Improving behavioral deficits induced by perinatal ethanol and stress exposure in adolescent male rat progeny via maternal melatonin treatment.

BACKGROUND AND AIM: Early-life stressful situations and binge drinking have been thus far acknowledged as two burdensome conditions that potentially give rise to negative outcomes and then synergistically affect brain development. In this context, the hippocampus, with the greatest number of glucocorticoid receptors (GCRs) in the brain, is responsible for regulating negative responses to stress. Prolonged glucocorticoid (GC) exposure can accordingly cause oxidative stress (OS), leading to cognitive and emotional dysfunction. Against this background, melatonin, as a powerful antioxidant and hypothalamus-pituitary-adrenal (HPA) axis regulator, was administered in this study to ameliorate cognitive impairments induced by perinatal ethanol and stress exposure in adolescent male rat progeny. METHODS: Wistar rat dams were exposed to ethanol (4 g/kg) and melatonin (10 mg/kg) from gestational day (GD) 6 to postnatal day (PND) 14 and then limited nesting material (LNS) from PND0 to PND14 individually or in combination. Maternal behavior was then investigated in mothers. Afterward, the plasma corticosterone (CORT) concentration, the OS marker, the corticotropin-releasing hormone receptor type 1 (CRHR1) expression, and the GCR and brain-derived neurotrophic factor (BDNF) levels were measured in the male pups. Moreover, behavioral tasks, including the elevated plus maze (EPM), the Morris water maze (MWM), the novel object recognition (NORT), and the object-location memory (OLM) tests were completed and assessed. RESULTS: The quantity and quality of maternal care significantly decreased in the mothers with dual exposure to ethanol and stress. The plasma CORT concentration in the progeny also dropped in the Ethanol + LNS group, but the risk-taking behavior elevated significantly. The ethanol and stress exposure further revealed a significant fall in the GCR and CRHR1 expression levels, compared with stress alone. The results of learning and memory tasks also indicated a significant reduction in spatial learning and memory among animals exposed to ethanol and stress. The BDNF mRNA levels correspondingly increased in the Ethanol + LNS group, compared with LNS alone. In the presence of ethanol and stress, the superoxide dismutase (SOD) and glutathione peroxidase (GPx) activities correspondingly declined. On the other hand, the malondialdehyde (MDA) levels augmented in the hippocampus of the animals with ethanol and LNS dual exposure, as compared with the control group. Melatonin treatment (MT) thus improved nursing behaviors in dams, prevented OS, enhanced the CRHR1 and GCR expression, and reduced the BDNF levels to the similar ones in the control group. The animals in the Ethanol + LNS + MT group ultimately showed an ameliorated performance at behavioral tasks, including the memory and risk-taking behavior. CONCLUSION: It was concluded that MT could prevent stress response and memory impairments arising from dual exposure to ethanol and stress by inhibiting OS.

Lateral Hypothalamus Corticotropin-releasing Hormone Receptor-1 Inhibition and Modulating Stress-induced Anxiety Behavior.

Introduction: Stress is a reaction to unwanted events disturbing body homeostasis and its pathways and target areas. Stress affects the brain through the lateral hypothalamic area (LHA), the orexinergic system that mediates the effect of corticotropin-releasing hormone (CRH) through CRH Receptor Type 1 (CRHr1). Therefore, this study explores the outcome of stress exposure on anxiety development and the involvement of the LHA through LHA-CRHr1. Methods: Male Wistar rats (220-250 g) implanted with a cannula on either side of the LHA received acute or chronic stress. Subsequently, exploratory behavior was examined using the Open Field (OF), and anxiety was tested by Elevated Plus Maze (EPM). Before sacrifice, the cerebrospinal fluid (CSF) and the blood were sampled. Nissl stain was performed on fixed brain tissues. Results: Acute stress reduced exploration in of and increased anxiety in EPM. LHA-CRHr1 inhibition reversed the variables to increase the exploration and decrease anxiety. In contrast, chronic stress did not show any effect on anxiety-related behaviors. Chronic stress decreased the cell population in the LHA, which was prevented by the CRHr1 inhibition. However, the CRHr1 inhibition could not reverse the chronic stress-induced increase in the CSF orexin level. Furthermore, plasma corticosterone levels increased through acute or chronic stress, impeded by the inhibition of CRHr1. Conclusion: Our results recognize LHA-CRHr1 as a capable candidate that modulates acute stress-induced anxiety development and chronic stress-induced changes in the cellular population of the region. Highlights: Acute stress, increased immobility of the rat in open field and elevated plus maze.Chronic stress, increased orexin production while decreasing neuronal survival.The anxiety and immobility were not developed in presence of CRHr1.CRHr1 blocking reversed the chronic stress changes in corticosterone and orexin. Plain Language Summary: Lateral Hypothalamus (LH) is a region involved in sleep and appetite regulation and recently known to play role in stress pathophysiology. The stress mediating function of the LH is performed through Corticotropin Releasing Hormone Receptor type-1 (CRHr1). This study explored the role of LH- CRHr1 in anxiety development and orexin production. Acute and chronic stress affected the behavior and molecular changes, differently. The acute stress increased the anxiety condition, while the chronic stress could only change the molecular criteria. Although we assumed that the inability of the chronic stress to develop anxiety may be attributable to habituation, the chronic stress could increase the plasma corticosterone and orexin level. All of the stress mal-changes in behavior and molecular level prevented by antagonising CRHr1 in the LH, indicating a gating function of LH-CRHr1 for stress development.

Hippocampal orexin-1 and endocannabinoid-1 receptors underlie the kainate-induced occlusion in theta-burst long- term potentiation.

INTRODUCTION: Seizures may result from the hyperexcitable neuronal activity of the brain. Multiple neurotransmitter receptors, including orexin (OX) and endocannabinoids interfere with forming the synaptic responses linked to the seizures. Therefore, this study investigates the involvement of OX-1 (OX1R) and endocannbinoid-1 (CB1R) receptors in the kainate- induced excitability in the synaptic field responses. MATERIAL AND METHODS: Theta pattern used to stimulate Schaffer collaterals and then metal microelectrodes to record the CA1 field excitatory postsynaptic potentials (fEPSPs). Input/ output stimulation and responses and paired- pulse (PP) stimuli employed to measure the state of synaptic activity in normal and kainate- induced seizure-like hyperexcitable activities and the slope of fEPSPs used as a measure of the change in the synaptic activity. Furthermore, agonists and antagonists of OX and endocannabinoids infused to investigate the involvement of their receptors. RESULT: The results showed that kainate application increased the fEPSP slope either in input stimuli with different intensities/output synaptic responses (I/O), or test pulse stimulated baseline synaptic responses (BSR) and, hence, increased the excitability of field responses in the CA1 region. However, neither kainate nor theta burst stimulation (TBS) could alter the PP stimuli -induced synaptic responses. TBS increased the fEPSP slope of the kainate-applied I/O and BSR, however, the increase was not high enough in BSR to be classified as long-term potentiation (LTP). The single-antagonist OX1R and CB1R administration prevented TBS- induced potentiation and partially recovered the effect by adding eCB or OX agonists in kainate-injected animals. In contrast, OX or combined eCB-OX antagonist application group demonstrated nearly full recovery of LTP induction. CONCLUSION: Our study concludes that blockade of OX1 or CB1 prevents the induction of LTP, and OX infusion or both receptor blockade recovers the LTP.

The Combined Effects of Perinatal Ethanol and Early-Life Stress on Cognition and Risk-Taking Behavior through Oxidative Stress in Rats.

Both prenatal ethanol and early-life stress have been shown to induce reduced risk-taking and explorative behavior as well as cognitive dysfunction in the offspring. In this study, we examined the effect of combined exposure to ethanol and early stress on maternal care, exploratory behavior, memory performances, and oxidative stress in male offspring. Pregnant rats were exposed to ethanol (4 g/kg) from gestational day (GD) 6-to postnatal day (PND) 14 and limited nesting material (LNS) from PND0-PND14 individually or in combination. Maternal behavior was evaluated during diurnal cycle. The level of corticosterone hormone and markers of oxidative stress were evaluated in the pups. Risk-taking and explorative behavior were assessed with the elevated-plus maze (EPM) test and cognitive behavior with the Morris water maze (MWM), novel object recognition (NORT), and object location memory (OLM) tests. In the mothers, perinatal alcohol or LNS either alone or in combination decreased maternal behavior. In the offspring, the combination of the two factors significantly increased the pup's plasma corticosterone concentration in comparison with ethanol and LNS alone. Reduced risk-taking behavior was observed in the ethanol, LNS and ethanol + LNS groups compared with the control group, and this was amplified in the co-exposure of ethanol and LNS groups. The MWM, NORT, and OLM tests revealed spatial and recognition memory impairment in the ethanol and LNS groups. This impairment was more profound in the co-exposure of ethanol and LNS. Also, we observed a significant decrease in superoxide dismutase (SOD) and glutathione peroxidase (GPx) activities and an increase in malondialdehyde (MDA) level in the hippocampus of ethanol and LNS co-exposed animals as compared with individual exposure of ethanol and LNS. While each factor independently produced similar outcomes, the results indicate that the dual exposure paradigm could significantly strengthen the outcomes.

Hippocampal orexin receptors: Localization and function.

Orexin (hypocretin) is secreted from the perifornical/lateral hypothalamus and is well known for sleep regulation. Orexin has two, orexin A and B, transcripts and two receptors, type 1 and 2 (OX1R and OX2R), located in the plasma membrane of neurons in different brain areas, including the hippocampus involved in learning, memory, seizures, and epilepsy, as physiologic and pathologic phenomena. OX1R is expressed in the dentate gyrus and CA1 and the OX2R in the CA3 areas. Orexin enhances learning and memory as well as reward, stress, seizures, and epilepsy, partly through OX1Rs, while either aggravating or alleviating those phenomena via OX2Rs. OX1Rs activation induces long-term changes of synaptic responses in the hippocampus, an age and concentration-dependent manner. Briefly, we will review the localization and functions of hippocampal orexin receptors, their role in learning, memory, stress, reward, seizures, epilepsy, and hippocampal synaptic plasticity.

Maternal ethanol exposure induces behavioral deficits through oxidative stress and brain-derived neurotrophic factor interrelation in rat offspring.

Alcohol consumption during pregnancy damages the central nervous system of developing fetus and results in persistent physical and neurobehavioral abnormalities, including learning and memory disorders. The hippocampus which is involved in learning and memory is highly susceptible to the ethanol neurotoxic effects. Oxidative stress is one of the mechanisms in alcohol-induced disorders. Ethanol also interferes with the brain-derived neurotrophic factors (BDNF) expression. Using vitamin E as a potent antioxidant, we studied the possible interrelation between oxidative stress and BDNF on cognition. Ethanol (4 g/kg) and vitamin E (100, 200, and 400 mg/kg) were given to pregnant Wistar rats on first day of gestation (GD) until weaning (28 days). Oxidative stress marker, BDNF expression, and cyclic AMP-response binding-protein (CREB) expression levels were measured on postnatal days (PND) 28. Object location memory (OLM) was evaluated on PND 34. Our results demonstrated that ethanol exposure significantly reduced glutathione peroxidase (GPx) activity, reduced glutathione (GSH), reduced/oxidized glutathione (GSH/GSSG) ratio, and increased superoxide dismutase (SOD) activity, malondialdehyde (MDA) levels, and carbonyl protein content in the hippocampus. Total BDNF, BDNF mRNA, and CREB expression significantly reduced in the hippocampus by ethanol exposure. Also, ethanol significantly reduced the discrimination index (DI) in the OLM test. In addition, vitamin E administration could reduce oxidative stress, increase significantly BDNF and CREB levels, and improve cognitive dysfunction induced by ethanol exposure. Collectively, results suggest that probably oxidative stress can interrelate with the BDNF system for modulating cognitive function in the ethanol-exposed rat.

Pharmacological upregulation of GLT-1 alleviates the cognitive impairments in the animal model of temporal lobe epilepsy.

It is known that hippocampal epileptogenesis is accompanied by hyperexcitability, glutamate-related neuronal dysfunctions and consequently cognitive deficits. However, the neuroprotective role of astrocytic glutamate uptake through the Glutamate Transporter-1 (GLT-1) remains to be unknown in these processes. Therefore, to assess the effect of glutamate uptake, pharmacological upregulation of GLT-1 using ceftriaxone administration (200 mg/kg/day, i.p, 5 days) was utilized in Li-PIL animal models of temporal lobe epilepsy (TLE). Glutamate concentration and glutamine synthetase activity were analyzed using biochemical assays. In addition, GLT-1 gene expression was assessed by RT-qPCR. Finally, cognitive function was studied using Morris water maze (MWM) test and novel object recognition task (NORT). Our results demonstrated that the acute phase of epileptogenesis (first 72 hours after Status Epilepticus) was accompanied by an increase in the hippocampal glutamate and downregulation of GLT-1 mRNA expression compared to controls. Ceftriaxone administration in epileptic animals led to a reduction of glutamate along with elevation of the level of glutamine synthetase activity and GLT-1 expression in the acute phase. In the chronic phase of epileptogenesis (4 weeks after Status Epilepticus), glutamate levels and GLT-1 expression were decreased compared to controls. Ceftriaxone treatment increased the levels of GLT-1 expression. Furthermore, impaired learning and memory ability in the chronic phase of epileptogenesis was rescued by Ceftriaxone administration. This study shows that astrocytic glutamate uptake can profoundly impact the processes of hippocampal epileptogenesis through the reduction of glutamate-induced excitotoxicity and consequently rescuing of cognitive deficits caused by epilepsy.

Piperine ameliorated memory impairment and myelin damage in lysolecethin induced hippocampal demyelination.

AIMS: We still do not have effective treatment for hippocampal demyelination and memory deficit, the two common comorbidities in multiple sclerosis (MS). This study aimed to assess the therapeutic effect of Piperine (the main alkaloid of black pepper) in an experimental model of demyelination. MAIN METHODS: Demyelination was induced in male Wistar rats by bilateral injection of lysolecithin (LPC) into the CA1 region of the hippocampus. Piperine (5, 10, 20 mg/kg) was daily injected intraperitoneally three days post LPC injection for ten days. The spatial memory was examined by the Morris water maze task. Demyelination and astrocyte activation were assessed by an immunohistological study. The gene expression analysis of TNF-α, IL1-ß, NF-κB, IL-10, Foxp3, iNOS, Nrf2, HO1, MBP, and BDNF was done using qPCR. The total antioxidant capacity of hippocampal tissue was measured using FRAP assay. KEY FINDINGS: Our results showed that piperine improved the memory performance and myelin repair in the hippocampal demyelination model. Piperine inhibited iNOS expression concomitant with enhanced expression levels of Nrf2, HO1 and the total antioxidant capacity in the hippocampal tissue. Piperine treatment significantly reduced the gene expression level of TNF-α, IL1-ß, NF-κB, and glial activation in the injured area; however, the mRNA level of IL-10, Foxp3, BDNF and MBP were significantly increased. SIGNIFICANCE: We found piperine to be an effective treatment for spatial memory impairment and myelin repair in the hippocampal demyelination model. However, further experimental evidence is needed to investigate the precise mechanisms underlying piperine as a promising therapeutic target in MS patients.

Investigating the role of the amygdala orexin receptor 1 in memory acquisition and extinction in a rat model of PTSD.

Understanding the mechanisms underlying memory is essential for the treatment of post-traumatic stress disorder (PTSD). Orexin, as a lateral hypothalamic (LH) neuropeptide, interferes with the stages of memory, primarily through the orexin receptor1 (Orx1R). The aim of this study was to evaluate the effects of amygdala Orx1R in the acquisition and extinction processes of PTSD modeled in animals. In three experiments, rats were divided into three groups: control (Naïve), shock (receiving a foot shock), and PTSD (experiencing Single prolonged stress (SPS) method). The first experiment aimed to evaluate LH activity in PTSD modeled rats. The second and third experiments aimed to evaluate the effects of Orx1R in the acquisition and extinction of fear memory in PTSD modeled animals. SB334867 (SB) or its solvent was microinjected into the amygdala and the rats were subjected to conditioning thereafter. In the second group, we used a single injection after conditioning. In the third group, we used three consecutive injections (one after each memory test). Some behaviors and Orx1R expression were evaluated. The freezing response was significantly longer in the PTSD group than on the control. Similarly, anxiety and sensitized fear were also intensified. CFos expression levels in LH was higher in the PTSD group. Inhibition of Orx1R in the amygdala significantly decreased memory acquisition, diminished anxiety, and decreased the sensitized fear in the SB group. Applying SB to the amygdala after each fear memory test significantly decreased freezing. Expression of Orx1R was significantly higher following fear conditioning. These results indicate a likely involvement of the orexin and amygdalar Orx1R in memory acquisition and in extinction of PTSD.

Folic Acid Protects Rat Cerebellum Against Oxidative Damage Caused by Homocysteine: the Expression of Bcl-2, Bax, and Caspase-3 Apoptotic Genes.

There is evidence that oxidative stress involves in homocysteine-induced pathogenesis. Considering the antioxidative properties of folic acid and its involvement as a cofactor for methionine synthase (MS) in the homocysteine-methionine cycle, the aim of this study was to evaluate the mechanism associated with homocysteine-induced toxicity and its prevention with folic acid supplementation. Male rat pups were divided into four groups including control, homocysteine (Hcy), Hcy + folic acid and folic acid groups. The Hcy group received Hcy 0.3-0.6 µmol/g body weight, while Hcy + folic acid group received folic acid orally as 0.011 µmol/g body weight along with Hcy on a postnatal day (PD) 4 until 25. The reduced and oxidized glutathione (GSH and GSSG) levels, GSH/GSSG ratio, protein carbonyl content, cystathionine ß synthase (CBS), and MS activities in the cerebellum were measured 25 days after birth. Levels of malondialdehyde (MDA), marker of lipid peroxidation were measured. Also, Bcl2, Bax, and caspase-3 expression levels were measured by real-time quantitative PCR. Furthermore, caspase-3 protein level assay was performed by the ELISA test. Results indicated that Hcy administration could promote both lipid and protein oxidation, which was associated with increased amounts of caspase-3 mRNA and protein levels and Bax mRNA expression level in this group. Cerebellar MS, CBS enzyme activity, GSH, GSSG, and GSH/GSH ratio did not change following Hcy administration. Folic acid significantly reduced MDA level, protein carbonyl content, Bax, the caspase-3 mRNA, and protein expression levels in the cerebellum of Hcy-treated group. Moreover, cerebellar MS, CBS enzyme activity, GSH, and GSH/GSH ratio increased following folic acid treatment. We conclude that Hcy might cause apoptosis in the cerebellum. We suggest that folic acid, in addition of having antioxidant properties, can protect cerebellum against homocysteine-mediated neurotoxicity via modulating the expression of proteins that are contributed in regulation of apoptosis in the rat's cerebellum.

Implications from early life stress on the development of mouse ovarian follicles: Focus on oxidative stress.

AIM: The early life stress has significant long-term effects on the development of the offspring. This study was undertaken to verify if maternal separation as a stressor agent affects the oxidative status and developmental competence of mouse pre-antral follicles (PF) during in vitro culture period. METHODS: Female litters of National Medical Research Institute mice were divided into two groups: maternally separated group (MS), separated from the mothers for 6 h per day from postnatal days 2-16; and the rest considered as the control group, which left undisturbed over the 14 days. The litters were sacrificed and the ovarian tissue was harvested to isolate the PF. The PF were in vitro cultured up to 12th day when ovulation was induced. The developmental parameters and oxidative status (i.e., total antioxidant capacity and Malondialdehyde levels, as well as the activities of superoxide dismutase, glutathione peroxidase and catalase) were assessed. RESULTS: The rates of survival, antrum formation, ovulation and oocyte maturation of PF derived from the MS group were significantly lower compared with those of the control group. Furthermore, the Malondialdehyde level of the MS group was significantly higher than that of the control group. By contrast, the total antioxidant capacity level was lower in the MS group with respect to the control group. Also, the activity of superoxide dismutase, glutathione peroxidase and catalase of PF, derived from the MS group, was significantly lower compared with those of the control group. CONCLUSION: Early life stress damages the developmental competence of mouse PF through induction of oxidative stress.

Hippocampal orexin receptor blocking prevented the stress induced social learning and memory deficits.

Stress as a homeostatic challenge leads to the malfunction of learning and memory processes, namely social learning and memory. The orexin system is involved in stress responses through connections to the hypothalamic-pituitary axis (HPA). In addition, the hippocampus, a structure vulnerable to stress-induced changes, expresses orexin receptors 1 and 2 (OXr1 and OXr2) in various sub-regions. The present study is aimed at assessing the effects of hippocampal orexin receptor blockade on social learning and memory impairments and anxiety development following stress. Male Wistar rats (220-250 g) underwent cannula implantation in the hippocampus. Acute (two mild electric shocks, 5.5 mA) and chronic stresses (ten days of restraint, 6 h daily) were applied with or without injection of orexin receptor antagonists (SB-334867 or TCS OX 29). Sociability and social novelty in animals were assessed in a three-chamber social maze at the end of stress application. Anxiety and exploratory behavior of animals were then examined, with 20 min intervals, using the open field (OF) and elevated plus maze (EPM) tests, respectively. Cisterna Magna cerebro-spinal fluid (CSF) was drained, before sacrifice, for orexin (OX) assay and trunk blood was collected to measure the plasma corticosterone (CRT). Neither the acute nor the chronic stress could affect the sociability. The acute but not chronic stress prevented the animal from sniffing the familiar caged rat in the novelty session, a response which was reversed following the blockade of both OXRs. Furthermore, acute but not chronic stress, led to increased anxiety and immobility behavior which were both impeded by blocking the orexin receptor (OXR). Conversely, OX content in CSF increased due to chronic restraint stress, an effect that was reversed by orexin blockade. Finally, elevated plasma CRT was recorded in response to both acute and chronic stresses. The observed increase in plasma CRT in chronically-stressed rats was abolished following inhibition of OXRs, however a similar effect was not seen in the acute-stress group. Our results identify hippocampal OXRs as potential candidates capable of preventing acute stress-induced impairments of social novelty and anxiety behavior, and chronic stress-induced plasma CRT and CSF orexin, changes. OXR manipulation may improve adaptation to stress pathophysiology.

Postnatal Administration of Homocysteine Induces Cerebellar Damage in Rats: Protective Effect of Folic Acid.

A widely held view suggests that homocysteine (Hcy) can contribute to neurodegeneration through promotion of oxidative stress. There is evidence that homocysteine is toxic to cerebellar Purkinje neurons in vitro; however, in vivo action of Hcy on Purkinje cell has not been investigated so far. Thus, this study was designed to evaluate the Hcy effects on neonatal rat cerebellum and cerebellar oxidative stress. We also evaluated the folic acid effects on biochemical alterations elicited by hyperhomocysteinemia (hHcy) in the cerebellum. Group I received normal saline, group II received Hcy subcutaneously twice a day at 8-h intervals (0.3-0.6 µmol/g body weight), group III received Hcy + folic acid (0.011 µmol/g body weight), and group IV received folic acid on postnatal day (PD) 4 until 25. On day 25, superoxide dismutase (SOD) and glutathione peroxidase (GPx) activities in the cerebellum and motor cortex were assayed. Malondialdehyde (MDA) levels were also evaluated as a marker of lipid peroxidation. Rotarod and locomotor activity tests were performed in PD 25-27. Our results indicated that administration of Hcy increased plasma, cortical, and cerebellar total Hcy levels; reduced GPx activity; and induced lipid peroxidation in the cerebellum. Hcy impaired performance on the rotarod in rats. However, treatment with folic acid significantly attenuated motor coordination impairment, GPx activity reduction, the lipid peroxidation process, and significantly reduced plasma total Hcy levels. Histological analysis indicated that Hcy could decrease Purkinje cell count and folic acid prevented this toxic effect. We conclude that Hcy can induce neurotoxicity and folic acid has neuroprotective effects against cerebellar Hcy toxicity.

Posterior hypothalamus glutamate infusion decreases pentylenetetrazol-induced seizures of male rats through hippocampal histamine increase.

OBJECTIVES: Seizures are epileptic manifestations that are intrinsically modulated through different neurotransmitters and receptor systems. Although glutamate increases excitation and hence seizures, it activates other systems which could potentially terminate seizures. Histamine originates from neurons of the posterior hypothalamus (PH) and can mediate anticonvulsant properties, but the effect of local PH glutamate on hippocampal histamine content is unknown. Therefore, in this study, the effect of PH glutamate and the involvement of hippocampal histamine in pentylenetetrazol (PTZ) induced seizure activity was studied. MATERIALS AND METHODS: OX2R antagonist (TCS OX2 29, 40nmol/1µl, intra-PH), AMPA/Kainate receptor antagonist (CNQX, 3mM, intra-PH) and glutamate (1mM) were injected bilaterally into PH using stereotaxic surgery. The intravenous PTZ infusion model was used to generate behavioral convulsions and the amount of hippocampal histamine content was then measured using a biochemical method. RESULTS: Administration of glutamate into PH decreased both seizure stage and the duration of tonic-clonic convulsion (TCC) with increasing TCC latency and hippocampal histamine content. Blocking OX2Rs alone or coinhibition of OX2Rs and AMPA/kainate receptors reversed these effects by increasing both seizure stage and TCC duration, and by decreasing both latency and consequent histamine content. CONCLUSIONS: Our findings suggest that glutamate administration into PH may control seizures (stages and duration) through increasing the hippocampal histamine content.

Evaluating the role of astrocytes on ß-estradiol effect on seizures of Pilocarpine epileptic model.

Epilepsy with periodic and unpredictable seizures is associated with hippocampal glutamate toxicity and tissue reorganization. Astrocytes play an important role in mediating the neuroprotective effects of estradiol and reducing seizure severity. Accordingly, the protective effects of low and high doses of estradiol on behavioral, astrocytic involvement and neuronal survival aspects of Pilocarpine-induced epilepsy were investigated. Lithium- Pilocarpine (30mg/kg) model was used to provoke epilepsy. Βeta-estradiol (2,40µg/µl) was injected subcutaneously from 48 before to 48h after seizure induction. Behavioral convulsions were then monitored and recorded on the day of induction. Four weeks later, glutamine synthetase (GS) activity and the astrocyte transporter GLT-1 expression of the hippocampus were measured. Moreover, hippocampal glutamate and GABA were evaluated to study excitability changes. Finally, neuronal counting in the hippocampus was also performed using Nissl staining. The latency for generalized clonic (GC) convulsions significantly increased while the rate of GC and death significantly reduced due to ß-estradiol treatment. GS activity and GLT-1 expression increased in the groups receiving the high dose of ß-estradiol and Pilocarpine. Furthermore, the amount of both GABA and glutamate content decreased due to high dose of estradiol, while only GABA increased in Pilocarpine treated rats. Finally, administration of ß-estradiol with low and high doses increased and improved the density of nerve cells. It is concluded that chronic administration ß-estradiol has anticonvulsant and neuroprotective properties which are plausibly linked to astrocytic activity.

Lateral hypothalamus orexinergic system modulates the stress effect on pentylenetetrazol induced seizures through corticotropin releasing hormone receptor type 1.

Stress is a trigger factor for seizure initiation which activates hypothalamic pituitary adrenal (HPA) axis as well other brain areas. In this respect, corticotropin releasing hormone (CRH) and lateral hypothalamus (LH) orexinergic system are involved in seizure occurrence. In this study, we investigated the role of LH area and orexin expression in (mediation of) stress effect on pentylenetetrazol (PTZ) -induced seizures with hippocampal involvement. Two mild foot shock stresses were applied to intact and adrenalectomized animals; with or without CRHr1 blocking (NBI 27914) in the LH area. Then, changes in orexin production were evaluated by RT-PCR. Intravenous PTZ infusion (25 mg/ml) -induced convulsions were scored upon modified Racine scale. Finally, hippocampal glutamate and GABA were evaluated to study excitability changes. We demonstrated that the duration and severity of convulsions in stress-induced as well as adrenalectomized group were increased. Plasma corticosterone (CRT) level and orexin mRNA expression were built up in the stress and/or seizure groups. Furthermore, glutamate and GABA content was increased and decreased respectively due to stress and seizures. In contrast, rats receiving CRHr1 inhibitor showed reduced severity and duration of seizures, increased GABA, decreased glutamate and corticosterone and also orexin mRNA compared to the inhibitor free rats. Stress and adrenalectomy induced augmenting effect on seizure severity and duration and the subsequent reduction due to CRHr1 blocking with parallel orexin mRNA changes, indicated the likely involvement of CRH1r induced orexin expression of the LH in gating stress effect on convulsions.

Melatonin ameliorates oxidative damage induced by maternal lead exposure in rat pups.

During the particular period of cerebellum development, exposure to lead (Pb) decreases cerebellum growth and can result in selective loss of neurons. The detection and prevention of Pb toxicity is a major international public health priorities. This research study was conducted to evaluate the effects of melatonin, an effective antioxidant and free radical scavenger, on Pb induced neurotoxicity and oxidative stress in the cerebellum. Pb exposure was initiated on gestation day 5 with the addition of daily doses of 0.2% lead acetate to distilled drinking water and continues until weaning. Melatonin (10mg/kg) was given once daily at the same time. 21 days after birth, several antioxidant enzyme activities including superoxide dismutase (SOD) and glutathione peroxidase (GPx) were assayed. Thiobarbituric acid reactive substance (TBARS) levels were measured as a marker of lipid peroxidation. Rotarod and locomotor activity tests were performed on postnatal days (PDs) 31-33 and a histological study was performed after completion of behavioral measurements on PD 33. The results of the present work demonstrated that Pb could induce lipid peroxidation, increase TBARS levels and decrease GPx and SOD activities in the rat cerebellum. We also observed that Pb impaired performance on the rotarod and locomotor activities of rats. However, treatment with melatonin significantly attenuated the motoric impairment and lipid peroxidation process and restored the levels of antioxidants. Histological analysis indicated that Pb could decrease Purkinje cell count and melatonin prevented this toxic effect. These results suggest that treatment with melatonin can improve motor deficits and oxidative stress by protecting the cerebellum against Pb toxicity.

Impact of the C1431T Polymorphism of the Peroxisome Proliferator Activated Receptor-Gamma (PPAR-γ) Gene on Fasted Serum Lipid Levels in Patients with Coronary Artery Disease.

BACKGROUND/AIMS: The C1431T polymorphism of peroxisome proliferator activated receptor-γ (PPAR-γ) gene is related to diabetes and metabolic-syndrome. However, studies have been inconclusive about its association with coronary artery disease (CAD) and there have been no studies analyzing the association of this polymorphism with fasted-serum-lipid levels in Iranian-individuals with CAD. We investigated the association of PPAR-γ C1431T-polymorphism with CAD and dyslipidaemia in 787 individuals. METHODS: Anthropometric-parameters and biochemical-measurements were evaluated, followed by genotyping. The association of the genetic-polymorphisms with CAD and lipid-profile was determined by univariate/multivariate-analyses. RESULTS: Patients with CT or CT+TT genotype were at an increased-risk of CAD relative to CC-carriers (adjusted odds ratio: 2.03; 95% confidence interval, 1.01-4.09; p = 0.046). However, in the larger population, CT genotype was present at a higher frequency in the group with a positive angiogram. Furthermore, CT+TT genotypes were associated with an altered fasted-lipid-profile in the initial population sample of patients with a positive angiogram, compared to the group with a negative-angiogram. The angiogram-positive patients carrying the T allele had a significantly higher triglyceride, serum C-reactive protein and fasting-blood-glucose. CONCLUSION: We have found the PPAR-γ C1431T polymorphism was significantly associated with fasted serum lipid profile in individuals with angiographically defined CAD. Since accumulating data support the role of PPAR-γ polymorphisms in CAD, further studies are required to investigate the association of this polymorphism with coronary artery disease.

Melatonin prevents oxidative damage induced by maternal ethanol administration and reduces homocysteine in the cerebellum of rat pups.

Chronic alcoholism leads to elevated plasma and brain homocysteine (Hcy) levels, as demonstrated by animal experiments. This study was designed to evaluate the alterations in offspring rat cerebellum following increase of plasma Hcy level induced by maternal exposure to ethanol and to investigate the possible protective role of melatonin administration upon cerebellar ethanol-induced neurotoxicity. The adult female rats were divided randomly into 4 groups, including one control and three experimental groups, after vaginal plagues. Group I received normal saline, group II received ethanol (4 g/kg), group III received ethanol+melatonin (10mg/kg) and group IV received melatonin on day 6 of gestation until weaning. 21 days after birth, plasma Hcy level, level of lipid peroxidation, the activities of several antioxidant enzymes including superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), and levels of bcl-2 and bax mRNA expression in cerebellum were determined. Our results demonstrated that ethanol could induce lipid peroxidation, and decrease antioxidants activities and increase plasma total Hcy level. We also observed that ethanol impaired performance on the rotarod and locomotor activities of rats. However, treatment with melatonin significantly attenuated motoric impairment, the lipid peroxidation process and restored the levels of antioxidant activities and significantly reduced plasma total Hcy levels. Moreover, melatonin reduced bax/bcl-2 ratio in the presence of ethanol. We conclude that these results provide evidence that ethanol neurotoxicity in part is related to increase of plasma Hcy levels and melatonin with reducing of plasma Hcy level has neuroprotective effects against ethanol toxicity in cerebellum.

Hippocampal orexin receptors inactivation reduces PTZ induced seizures of male rats.

INTRODUCTION: Orexin has been shown to be involved in a number of physiological and behavioral processes including, feeding and metabolism, reward, nociception, and anxiety. Furthermore, orexin can cause increased neuronal excitability that gives rise to epileptic activity. The distribution of orexin receptor expression in the hippocampus, suggests a possible importance of orexin in the control of seizures in the temporal lobe epilepsy. Therefore, in this study, the effect of hippocampal orexin 1 and 2 receptors on seizure and glutamate and GABA (gamma-aminobutyric acid) contents was explored. MATERIALS AND METHODS: Orexin 1 receptor (OX1R) antagonist (SB) and OX2R antagonist (TCS) were administrated bilaterally through separate cannulae into both hippocampi. Behavioral convulsions were provoked by intravenous pentylenetetrazol (PTZ) application model. The amount of total hippocampal glutamate and GABA contents was then measured by a biochemical method. RESULTS: SB (50 nmol) infusion reduced seizure stage, duration and decreased glutamate while GABA content was increased. SB (200 nmol) also reduced seizure stage, duration and glutamate content, without change of GABA content. TCS (20 nmol) infusion reduced seizure stage and duration without concomitant change in glutamate and GABA contents. Further, TCS (40 nmol) did neither affect the seizure nor the GABA, while decreased glutamate content. Co-administration of SB (50 nmol) with TCS (40 nmol) and also SB (200 nmol) with TCS (40 nmol) reduced seizure stage, duration and glutamate, but increased GABA content. CONCLUSION: It is concluded that OX1R and OX2R antagonists reduce convulsive intensity, partially through alterations of hippocampal glutamate and GABA contents.