Ethanol-induced alterations of the antioxidant defense system in rat kidney

We report here the effects of chronic ethanol consumption on the antioxidant defense system in rat kidney. Thirty-two male Wistar rats were randomly divided in two identical groups and were treated as follows: control group (water for fluid) and the ethanol-fed group (2 g/kg body weight/24 h). The animals were sacrificed after 10 weeks, and respectively 30 weeks of ethanol consumption, and the renal tissue was isolated and analyzed. Results revealed that kidney alcohol dehydrogenase activities increased significantly after ethanol administration, but the electrophoretic pattern of alcohol dehydrogenase isoforms was unmodified. The SDS polyacrylamidegel electrophoretic study of kidney proteins has revealed the appearance of two new protein bands after long-term ethanol consumption. The kidney reduced glutathione/oxidized glutathione ratio decreased, indicating an oxidative stress response due to ethanol ingestion. The malondialdehyde contents and xanthine oxidase activities were unchanged. The antioxidant enzymatic defense system showed a different response during the two periods of ethanol administration. After 10 weeks, catalase, glutathione peroxidase, glutathione reductase, and glucose-6-phosphate dehydrogenase were activated, while superoxide dismutase, glutathione transferase, and gamma-glutamyltranspeptidase levels were stationary. After 30 weeks, superoxide dismutase and glutathione peroxidase activities were unmodified, but catalase, glutathione transferase, gamma-glutamyltranspeptidase, glutathione reductase, and glucose-6-phosphate dehydrogenase activities were significantly increased. Remarkable changes have been registered after 30 weeks of ethanol administration for glutathione reductase and glucose-6-phosphate dehydrogenase activities, including an increase by 106 and 216' of control values, respectively. These results showed specific changes in rat kidney antioxidant system and glutathione status as a consequence of long-term ethanol administration.     

Glutathione concentration and peptidase activity in the lens after exposure to microwaves

This study was undertaken for observation of early changes in glutathione concentration and the activity of carboxypeptidase A and aminopeptidase in the cortex and core of the lens as well as for determination of the cumulating effect of microwave energy after repeated exposures to microwaves. Experiments were carried out on New Zealand rabbits. The control group was compared to experimental groups exposed every day for 5 minutes to microwave irradiation of the eyeballs at power densities of 5 X 10(-3) W/cm2 and 10 X 10(-3) W/cm2 during 10, 20 and 30 days. Differences were found between the control group and the groups of animals exposed to microwaves in which the glutathione concentration in the cortex and core of the lens was decreasing with time in proportion to the number of exposures. Parallelly to the number of days of exposure to microwaves the enzymatic activity of carboxypeptidase A and aminopeptidase increased in the cortex of the lens. The observed changes demonstrate cumulation of the absorbed microwave energy leading to changes in the permeability of the capsule and membranes of lenticular fibres which lead to secondary metabolic disturbances in the lens of the eye.     

The effect of environmental factors of the coking plant on the enzymatic activity of rabbit oral mucosa.

Histochemical methods were used to investigate the activities of some intracellular enzymes in the oral mucosa of the rabbits which had been kept in selected work-stands of a coking-plant for 3 months. The findings were compared with the results for the enzymatic activity of the oral mucosa of control rabbits. The epithelium of the oral mucosa of the experimental rabbits was found to be proliferated acanthotically; moreover, there occurred some other morphological changes of the mucosa which often resembled precancerous states of leukoplakia type. In comparison with the control group, the activities of the studied enzymes, i.e. reduced NAD dehydrogenase (NADD), glucose-6-phosphate dehydrogenase (G6PD), lactate dehydrogenase (LDS), succinate dehydrogenase (SD), cytochrome oxidase (CO), and phosphorylase a+b in the epithelium and the connective-tissue cells of the studied mucosa of the experimental animals were as a rule markedly lowered, this decline being especially pronounced for the three last-mentioned enzymes. It was only the proliferating stratum basale of the experimental rabbit epithelium that frequently exhibited enhanced activities of NADD and LDS. Besides, the activities of NADD, G6PD and LDS were of a markedly diverse intensity in the cells of the chaotically proliferating stratum spinosum of the experimental rabbit mucosa. The results point to the noxious modifying effect of chemical and physical agents of the investigated environment on the oral mucosa of the animals studied.     

The morphofunctional characteristics of the neurons in the sensorimotor cortex of old rabbits during the trace assimilation of rhythm]

Extracellular neuronal activity was recorded from 460 neurons from alert young (5-7 months), middle-aged (54-65 months) and old (66-85 months) rabbits. Trace rhythmic activity of sensorimotor cortical neurons was examined after long-lasting (10-20 min) rhythmic (0.5-2 Hz) electrocutaneous stimulation of the contralateral forelimb. Spectral analysis of spike activity showed age-related differences in capability of producing a rhythm of previous stimulation in spontaneous neuronal activity. In young animals propriate rhythmic fluctuations of firing rate appeared after the first or second sessions of stimulations (on the first experimental day), in middle-aged ones--after 2-4 sessions (on the second or third days); cortical neurons in old rabbits did not exhibit trace rhythmic activity. Significant morphological changes in glial and neuronal cells were observed in sensorimotor cortex of old rabbits. It is proposed that morphological deteriorations may be the reason of the impairement of trace processes during aging.     

Oral ketoconazole therapy for experimental candida albicans keratitis in rabbits

Oral ketoconazole (100 mg daily for 3 weeks) markedly reduced the severity of experimental Candida albicans keratitis in a group of 10 rabbits. Clinical scores of affected eyes were statistically significantly lower in the treated group than in a control group of 10 untreated rabbits. All cultures of corneal scrapings in the treated eyes were negative on the 15th day after the inoculation, whilst three positive cultures were still obtained on the 21st day in the control animals. Histopathological examination of eyes from treated and untreated rabbits showed great differences in the intensity of inflammatory changes in the two groups.     

Ethanol‐induced alterations of the antioxidant defense system in rat kidney

We report here the effects of chronic ethanol consumption on the antioxidant defense system in rat kidney. Thirty‐two male Wistar rats were randomly divided in two identical groups and were treated as follows: control group (water for fluid) and the ethanol‐fed group (2 g/kg body weight/24 h). The animals were sacrificed after 10 weeks, and respectively 30 weeks of ethanol consumption, and the renal tissue was isolated and analyzed. Results revealed that kidney alcohol dehydrogenase activities increased significantly after ethanol administration, but the electrophoretic pattern of alcohol dehydrogenase isoforms was unmodified. The SDS polyacrylamidegel electrophoretic study of kidney proteins has revealed the appearance of two new protein bands after long‐term ethanol consumption. The kidney reduced glutathione/oxidized glutathione ratio decreased, indicating an oxidative stress response due to ethanol ingestion. The malondialdehyde contents and xanthine oxidase activities were unchanged. The antioxidant enzymatic defense system showed a different response during the two periods of ethanol administration. After 10 weeks, catalase, glutathione peroxidase, glutathione reductase, and glucose‐6‐phosphate dehydrogenase were activated, while superoxide dismutase, glutathione transferase, and γ‐glutamyltranspeptidase levels were stationary. After 30 weeks, superoxide dismutase and glutathione peroxidase activities were unmodified, but catalase, glutathione transferase, γ‐glutamyltranspeptidase, glutathione reductase, and glucose‐6‐phosphate dehydrogenase activities were significantly increased. Remarkable changes have been registered after 30 weeks of ethanol administration for glutathione reductase and glucose‐6‐phosphate dehydrogenase activities, including an increase by 106 and 216' of control values, respectively. These results showed specific changes in rat kidney antioxidant system and glutathione status as a consequence of long‐term ethanol administration. © 2005 Wiley Periodicals, Inc. J Biochem Mol Toxicol 19:386‐395, 2005; Published online in Wiley InterScience ( www.interscience.wiley.com ). DOI 10.1002/jbt.20101     

Effect of prenatal hypobaric hypoxia on activity of the rat brain phosphoinositide system

Activity of the phosphoinositide system of intracellular signalization was studied in offspring of rats exposed to severe hypobaric hypoxia at the 14–16th (group 1) or the 18–20th day (group 2) of prenatal hypoxia. At the age of 15 days, in animals of both experimental groups the basal level of triphosphoinositides in the brain cortex was shown to be elevated as compared with control. In the group 1, this parameter also remains elevated in adult animals. Application of glutamate produces a more pronounced increase of the inositephosphates in brain slices of the 15-day old rats of the group 1 than in slices of animals of the control group. In the 15-day old rats of the group 2, as compared with control, the phosphoinositide response to glutamate application was reduced. No changes in the inositephosphate levels were revealed after application of glutamate upon slices of adult (the 90-day old) control animals and of adult rats of the group 2. In slices of adult rats of the group 1, on the contrary, the glutamate application produced an increase of the inositephosphate content. The obtained data indicate essential changes of the phosphoinositide metabolism in the brain of rats exposed to action of hypoxia at the period of prenatal development. The character and the severity of these changes depend on the period of development when action of hypoxia occurs.     

A model for demonstration of reversal of impairment of endothelium-dependent relaxation in the cholesterol-fed rabbit.

This investigation was undertaken to determine whether it was possible to restore endothelium-dependent relaxation (EDR) in the cholesterol-fed rabbit model of atherosclerosis following discontinuation of the cholesterol. New Zealand white rabbits, approximately 8 weeks of age, were randomized into (i) control group (9 animals fed a standard rabbit diet) and (ii) experimental group (27 animals: fed the same diet supplemented with 2.5% cholesterol). The experimental animals were restored to the standard diet after 3 weeks. EDR to acetylcholine (-9.0 to -5.0 log mol/L) was examined in the experimental animals at 3, 7, and 15 weeks after commencement of the study (n = 9 at each stage) and the nine control animals examined after 7 weeks. At the end of 7 weeks, EDR to acetylcholine (-6.0 log mol/L) was significantly (p less than 0.05) impaired in the experimental group (34.3 +/- 3.8%) compared with that in the control group (79.8 +/- 3.0%). The loss of EDR was not apparent in the experimental group at 3 weeks (relaxation: 81.7 +/- 4.7%). At the end of 15 weeks, the EDR was significantly restored in the experimental group (relaxation: 63.6 +/- 5.1%). These findings demonstrate that it is possible to reverse the loss of EDR that occurs with cholesterol feeding in the rabbit by limiting the period of exposure to a high cholesterol diet.     

Radiation-induced histochemical and ultrastructural changes in the enterocytes in the rabbit.

Ultrastructural changes and enzyme activities in the cell organelles of rabbits were studied within 1, 3, 6, 9, 15 and 30 days of the irradiation with 550 rads of gamma-radiation. Between the 1st and 3rd day after irradiation there was a fall in the succinate dehydrogenase activity in the swollen and frequently tigroidal mitochondria whose number distinctly diminished. By the 15th day these changes disappeared. In the hyaloplasm there was weakening of the reaction for lactate dehydrogenase after 1 day, and an increase in number of polysomes, glycogen granules and smooth vesicles after 3-9 days after irradiation. The glucose-6-phosphatase activity was unchanged and so was the shape of the Golgi apparatus. The activity of lysosomal enzymes and the number of lysosomes in the experimental groups was approximately normal. By the 6th day the activity of alkaline phosphatase in the striated border was lowered, subsequently normal. On the whole, the intensity of postradiation changes in the intestinal mucosal epithelium is correlated with the rhythm of proliferation and shedding of epithelial cells, although some signs of injury persist for longer time periods.     

Intravenous miconazole therapy for experimental keratomycosis in rabbits

Intravenous miconazole (90 mg daily for 3 weeks) reduced the intensity of experimental fungal keratitis due to Candida albicans in a group of 10 rabbits. Clinical scores of affected eyes were statistically significantly lower in the treated group than in a control group of 10 untreated rabbits. All cultures of corneal scrapings were negative on 18th day after inoculation in the treated group, but four cultures were still positive on the 21st day in the control animals. Histopathological examination of eyes from treated and untreated rabbits showed great differences in the severity of inflammatory changes in the two groups.     

Influence of alternating extremely low frequency ELF magnetic field on structure and function of pancreas in rats.

The aim of this study was to estimate the influence on ultrastructure and function of endocrine and excretoric part of pancreas in rats of extremely low frequency alternating magnetic fields with parameters used in therapy in humans. The animals from the two experimental groups were exposed to a rectangular magnetic field waveform at a frequency of 10 Hz and induction of 1.8-3.8 mT--(group P) or a sinusoidal magnetic field at a frequency of 40 Hz and induction of 1.3-2.7 mT--(group S), respectively. The control rats were subjected to sham exposure. The cycle of 1, 3, 6, 9, and 14 daily exposure sessions lasting 30 min was made in all groups. Some of rats after finishing the cycle of 14 exposures were left in the same conditions except for the magnetic field for 3 or 10 days. In both groups of rats exposed to magnetic field, a distinct tendency to decrease glucose concentration, compared to control group, was observed during the exposure cycle. Serum glucose became normal after the end of exposure sessions. The concentrations of insulin in both groups of rats exposed to magnetic field were significantly higher, compared to the controls, during the exposure cycle. After the end of exposure cycle the concentration of insulin in group S became normal. In contrast, in group P the concentration of insulin decreased significantly on the last day of exposure, with a subsequent increase in the following days. The activity of alpha-amylase and lipase in the serum of experimental and control rats was not affected. In both groups of exposed rats, reversible changes of ultrastructure of the pancreatic islets, including expansion of the Golgi apparatus, extension of rough endoplasmatic reticulum, mitochondrial swelling, expansion of beta-granules and increase in number of empty vesicles in beta cells, occurred during the exposure. In acinar cells of exposed animals, a slight extension of rough endoplasmatic reticulum and mitochondrial swelling as transitory changes were observed. The structural and functional changes in pancreas are probably adaptative ones.     

Structural changes in the central nervous system induced by a constant magnetic field

The effect of the permanent magnetic field (PMF) on the brain cortex of rats has been assessed. The study was performed on neurons, astrocytes and synapses of post- and precentral brain cortical zones. CNS sensitivity to PMF was determined. When the animals were exposed to PMF once or 15 times the changes in microstructures were of compensatory-adaptive character, indicative of the enhancement in their functional activity. When the animals were exposed to PMF 30 times, dystrophic changes underlying the impairment of CNS functional activity developed, 10 days after the exposure to PMF was discontinued regenerative intracellular processes were revealed.     

Cytochemical research on the effect of a synthetic enkephalin analog on the protein content and enzyme activity of neurons

Using quantitative cytochemical technique a study was made of the effect of the synthetic analog of the Tyr-D-Ala-Gly-Phe-NH2 on the content and concentration of proteins and on the activity of enzymes (aminopeptidase, glutamate dehydrogenase and acid phosphatase) in neurons of the brain motor cortex and nucleus caudatus of rabbits and rats. The essential changes of the parameters used were registered 3 days after neuropeptide injection. A 30 minutes effects of the synthetic analog of enkephalins in protein metabolism was not so pronounced as a 3 days effect, the former being observed only in neurons of the brain motor cortex. Long-lasting effects of the neuropeptide Tyr-D-Ala-Gly-Phe-NH2 on the metabolism in brain are discussed.     

Electrical activity of the visual cortex under conditions of altered monoamine levels in the brain of animals

In experiments on 8 rabbits and 12 rats changes in electrograms of the visual cortex of alert animals were studied under photic stimulation in conditions of pharmacological action on monoamine (MA) brain systems. After injection of MA precursors (5-oxitriptophane and d, 1-dioxiphenylalanine) following phenomena were observed: a) decrease of the amplitude of the averaged evoked potentials to rhythmic photic stimuli (1-20 imp. sec.-1); b) an enhancement of fast (15-25 Hz) oscillations in the cortical spontaneous electrical activity and weakening and modification of the effects of the blockader of synthesis of MA-alpha-methyl-dioxiphenylalanine. Under light stimulation potentiation of MA precursors effects was observed in the frequency spectra of electrocorticograms. In the same conditions the specificity of action of cathecholamines precursor was revealed in the form of an increase of power of rhythms of 5-7 Hz and it; decrease in 2-3 Hz. Possible mechanisms of the revealed phenomena are discussed.     

Chemical preconditioning with 3-nitropropionic acid in hearts: role of mitochondrial K(ATP) channel

We investigated the cardioprotective effect of 3-nitropropionic acid (3-NPA), an inhibitior of mitochondrial succinate dehydrogenase, and we wanted to show whether this protection is mediated by of opening mitochondrial ATP-sensitive potassium (K(ATP)) channels. Adult rabbits were treated with either 3-NPA (3 mg/kg iv) or saline (n = 6 rabbits/group). After 30 min (for early phase) or 24 h (for late phase) of the treatment, the animals were subjected to 30 min of ischemia and 3 h of reperfusion (ischemia-reperfusion). 5-Hydroxydecanoate (5-HD, 5 mg/kg iv),the mitochondrial K(ATP) channel blocker, was administered 10 min before ischemia-reperfusion in the saline- and 3-NPA-treated rabbits. 3-NPA caused a decrease in the infarct size from 27.8 +/- 4.2% in the saline group to 16.5 +/- 1.0% in the 3-NPA-treated rabbits during early phase and from 30.4 +/- 4.2% in the saline group to 17.6 +/- 1.05 in the 3-NPA group during delayed phase (P < 0.05, % of risk area). The anti-infarct effect of 3-NPA was blocked by 5-HD as shown by an increase in infarct size to 33 +/- 2.7% (early phase) and 31 +/- 2.4% (delayed phase) (P < 0.05 vs. 3-NPA groups). 5-HD had no proischemic effect in control animals. Also, 3-NPA had no effect on systemic hemodynamics. We conclude that 3-NPA induces long-lasting anti-ischemic effects via opening of mitochondrial K(ATP) channels.     

Callus stimulation in distraction osteogenesis

Distraction osteogenesis has been described as in vivo tissue engineering. The ability to stimulate this process for the repair of bony defects or lengthening of congenitally shortened facial structures is likely to significantly impact the field of craniofacial surgery. The purpose of this study was to determine whether mechanical stimulation of the distracted rabbit mandible would accelerate the maturation of the bony callus when applied during the early consolidation period. Twenty adult New Zealand White rabbits underwent unilateral mandibular osteotomy. A uni-directional internal distractor device (Synthes, Paoli, Pa.) was positioned along a plane perpendicular to the line of osteotomy. After a 7-day latency period, distraction was commenced at a rate of 1.0 mm/day for 12 days in all animals. In a control group of 10 rabbits, a consolidation period of 8 weeks was observed before they were killed. In the experimental group of 10 rabbits, daily alternate compression and distraction of 1 mm (sequential compression and distraction) was performed for 3 weeks followed by a 5-week period of rigid fixation. Each animal received a dose of a fluorescent label at three different time points during the study: at the end of the distraction period, 3 weeks after the completion of the distraction phase, and 3 days before it was killed. All animals were killed 8 weeks after the completion of the distraction phase. Undecalcified histologic analysis and 3-point bending tests to failure were performed on the extracted mandibles. The results of the experimental and control groups were compared.Four animals in the control group and three animals in the experimental group were excluded from the study because of screw loosening resulting in distractor dislodgment or because of infection. On histologic analysis, cortical thickness at the center of the callus was found to be significantly greater in the experimental group compared with the control group when normalized to the contralateral hemimandible (83 percent versus 49 percent, respectively; p < 0.007). The ratio of cortical to cancellous bone in the distracted callus was uniformly found to be greater in the experimental specimens. The mineral apposition rate was calculated by using fluorescence microscopy and found to be significantly greater in the experimental group both during the period of sequential compression and distraction (3.2 microm/day versus 2.1 microm/day, p = 0.02) and after the period of sequential compression and distraction (1.4 microm/day versus 1.1 microm/day, p = 0.006). Mechanical testing revealed no significant differences in bending strength or stiffness between experimental or control groups (p = 0.54 and 0.47, respectively). This study has demonstrated that daily alternating compression and distraction of 1 mm amplitude during the early consolidation period has a stimulatory impact on callus formation with respect to osteoblastic activity, remodeling, and maturation of bone. Optimal timing and amplitude of sequential movement, long-term biomechanical differences, and molecular pathways have yet to be elucidated.     

Regression of the changes developed in the adrenal cortex in hypercholesterolemia

Administration of cholesterin to rabbits for 2 months results in its increased concentration in blood and in adrenal cortex. Adrenocorticocytes demonstrate a decreased nuclear-cytoplasmic ratio, decreased concentration of protein and RNA in cytoplasm, lowered succinate dehydrogenase and 3 beta-ol-++steroid dehydrogenase activity. In 2 months after abolishing cholesterin, a considerable part of the changes mentioned remains. If during this period dexamethasone, inhibiting the gland activity, is administered, the regression of the changes decelerates. Administration of the adrenal cortex activator--corticotropin--facilitates normalization of most of the parameters, altered in the gland under the effect of cholesterin administration.     

Neuronal specialization of the motor cortex in normal rabbits and following destruction of the visual cortex

The activity of neurones of the anterolateral part of the motor cortex in food-acquisition behaviour was compared in two control rabbits and in three rabbits after the operation of bilateral ablation of the striatal cortex. In two of three operated rabbits the pattern of behavioural specialization lost considerably the specificity peculiar to the motor cortex (predominance of G-neurones activated in grasping of food), approaching (but not becoming identical) the pattern of specialization of the visual cortex neurones: the number of G-neurones decreased in a half, and the number of L-neurones (activated in connection with the acts of instrumental food-acquisition behaviour which animals were trained to in the experimental cage) was doubled. Changes of the activity were significantly less expressed in the third operated rabbit. The number of the neurones activated in food-acquisition behaviour in operated rabbits in comparison with the control ones was reduced in the upper layers of the cortex and increased in the lower layers. The resemblance is discussed of the basic processes of animals learning and behaviour recovery.     

The role of serotonin-modulated anticonsolidation protein in memory formation in rats in a shuttle box

Two series of experiments were carried out in Wistar male rats. In the first series, rats were trained to acquire conditioning in a shuttle box to 50% and 80% learning criteria. In the animals of the experimental group that achieved 50% learning criterion, a significant decrease in the levels of serotonin-modulated anticonsolidation protein (SMAP) (solid phase, indirect ELISA-test) was observed in the temporal cortex as compared to the animals of the active control group. In the animals of the experimental group that achieved 80% learning criterion, such a decrease was found in the occipital and temporal cortex. In the second series of the experiments, animals of the experimental group were injected with SMAP in saline at a concentration of 1.5 mg/ml in a volume of 10 microl through the cannula implanted into the left lateral ventricle of the brain. Control animals were administered with heating-inactivated SMAP in the same amount. The substances were injected to the animals under light ether anesthesia daily 40 min prior to learning sessions. Learning sessions were carried out in the shuttle box for several days to 50% learning criterion. The experimental rats achieved learning criterion within 7-8 days, whereas intact and control animals reached the same criterion within 4 days. Furthermore, the experimental group of animals differed in increased levels of fear, anxiety and aggression which did not decline throughout the whole learning period. The conclusion was made that SMAP participated in negative regulation of the memory trace formation.     

Ozone oxidative preconditioning: a protection against cellular damage by free radicals.

There is some anecdotal evidence that oxygen-ozone therapy may be beneficial in some human diseases. However so far only a few biochemical and pharmacodynamic mechanisms have been elucidated. On the basis of preliminary data we postulated that controlled ozone administration would promote an oxidative preconditioning preventing the hepatocellular damage mediated by free radicals. Six groups of rats were classified as follows: (1) negative control, using intraperitoneal sunflower oil; (2) positive control using carbon tetrachloride (CCl4) as an oxidative challenge; (3) oxygen-ozone, pretreatment via rectal insufflation (15 sessions) and after it, CCl4; (4) oxygen, as group 3 but using oxygen only; (5) control oxygen-ozone, as group 3, but without CCl4; group (6) control oxygen, as group 5, but using oxygen only. We have evaluated critical biochemical parameters such as levels of transaminase, cholinesterase, superoxide dismutase, catalase, phospholipase A, calcium dependent ATPase, reduced glutathione, glucose 6 phosphate dehydrogenase and lipid peroxidation. Interestingly, in spite of CCl4 administration, group 3 did not differ from group 1, while groups 2 and 4 showed significant differences from groups 1 and 3 and displayed hepatic damage. To our knowledge these are the first experimental results showing that repeated administration of ozone in atoxic doses is able to induce an adaptation to oxidative stress thus enabling the animals to maintain hepatocellular integrity after CCl4 poisoning.