Gülay Üzüm

Changes in the permeability of the blood-brain barrier in acute hyperammonemia. Effect of dexamethasone.

This study was designed to determine the contribution of elevated plasma ammonia levels to blood-brain barrier (BBB) abnormalities in the presence of intact liver. The permeability changes of the BBB were investigated grossly with Evans blue (EB) and quantitatively by measuring the blood-to-brain transfer content for alpha-aminoisobutyric acid (AIB) in normal rats and rats subjected to sublethal doses of ammonium acetate (NH4OAc) (750 and 600 mg/kg ip; at 30-min intervals). Some rats were pretreated with dexamethasone (DXN). Injection of NH4OAc increased both plasma and brain ammonia concentrations about 16-and 5-fold, respectively, above the control level. In rats receiving NH4OAc injection, the blood-to-brain transfer constant (Ki) for AIB was increased 3- to 11-fold. The elevated Ki values were limited to certain gray matter areas and less pronounced permeability changes were detected in white matter. Extravasation sites of EB were more restricted and were especially observed in thalamus and cerebellum, whereas cortex and white matter were unaffected. Dexamethasone pretreatment for 3 d reduced both leakage of EB and the Ki for AIB in NH4OAc injected animals, whereas acute treatment appeared ineffective. Dexamethasone did not prevent the development of coma but slightly decreased the ammonia concentration in plasma and brain. The results obtained indicate that hyperammonemia may disrupt BBB integrity not only to AIB and EB but also enhance the transport of other solutes.

Long term consequences on spatial learning-memory of low-calorie diet during adolescence in female rats; hippocampal and prefrontal cortex BDNF level, expression of NeuN and cell proliferation in dentate gyrus.

Calorie restriction (CR) is argued to positively affect general health, longevity and normally occurring age-related reduction of cognition. Obesity during adolescence may adversely affect cognition in adulthood but, to date effects of CR have not been investigated. We hypothesized that feeding with as low as 15% low-calorie diet (LCD) during adolescence would increase hippocampal and prefrontal BDNF (Brain-derived neurotrophic factor) levels, proliferative cells and neuron numbers in dentate gyrus (DG), thus positively affecting spatial memory in adulthood. Spatial learning-memory function was improved in adult female Sprague-Dawley rats fed with LCD during adolescence. PCNA (Proliferating cell nuclear antigen-cell proliferation marker) expressing cells and NeuN (Neuronal nuclear antigen-neuron marker) expressing cells in hippocampus DG which are critically involved in memory were increased. Hippocampus and prefrontal cortex BDNF levels were increased while serum glucose levels and level of lipid peroxidation indicator malondialdehyde in serum and hippocampus were reduced. Our unique results suggest that improved cognition in adult rats with LCD feeding during adolescence may result from the increase of neurogenesis and BDNF. These findings reveal the importance of nutrition in adolescence for cognitive function in adulthood. Our results may be useful for further studies aiming to treat age-related cognitive impairments.

NITRIC OXIDE INVOLVEMENT IN SEIZURES ELICITED BY PENTYLENTETRAZOL AND SEX DEPENDENCE

It has been known that susceptibility to some types of epilepsy is affected by sex. In addition, the role of NO in epileptogenesis is still unclear; NO has been suggested to be either an anticonvulsive or a proconvulsive agent. In an attempt to elucidate both the role of NO and sex differences in sensitivity to seizures, male and female Wistar rats were treated intraperitoneally (i.p.) by pentylentetrazol (PTZ)(80 mg/kg) and by a nitric oxide synthase(NOS) inhibitor N-omega-nitro-L-arginine-mthylester(L-NAME)(50mg/kg) and a NO precursor sodium-nitroprusside(SNP)(2.5mg/kg)- applied 15 min. before PTZ injection. Latency, frequency, severity, and duration of generalized clonic and clonic-tonic convulsions were recorded. Furthermore, alterations in severity, latency, frequency, and duration of convulsions were observed to correlate with NO. Both sexes, injected with PTZ, showed repetitive seizure patterns. Seizures were found to be more severe in females. L-NAME and SNP pretreatment produced paradoxical effects on PTZ-induced seizures in both sexes. L-NAME completely prevented PTZ-induced seizures in male rats, whereas increased severity, frequency, duration, and significantly shortened the latency in female rats. Unexpectedly, SNP increased convulsion severity, frequency, duration, and shortened latencies in male, whereas it decreased convulsion severity, frequency, and duration and prolonged latency in females. These results indicate that endogenous NO is involved in the regulation of convulsive action suggesting a role depending on sex.

NICOTINE IMPROVES LEARNING AND MEMORY IN RATS: MORPHOLOGICAL EVIDENCE FOR ACETYLCHOLINE INVOLVEMENT

It has been suggested that nicotine improves rapid information processing (learning and memory) tasks. However, it is not clear which aspects of cognition actually underlie these improvements because relatively less attention has been given to nicotinic cholinergic systems compared to muscarinic systems. The authors therefore studied the effects of nicotine on the learning and memory performance by a step-through passive avoidance task. Nicotine (0.4 mg/kg) was administered s.c. single dose (acute group), once a day for 3 days (subchronic group) or 21 days (chronic group). Nicotine treated and control rats were trained in one trial learning step-through passive avoidance task, where retention latencies were carried out 1 h, 24 h, and 3 days after learning trial. Treatment with nicotine before training session prolonged the latencies significantly (p < .01). Control group, acute, subacute and chronic nicotine treatment groups showed latencies 4.75 ± 0.6, 69.4 ± 14, 116.2 ± 30, and 118.5 ± 23 s, respectively. In addition, to prove the actual contribution of nicotinic cholinergic system in improvement of learning and memory processing, histological methods that permit the visualization and quantification of ACh levels were used. Electron microscopic evaluation revealed increased numbers of Ach-containing vesicles especially in hippocampus in chronic nicotine-treated rats; although frontal and temporal cortex in addition to hippocampus showed increment in Ach vesicles in a lesser extent in all nicotine treatment groups. These results indicate that long-term nicotine treatment can be important for improving cognitive function in regard to increased cholinergic activity.

Erythropoietin pretreatment suppresses seizures and prevents the increase in inflammatory mediators during pentylenetetrazole-induced generalized seizures

Erythropoietin (EPO) suppresses epileptic seizures, but the mechanism is unclear. The search for novel targets in the therapy of epilepsy has focused recently on brain inflammation since brain inflammation and the associated blood–brain barrier (BBB) damage appears to be an integral part of epilepsy pathophysiology. We examined the effects of EPO on proinflammatory mediators in brain and serum in PTZ-induced generalized seizure model. The inflammation markers (IL-1β, TNF-α, IL-6, IL-10), BBB and neuron damage markers (S100B, Neuron specific enolase; NSE, respectively) in serum and brain of Sprague–Dawley male rats were examined with the ELISA method. Nitric oxide synthase (NOS) isoforms were investigated immunohistochemically in hippocampus. EPO treatment 4 h and 24 h before PTZ administration had diverse effects. EPO treatment 4 h before PTZ administration elongated the seizure latency, decreased the inflammation and damage markers in serum and brain significantly, whereas EPO treatment 24 h before PTZ administration lowered inflammation and damage markers to control levels and decreased the seizure stage. PTZ-induced seizures increased inducible NOS (iNOS) activity and decreased endothelial NOS (eNOS) activity in hippocampus. Both EPO pretreatments reversed these effects. These findings, i.e., decreased iNOS activity and increased eNOS activity by EPO suggest the first time that the favorable effect of EPO pretreatment on inflammatory mediators triggered by PTZ-induced seizures. This can provide further insight into epilepsy treatment and new prophylactic strategies against epilepsy risk.

The effects of atorvastatin on memory deficit and seizure susceptibility in pentylentetrazole-kindled rats

Deficits in memory function have been observed in pentylentetrazole (PTZ)-kindled rats. In the present study we examined the effects of atorvastatin ((3-hydroxy-3-methylglutaryl-coenzyme A [HMG-CoA] reductase inhibitor) on PTZ kindling and related memory deficits in rats trained with the passive avoidance test. Subconvulsive PTZ doses rendered a gradual increase in seizure activity. PTZ kindling caused long-term memory to deteriorate. Atorvastatin per se and in PTZ-kindled rats improved learning and memory functions. It also prolonged latency (time to appearance of spike potentials) and diminished the amplitude and frequency of spike potentials, which indicate epileptic discharges. These novel findings suggest that the favorable effect of the atorvastatin on memory deficits provoked by PTZ kindling might be of clinical utility.

Effects of Luteolin on Liver, Kidney and Brain in Pentylentetrazol-Induced Seizures: Involvement of Metalloproteinases and NOS Activities.

OBJECTIVE Flavonoids are an important group of recognized antioxidants in plants. Luteolin (LUT) is a natural flavonoid in the plant kingdom. This study was aimed to investigate the effects of the LUT in the liver, kidney and brain of pentylentetrazol (PTZ)-induced seizure and the relationship between nitric oxide synthases (iNOS, eNOS) and matrix metalloproteinases (MMP2, MMP9). MATERIALS AND METHODS LUT (10 mg/kg) was given intraperitoneally during two weeks prior to seizure induction. A single dose PTZ 80 mg/kg i.p. was administered and seizures were observed and evaluated with regard to latency, frequency and stage for one hour. RESULTS Seizure frequen cy after PTZ administration was significantly decreased in LUT pretreated rats (p<0.05). An increase of immunhistochemical reactions of iNOS and MMP2, but a decrease of eNOS activity, were observed in rat hippocampus and peripheral tissues during the PTZ induced seizures. LUT pretreatment reversed the iNOS and MMP2 activity to the control levels and significantly increased the eNOS activity (p<0.001). CONCLUSION LUT seems to have an effective role in reducing the seizure frequency and a protective role on peripheral organ injury in animal models of seizure. The protective effect of LUT in seizures and the seizure induced peripheral tissue damage warrant further investigations.

The effects of 17β-estradiol on blood brain barrier integrity in the absence of the estrogen receptor alpha; an in-vitro model

The blood-brain barrier (BBB), which saves the brain from toxic substances, is formed by endothelial cells. It is mainly composed of tight junction (TJ) proteins existing between endothelial cells. Estrogen is an important regulatory hormone of BBB permeability. It protects the BBB before menopause, but may increase BBB permeability with aging. In addition, nitric oxide modulates BBB permeability. Alcohol impairs the integrity of the BBB with oxidants and inflammatory mediators such as iNOS. We investigated the effects of estrogen on BBB integrity in an in vitro BBB model created with ERα-free HUVEC (human umbilical vein endothelial-like cells) to mimics the menopausal period. In vitro BBB model is created with HUVEC/C6 (rat glioma cells) co-culture. The effect of 17β-estradiol on ethanol-induced BBB disruption and change/or increase of iNOS activity, which modulate BBB integrity, were evaluated. Inducibility and functionality of BBB were investigated using transendothelial electrical resistance (TEER) and the expression of proteins TJ proteins (occludin and claudin-1) and iNOS activity by immunostaining. Our results revealed that 17β-estradiol treatment before and after ethanol decrease expression of occludin and claudin-1 and value of TEER which are BBB disrupt indicators. In addition, ethanol and 17β-estradiol separately and pre- and post-ethanol 17β-estradiol treatment increased iNOS expression. Thus our study suggests caution in the use of 17β-estradiol after menopause because 17β-estradiol at this time may both increase the inflammatory process as well as damage the BBB. We think that beneficial effects of 17β-estradiol may be through ERα but it needs further studies.

Pre- and post-estrogen administration in global cerebral ischemia reduces blood-brain barrier breakdown in ovariectomized rats

The aim of present study was to determine the effect of estrogen treatment on blood-brain barrier permeability in rats with induced global cerebral ischemia. The study included six-month-old female Sprague-Dawley rats which were divided into the following groups: Control-Ischemia-Reperfusion (C + I-R); Ovariectomy-Ischemia-Reperfusion (Ovx + I-R); Ovariectomy + Estrogen + Ischemia-Reperfusion (Ovx + E + I-R); Ovariectomy + Ischemia-Reperfusion + Estrogen (Ovx + I-R + E). Ischemia-reperfusion was induced by clamping two carotid arteries, then opening the clamp. Blood-brain barrier permeability was visualized by Evans Blue extravasation and quantified by spectrophotometry. Our results indicate that following ischemia-reperfusion the BBB permeability is increased in ovariectomized rats (Evans Blue extravasation) compared to the control group in the cortex, thalamus, hippocampus, cerebellum and brain stem, while in the midbrain no significant increase was detected. In contrast, BBB permeability in the groups treated with estrogen, administered either before or after ischemia-reperfusion, was significantly lower than in ovariectomized animals. In conclusion, the increase in BBB permeability resulting from experimentally induced cerebral ischemia was prevented by exogenous estrogen treatment. The study results indicate that estrogen may be used for therapeutic purposes in ischemia-reperfusion.

EFFECTS OF SPERMIDINE TREATMENT ON NEUROBEHAVIORAL DEVELOPMENT IN INTRAUTERINE GROWTH RETARDED (IUGR) RATS

It was previously shown that polyamine treatment could induce precocious development of several somatic and neurobehavioral functions in newborn rats. This study investigates the effects of daily injections of spermidine (SPMD) 50 μl/10 g s.c. on neurobehavioral development of newborn rats experiencing undernutrition. Neurobehavioral development was assessed by measurements of gripping and righting reflexes. SPMD treated intrauterine growth retarded (IUGR) rats reached righting reflex control values at 30 days postnatal (1.87±0.78 s vs 1.75±0.66 s). Beginning from 7 days postnatal, gripping reflex values of SPMD treated IUGR rats declined, reaching that of controls at 30 days postnatal (1.77±91° vs 1.82±65°). These results suggest the utility of exogenous SPMD in rats experiencing undernutrition, thus indicating a clinical relevance.