Taner Dagci

Effects of nitric oxide synthase inhibition on spatial discrimination learning and central DA2 and mACh receptors.

Cholinergic and dopaminergic systems are involved in spatial memory and are modulated by nitric oxide (NO); NO has well documented effects on place learning in rodents. The aim of the present study was to investigate the effect of NOS inhibition on place learning in the water maze and to evaluate the relationships between NOS inhibition, learning performance, dopamine (DA) D2 and muscarinic acetylcholine (mACh) receptors. Male Sprague-Dawley rats received the NOS inhibitor Nomega-Nitro-l-Arginine (l-NA), or saline and were trained in the water maze. Rats that were not trained, but received the same treatments were also included. Following treatments with or without water maze training, [3H]-QNB and [3H]-spiperone binding in cortex, striatum and hippocampus were determined to assess the effects of NOS inhibition and/or learning on DA D2 and mACh receptor regulation. The overall results of the present study showed that: (1) NOS inhibition impairs performance in the MWM; (2) NOS inhibition does not affect specific binding to DA D2 (striatum and hippocampus) and mACh (cortex and hippocampus) receptors; (3) MWM training lowers D2 and mACh receptor binding in cortical regions.

The effect of hyperbaric oxygen on neuroregeneration following acute thoracic spinal cord injury

AIMS Although hyperbaric oxygen (HBO) treatment following spinal cord injury (SCI) have been studied in terms of neurological function and tissue histology, there is a limited number studies on spinal cord tissue enzyme levels. MAIN METHODS The effect of HBO treatment in SCI was investigated by measuring superoxide dismutase (SOD), catalase, glutathione peroxidase (GPx), nitric oxide synthase (NOS) and nitric oxide (NO) activity in the injured tissue. SCI was induced by applying an aneurysm clip extradurally at the level of T9-T11 vertebrae. Preoperative HBO (preopHBO) treatment was applied for 5days and postoperative HBO (postopHBO) for 7days. KEY FINDINGS In the preopHBO group, a significant decrease was observed in NOS and NO compared to the SCI group. There was a decrease in SOD, NOS and NO in the postopHBO group when compared to the SCI group. In the pre-postHBO group SOD, GPx, NOS and NO decreased significantly. There was a decrease in SOD in postopHBO compared to preopHBO. In the prepostopHBO, SOD decreased significantly compared to that in the preopHBO group. The prepostopHBO presented a significant decrease in GPx compared to postopHBO (p<0.05 for all parameters). No significant difference was observed for catalase for all groups. Significant improvement was found in BBB scores for both postopHBO and prepostHBO groups when compared to the SCI group (p<0.05). SIGNIFICANCE HBO treatment was found to be beneficial following SCI in terms of biochemical parameters and functional recovery in the postoperative period.

Forced swim stress elicits region-specific changes in CART expression in the stress axis and stress regulatory brain areas

CART mRNA and peptides are highly expressed in the anatomical structures composing the hypothalamo-pituitary-adrenal (HPA) axis and sympatho-adrenal system. Anatomical and functional studies suggest that CART peptides may have a role in the regulation of the neuroendocrine and autonomic responses during stress. Our previous study showed that CART peptides increased significantly in the male hypothalamus and amygdala 10min after the forced swim stress. The present study aimed to examine the effect of forced swim stress on CART peptide expression in selected brain regions, including those where CART peptide expression has not been reported before (frontal cortex, pons, medulla oblongata), as well as in endocrine glands related to stress in male Sprague Dawley rats. A total of 16 (n=8) animals were used, including control groups. Rats were subjected to forced swim on two consecutive days, and sacrificed on the second day, 2h after the termination of the stress procedure. Frontal cortex, pons, medulla oblongata, hypothalamus, pituitary and adrenal glands were dissected and homogenized. CART peptide expression in these tissues was measured by Western Blotting and six different CART peptide fragments were identified. Our results showed that forced swim stress elicited region-specific changes in CART peptide expression. CART was upregulated in the frontal cortex, hypothalamus, medulla oblongata and adrenal gland while there was no change in the pons and pituitary. Enhanced CART peptide fragments in these brain regions and adrenal glands may have a role in the regulation of the HPA and sympatho-adrenal axis activity during stress response.

Effects of resveratrol on hydrogen peroxide-induced oxidative stress in embryonic neural stem cells

Resveratrol, a natural phenolic compound, has been shown to prevent cardiovascular diseases and cancer and exhibit neuroprotective effects. In this study, we examined the neuroprotective and antioxidant effects of resveratrol against hydrogen peroxide in embryonic neural stem cells. Hydrogen peroxide treatment alone increased catalase and glutathione peroxidase activities but did not change superoxide dismutase levels compared with hydrogen peroxide + resveratrol treatment. Nitric oxide synthase activity and concomitant nitric oxide levels increased in response to hydrogen peroxide treatment. Conversely, resveratrol treatment decreased nitric oxide synthase activity and nitric oxide levels. Resveratrol also attenuated hydrogen peroxide-induced nuclear or mitochondrial DNA damage. We propose that resveratrol may be a promising agent for protecting embryonic neural stem cells because of its potential to decrease oxidative stress by inducing higher activity of antioxidant enzymes, decreasing nitric oxide production and nitric oxide synthase activity, and alleviating both nuclear and mitochondrial DNA damage.

Bladder Function Recovery in Rats With Traumatic Spinal Cord Injury After Transplantation of Neuronal-Glial Restricted Precursors or Bone Marrow Stromal Cells

PURPOSE We investigated functional recovery of the lower urinary system in rats with spinal cord injury after transplanting neuronal restricted precursors/glial restricted precursors or neural cells derived from bone marrow stromal cells into the injured area of the spinal cord. MATERIALS AND METHODS A total of 30 rats underwent experimentation in 4 groups, including group 1--sham operation, group 2--spinal cord injury plus neuronal restricted precursor/glial restricted precursor transplantation, group 3--spinal cord injury plus bone marrow stromal cell transplantation and group 4--spinal cord injury control. All rats in the 4 groups were investigated urodynamically and sacrificed on day 28 after transplantation. The cells transplanted into the injured spinal cord underwent histological investigation. RESULTS Transplanted cells (neuronal and glial restricted precursors, and bone marrow stromal cells) were found to maintain a presence in the injured spinal cord area. Baseline pressure, maximum capacity, mean uninhibited contraction amplitude, mean voiding pressure, voided volume and post-void residual volume were significantly better in groups 2 and 3 than in group 4, while baseline pressure in group 2 was better than that in group 3. We found no significant difference among the groups according to mean uninhibited contraction frequency. CONCLUSIONS Although neuronal/glial restricted precursor transplanted rats seemed to have more improvement, all rats in groups 2 and 3 showed some significant improvement in lower urinary system function. On the other hand, the level of this improvement was far from complete functional recovery.

THE EFFECT OF OCTREOTIDE ON KAINATE-INDUCED WET DOG SHAKES AND SEIZURE ACTIVITY IN MALE AND FEMALE RATS

Systemic kainic acid (KA) administration to rats triggers wet dog shakes (WDS) followed by epileptic seizures. Although WDS are often associated with the occurrence of seizures, we have recently shown that following nitric oxide (NO) synthesis inhibition, the number of WDS decreased; subsequently the onset of seizure activity was shortened, and the number of convulsions was increased. Somatostatin (SS), whose release appears to be controlled by NO, inhibits seizure activity. There are sex differences in seizure susceptibility as well as in SS and NO activities in brain. The present study was undertaken to assess the effect of octreotide (OC), a stable SS analogue, on KA-induced WDS and seizures in rats, with emphasis on possible sex differences. WDS and seizures were induced by KA in male and female (proestrus) Sprague Dawley rats; OC or saline was injected 30 min before KA and the behavior was monitored for 120 min after KA. Octreotide increased the number of WDS and decreased the number of convulsions; this effect was more pronounced in males. Onset of KA-induced seizure ac tivity was earlier in females than males; however, there was no effect of OC on seizure latency. Seizure activity started after the termination of WDS. These results show OC has opposite effects on WDS and convulsions, in that it stimulates the former and inhibits the latter. These results support our previous findings that WDS and seizure activity involve separate mechanisms and suggest that WDS may have an inhibitory effect on limbic seizures.

Effects of neuronal and glial restricted precursor cells transplantation on erectile function after experimentally induced spinal cord injury.

INTRODUCTION Erectile dysfunction is common among patients with spinal cord injury (SCI). AIM This study aims to investigate the recovery of penile erectile functions of the rats with spinal cord injury (SCI) following transplantation of endogenous neuronal precursors cell (neuronal restricted precursors [NRP]/glial restricted precursors [GRP]) into the injured area of spinal cord. METHODS Twenty-two rats were experimented in three groups. Group 1 (N = 6): Sham; Group 2 (N = 10): SCI + NRP/GRP transplanted in day 9 after operation; Group 3 (N = 6): SCI + culture medium transplanted in day 9 after operation. Analysis of penile reflexes and cavernosal nerve stimulation studies were performed in day 28 after transplantation for each group. All rats in three groups were then sacrificed and the injured regions of spinal cords underwent histological investigation. MAIN OUTCOME MEASURES These results show improvements to some extent in locomotor and erectile functions although these improvements are far from full functional recovery. RESULTS Cavernosal nerve stimulation resulted in significantly higher intracavernosal pressure in Group 3 (SCI) although there was no difference between Group 1 (sham) and Group 2 (SCI + NRP/GRP). Number of clusters was similar between groups. Number of erections was higher in Group 3 (SCI) than Groups 1 and 2, and number of cups was higher in Group 2 (SCI + NRP/GRP) than the other two groups. Number of flips was similar in Groups 1 and 2 but lower in Group 3. Number of long flips was highest in Group 1 and lowest in Group 3. The differences between groups were significant. CONCLUSION This study emphasized the healing potential of NRP/GRP transplantation following experimental SCI. However, further experimental and clinical studies are required to advance this treatment modality.

Tideglusib protects neural stem cells against NMDA receptor overactivation

BACKGROUND N-methyl-d-aspartate (NMDA) receptors are major pharmacological targets to prevent or reduce the progression of neurodegenerative diseases. Successful therapy with NMDA receptor antagonists in humans has been limited by the severe side effects of complete receptor blockade. The aim of the present study was to investigate the possible protective effects of tideglusib against NMDA receptor overactivation in neural stem cells. METHODS We measured the alteration in membrane integrity, free radical generation, intracellular Ca(2+) accumulation, mitochondrial membrane potential (MMP)/mitochondrial morphology and glycogen synthase kinase-3 (α/β isoforms and phospho-GSK-3α/β) protein expression levels following treatments. RESULTS NMDA treatment, with or without d-serine, significantly increased LDH leakage and triggered cell death in neural stem cells. Reactive oxygen species (ROS) formation and intracellular Ca(2+) levels were increased following NMDA receptor overactivation. The significant reduction in MMP was found in NMDA/d-serine-treated cells. Tideglusib significantly decreased ROS production and membrane degradation, but did not change intracellular Ca(2+) levels following NMDA receptor activation. Both in the presence or in the absence of NMDA/d-serine, tideglusib increased MMP and the levels of phospho-GSK-3β in NSCs. Moreover, GW9662 (a peroxisome-proliferator-activated receptor gamma (PPARγ) antagonist) treatment significantly inhibited the protective effect of tideglusib in NMDA/d-serine-treated cells. CONCLUSION Our study provides the evidence that GSK-3β and PPARγ may be directly involved in pathways leading to NMDA receptor-induced cell death and that the inhibitors including tideglusib may exert neuroprotective effect against these receptor overactivation.

NADPH-d and Fos reactivity in the rat spinal cord following experimental spinal cord injury and embryonic neural stem cell transplantation

AIMS The aim of this study is to determine the role of nitric oxide (NO) in neuropathic pain and the effect of embryonic neural stem cell (ENSC) transplantation on NO content in rat spinal cord neurons following spinal cord injury (SCI). MAIN METHODS Ninety adult male Sprague-Dawley rats were divided into 3 groups (n=30, each): control (laminectomy), SCI (hemisection at T12-T13 segments) and SCI+ENSC. Each group was further divided into sub-groups (n=5 each) based on the treatment substance (L-NAME, 75 mg/kg/i.p.; L-arginine, 225 mg/kg/i.p.; physiological saline, SF) and duration (2h for acute and 28 days for chronic groups). Pain was assessed by tail flick and Randall-Selitto tests. Fos immunohistochemistry and NADPH-d histochemistry were performed in segments 2 cm rostral and caudal to SCI. KEY FINDINGS Tail-flick latency time increased in both acute and chronic L-NAME groups and increased in acute and decreased in chronic L-arginine groups. The number of Fos (+) neurons decreased in acute and chronic L-NAME and decreased in acute L-arginine groups. Following ENSC, Fos (+) neurons did not change in acute L-NAME but decreased in the chronic L-NAME groups, and decreased in both acute and chronic L-arginine groups. NADPH-d (+) neurons decreased in acute L-NAME and increased in L-arginine groups with and without ENSC transplantation. SIGNIFICANCE This study confirms the role of NO in neuropathic pain and shows an improvement following ENSC transplantation in the acute phase, observed as a decrease in Fos(+) and NADPH-d (+) neurons in spinal cord segments rostral and caudal to injury.

Medium modification with bone morphogenetic protein 2 addition for odontogenic differentiation

The aim of this study was to evaluate whether medium modification improves the odontogenic differentiation of human dental pulp stem cells (DPSC) in vitro and in vivo. DPSC isolated from human impacted third molar teeth were analysed for clusters of differentiation with flow cytometry. Odontogenic differentiation was stimulated by medium modification with the addition of bone morphogenetic protein 2 (BMP2). The expression of dentin sialophosphoprotein, dentin matrix protein 1, enamelysin/matrix metalloproteinase 20 and the phosphate-regulating gene with homologies to endopeptidases on the X chromosome of the cells were analysed with RT-PCR at 7, 14 and 21 days. Then, DPSC were transplanted on the back of immunocompromised mice via a hydroxyapatite tricalcium phosphate scaffold, and the structure of the formed tissue was investigated. The cells were identified as mesenchymal stem cells with a 98.3% CD73 and CD90 double-positive cell rate. The increase in mineralization capacity and expression of human enamel-dentin specific transcripts proportional to the culture period were determined after differentiation. Six weeks after transplantation, an osteo-dentin matrix was formed in the group in which odontogenic differentiation was stimulated, and the odontogenic characteristics of the matrix were confirmed by histological examination and RT-PCR analysis. Odontogenic differentiation of the isolated and characterized human DPSC was improved with medium modification by the addition of BMP2 in vitro and in vivo. The defined medium and applied technique have a potential use for forming reparative dentin in the future, but the effects of the method should be investigated in long-term studies.