Influence of treadmill training on motor performance and organization of exploratory behavior in Meriones unguiculatus with unilateral ischemic stroke: Histological correlates in hippocampal CA1 region and the neostriatum

This study examined the effects of motor stimulation via treadmill on the behavior of male gerbils after external carotid ischemic brain lesion. The animals were assigned to five groups; ischemic with no stimulation (SIG), ischemic with stimulation (SIG 12/24/48/72 It after surgery), non-ischemic with no stimulation (CC), non-ischemic with stimulation (CE) and sham, surgery without occlusion with no stimulation (SH). All the animals were tested in the open-field (OF) and rotarod (RR), 4 days after surgery in order to evaluate exploratory behaviors and motor performance. Data were submitted to one-way variance (ANOVA) and Dunnett`s post hoc comparisons. SIG and SIG 12 groups showed a significant decrease in motor response (crossing) when compared to the control group (CC) (F = 20.65, P < 0.05) in the OF. SIG 12 group showed an increase in grooming behavior (F = 23.136, P < 0.05) and all ischemia groups (SIG, SIG 12/24/48/72) spent less time on the RR (F = 10.40, P < 0.05), when compared to the control group (CC). Histological analyses show extensive lesions in the hippocampus and neostriatum for all groups with ischemia (SIG, SIG 12/24/48/72), which are structures involved in the organization of motor behavior. Interestingly, the most pronounced damage was found in animals submitted to motor stimulation 12 h after ischemia which can be correlated to the increased number of grooming behavior showed by them in the OF. These findings suggest that motor stimulation through treadmill training improve motor behavior after ischemia, except when it starts 12h after surgery. (c) 2007 Elsevier Ireland Ltd. All rights reserved.

Behavioural alterations are independent of sickness behaviour in chronic experimental Chagas disease

The existence of the nervous form of Chagas disease is a matter of discussion since Carlos Chagas described neurological disorders, learning and behavioural alterations in Trypanosoma cruzi-infected individuals. In most patients, the clinical manifestations of the acute phase, including neurological abnormalities, resolve spontaneously without apparent consequence in the chronic phase of infection. However, chronic Chagas disease patients have behavioural changes such as psychomotor alterations, attention and memory deficits, and depression. In the present study, we tested whether or not behavioural alterations are reproducible in experimental models. We show that C57BL/6 mice chronically infected with the Colombian strain of T. cruzi (150 days post-infection) exhibit behavioural changes as (i) depression in the tail suspension and forced swim tests, (ii) anxiety analysed by elevated plus maze and open field test sand and (iii) motor coordination in the rotarod test. These alterations are neither associated with neuromuscular disorders assessed by the grip strength test nor with sickness behaviour analysed by temperature variation sand weight loss. Therefore, chronically T. cruzi-infected mice replicate behavioural alterations (depression and anxiety) detected in Chagas disease patients opening an opportunity to study the interconnection and the physiopathology of these two biological processes in an infectious scenario.

Voluntary alcohol consumption is controlled via the neuropeptide Y Y1 receptor.

We have shown previously that voluntary ethanol consumption and resistance to ethanol-induced sedation are inversely related to neuropeptide Y (NPY) levels in NPY-knock-out (NPY(-/-)) and NPY-overexpressing mice. In the present report, we studied knock-out mice completely lacking the NPY Y1 receptor (Y1(-/-)) to further characterize the role of the NPY system in ethanol consumption and neurobiological responses to this drug. Here we report that male Y1(-/-) mice showed increased consumption of solutions containing 3, 6, and 10% (v/v) ethanol when compared with wild-type (Y1(+/+)) control mice. Female Y1(-/-) mice showed increased consumption of a 10% ethanol solution. In contrast, Y1(-/-) mice showed normal consumption of solutions containing either sucrose or quinine. Relative to Y1(+/+) mice, male Y1(-/-) mice were found to be less sensitive to the sedative effects of 3.5 and 4.0 gm/kg ethanol as measured by more rapid recovery from ethanol-induced sleep, although plasma ethanol levels did not differ significantly between the genotypes. Finally, male Y1(-/-) mice showed normal ethanol-induced ataxia on the rotarod test after administration of a 2.5 gm/kg dose. These data suggest that the NPY Y1 receptor regulates voluntary ethanol consumption and some of the intoxicating effects caused by administration of ethanol.

Silencing mutant ataxin-3 rescues motor deficits and neuropathology in machado-joseph disease transgenic mice.

Machado-Joseph disease (MJD) or spinocerebellar ataxia type 3 (SCA3) is an autosomal dominantly-inherited neurodegenerative disorder caused by the over-repetition of a CAG codon in the MJD1 gene. This expansion translates into a polyglutamine tract that confers a toxic gain-of-function to the mutant protein - ataxin-3, leading to neurodegeneration in specific brain regions, with particular severity in the cerebellum. No treatment able to modify the disease progression is available. However, gene silencing by RNA interference has shown promising results. Therefore, in this study we investigated whether lentiviral-mediated allele-specific silencing of the mutant ataxin-3 gene, after disease onset, would rescue the motor behavior deficits and neuropathological features in a severely impaired transgenic mouse model of MJD. For this purpose, we injected lentiviral vectors encoding allele-specific silencing-sequences (shAtx3) into the cerebellum of diseased transgenic mice expressing the targeted C-variant of mutant ataxin-3 present in 70% of MJD patients. This variation permits to discriminate between the wild-type and mutant forms, maintaining the normal function of the wild-type allele and silencing only the mutant form. Quantitative analysis of rotarod performance, footprint and activity patterns revealed significant and robust alleviation of gait, balance (average 3-fold increase of rotarod test time), locomotor and exploratory activity impairments in shAtx3-injected mice, as compared to control ones injected with shGFP. An important improvement of neuropathology was also observed, regarding the number of intranuclear inclusions, calbindin and DARPP-32 immunoreactivity, fluorojade B and Golgi staining and molecular and granular layers thickness. These data demonstrate for the first time the efficacy of gene silencing in blocking the MJD-associated motor-behavior and neuropathological abnormalities after the onset of the disease, supporting the use of this strategy for therapy of MJD.

New evidence of neuroprotection by lactate after transient focal cerebral ischaemia: extended benefit after intracerebroventricular injection and efficacy of intravenous administration.

BACKGROUND: Lactate protects mice against the ischaemic damage resulting from transient middle cerebral artery occlusion (MCAO) when administered intracerebroventricularly at reperfusion, yielding smaller lesion sizes and a better neurological outcome 48 h after ischaemia. We have now tested whether the beneficial effect of lactate is long-lasting and if lactate can be administered intravenously. METHODS: Male ICR-CD1 mice were subjected to 15-min suture MCAO under xylazine + ketamine anaesthesia. Na L-lactate (2 µl of 100 mmol/l) or vehicle was administered intracerebroventricularly at reperfusion. The neurological deficit was evaluated using a composite deficit score based on the neurological score, the rotarod test and the beam walking test. Mice were sacrificed at 14 days. In a second set of experiments, Na L-lactate (1 µmol/g body weight) was administered intravenously into the tail vein at reperfusion. The neurological deficit and the lesion volume were measured at 48 h. RESULTS: Intracerebroventricularly injected lactate induced sustained neuroprotection shown by smaller neurological deficits at 7 days (median = 0, min = 0, max = 3, n = 7 vs. median = 2, min = 1, max = 4.5, n = 5, p < 0.05) and 14 days after ischaemia (median = 0, min = 0, max = 3, n = 7 vs. median = 3, min = 0.5, max = 3, n = 7, p = 0.05). Reduced tissue damage was demonstrated by attenuated hemispheric atrophy at 14 days (1.3 ± 4.0 mm(3), n = 7 vs. 12.1 ± 3.8 mm(3), n = 5, p < 0.05) in lactate-treated animals. Systemic intravenous lactate administration was also neuroprotective and attenuated the deficit (median = 1, min = 0, max = 2.5, n = 12) compared to vehicle treatment (median = 1.5, min = 1, max = 8, n = 12, p < 0.05) as well as the lesion volume at 48 h (13.7 ± 12.2 mm(3), n = 12 vs. 29.6 ± 25.4 mm(3), n = 12, p < 0.05). CONCLUSIONS: The beneficial effect of lactate is long-lasting: lactate protects the mouse brain against ischaemic damage when supplied intracerebroventricularly during reperfusion with behavioural and histological benefits persisting 2 weeks after ischaemia. Importantly, lactate also protects after systemic intravenous administration, a more suitable route of administration in a clinical emergency setting. These findings provide further steps to bring this physiological, commonly available and inexpensive neuroprotectant closer to clinical translation for stroke.

Effects of Enriched Physical and Social Environments on Motor Performance, Associative Learning, and Hippocampal Neurogenesis in Mice.

We have studied the motor abilities and associative learning capabilities of adult mice placed in different enriched environments. Three-month-old animals were maintained for a month alone (AL), alone in a physically enriched environment (PHY), and, finally, in groups in the absence (SO) or presence (SOPHY) of an enriched environment. The animals' capabilities were subsequently checked in the rotarod test, and for classical and instrumental learning. The PHY and SOPHY groups presented better performances in the rotarod test and in the acquisition of the instrumental learning task. In contrast, no significant differences between groups were observed for classical eyeblink conditioning. The four groups presented similar increases in the strength of field EPSPs (fEPSPs) evoked at the hippocampal CA3-CA1 synapse across classical conditioning sessions, with no significant differences between groups. These trained animals were pulse-injected with bromodeoxyuridine (BrdU) to determine hippocampal neurogenesis. No significant differences were found in the number of NeuN/BrdU double-labeled neurons. We repeated the same BrdU study in one-month-old mice raised for an additional month in the above-mentioned four different environments. These animals were not submitted to rotarod or conditioned tests. Non-trained PHY and SOPHY groups presented more neurogenesis than the other two groups. Thus, neurogenesis seems to be related to physical enrichment at early ages, but not to learning acquisition in adult mice.

Antinociceptive effect of novel pyrazolines in mice

The antinociceptive effect of six novel synthetic pyrazolines (3-ethoxymethyl-5-ethoxycarbonyl-1H-pyrazole (Pz 1) and its corresponding 1-substituted methyl (Pz 2) and phenyl (Pz 3) analogues, and 3-(1-ethoxyethyl)-5-ethoxycarbonyl-1H-pyrazole (Pz 4) and its corresponding 1-substituted methyl (Pz 5) and phenyl (Pz 6) analogues) was evaluated by the tail immersion test in adult male albino mice. The animals (N = 11-12 in each group) received vehicle (5% Tween 80, 10 ml/kg, sc) or 1.5 mmol/kg of each of the pyrazolines (Pz 1-Pz 6), sc. Fifteen, thirty and sixty minutes after drug administration, the mice were subjected to the tail immersion test. Thirty minutes after drug administration Pz 2 and Pz 3 increased tail withdrawal latency (vehicle = 3.4 ± 0.2; Pz 2 = 5.2 ± 0.4; Pz 3 = 5.9 ± 0.4 s; mean ± SEM), whereas the other pyrazolines did not present antinociceptive activity. Dose-effect curves (0.15 to 1.5 mmol/kg) were constructed for the bioactive pyrazolines. Pz 2 (1.5 mmol/kg, sc) impaired motor coordination in the rotarod and increased immobility in the open-field test. Pz 3 did not alter rotarod performance and spontaneous locomotion, but increased immobility in the open field at the dose of 1.5 mmol/kg. The involvement of opioid mechanisms in the pyrazoline-induced antinociception was investigated by pretreating the animals with naloxone (2.75 µmol/kg, sc). Naloxone prevented Pz 3- but not Pz 2-induced antinociception. Moreover, naloxone pretreatment did not alter Pz 3-induced immobility. We conclude that Pz 3-induced antinociception involves opioid mechanisms but this is not the case for Pz 2.

Physical exercise prevents motor disorders and striatal oxidative imbalance after cerebral ischemia-reperfusion

Stroke is the third most common cause of death worldwide, and most stroke survivors present some functional impairment. We assessed the striatal oxidative balance and motor alterations resulting from stroke in a rat model to investigate the neuroprotective role of physical exercise. Forty male Wistar rats were assigned to 4 groups: a) control, b) ischemia, c) physical exercise, and d) physical exercise and ischemia. Physical exercise was conducted using a treadmill for 8 weeks. Ischemia-reperfusion surgery involved transient bilateral occlusion of the common carotid arteries for 30 min. Neuromotor performance (open-field and rotarod performance tests) and pain sensitivity were evaluated beginning at 24 h after the surgery. Rats were euthanized and the corpora striata was removed for assay of reactive oxygen species, lipoperoxidation activity, and antioxidant markers. Ischemia-reperfusion caused changes in motor activity. The ischemia-induced alterations observed in the open-field test were fully reversed, and those observed in the rotarod test were partially reversed, by physical exercise. Pain sensitivity was similar among all groups. Levels of reactive oxygen species and lipoperoxidation increased after ischemia; physical exercise decreased reactive oxygen species levels. None of the treatments altered the levels of antioxidant markers. In summary, ischemia-reperfusion resulted in motor impairment and altered striatal oxidative balance in this animal model, but those changes were moderated by physical exercise.

Down-regulation of cerebellar 5-HT2C receptors in pilocarpine-induced epilepsy in rats: Therapeutic role of Bacopa monnieri extract

Epilepsy is a syndrome of episodic brain dysfunction characterized by recurrent unpredictable, spontaneous seizures. Cerebellar dysfunction is a recognized complication of temporal lobe epilepsy and it is associated with seizure generation, motor deficits and memory impairment. Serotonin is known to exert a modulatory action on cerebellar function through 5HT2C receptors. 5-HT2C receptors are novel targets for developing anticonvulsant drugs. In the present study, we investigated the changes in the 5-HT2C receptors binding and gene expression in the cerebellum of control, epileptic and Bacopa monnieri treated epileptic rats. There was a significant down regulation of the 5-HT content (pb0.001), 5-HT2C gene expression (pb0.001) and 5-HT2C receptor binding (pb0.001) with an increased affinity (pb0.001). Carbamazepine and B. monnieri treatments to epileptic rats reversed the down regulated 5-HT content (pb0.01), 5-HT2C receptor binding (pb0.001) and gene expression (pb0.01) to near control level. Also, the Rotarod test confirms the motor dysfunction and recovery by B. monnieri treatment. These data suggest the neuroprotective role of B. monnieri through the upregulation of 5-HT2C receptor in epileptic rats. This has clinical significance in the management of epilepsy

Glutamate Receptor Gene Expression: IP3, cAMP, cGMP Functional Regulation in Hypoglycaemic and Diabetic Rats

Department of Biotechnology, Cochin University of Science and Technology

Effects of the essential oil from Citrus aurantium L. in experimental anxiety models in mice

Citrus aurantium L. is popularly used to treat anxiety, among other indications suggesting central nervous system action. Previous studies showed anxiolytic effect in the essential oil from peel in mice evaluated on the elevated plus maze [Carvalho-Freitas, M.I.R., Costa, M., 2002. Anxiolytic and sedative effects of extracts and essential oil from Citrus aurantium L. Biological and Pharmaceutical Bulletin 25, 1629-1633.]. In order to better characterize the activity of the essential oil, it was evaluated in two other experimental models: the light-dark box and the marble-burying test, respectively related to generalized anxiety disorder and to obsessive compulsive disorder. Mice were treated acutely by oral route 30 min (single dose) or once a day for 15 days (repeated doses) before experimental procedures. In light-dark box test, single treatment with essential oil augmented the time spent by mice in the light chamber and the number of transitions between the two compartments. There were no observed alterations in the parameters evaluated in light-dark box after repeated treatment. Otherwise, single and repeated treatments with essential oil were able to suppress marble-burying behavior. At effective doses in the behavioral tests, mice showed no impairment on rotarod procedure after both single and repeated treatments with essential oil, denoting absence of motor deficit. Results observed in marble-burying test, related to obsessive compulsive disorder, appear more consistent than those observed in light-dark box. (c) 2005 Elsevier B.V. All rights reserved.

Developmental lead exposure: behavioral alterations in the short and long term

Wistar dams were exposed to 500 ppm of Pb, as Ph acetate, or 660 ppm Na acetate in drinking water during pregnancy and lactation. Male pups at 23 (weaned) or 70 days (adult) of age were submitted to behavioral evaluation and Pb determination. The behaviors evaluated were: locomotor activity (open-field test), motor coordination (rotarod test), exploratory behavior (holeboard test), anxiety (elevated plus maze and social interaction tests), and learning and memory (shuttle box). Ph levels were measured in the blood and cerebral regions (hippocampus and striatum) of dams and pups. The results of the present report demonstrated that exposure to Ph during pregnancy and lactation induces in weaned pups hyperactivity, decreased exploratory behavior, and impairment of learning and memory. These alterations were observed at blood Ph levels in the range that may be attained in children chronically exposed to low levels of Pb (21 +/- 3 mug/dl). Regarding adults, the results demonstrated that the regimen of exposure adopted induces anxiety in these animals at nondetectable blood Ph levels. (C) 2001 Elsevier B.V. All rights reserved.

Sedation and need-free salt intake in rats treated with clonidine

The effect of intraperitoneal injection of clonidine (9-72 mu g/kg) on need-free 1.5% NaCl intake and on performance (defined as percent of a complete trial) in the rotarod test, was studied in normovolemic adult male rats. Clonidine (18 and 36 mu g/kg) inhibited the 1.5% NaCl intake in a 2-h test at doses that did not alter the performance in the rotarod test. The dose of 36 mu g/kg did not inhibit 10% sucrose intake. Only the highest dose (72 mu g/kg) of clonidine inhibited the 1.5% NaCl intake and the performance in the rotarod test, and produced signs of sedation. Sedation was determined either by change in posture (immobility or lack of postural tonus) of the animals during the ingestive test or by their performance in the rotarod test. The results suggest that sedation is not a determinant effect on the inhibition of 1.5% NaCl intake induced by clonidine. (C) 1999 Elsevier B.V.

Gabaergic-benzodiazepine system is involved in the crotoxin-induced anxiogenic effect

The behavioral effects of crotoxin (CTX), the major component of Crotalus durissus terrificus venom, were studied in rats submitted to the open field, holeboard, and social interaction tests. CTX (100, 250, and 500 mu g/kg, IP) was administered 2 h before the tests. In the open field, CTX reduced ambulation (250 mu g/kg) and rearing (250 and 500 mu g/kg) and increased grooming (100 and 250 mu g/kg) and freezing (250 mu g/kg). In the holeboard and social interaction, all the CTX doses evaluated decreased, respectively, head dip and head dipping, and social interaction time. The CTX-induced behavioral alterations could be attributed to its neuromuscular transmission blockade, but this possibility was ruled out because CTX (250 and 500 mu g/kg, IP, 2 h before the rotarod test) was unable to modify the rotarod performance of rats. The involvement of the benzodiazepine receptor in the CTX-induced behavioral alterations was investigated through the pretreatment (30 min before the tests, IP) of the animals with diazepam (1.2 mg/kg), or flumazenil (4 and 10 mg/kg). Both diazepam and flumazenil antagonized the CTX induced behavioral alterations in the open field, holeboard, and social interaction tests. This study demonstrated that: (1) CTX is an anxiogenic compound; and (2) the gabaergic-benzodiazepine system may play a role in the CTX-induced anxiogenic effect. (C) 1999 Elsevier B.V.

Citrus aurantium L. essential oil exhibits anxiolytic-like activity mediated by 5-HT1A-receptors and reduces cholesterol after repeated oral treatment

Background: The current treatments for anxiety disorders and depression have multiple adverse effects in addition to a delayed onset of action, which has prompted efforts to find new substances with potential activity in these disorders. Citrus aurantium was chosen based on ethnopharmacological data because traditional medicine refers to the Citrus genus as useful in diminishing the symptoms of anxiety or insomnia, and C. aurantium has more recently been proposed as an adjuvant for antidepressants. In the present work, we investigated the biological activity underlying the anxiolytic and antidepressant effects of C. aurantium essential oil (EO), the putative mechanism of the anxiolytic-like effect, and the neurochemical changes in specific brain structures of mice after acute treatment. We also monitored the mice for possible signs of toxicity after a 14-day treatment.Methods: The anxiolytic-like activity of the EO was investigated in a light/dark box, and the antidepressant activity was investigated in a forced swim test. Flumazenil, a competitive antagonist of benzodiazepine binding, and the selective 5-HT1A receptor antagonist WAY100635 were used in the experimental procedures to determine the mechanism of action of the EO. To exclude false positive results due to motor impairment, the mice were submitted to the rotarod test.Results: The data suggest that the anxiolytic-like activity observed in the light/dark box procedure after acute (5 mg/kg) or 14-day repeated (1 mg/kg/day) dosing was mediated by the serotonergic system (5-HT1A receptors). Acute treatment with the EO showed no activity in the forced swim test, which is sensitive to antidepressants. A neurochemical evaluation showed no alterations in neurotransmitter levels in the cortex, the striatum, the pons, and the hypothalamus. Furthermore, no locomotor impairment or signs of toxicity or biochemical changes, except a reduction in cholesterol levels, were observed after treatment with the EO.Conclusion: This work contributes to a better understanding of the biological activity of C. aurantium EO by characterizing the mechanism of action underlying its anxiolytic-like activity. © 2013 Costa et al; licensee BioMed Central Ltd.