Delayed protection by MK-801 and tetrodotoxin in a rat organotypic hippocampal culture model of ischemia

BACKGROUND AND PURPOSE: The hippocampus demonstrates a regional pattern of vulnerability to ischemic injury that depends on its characteristic differentiation and intrinsic connections. We now describe a model of ischemic injury using organotypic hippocampal culture, which preserves the anatomic differentiation of the hippocampus in long-term tissue culture. METHODS: Ischemic conditions were modeled by metabolic inhibition. Cultures were briefly exposed to potassium cyanide to block oxidative phosphorylation and 2-deoxyglucose to block glycolysis. The fluorescent dye propidium iodide was used to observe membrane damage in living cultures during recovery. RESULTS: 2-Deoxyglucose/potassium cyanide incubation resulted in dose-dependent, regionally selective neuronal injury in CA1 and the dentate hilus, which began slowly after 2 to 6 hours of recovery. Subsequent histological examination of cultures after 1 to 7 days of recovery demonstrated neuronal pyknosis that was correlated with the early, direct observation of membrane damage with propidium. Both propidium staining and histological degeneration were prevented by the noncompetitive N-methyl-D-aspartate (NMDA) receptor antagonist MK-801 when administered 30 minutes after the end of the exposure to 2-deoxyglucose and potassium cyanide. Tetrodotoxin, which blocks voltage-dependent sodium channels, had protective effects that were greatest during the period of 2-deoxyglucose and potassium cyanide incubation but also produced protection against the mildest conditions of metabolic inhibition when administered after 30 minutes of recovery. CONCLUSIONS: This in vitro model reproduced elements of the time course, regional vulnerability, and pharmacologic sensitivities of in vivo ischemic hippocampal injury. Inhibition of metabolism in organotypic culture provides a rapid, easily controlled injury and reproduces the in vitro pattern of hippocampal regional vulnerability to ischemia. It is the first in vitro model of ischemia to exhibit complete protection by delayed administration of an NMDA receptor antagonist during recovery from a brief insult. The protective effects of tetrodotoxin suggest that an early period of sodium entry into cells during and after ATP depletion may be responsible for the more prolonged period of toxic NMDA receptor activation.     

Neurotoxic effects of sodium nitroprusside in rat hippocampal slices

The effect of a permanent transection on myelin gene expression in a regenerating sciatic nerve and in an adult sciatic nerve was compared to establish the degree of axonal control exerted upon Schwann cells in each population. First, the adult sciatic nerve was crushed, and the distal segment allowed to regenerate. At 12 days post-crush, the sciatic nerve was transected distal to the site of crush to disrupt the Schwann cell-axonal contacts that had reformed. Messenger RNA (mRNA) levels coding for five myelin proteins were assayed in the distal segment of the crush-transected nerve after 9 days and were compared to corresponding levels in the distal segments of sciatic nerves at 21 days post-crush and 21 days post-transection using Northern blot and slot-blot analysis. Levels of mRNAs found in the distal segment of the transected and crush-transected nerve suggested that Schwann cells in the regenerating nerve and in the mature adult nerve are equally responsive to axonal influences. The crush-transected model allowed the genes that were studied to be classified according to their response to Schwann cell-axonal contact. The levels of mRNAs were 1) down-regulated to basal levels (P0 and MBP mRNAs), 2) down-regulated to undetectable levels (myelin-associated glycoprotein mRNAs), 3) upregulated (mRNAs encoding 2'3'-cyclic nucleotide phosphodiesterase and beta-actin), or 4) not stringently controlled by the removal of Schwann cell-axonal contact (proteolipid protein mRNAs). This novel experimental model has thus provided evidence that the expression of some of the important myelin genes during peripheral nerve regeneration is dependent on continuous signals from the ingrowing axons.     

Immediate and long-term effects of neonatal MK-801 treatment on nonspatial learning

These experiments observed the immediate and long-term effects of neonatal treatment with MK-801 on patterned single alternation (PSA), a form of nonspatial, memory-based learning. Rat pups were injected daily on postnatal days (PND) 7-19, with MK-801 (MK+) or the less active isomer of MK-801 (MK-) (0.25 mg/kg), and trained at either PND 22 or 60. Rats treated with MK+ or MK- and trained on PND 22 were significantly impaired in PSA when compared with the saline control. Beyond the learning impairment, MK+ rats showed an overall decreased running speed during training. They also presented an array of abnormal behaviors and significant weight loss. These nonassociative variables were determined for several doses (0.025, 0. 05, 0.1, 0.15, and 0.20 mg/kg) through PND days 22-25. Rats that received the threshold dose for secondary effects (0.025 mg/kg) also showed an overall decrease in running speed, but failed to show a significant nonspatial learning impairment on PSA. The PSA learning impairment was found to be not long lasting; rats trained at PND 60, after neonatally receiving the original high dose of MK-801, did not differ from controls.     

MK-801 has neuroprotective and antiproliferative effects in retinal laser injury

PURPOSE: Treatment of the retina by laser photocoagulation often is complicated by an immediate side effect of visual impairment, caused by unavoidable, laser-induced destruction of healthy tissue adjacent to the lesion. A neuroprotective therapy that salvages this healthy tissue might enhance the benefit obtained from the treatment. This study was proposed to determine whether glutamate-receptor blockers can provide adjuvant neuroprotection during laser photocoagulation. The effect of MK-801, an NMDA-receptor antagonist, on laser-induced retinal injury was examined, in a rat model. METHODS: Argon laser lesions were created in the retinas of 36 DA rats, and were followed immediately by intraperitoneal injections of MK-801 (2 mg/kg) or saline. The animals were killed after 3, 20, or 60 days and the retinal lesions were evaluated histologically and morphometrically. RESULTS: Photoreceptor-cell loss was significantly less in MK-801-treated rats than in control animals. The proliferative membrane composed of retinal pigment epithelial cells and neovascular blood vessels, which was seen at the base of the lesion in control group retinas, was smaller in the MK-801-treated retinas. In rats treated with a higher dose of MK-801, the lesions showed almost no proliferative reaction. CONCLUSIONS: A potent noncompetitive NMDA-receptor blocker, MK-801 exhibits neuroprotective and antiproliferative properties in the retina. Glutamate-receptor blockers should be investigated further as potential adjuvant therapy in retinal photocoagulation treatments.     

Modeling the effects of the NMDA receptor antagonist MK-801 on timing in rats.

The NMDA receptor antagonist MK-801 produces different effects on timing tasks. In particular, MK-801 produces an underestimation of duration when animals are tested with the differential reinforcement of low rate of responding (DRL) schedule and an overestimation of duration when animals are tested with the peak-interval (PI) procedure. The goal of this study was to develop a model-based explanation for this discrepancy. Two computer simulations were conducted via an implementation of scalar expectancy theory (SET). In Simulation 1, SET was used to provide a quantitative account of PI timing data. Simulation 2 used parameter estimates from Simulation 1 to predict effects of MK-801 on the DRL task. DRL predictions provided a close match to previous empirical data. Results of the simulations suggest that differences in the literature are likely due to inherent differences between PI and DRL tasks, rather than fundamental differences in timing. Overall, the role of NMDA receptors in timing appears to be multifaceted, impacting perception, memory, and decision processes. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Conditioned stimulus familiarity determines effects of MK-801 on fear extinction.

Six experiments studied the role of conditioned stimulus (CS) familiarity in determining the effects of the N-methyl-d-aspartate (NMDA) receptor antagonist MK-801 on fear extinction. Systemic administration of MK-801 (0.1 mg/kg) impaired initial extinction but not reextinction learning. MK-801 impaired reextinction learning when the CS was relatively novel during reextinction training but not initial extinction learning when the CS was relatively familiar during initial extinction training. A context change failed to reinstate the sensitivity of initial fear extinction learning about a relatively familiar CS to MK-801. These experiments show that CS familiarity is an important determinant of effects of MK-801 on fear extinction learning: MK-801 impaired extinction learning about novel stimuli but spared extinction learning about familiar stimuli. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Impact of metyrapone on MK-801-induced alterations in the rat dopamine D1 receptors

The control of cytomegalovirus infection and disease continues to be a major problem in transplantation and different strategies have been developed to reduce its incidence. Early diagnosis of infection soon after transplantation, using molecular tools such as the polymerase chain reaction, have resulted in successful clinical trials using the strategy of pre-emptive therapy. Adoptive transfer of immune cells, which are predominantly the cytomegalovirus-specific cytotoxic T lymphocytes, into transplant recipients has been shown to restore effective immunity. A vaccine preparation has been in development aimed at preventing primary infections in allograft recipients though effective protection has yet to be shown. The mechanisms of viral pathogenesis in chronic graft rejection remain unclear; however, recent contributions from the field of cell biology have increased our understanding of possible processes which have the potential for application in the field of gene therapy for the treatment of disease.     

The effect of dizocilpine (MK-801) on conditional discrimination learning in the rat

OBJECTIVES: To report the effects of the anti-oestrogen tamoxifen on biochemical and haematological parameters. DESIGN: Randomized, double-blind comparison of tamoxifen 20 mg per day and placebo, over two years. SETTING: A university hospital. SUBJECTS: Forty-six healthy late-postmenopausal women (mean, SD time since menopause; 11, seven years). MAIN OUTCOME MEASURES: Blood specimens were drawn in the fasting state at baseline, six months and two years for measurement of haemoglobin, haematocrit, erythrocyte mean cell volume, mean erythrocyte haemoglobin, leucocyte count, platelet count, urea, electrolytes, creatinine and albumin. RESULTS: There was a significant decline in the haemoglobin concentration in the tamoxifen group (-4.4, 1.2 g/L; mean, SE) and its levels were lower in this group than in those receiving placebo (P = 0.004). Similarly, haematocrit, erythrocyte count and total leucocyte count were lower in those on placebo (P = 0.002, P = 0.01 and P = 0.01, respectively) and platelet count showed a similar trend (P = 0.08). In the tamoxifen group, the level of serum albumin fell significantly (-2.2, 0.4 g/L) and was lower throughout the study than that in the placebo group (P = 0.006). That of serum urea tended to fall (-0.4, 0.2 mmol L) but the between-groups comparison was not significant (P = 0.18). CONCLUSIONS: These data suggest that tamoxifen exerts a haemodilutory effect in normal postmenopausal women. Since a similar effect has been reported in response to postmenopausal oestrogen therapy, it is likely that the observed changes represent another oestrogenic effect of tamoxifen in postmenopausal women. Haemodilution may contribute to the reduced incidence of cardiovascular disease reported in tamoxifen-treated women, and, therefore, its assessment in the new oestrogen agonists/antagonists being developed for cardiovascular disease prevention may be important.     

Is MK-801 neuroprotection mediated by systemic hypothermia in the immature rat?

Hypothermia after hypoxia-ischaemia (HI) confounds the interpretation of the effects of neuroprotective drug intervention. The effect of 0.5 mg/kg of dizocilpine (MK-801) administered after HI on rectal temperature at 2-36 h and on brain damage 2 weeks after the insult was evaluated in the immature rat. In pups kept at an ambient temperature of 21 degrees C, MK-801 lowered the temperature by 1.1 degrees C and reduced the brain damage by 45%. In pups held at an ambient temperature of 33 degrees C, MK-801 treatment afforded a 34% reduction of brain damage without lowering the rectal temperature. In conclusion, the neuroprotection offered by MK-801 does not depend on systemic hypothermia in this model.     

The effects of adenine dinucleotides on epileptiform activity in the CA3 region of rat hippocampal slices

Alpha, omega-adenine dinucleotides (Ap(n)A) consist of two adenosine molecules linked at the 5' position by phosphate groups, the number of which is denoted by n and can range from 2 to 6. The aim of this study was to investigate the effect of Ap4A and Ap5A on the rate of epileptiform activity. Hippocampal slices (450 microm), when perfused with a medium containing no added magnesium and 4-aminopyridine (50 microM), generate epileptiform activity of an interictal nature. Ap4A and Ap5A at 1 microM depressed the discharge rate to a significant extent. At this concentration adenosine (1 microM) did not produce any effect. However at 10 microM adenosine, Ap4A and Ap5A all decreased the burst frequency. Adenosine deaminase (0.2 U/ml) totally annulled the inhibition of epileptiform activity produced by 10 microM adenosine or 1 microM Ap4A and Ap5A. Adenosine deaminase did not significantly change the maximum depression of activity produced by 10 microM Ap4A and Ap5A. 8-cyclopentyl-1,3-dimethylxanthine, an A1, receptor antagonist, increased the basal rate of epileptiform activity and prevented the depression of burst discharges by Ap4A. 5'-adenylic acid deaminase converts AMP into IMP which is inactive. 5'-adenylic acid deaminase did not prevent the inhibitory effects of Ap4A. The results suggests that in the CA3 region of the hippocampus, Ap4A and Ap5A act partly by stimulating xanthine-sensitive receptors directly and partly through the formation of the metabolite, adenosine.     

Bivalent effects of MK-801 on ethanol-induced sensitization do not parallel its effects on ethanol-induced tolerance.

The relationship between the effects of the N-methyl-D-aspartate (NMDA) antagonist MK-801 on acute responses to ethanol and its ability to block ethanol sensitization and tolerance was examined in DBA/2J mice. Cross-sensitization between these drugs was also studied. Repeated administration of 0.1 mg/kg MK-801 with ethanol potentiated, whereas 0.25 mg/kg attenuated, sensitization to ethanol's locomotor stimulant effects; rearing was similarly affected. There was evidence for cross-sensitization between ethanol and 0.25 mg/kg MK-801. MK-801 potentiated ethanol's ataxic effects in the grid test, but had no effect on tolerance to this effect. MK-801's effects on ethanol sensitization appeared to be related to its own behavioral effects, rather than NMDA receptor blockade per se. Further, these studies demonstrate dissociation between ethanol sensitization and tolerance. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Activity-induced enhancement of HB-GAM expression in rat hippocampal slices

Heparin-binding growth-associated molecule (HB-GAM) is a developmentally regulated secretory protein with neurite outgrowth-promoting activity. High-frequency stimulation leading to induction of long-term potentiation (LTP) resulted in increased expression of HB-GAM in rat hippocampal area CA1. When tetanization was given in the presence of antagonists of the N-methyl-D-aspartate (NMDA) receptor and postsynaptic voltage-gated calcium channels, the mRNA level was comparable to control levels. The results indicate that high frequency stimulation inducing LTP results in calcium-dependent enhancement in HB-GAM expression, and imply a role for this extracellular protein in the modulation of synaptic function in the hippocampus.     

Examination of persistent effects of repeated administration of pentylenetetrazol on rat hippocampal CA1: evidence from in vitro study on hippocampal slices

The early and long-lasting effects of pentylenetetrazol-kindling on hippocampal CA1 synaptic transmission were investigated. Experiments were carried out in the hippocampal slices from control and kindled rats at two post-kindling periods, i.e. 48-144 h (early phase) and 30-33 days (long-lasting phase). Field potentials, i.e. population excitatory postsynaptic potential (pEPSP) and population spike (PS) were recorded at the stratum pyramidale following stimulation of the stratum radiatum. Kindling-induced changes in synaptic transmission were assessed by stimulus-response functions and paired-pulse responses. The results showed that 48-144 h after kindling, the PS amplitude in the CA1 of kindled slices enhanced, and a second PS appeared compared to control slices. But at 30-33 days after kindling, the pEPSP slope in the CA1 of kindled slices enhanced without any change in the PS compared with those in the control slices. Evaluation of paired-pulse responses showed a significant reduction in paired-pulse inhibition for PS 48-144 h after kindling and a significant increase in paired-pulse inhibition for pEPSP 30-33 days after kindling. Our results suggest that pentylenetetrazol-kindling is accompanied by enhanced excitability and a reduction of paired-pulse inhibition in hippocampal CA1. The increased paired-pulse inhibition one month after kindling, may be interpreted as an adaptive process to cope with subsequent seizures.     

Subchronic MK-801 treatment to juvenile rats attenuates environmental effects on adult spatial learning

Forty eight primary and 20 secondary chondrosarcomas were treated surgically 1975 through 1991. An evaluation of the data of the Bone Tumor Register of the Semmelweis Medical University proved that the incidence of the malignant transformation and development of secondary chondrosarcomas is 3% and 2.6% among solitary osteochondromas. The authors summarize the clinicopathological characteristics of the malignant transformation. A retrospective evaluation of the histological grade of the malignancy proved that 67% of tumors were classified as grade I; 18% as grade II and 15% as grade III. The survival of the patients was mainly determined by the grade of the malignancy. A 95% 5-years survival was found in the grade I group and a 10% survival only in the grade II and III groups. In the cases of highly malignant chondrosarcomas radical surgical intervention i.e. amputation is recommended, considering the most often extra-compartmental location of the tumors. Low malignant and intra-compartmental highly malignant chondrosarcomas should be treated, however, by limb saving surgery. In the cases of large inresecable but low malignant chondrosarcomas debulking surgery is also acceptable.     

Thapsigargin inhibits bicuculline-induced epileptiform excitability in rat hippocampal slices

Evoked field potentials were recorded in the CA3 region of rat hippocampal slices to detect whether intracellular Ca2+ stores are involved in the epileptiform effects of the two prototypic GABA(A) antagonists, bicuculline methiodide (BMI) and gabazine (SR-95531; GBZ). Field population spikes gradually increased and became repetitive (epileptiform bursting) in the presence of either BMI (5 microM), or GBZ (5 microM). Thapsigargin (2 microM), a depletor of intracellular Ca2+ stores, reduced the epileptiform effect of BMI, but had no significant effect on the GBZ-induced hyperexcitability. These data suggest that Ca2+ release from intracellular stores participates in the epileptiform response of hippocampal CA3 neurons to BMI, but not in the response to GBZ.     

Chronic treatment with MK-801 decreases D2 dopamine receptor function in rat striatum

Present results show that a single treatment with dizocilpine (MK-801, 0.25 mg/kg IP) failed to modify the specific binding to D1 or D2 DA receptors. In contrast, repeated administrations for 3 weeks resulted in a statistically significant decrease of [3H]Spiroperidol binding to cortical or striatal membranes but did not change the number or the apparent affinity of [3H]MK-801 binding in well-washed cortical membranes. Consistent reduction in specific D2 receptor mediated behavior was obtained. The data suggest that the changes in DAergic function following repeated administrations with MK-801 could be suggestive of potential therapeutic uses of negative allosteric drugs in some DA related dysfunctions.     

Biphasic effect of hydrogen peroxide on field potentials in rat hippocampal slices

In the CA1 region of rat hippocampal slices, H2O2 (0.294-2.94 mM) caused initial augmentation, and subsequent long-lasting depression, of population spikes and excitatory postsynaptic potentials. The effect of H2O2 may not be mediated by its degradation product, hydroxyl radicals, because an iron chelator deferoxamine did not block the effect. A catalase inhibitor 3-amino-1,2,4-triazole only modestly attenuated the initial augmentation, suggesting that the effect of H2O2 is not attributable to catalase-dependent O2 generation, either. An N-methyl-D-aspartate receptor antagonist DL-2-amino-5-phosphonovaleric acid had no influence on the effect of H2O2, whereas a gamma-aminobutyric acid type A receptor channel blocker picrotoxin attenuated long-lasting depression, indicating that gamma-aminobutyric acid-mediated inhibition is altered during the depression phase. The initial augmentation but not subsequent depression was attenuated by a phospholipase A2/C inhibitor 4-bromophenacyl bromide, suggesting the involvement of lipid signaling molecule(s) in the enhancement of excitatory synaptic transmission. These results suggest that H2O2 regulates hippocampal synaptic transmission via multiple mechanisms.     

Effects of lead on the release of glutamate neurotransmitter in rat hippocampal slices]

The authors describe their case of the extremely rare multiple metastases of cutan melanoma malignum in the upper urinary tracts as it was treated with operations. In the case of their 17-year old patient first percutan resection of the right side of the renal pelvis-wall was done then one and half months later ureteronephrectomia was carried out on the right side because of metastases in the renal pelvis and the ureter. The primary tumour had been removed from the hairy area of the head 9 months earlier in a dermatological department. The authors have met only 14 similar cases in the international literature. In Hungary no review of any similar case has been found.     

Effects of no precursor and donor on neuronal activity of rat hippocampal slices

Human N-acetylmuramyl-L-alanine amidase (EC 3.5.1.28) degrades peptidoglycan, a major component of bacterial cell walls with potent pro-inflammatory cytokine-inducing properties. We postulate that degradation of peptidoglycan by N-acetylmuramyl-L-alanine amidase is important for the inactivation of inflammatory peptidoglycan products in human tissues. The inflammatory activities of peptidoglycan digested by lysozyme and/or amidase were investigated using two properties of peptidoglycan: its capacity to induce the release of the inflammatory cytokines IL-1, IL-6 and TNF-alpha in vivo and in vitro and its capacity to induce arthritis in Lewis rats. The results show that after subsequent treatment with both lysozyme and amidase, the peptidoglycan products were unable to induce arthritis in Lewis rats. The production of pro-inflammatory cytokines in mice after intravenous injection of cell wall fragments was lower after in vitro degradation of the cell wall fragments by amidase. These in vivo results were confirmed with whole blood assays in which the production of pro-inflammatory cytokines was measured after stimulation with lysozyme- and amidase-treated peptidoglycan. The results show that human N-acetylmuramyl-L-alanine amidase possesses an enzymatic activity capable of inactivating inflammatory peptidoglycan by lowering its cytokine-inducing properties.