of the hepatic metabolism, hormone levels (testosterone, ACTH, corticosterone), lipoperoxidation markers (malondialdehyde and conjugated diens in forebrain cortex and in cerebellum) and advanced glycation end-products were analysed. Tissue samples underwent histological examination. Histological outcomes showed a protective effect of antioxidants on hepatic and cerebellar injury caused by chronic ethanol intake. Anabolic steroids protected especially the central nervous tissue against the toxicity of alcohol. Both, antioxidant vitamins and anabolic steroids protect against the ethanol-induced toxicity, however, this effect is tissue specific.     

百草枯对小鼠神经行为发育及学习记忆功能的影响

目的观察百草枯(PQ)对发育期C57BL/6J小鼠神经发育的毒性作用,并探讨百草枯对小鼠学习记忆的影响。方法 80只出生21日龄的仔鼠分为对照组(生理盐水)、1.25、2.5、5、10 mg/(kg·d)五组,灌胃染毒百草枯,每天一次,连续30 d。观察小鼠的一般生理和神经行为发育情况,并在染毒结束后进行Morris水迷宫实验和避暗实验,测试小鼠的学习记忆功能。神经行为学测试结束后取小鼠大脑,称重并进行病理检查,同时利用透射电镜观察各组小鼠中脑黑质部超微结构。结果染毒期间小鼠一般状况没有明显变化,染毒结束后各组体重没有统计学差异;在Morris水迷宫测试中,各组差异没有统计学意义,而避暗实验中与对照组相比,高剂量组的避暗潜伏期延长,差异有显著性(P<0.05);在病理切片和透射电镜观察中,在高剂量组分别观察到黑质细胞减少和神经元细胞凋亡。结论百草枯暴露对发育期小鼠成年后神经行为有影响,同时会使小鼠成年后出现脑组织的病理变化,发生器质性的病变。     

Loss of lysosomal membrane protein NCU-G1 in mice results in spontaneous liver fibrosis with accumulation of lipofuscin and iron in Kupffer cells

Human kidney predominant protein, NCU-G1, is a highly conserved protein with an unknown biological function. Initially described as a nuclear protein, it was later shown to be a bona fide lysosomal integral membrane protein. To gain insight into the physiological function of NCU-G1, mice with no detectable expression of this gene were created using a gene-trap strategy, and Ncu-g1^gt/gt mice were successfully characterized. Lysosomal disorders are mainly caused by lack of or malfunctioning of proteins in the endosomal-lysosomal pathway. The clinical symptoms vary, but often include liver dysfunction. Persistent liver damage activates fibrogenesis and, if unremedied, eventually leads to liver fibrosis/cirrhosis and death. We demonstrate that the disruption of Ncu-g1 results in spontaneous liver fibrosis in mice as the predominant phenotype. Evidence for an increased rate of hepatic cell death, oxidative stress and active fibrogenesis were detected in Ncu-g1^gt/gt liver. In addition to collagen deposition, microscopic examination of liver sections revealed accumulation of autofluorescent lipofuscin and iron in Ncu-g1^gt/gt Kupffer cells. Because only a few transgenic mouse models have been identified with chronic liver injury and spontaneous liver fibrosis development, we propose that the Ncu-g1^gt/gt mouse could be a valuable new tool in the development of novel treatments for the attenuation of fibrosis due to chronic liver damage.KEY WORDS: NCU-G1, Lysosome, Fibrosis     

Effect of L-asparaginase on retention of maze learning in mice

Long-term memory impairment has been described previously in mice receiving inhibitors of protein synthesis. In the present work, the enzyme L-asparaginase was injected into mice by an intrathecal or by an intraperitoneal route and produced a significant impairment of memory. Glutamine and asparagine prevented the effect of asparaginase when injected by the intraperitoneal route.