Dexmedetomidine protects neurons from kainic acid-induced excitotoxicity by activating BDNF signaling.

Glutamatergic excitotoxicity is crucial in the pathogenesis of epileptic seizures. Dexmedetomidine, a potent and highly selective α2 adrenoceptor agonist, inhibits glutamate release from nerve terminals in rat cerebrocortical nerve terminals. However, the ability of dexmedetomidine to affect glutamate-induced brain injury is still unknown. Therefore, the present study evaluated the protective effect of dexmedetomidine against brain damage by using a kainic acid (KA) rat model, a frequently used model for temporal lobe epilepsy. Rats were treated with dexmedetomidine (1 or 5 µg/kg, intraperitoneally) 30 min before the KA (15 mg/kg) intraperitoneal injection. KA-induced seizure score and elevations of glutamate release in rat hippocampi were inhibited by pretreatment with dexmedetomidine. Histopathological and TUNEL staining analyzes showed that dexmedetomidine attenuated KA-induced neuronal death in the hippocampus. Dexmedetomidine ameliorated KA-induced apoptosis, and this neuroprotective effect was accompanied by inhibited the KA-induced caspase-3 expression as well as MAPKs phosphorylation, and reversed Bcl-2 down-expression, coupled with increased Nrf2, BDNF and TrkB expression in KA-treated rats. The results suggest that dexmedetomidine protected rat brains from KA-induced excitotoxic damage by reducing glutamate levels, suppressing caspase-3 activation and MAPKs phosphorylation, and enhancing Bcl-2, Nrf2, BDNF and TrkB expression in the hippocampus. Therefore, dexmedetomidine may be beneficial for preventing or treating brain disorders associated with excitotoxic neuronal damage. In conclusion, these data suggest that dexmedetomidine has the therapeutic potential for treating epilepsy.

Houttuyniae Herba Attenuates Kainic Acid-Induced Neurotoxicity via Calcium Response Modulation in the Mouse Hippocampus.

Epilepsy is a complex neurological disorder characterized by the repeated occurrence of electrical activity known as seizures. This activity induces increased intracellular calcium, which ultimately leads to neuronal damage. Houttuyniae Herba, the aerial part of Houttuynia cordata, has various pharmacological effects and is widely used as a traditional herb. In the present study, we evaluated the protective effects of Houttuyniae Herba water extract on kainic acid-induced neurotoxicity. Kainic acid directly acts on calcium release, resulting in seizure behavior, neuronal damage, and cognitive impairment. In a rat primary hippocampal culture system, Houttuyniae Herba water extract significantly protected neuronal cells from kainic acid toxicity. In a seizure model where mice received intracerebellar kainic acid injections, Houttuyniae Herba water extract treatment resulted in a lower seizure stage score, ameliorated cognitive impairment, protected neuronal cells against kainic acid-induced toxicity, and suppressed neuronal degeneration in the hippocampus. In addition, Houttuyniae Herba water extract regulated increases in the intracellular calcium level, its related downstream pathways (reactive oxygen species production and mitochondrial dysfunction), and calcium/calmodulin complex kinase type II immunoreactivity in the mouse hippocampus, which resulted from calcium influx stimulation induced by kainic acid. These results demonstrate the neuroprotective effects of Houttuyniae Herba water extract through inhibition of calcium generation in a kainic acid-induced epileptic model.

Nitric oxide synthase inhibition reverts muscarinic receptor down-regulation induced by pilocarpine- and kainic acid-evoked seizures in rat fronto-parietal cortex.

We investigated how nitric oxide (NO) synthase inhibitor modulates muscarinic receptor expression in epileptic rats. We found that subchronic treatment (4 days) with Nω-nitro-l-arginine reduced the down-regulation of muscarinic receptors induced by pilocarpine and kainic acid in rat fronto-parietal cortex, notwithstanding the dramatic potentiation of seizures induced by both convulsants. Furthermore, functional experiments in fronto-parietal cortex slices, showed that Nω-nitro-l-arginine reduces the down-regulating effect of pilocarpine on carbachol-induced phosphoinositol hydrolysis. Finally, Nω-nitro-l-arginine greatly potentiated the induction of basic fibroblast growth factor (FGF2) by pilocarpine. These data suggest a potential role of NO in a regulatory feedback loop to control muscarinic receptor signal during seizures. The dramatic potentiation of convulsions by NO synthase inhibitors in some animal models of seizures could derive from preventing this feedback loop.

Administration of docosahexaenoic acid before birth and until aging decreases kainate-induced seizures in adult zebrafish.

Docosahexaeonic acid (DHA) is the final compound in the omega-3 polyunsaturated fatty acids (PUFA) synthetic pathway and the most abundant PUFA found in the brain. DHA plays an essential role in the development of the brain, and the intakes in pregnancy and early life affect growth and cognitive performance later in childhood. Recently, it has been proposed that dietary intake of DHA could be a non-pharmacological interventional strategy for the treatment of seizures in humans. However, to date, the experimental approaches to study the antiepileptic effect of DHA have been exclusively restricted to rodent models during short-to-medium periods of treatment. The purpose of the present study was to test the chronic anticonvulsivant effects of DHA supplementation in zebrafish from the pre-spawning stage to aging, taking advantage of our recently described kainate-induced seizure model using this animal. To that end, two groups of adult female zebrafish were fed with standard or 200mg/kg DHA-enriched diets during 1 month previous to the spawning, and offspring subdivided in two categories, and subsequently fed with standard or DHA diets, generating 4 groups of animals that were aged until 20 months. Afterward, KA was intraperitoneally administered and epileptic score determined. All the DHA-enriched groups presented antiepileptic effects compared to the control group, showing that DHA presents an anticonvulsant potential. Among the studied groups, zebrafish fed with DHA from the pre-spawning stage to aging presented the best antiepileptic profile. These results show a neuroprotective benefit in zebrafish fed with DHA-enriched diet before birth and during the whole life.

Pu-Erh tea and GABA attenuates oxidative stress in kainic acid-induced status epilepticus.

BACKGROUND: Pu-Erh tea is one of the most-consumed beverages due to its taste and the anti-anxiety-producing effect of the gamma-aminobutyric acid (GABA) if contains. However the protective effects of Pu-Erh tea and its constituent, GABA to kainic acid (KA)-induced seizure have not been fully investigated. METHODS: We analyzed the effect of Pu-Erh tea leaf (PETL) and GABA on KA-induced neuronal injury in vivo and in vitro. RESULTS: PETL and GABA reduced the maximal seizure classes, predominant behavioral seizure patterns, and lipid peroxidation in male FVB mice with status epilepticus. PETL extracts and GABA were effective in protecting KA-treated PC12 cells in a dose-dependent manner and they decreased Ca(2+) release, ROS production and lipid peroxidation from KA-stressed PC12 cells. Western blot results revealed that mitogen-activated protein kinases (MAPKs), RhoA and cyclo-oxygenase-2 (COX-2) expression were increased in PC12 cells under KA stress, and PETL and GABA significantly reduced COX-2 and p38 MAPK expression, but not that of RhoA. Furthermore, PETL and GABA reduced PGE(2) production from KA-induced PC12 cells. CONCLUSIONS: Taken together, PETL and GABA have neuroprotective effects against excitotoxins that may have clinical applications in epilepsy.

Increased sensitivity to kainic acid in a genetic model of reduced NMDA receptor function.

The pathophysiology of schizophrenia may involve reduced NMDA receptor function and experimental models of NMDA receptor hypofunction have proven useful for characterizing neurobiological abnormalities potentially relevant to schizophrenia. The present study assessed behavioral responses and induction of Fos after administration of kainic acid to wild type mice (NR1(+/+)) and mice with genetically reduced NMDA receptor expression (NR1(neo/neo)). At a dose of 20 mg/kg, kainic acid induced lethal seizures in 100% of the NR1(neo/neo) mice tested but produced no lethal seizures in the wild type mice. The NR1(neo/neo) mice also exhibited enhanced behavioral responses to kainic acid at a dose of 15 mg/kg but no lethal seizures were produced by this dose. A greater induction of Fos was observed in neocortical and limbic cortical regions of the NR1(neo/neo) compared to NR1(+/+) mice after administration of 15 mg/kg kainic acid. In contrast, there were no differences between the genotypes in kainic acid induced Fos in the amygdala, hippocampus, lateral septum, and nucleus accumbens. In order to determine if altered behavioral phenotypes of the NR1(neo/neo) mice could be related to increased sensitivity of kainate receptors to endogenous glutamate, effects of the highly selective kainate antagonist LY382884 were examined. The kainate antagonist reduced the exaggerated acoustic startle responses, deficits in prepulse inhibition of acoustic startle, and motor hyperactivity in the NR1(neo/neo) mice. These findings suggest that selective kainate receptor antagonists could be novel therapeutic candidates for schizophrenia.

NF-kappaB modulators from Valeriana officinalis.

Valeriana officinalis (Valerianaceae) has been of great interest for its therapeutic uses for treating mild nervous tension and temporary sleeping problems. In traditional European medicine it has been also reported as an antiinflammatory remedy. This study reports that the EtOAc extract of the underground parts of V. officinalis showed inhibitory activity against NF-kappaB at 100 microg/mL in the IL-6/Luc assay on HeLa cells and provided protection against excitotoxicity in primary brain cell cultures at micromolar concentrations. Bioassay-guided fractionation of the EtOAc extract led to the isolation of three known sesquiterpenes: acetylvalerenolic acid (1), valerenal (2) and valerenic acid (3), 1 and 3 were active as inhibitors of NF-kappaB at a concentration of 100 microg/mL. Acetylvalerenolic acid (1) reduced NF-kappaB activity to 4%, whereas valerenic acid (3) reduced NF-kappaB activity to 25%.

Neuroprotective effects of butterbur and rough aster against kainic Acid-induced oxidative stress in mice.

The separate and combined neuroprotective effects of rough aster (Aster scaber) and butterbur (Petasite japonicus) extracts against oxidative damage in the brain of mice challenged with kainic acid were examined by comparing behavioral changes and biochemical parameters of oxidative stress. Rough aster butanol extract (400 mg/kg) and/or butterbur butanol extract (150 or 400 mg/kg) were administered to male ICR mice, 6-8 weeks old, through a gavage for 4 days consecutively, and on day 4, kainic acid (50 mg/kg) was administered intraperitoneally. Compared with the vehicle-treated control, no significant changes in body and brain weight were observed in mice administered rough aster or butterbur butanol extract. Administration of kainic acid only, causing a lethality of approximately 54%, resulted in a significant decrease of total glutathione level and increase of thiobarbituric acid-reactive substances (TBARS) value in brain tissue. The administration of butterbur or rough aster extract (400 mg/kg) decreased the lethality (50%) of kainic acid to 25%, alleviated the behavioral signs of neurotoxicity, restored the cytosolic glutathione level of brain homogenate to approximately 80% (P < .05), and reduced kainic acid-induced increases in TBARS values. In contrast to no significant neuroprotection by butterbur extract at a low dose (150 mg/kg), the combination of rough aster extract and butterbur extract reduced the lethality to 12.5%. Moreover, the combination delayed the onset time of behavioral signs by twofold, and significantly preserved the level of cytosolic glutathione peroxidase and glutathione reductase activities. However, the other biochemical parameters were not altered significantly by the combination. Thus, the combination of two vegetable extracts significantly increased the neuroprotective action against kainic acid-induced neurotoxicity. Based on these findings, the combination of butterbur extract and rough aster extract contains a functional agent or agents that protect against oxidative stress in the brain of mice.

Acute and late effects on induction of allodynia by acromelic acid, a mushroom poison related structurally to kainic acid.

1. Ingestion of a poisonous mushroom Clitocybe acromelalga is known to cause severe tactile pain (allodynia) in the extremities for a month and acromelic acid (ACRO), a kainate analogue isolated from the mushroom, produces selective damage of interneurons of the rat lower spinal cord when injected either systemically or intrathecally. Since ACRO has two isomers, ACRO-A and ACRO-B, here we examined their acute and late effects on induction of allodynia. 2. Intrathecal administration of ACRO-A and ACRO-B provoked marked allodynia by the first stimulus 5 min after injection, which lasted over the 50-min experimental period. Dose-dependency of the acute effect of ACRO-A on induction of allodynia showed a bell-shaped pattern from 50 ag x kg(-1) to 0.5 pg x kg(-1) and the maximum effect was observed at 50 fg x kg(-1). On the other hand, ACRO-B induced allodynia in a dose-dependent manner from 50 pg x kg(-1) to 50 ng x kg(-1). 3. N-methyl-d-aspartate (NMDA) receptor antagonists and Joro spider toxin, a Ca(2+)-permeable AMPA receptor antagonist, inhibited the allodynia induced by ACRO-A, but not by ACRO-B. However, other AMPA/kainate antagonists did not affect the allodynia induced by ACRO. 4. Whereas no neuronal damage was observed in the spinal cord in ACRO-A-treated mice, induction of allodynia by ACRO-A (50 fg x kg(-1)) and ACRO-B (50 ng x kg(-1)) was selectively lost 1 week after i.t. injection of a sublethal dose of ACRO-A (50 ng x kg(-1)) or ACRO-B (250 ng x kg(-1)). Higher doses of ACRO-A, however, could evoke allodynia dose-dependently from 50 pg x kg(-1) to 500 ng x kg(-1) in the ACRO-A-treated mice. The allodynia induced by ACRO-A (500 ng x kg(-1)) was not inhibited by Joro spider toxin or NMDA receptor antagonists. These properties of the late allodynia induced by ACRO-A were quite similar to those of the acute allodynia induced by ACRO-B. 5. ACRO-A could increase [Ca(2+)](i) in the deeper laminae, rather than in the superficial laminae, of the spinal cord. This increase was not blocked by the AMPA-preferring antagonist GYKI52466 and Joro spider toxin. 6. Taken together, these results demonstrate the stereospecificity of ACRO for the induction of allodynia and suggest the presence of a receptor specific to ACRO.

Protective effect of ginsenosides, active ingredients of Panax ginseng, on kainic acid-induced neurotoxicity in rat hippocampus.

Ginsenosides are known to attenuate glutamate-induced cell injuries in vitro. We investigated the in vivo effect of ginsenosides on kainic acid (KA)-induced neurotoxicity in rat hippocampus using the methods of acid fuchsin (AF) staining and heat-shock protein-70 (HSP-70) immunoreactivity to detect neuronal death and stress, respectively. Pretreatment of ginsenosides (50 or 100 mg/kg for 7 days) via intraperitoneal (i.p.) administration significantly attenuated KA (10 mg/kg i.p.)-induced cell death by decreasing AF-positive neurons in both CA1 and CA3 regions of rat hippocampus compared with KA treatment alone. Pretreatment of ginsenosides (50 or 100 mg/kg for 7 days) via i.p. administration also significantly suppressed KA-induced induction of HSP-70 in both regions of rat hippocampus. These results show that ginsenosides are effective in protecting hippocampal CA1 and CA3 cells against KA-induced neurotoxicity.

Protection by an adenosine analogue against kainate-induced extrahippocampal neuropathology.

1. The glutamate analogue kainic acid produces neuronal damage in the central nervous system. We have reported that analogues of adenosine, such as R-N6-phenylisopropyladenosine (R-PIA) can, at doses as low as 10 microg/kg IP, prevent the hippocampal damage that follows the systemic administration of kainate. The present work was designed to examine purine protection against kainate in extrahippocampal regions by using histological methods. 2. The results show that R-PIA, at a dose of 25 microg/kg IP in rats, can protect against the neuronal damage caused by kainate in the basolateral amygdaloid nuclei, the pyriform cortex and around the rhinal fissure. This protection could be prevented by the simultaneous administration of the A1 adenosine receptor antagonist 1,3-dipropyl-8-cyclopentylxanthine, confirming that the protection involved adenosine A1 receptors. No protection was observed in the posterior amygdaloid nuclei or the entorhinal cortex, suggesting the absence of relevant adenosine receptors or a different mechanism of excitotoxicity.

Blockade of pilocarpine- or kainate-induced mossy fiber sprouting by cycloheximide does not prevent subsequent epileptogenesis in rats.

Post-injury sprouting of hippocampal mossy fibers has been suggested to be a causal mechanism underlying the development of temporal lobe epilepsy. However, this hypothesis rests entirely on indirect correlational evidence. Here we demonstrate that cycloheximide, a protein synthesis inhibitor, blocked pilocarpine- and kainate-induced mossy fiber sprouting in rats, but did not prevent the subsequent development of spontaneous seizures or affect their frequency. These results provide direct evidence against a causal role for mossy fiber sprouting in temporal lobe epileptogenesis.

Antiseizure activity of insulin: insulin inhibits pentylenetetrazole, penicillin and kainic acid-induced seizures in rats.

The present study was undertaken to evaluate the antiseizure activity spectrum of insulin against various behavioral seizure models in rats. Insulin was injected intraperitoneally (i.p.) at a test dose of 1 U/kg. Dextrose (3 g/kg) was administered simultaneously with insulin to counteract its hypoglycemic effect and induce a normoglycemic state. Insulin was found to significantly decrease the incidence, intensity and mortality rate and prolong the latency of generalized tonic-clonic convulsions induced by pentylenetetrazole (60 mg/kg i.p.) and significantly decrease the intensity and mortality rate and prolong the latency of generalized tonic-clonic convulsions induced by penicillin (2000 U/intracerebrocortical). Insulin was not only found to prolong the latency of all the seizure components but was found to reduce the incidence of focal myoclonic twitches and generalized tonic-clonic convulsions induced by kainic acid (12 mg/kg i.p.) as well. Insulin was shown to be ineffective to suppress ouabain (5 micrograms/intracerebroventricular) induced seizures. These findings indicate that insulin possesses a broad spectrum of antiseizure activity in rats. Interaction with brain Na(+)-K(+)-ATPase has been discussed as a possible mechanism of action.

Inhibition of kainic acid binding to glutamate receptors by extracts of Gastrodia.

S-(4-hydroxybenzyl)glutathione was isolated as the major principle responsible for the inhibition of the in vitro binding of kainic acid to brain glutamate receptors by water extracts of the plant Gastrodia elata. The affinity (IC50 value) of the compound is slightly lower compared to glutamate and glutathione.

Differential effects of neuroexcitatory amino acids on corticotropin-releasing hormone-41 and vasopressin release from rat hypothalamic explants.

We have investigated the direct effects of different neuroexcitatory amino acids (EAA) on the secretion of CRH-41 and arginine vasopressin (AVP) from the rat hypothalamus maintained in vitro. CRH-41 and AVP released in the medium were assayed by RIA before and after incubation with N-methyl-D-aspartate (NMDA), N-methyl-D,L-aspartic acid, kainate (KA), and quisqualate in the concentration range 1 nM to 1 mM in either the absence or the presence of 1 mM Mg2+ in the medium. In the case of NMDA, the effect of the addition of glycine (1 and 10 microM) to the incubation medium was also studied. Finally, we investigated whether different periods of exposure (up to 100 min) of hypothalamic explants to NMDA and KA would affect CRH-41 release. While no EAA was able to induce CRH-41 release under any of the above conditions, 20-min incubations with NMDA in the dose range of 1 nM to 1 mM in the absence of added Mg2+ significantly stimulated AVP release in a dose-related fashion; the maximum effect occurred at a concentration of 1 mM [ratio of stimulated collection/basal collection: NMDA, 1.51 +/- 0.10, controls, 0.86 +/- 0.05 (mean +/- SEM); P < 0.001]. KA also showed a dose-related stimulatory effect in the dose range of 1 nM to 1 mM, with maximal AVP stimulation at 10 microM (KA, 1.91 +/- 0.28; controls, 0.90 +/- 0.03; P < 0.01). The effects of both NMDA and KA on AVP were completely reversed by the competitive antagonists D,L-2-amino-5-phosphonovaleric acid and 6-cyano-7-nitroquinoxaline-2,3 dione, respectively, at doses 10 times higher than those of the agonists. N-Methyl-D,L-aspartic acid stimulated AVP secretion only at a dose of 10 mM (P < 0.01), whereas quisqualate was ineffective at any concentration. The addition of 1 mM Mg2+ to the medium blocked the effect of NMDA, while attenuating AVP stimulation induced by KA. The stimulatory effect of KA on AVP was significantly reduced by D-L-2-amino-5-phosphonovaleric acid (P < 0.05), suggesting that KA may also act through NMDA receptors. Moreover, the presence of glycine in the incubation medium did not result in any effect of NMDA on CRH-41 secretion, nor did it appear to potentiate NMDA-induced AVP release.(ABSTRACT TRUNCATED AT 400 WORDS)

Effect of nifedipine and anticonvulsants on kainic acid-induced seizures in mice.

The calcium-channel inhibitor nifedipine and several anticonvulsant drugs were evaluated for effects on seizures induced by intracerebroventricular injection of 0.14 microgram of kainic acid. These seizures were markedly exacerbated by valproic acid and moderately inhibited by diazepam. Nifedipine decreased the duration of each individual seizure episode, but did not block the development of seizures. It is concluded that nifedipine prevents the maintenance or propagation of kainate-induced seizures.