Age-related decline of various cognitive functions in well-experienced male rats treated with the putative anti-aging compound (2R)-1-(1-benzofuran-2-yl)-N-propylpentane-2-amine ((-)BPAP).

Aging-associated cognitive disorders lack proper medication. To meet this need translation-wise, modification of the animal models is also required. In the present study, effect of the putative anti-aging compound (2R)-1-(1-benzofuran-2-yl)-N-propylpentane-2-amine ((-)BPAP, a deprenyl derivative) on age-related cognitive decline was investigated in experienced, aged Long-Evans rats. During their lifetime, animals had acquired knowledge in various cognitive assays. Their performance in these tests was then parallel followed from the age of 27 months until their death meanwhile half of them were treated with BPAP. Cognitive performance in various tasks showed different sensitivities/resistances to age-related impairment. Pot jumping performance (motor skill-learning) started to impair first, at 21 months of age, followed by decreasing performance in five-choice serial reaction time task (attention) at 26 months. Navigation performance in Morris water maze (spatial learning) started to decline at 31 months. Performance in a cooperation task (social cognition) started to decline the latest, at 34 months. Our findings suggest that in this process, the primary factor was the level of motivation to be engaged with the task and not losing the acquired knowledge. The average lifespan of the tested rat population was 36 months. BPAP could not improve the cognitive performance; neither could it prolong lifespan. A possible reason might be that dietary restriction and lifelong cognitive engagement had beneficial effects on cognitive capabilities and lifespan creating a "ceiling effect" for further improvement. The results confirmed that experienced animals provide a translationally relevant model to study age-related cognitive decline and measure the effect of putative anti-aging compounds.

Vasopressin as Possible Treatment Option in Autism Spectrum Disorder.

Autism spectrum disorder (ASD) is rather common, presenting with prevalent early problems in social communication and accompanied by repetitive behavior. As vasopressin was implicated not only in salt-water homeostasis and stress-axis regulation, but also in social behavior, its role in the development of ASD might be suggested. In this review, we summarized a wide range of problems associated with ASD to which vasopressin might contribute, from social skills to communication, motor function problems, autonomous nervous system alterations as well as sleep disturbances, and altered sensory information processing. Beside functional connections between vasopressin and ASD, we draw attention to the anatomical background, highlighting several brain areas, including the paraventricular nucleus of the hypothalamus, medial preoptic area, lateral septum, bed nucleus of stria terminalis, amygdala, hippocampus, olfactory bulb and even the cerebellum, either producing vasopressin or containing vasopressinergic receptors (presumably V1a). Sex differences in the vasopressinergic system might underline the male prevalence of ASD. Moreover, vasopressin might contribute to the effectiveness of available off-label therapies as well as serve as a possible target for intervention. In this sense, vasopressin, but paradoxically also V1a receptor antagonist, were found to be effective in some clinical trials. We concluded that although vasopressin might be an effective candidate for ASD treatment, we might assume that only a subgroup (e.g., with stress-axis disturbances), a certain sex (most probably males) and a certain brain area (targeting by means of virus vectors) would benefit from this therapy.

Lack of general learning ability factor in a rat test battery measuring a wide spectrum of cognitive domains.

Objective: In the framework of a larger project aiming to test putative cognitive enhancer drugs in a system with improved translational validity, we established a rodent test battery, where different, clinically relevant cognitive domains were investigated in the same animal population. The aim of the current study was to check whether performances in the different tasks representing different cognitive functions are assay-specific or may originate in an underlying general learning ability factor. Methods: In the experiments 36 Long-Evans and 36 Lister Hooded rats were used. The test battery covered the following cognitive domains: attention and impulsivity (measured in the 5-choice serial reaction time task), spatial memory (Morris water-maze), social cognition (cooperation task), cognitive flexibility (attentional set shifting test), recognition memory (novel object recognition) and episodic memory (water-maze based assay). The outcome variables were analyzed by correlation analysis and principal component analysis (PCA). The datasets consisted of variables measuring learning speed and performance in the paradigms. From the raw variables composite variables were created for each assay, then from these variables a composite score was calculated describing the overall performance of each individual in the test battery. Results: Correlations were only found among the raw variables characterizing the same assay but not among variables belonging to different tests or among the composite variables. The PCAs did not reduce the dimensionality of the raw or composite datasets. Graphical analysis showed variable performance of the animals in the applied tests. Conclusions: The results suggests the assay outcomes (learning performance) in the system are based on independent cognitive domains.

Translational Difficulties in Querying Rats on "Orientation".

The aim of this study was to translate the "orientation" query of the ADAS-Cog inventory to rats and to investigate whether they can determine which time of the day they are. For this purpose, we established a modified Morris water-maze navigation task where the escape platform was placed onto various locations at different times of the day: "morning", "noon" and "evening". In each of these sessions rats swam a "query" trial and a "confirmatory" trial, 30 min apart. Lister Hooded rats randomly chose among the three possible target locations, while Long Evans rats partly followed a win-stay strategy by preferring to visit first to the platform position of the previous session. Despite simplifying the task to a morning-evening discrimination, Lister Hooded rats continued searching by chance, while Long Evans rats switched to the mentally less demanding random strategy. We then inserted a board into the pool which required longer swimming path from the animals when they were correcting an initial wrong choice, but this modification did not result in a change in the above strategies. Lastly, in a separate group of Long-Evans rats, the training conditions were modified inasmuch an incorrect choice was definitely punished by impeding the animals to correct it and confining them to a platform-free part of the maze for the whole trial period. However, even these stricter conditions were not sufficient to make the rats distinguish times of the day. The observed lack of time discrimination may source from an evolutionary built in mechanism characteristic for the rat species or this ability may have only been lost in laboratory rats.

l-Alpha Glycerylphosphorylcholine as a Potential Radioprotective Agent in Zebrafish Embryo Model.

This work establishes the zebrafish embryo model for ionizing radiation (IR) modifier research and also evaluates the protective effect of l-alpha glycerylphosphorylcholine (GPC). Embryos were exposed to a single-fraction whole-body gamma irradiation (5, 10, 15, and 20 Gy) at different postfertilization time points and were serially assessed for viability and macro- and micromorphologic abnormalities. After toxicity evaluation, 194 µM of GPC was added for certain groups with 3-h incubation before the radiation. Nuclear factor kappa B (NF-κB) and interleukin-1ß (IL-1ß) expression changes were measured using quantitative real-time polymerase chain reaction. A higher sensitivity could be observed at earlier stages of the embryogenesis. The lethal dose (LD50) for 6 hours postfertilization (hpf) embryos was 15 Gy and for 24 hpf was 20 Gy on day 7, respectively. GPC administration resulted in a significant improvement in both the distortion rate and survival of the 24 hpf embryos. Qualitative evaluation of the histological changes confirmed the protective effect of GPC. IL-1ß and NF-κB overexpression due to 10 Gy irradiation was also reduced by GPC. GPC exhibited promising radioprotective effects in our zebrafish embryo model, decreasing the irradiation-induced morphological damage and lethality with significant reduction of IR-caused pro-inflammatory activation.

Novel real-time cell analysis platform for the dynamic monitoring of ionizing radiation effects on human tumor cell lines and primary fibroblasts.

Translational research in radiation oncology is important for the detection of adverse radiation effects, cellular responses, and radiation modifications, and may help to improve the outcome of radiation therapy in patients with cancer. The present study aimed to optimize and validate a real­time label­free assay for the dynamic monitoring of cellular responses to ionizing radiation. The xCELLigence system is an impedance­based platform that provides continuous information on alterations in cell size, shape, adhesion, proliferation, and survival. In the present study, various malignant human primary fibroblast cells (U251, GBM2, MCF7, A549, HT­29) were exposed to 0, 5 and 10 Gy of Cobalt60 radiation. As well as the xCELLigence system, cell survival and proliferation was evaluated using the following conventional end­point cell­based methods: Clonogenic, MTS, and lactate dehydrogenase assays, and apoptosis was detected by fluorescence­activated cell sorting. The effects of ionizing radiation were detected for each cell line using impedance monitoring. The real­time data correlated with the colony forming assay results. At low cell densities (1,000­2,000 cells/well) the impedance­based method was more accurate at monitoring dose­dependent changes in the malignant human primary fibroblast cell lines, as compared with the end­point assays. The results of the present study demonstrated that the xCELLigence system may be a reliable and rapid diagnostic method for the monitoring of dynamic cell behavior following radiation. In addition, the xCELLigence system may be used to investigate the cellular mechanisms underlying the radiation response, as well as the time­dependent effects of radiation on cell proliferation and viability.

Single dose irradiation of defined region of rat brain with stereotactic brainlab system.

UNLABELLED: Background and purpose of our study was to develop a precise dose delivery technique for partial brain irradiation of two rats simultaneously. METHODS: Using a self-developed frame stereotactic radiotherapy with single doses of 30-90 Gy was delivered to the frontal lobe of 22 animals. Tolerability and reproducibility of the method were evaluated and dosimetric measurements were conducted to verify the treatment plans. 2, 4 and 6 months after the irradiation magnetic resonance imaging (MRI) scans and histopathological examinations were performed to detect late radiation induced biological changes. RESULTS: Immobilization device provided excellent reproducibility and tolerability. Dosimetry revealed good correspondence with planned dose distribution, but the measured absorbed dose was 30% lower than the planned dose. During the 6 months follow-up period the procedure related death of subject animals after 30 Gy, 70 Gy and 90 Gy were 0%, 20% and 100% respectively. T2 signal and structural changes on MRI scans found to be dose and time dependent. While 30 Gy caused no detectable structural changes, 70 Gy lead to cystic necrosis in 2 cases after 4 month. Histopathology revealed signs of necrosis on macroscopic examination after 70 Gy in the high dose region involving both frontal lobes, and no obvious microscopic changes in the surrounding area were detectable. CONCLUSION: Our technique of rat cranial irradiation using human stereotactic system provided high accuracy of single dose delivery for a pair of small animals, resulting in brain injury in the defined area. This method proved to be a reproducible model for preclinical studies on radiation effects.

Radio-neuroprotective effect of L-alpha-glycerylphosphorylcholine (GPC) in an experimental rat model.

Ionizing radiation plays a major role in the treatment of brain tumors, but side-effects may restrict the efficacy of therapy. In the present study, our goals were to establish whether the administration of L-alpha-glycerylphosphorylcholine (GPC) can moderate or prevent any of the irradiation-induced functional and morphological changes in a rodent model of hippocampus irradiation. Anesthetized adult (6-weeks-old) male Sprague-Dawley rats were subjected to 40 Gy irradiation of one hemisphere of the brain, without or with GPC treatment (50 mg/kg bw by gavage), the GPC treatment continuing for 4 months. The effects of this partial rat brain irradiation on the spatial orientation and learning ability of the rats were assessed with the repeated Morris water maze (MWM) test. Histopathologic (HP) evaluation based on hematoxylin-eosin and Luxol blue staining was performed 4 months after irradiation. The 40 Gy irradiation resulted in a moderate neurological deficit at the levels of both cognitive function and morphology 4 months after the irradiation. The MWM test proved to be a highly sensitive tool for the detection of neurofunctional impairment. The site navigation of the rats was impaired by the irradiation, but the GPC treatment markedly decreased the cognitive impairment. HP examination revealed lesser amounts of macrophage density, reactive gliosis, calcification and extent of demyelination in the GPC-treated group. GPC treatment led to significant protection against the cognitive decline and cellular damage, evoked by focal brain irradiation at 40 Gy dose level. Our study warrants further research on the protective or mitigating effects of GPC on radiation injuries.

Peripheral inflammatory activation after hippocampus irradiation in the rat.

PURPOSE: To detect the possible biochemical signs of inflammatory activation in the peripheral circulation in a rodent model of hippocampus irradiation, and to examine the effects of L-alpha-glycerylphosphorylcholine (GPC) in this experimental protocol. MATERIALS AND METHODS: Anesthetized Sprague-Dawley rats were subjected to 40 Gy cobalt irradiation of both hemispheres of the hippocampus, with or without GPC treatment (50 mg/kg intravenously (i.v.), 5 min before the irradiation, n = 6, each). A third group (n = 6) served as saline-treated control. Blood samples were obtained 3 h after the end of irradiation in order to examine the changes in plasma histamine, tumor necrosis factor-alpha (TNF-α), interleukin 1-beta, interleukin 6 (IL-6) and interleukin 10 (IL-10); liver tissue samples were taken to determine adenosine triphosphate (ATP) concentrations. RESULTS: The hepatic ATP levels were significantly declined, while plasma concentrations of circulating TNF-α, IL-6, IL-10 and histamine were significantly increased after hippocampus irradiation. GPC treatment significantly reduced the irradiation-induced release of cytokines and histamine, and the liver ATP level was maintained at the control value. CONCLUSIONS: Targeted brain irradiation produced measurable pro- and anti-inflammatory cytokine changes in the systemic circulation. GPC supplementation provides significant protection against irradiation-induced peripheral pro-inflammatory activation and ATP depletion.

Association of vitamin D receptor gene polymorphisms and Parkinson's disease in Hungarians.

Vitamin D receptor (VDR) gene encodes a transcription factor that influences calcium homeostasis and immunoregulation, and may play a role in neurological disorders including Parkinson's disease (PD). The investigations of the association between VDR and PD in different populations revealed various results. In a present study 100 PD patients and 109 healthy controls from the Hungarian population were genotyped for four polymorphic sites (BsmI, ApaI, FokI and TaqI) in the VDR gene. The polymorphisms were determined by polymerase chain reaction and restriction fragment length polymorphism (PCR-RFLP). Our results demonstrate an association between the FokI C allele and PD; the frequency of the C allele was significantly higher in PD patients than in controls, suggesting that this polymorphism may have a role in the development of PD in these patients.

Development of a small-animal focal brain irradiation model to study radiation injury and radiation-injury modifiers.

PURPOSE: Our aim was to establish an effective small-animal focal brain radiation model for research on brain injuries. MATERIAL AND METHODS: Groups of up to six rats were exposed to a range of doses from 120-40 Gy, at 10 intervals of a 6 MeV electron beam. Open-field motor functions and water maze learning-memory tests were performed after the irradiation at two-week intervals. Morphological changes were detected through repeated magnetic resonance imaging (MRI) monthly and were compared with the histopathological findings to determine if they predicted late microscopic changes. RESULTS: The development of necrosis proved to be dose-dependent. 120 Gy resulted in serious deterioration within 4 weeks in all rats. Localized necrosis in one hemisphere was detected 2 months after the irradiation with ≥ 70 Gy, and 3 months after 40-60 Gy consistent for all animals. The Morris water maze (MWM) tests proved to be the most sensitive tool for the early detection of a brain functional impairment. MRI screening provided useful information on the development of radiation necrosis, which defined the time point for histological examinations. CONCLUSIONS: The described method permits accurate dose delivery to a definite part in one hemisphere of the brain for six rats at a time. Following complex examinations, a dose of 40 Gy and a follow-up time of 4 months are proposed for investigations on neuroradiation modifiers.

Neuropathology of partial PGC-1α deficiency recapitulates features of mitochondrial encephalopathies but not of neurodegenerative diseases.

BACKGROUND: Deficient peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α) function is one component of mitochondrial dysfunction in neurodegenerative diseases. Current molecular classification of such diseases is based on the predominant protein accumulating as intra- or extracellular aggregates. Experimental evidence suggests that mitochondrial dysfunction and impaired protein processing are closely interrelated. In vitro findings further indicate that PGC-1α dysfunction may contribute to protein misfolding in neurodegeneration. OBJECTIVE: To systematically evaluate the neuropathological alterations of mice lacking the expression of the full-length PGC-1α protein (FL-PGC-1α) but expressing an N-truncated fragment. METHODS: To assess the pattern of neurodegeneration-related proteins, we performed immunostaining for Tau, pTau, α-synuclein, amyloid-ß, amyloid precursor protein, prion protein, FUS, TDP-43 and ubiquitin. Using hematoxylin and eosin, Klüver-Barrera and Bielschowsky silver stainings and anti-GFAP immunohistochemistry, we performed an anatomical mapping to provide a lesion profile. RESULTS: The immunohistochemical pattern of neurodegeneration-related proteins did not differ between FL-PGC-1α knockout and wild-type animals, and there was a complete lack of protein deposits or ubiquitin-positive inclusions. The analysis of neuropathological alterations revealed widespread vacuolation predominating in the cerebral white matter, caudate-putamen, thalamus and brainstem, and reactive astrogliosis in the brainstem and cerebellar nuclei. This morphological phenotype was thus reminiscent of human mitochondrial encephalopathies, especially the Kearns-Sayre syndrome. CONCLUSION: We conclude that the lack of FL-PGC-1α per se is insufficient to recapitulate major features of neurodegenerative diseases, but evokes a pathology seen in mitochondrial encephalopathies, which makes PGC-1α-deficient mice a valuable model for this yet incurable group of diseases.

Assessment of the role of multidrug resistance-associated proteins in MPTP neurotoxicity in mice.

GOALS: The available scientific data indicate that the pathomechanism of Parkinson's disease (PD) involves the accumulation of endogenous and exogenous toxic substances. The disruption of the proper functioning of certain transporters in the blood-brain barrier and in the blood-cerebrospinal fluid barrier in PD would accompany to that accumulation. Although there is an emerging role of the dysfunction of multidrug resistance-associated proteins (MRPs), members of ATP-b nding cassette (ABC) transporter superfamily, in neurodegenerative disorders, there is only a few available data as regards PD. So the aim of our study was the assessment of the role of certain MRPs (1 ,2, 4 and 5) in neurotoxicity induced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine METHODS: Following the intraperitoneal administration of silymarin (with MRP1, 2, 4 and 5 inhibitory effects), naringenin (with MRP1, 2 and 4 stimulatory effects), sulfinpyrazone (with MRP1, 4 and 5 inhibitory and MRP2 stimulatory effects) and allopurinol (with MRP4 stimulatory effect in doses of 100 mg/kg, 100 mg/kg, 100 mg/kg and 60 mg/kg, respectively, for one week before and after the administration of MPTP in C57B/6 mice in acute dosing regimen the striatal concentrations of dopamine, 3,4-dihydroxyphenylacetic acid and homovanillic acid has been measured using high-performance liquid chromatography. RESULTS: Although the results of these experiments showed that neither of these substances exerted significant influence on MPTP-induced striatal depletion of dopamine and its metabolites, naringenin exerted a slight prevention of dopamine decrease, while allopurinol considerably enhanced the MPTP-induced lethality in mice. The explanation of these findings would be that the stimulation of MRP1- and MRP2-mediated transport of glutathione conjugates of toxic substances may have slight beneficial effects, while stimulation of MRP4-mediated efflux of brain urate, which has an important antioxidant potency, may worsen the effects of oxidative stress.

Kynurenines in cognitive functions: their possible role in depression.

Depression can originate from changes in tryptophan availability, caused by activation of the kynurenine pathway (KP) as a result of inflammation. Alterations in the KP and the changing levels of its metabolites have recently been considered to be factors contributing to the pathogenesis of depression. The key molecular mediator which induces the conversion of tryptophan into kynurenine is indoleamine-2,3-dioxygenase. Following its activation, both the production of neurotoxic compounds and the diminished peripheral accessibility of tryptophan are regarded as essential steps in the pathophysiological processes. The aim of this review is to survey the role of the KP in depression and its relationships with cognitive functions.

Targeting the kynurenine pathway-related alterations in Alzheimer's disease: a future therapeutic strategy.

Alzheimer's disease (AD) is a chronic neurodegenerative disorder associated with dementia as a main feature. Despite decades of thorough research in the field of AD, the pathomechanism is still not fully understood. The development of novel experimental models can help us in the discovery of both genetic and non-genetic components of disease pathogenesis. As currently available therapies in AD can provide merely moderate or only temporary symptomatic relief, there is a great demand for the development of new drugs with higher therapeutic potential. Some of the candidates would be those targeting the kynurenine pathway, the neuroactive metabolites of which are surely involved in both neurodegeneration and neuroprotection, mainly in relation with glutamate excitotoxicity and oxidative stress. Both analogs of the neuroprotective kynurenic acid and small molecule enzyme inhibitors preventing the formation of neurotoxic compounds may have potential therapeutic significance. However, there is a great need for new strategies to improve efficacy, transport across the blood-brain barrier and bioavailability, naturally with simultaneous minimization of the adverse side-effects.

Endogenous neuroprotection in chronic neurodegenerative disorders: with particular regard to the kynurenines.

Parkinson's disease (PD) and Huntington's disease (HD) are progressive chronic neurodegenerative disorders that are accompanied by a considerable impairment of the motor functions. PD may develop for familial or sporadic reasons, whereas HD is based on a definite genetic mutation. Nevertheless, the pathological processes involve oxidative stress and glutamate excitotoxicity in both cases. A number of metabolic routes are affected in these disorders. The decrease in antioxidant capacity and alterations in the kynurenine pathway, the main pathway of the tryptophan metabolism, are features that deserve particular interest, because the changes in levels of neuroactive kynurenine pathway compounds appear to be strongly related to the oxidative stress and glutamate excitotoxicity involved in the disease pathogenesis. Increase of the antioxidant capacity and pharmacological manipulation of the kynurenine pathway are therefore promising therapeutic targets in these devastating disorders.

Cox-2 inhibitor attenuates NO-induced nNOS in rat caudal trigeminal nucleus.

OBJECTIVE: The aim of the present study was to determine which isoform of the cyclooxygenase (COX) enzyme plays a role in the neuronal nitric oxide synthase (nNOS) activation caused by nitroglycerin (NTG), in the most caudal part of the trigeminal caudal nucleus (TNC) of the rat. BACKGROUND: Nitric oxide donor, NTG, can trigger migraine attack in migraineurs, but not in healthy persons. In rats, subcutaneous administration of NTG (10 mg/kg) increases significantly the number of nNOS-immunoreactive neurons in the TNC after 4 hours, which could be attenuated by acetyl-salicylate (Aspirin), a nonselective COX-inhibitor. METHODS: SPRD rats were divided into 3 groups: (1) control group (no drug administration), (2) NS398 (selective COX-2 inhibitor) administration (1, 3, or 5 mg/kg), and (3) SC560 (selective COX-1 inhibitor) administration (1, 5, or 10 mg/kg). Thirty minutes after drug administration, the animals received NTG (10 mg/kg) or placebo injection. Four hours later the animals were transcardially perfused and the cervical part of the TNC was removed for immunohistochemistry. Results.-The selective COX-2 inhibitor NS398 in contrast to the selective COX-1 inhibitor SC560 attenuates the NTG-induced nNOS expression dose-dependently. CONCLUSION: These findings suggest that metabolites deriving from COX-2 (but not COX-1) may be the most important factors in the NTG-induced nNOS expression. These data could help to better understand the pathogenesis of headaches and the action of antimigraine drugs.