Alterations in cytochrome c oxidase activity and energy metabolites in response to kainic acid-induced status epilepticus

The effects of kainic acid (KA)-induced limbic seizures have been investigated on cytochrome c oxidase (COx) activity, COx subunit IV mRNA abundance, ATP and phosphocreatine (PCr) levels in amygdala, hippocampus and frontal cortex of rat brain. Rats were killed either 1 h, three days or seven days after the onset of status epilepticus (SE) by CO2 and decapitation for the assay of COx activity and by head-focused microwave for the determination of ATP and PCr. Within 1 h COx activity and COx subunit IV mRNA increased in all brain areas tested between 120% and 130% of control activity, followed by a significant reduction from control, in amygdala and hippocampus on day three and seven, respectively. In amygdala, ATP and PCr levels were reduced to 44% and 49% of control 1 h after seizures. No significant recovery was seen on day three or seven. Pretreatment of rats with the spin trapping agent N-tert-butyl-alpha-phenylnitrone (PBN, 200 mg kg(-1), i.p.) 30 min before KA administration had no effect on SE, but protected COx activity and attenuated changes in energy metabolites. Pretreatment for three days with the endogenous antioxidant vitamin E (Vit-E, 100 mg/kg, i.p.) had an even greater protective effect than PBN. Both pretreatment regimens attenuated KA-induced neurodegenerative changes, as assessed by histology and prevention of the decrease of COx subunit IV mRNA and COx activity in hippocampus and amygdala, otherwise seen following KA-treatment alone. These findings suggest a close relationship between SE-induced neuronal injury and deficits in energy metabolism due to mitochondrial dysfunction.     

Involvement of nitric oxide in kainic acid-induced excitotoxicity in rat brain

The involvement of nitric oxide (NO) in kainic acid (KA)-induced excitotoxicity was studied in rat brain. With the onset of KA (15 mg kg(-1), s.c.)-induced seizures (convulsions) 30 min after injection, increases in NO, as measured by the formation of citrulline, were seen in cortex (302%), amygdala (171%) and hippocampus (203%). The highest increases were determined 90 min after onset of seizures (120 min after KA injection) with 633%, 314% and 365%, respectively. These changes in NO preceded significant decreases in ATP and phosphocreatine (PCr) ranging from 44 to 53% for ATP and from 40 to 52% for PCr in the respective brain areas. With the exception of the cortex, normal citrulline values were restored within 24 h. Pretreatment with the spin trapping agent N-tert-butyl-alpha-phenylnitrone (PBN, 200 mg kg(-1), i.p.) or the antioxidant vitamin E (Vit-E, 100 mg kg(-1) per day for 3 days) prevented the increase in citrulline and significantly attenuated the loss in ATP and PCr without affecting seizure activity. It is concluded that seizures induced by KA produced a marked increase in the free radical NO, causing oxidative stress and leading to depletion of energy stores. The prevention of the increase in NO and preservation of ATP and PCr levels by PBN and Vit-E suggests the involvement of NO and other related free radicals, such as peroxynitrite (ONOO(-)). The lack of effect of PBN and Vit-E on seizure activity, suggests that NO is not involved in mechanisms regulating KA seizure generation and propagation. PBN and Vit-E or similar compounds may be important protective agents against status epilepticus-induced neuronal degeneration.     

Depletion of energy metabolites following acetylcholinesterase inhibitor-induced status epilepticus: protection by antioxidants

Status epilepticus (SE)-induced neuronal injury may involve excitotoxicity, energy impairment and increased generation of reactive oxygen species (ROS). Potential treatment therefore should consider agents that protect mitochondrial function and ROS scavengers. In the present study, we examined whether the spin trapping agent N-tertbutyl-alpha-phenylnitrone (PBN) and the antioxidant vitamin E (DL-alpha-tocopherol) protect levels of high-energy phosphates during SE. In rats, SE was induced by either of two inhibitors of acetylcholinesterase (AChE), the organophosphate diisopropylphosphorofluoridate (DFP, 1.25 mg/kg, sc)- or the carbamate carbofuran (1.25 mg/kg, sc). Rats were sacrificed 1 h or 3 days after onset of seizures by head-focused microwave (power, 10 kW; duration 1.7 s) and levels of the energy-rich phosphates adenosine triphosphate (ATP) and phosphocreatine (PCr) and their metabolites adenosine diphosphate (ADP) and adenosine monophosphate (AMP), and creatine (Cr), respectively, were determined in the cortex, amygdala and hippocampus. Within 1 h of seizure activity, marked declines were seen in ATP (34-60%) and PCr (25-52%). Total adenine nucleotides (TAN = ATP + ADP + AMP) and total creatine compounds (TCC = PCr + Cr) were also reduced (TAN 38-60% and TCC 25-47%). No changes in ATP/AMP ratio were seen. Three days after the onset of seizures, recovery of ATP and PCr was significant in the amygdala and hippocampus, but not in the cortex. Pretreatment of rats with PBN (200 mg/kg, ip, in a single dose), 30 min before DFP or carbofuran administration, prevented induced seizures and partially prevented depletion of high-energy phosphates. Pretreatment with the natural antioxidant vitamin E (100 mg/kg, ip/day for 3 days), partially prevented loss of high energy phosphates without affecting seizures. In controls, citrulline, a product of nitric oxide synthesis, was found to be highest in the amygdala, followed by hippocampus, and lowest in the cortex. DFP- or carbofuran-induced seizures caused elevation of citrulline levels seven- to eight-fold in the cortex and three- to four-fold in the amygdala and hippocampus. These results suggest a close relationship between SE, excitotoxicity and energy metabolism. The involvement of oxidative stress is supported by the findings that DFP and carbofuran trigger an excessive nitric oxide (NO) production in the seizure relevant regions of the brain.     

Nitrone spin trapping compound N-tert-butyl-α-phenylnitrone prevents seizures induced by anticholinesterases

The neuroprotection afforded by spin trapping agents such as N-tert-butyl-α-phenylnitrone (PBN) has lent support to the hypothesis that increased production of reactive oxygen species (ROS) is a major contributing factor to excitotoxicity, aging and cognitive decline. Little is known, however, about the pharmacological properties of PBN. We have compared the acute effects of PBN on the development of seizures induced by the irreversible acetylcholinesterase (AChE) inhibitor diisopropylphosphorofluoridate (DFP), the reversible AChE inhibitor physostigmine (PHY), the muscarinic cholinergic receptor agonist pilocarpine (PIL) and the glutamatergic receptor agonist kainic acid (KA). Rats were sacrificed 90 min after the injection of seizure-inducing agents. In situ hybridization was used to detect the induction of immediate early gene (IEG) c-fos and c-jun mRNA's and the levels of AChE mRNA. The activity of AChE was visualized by AChE staining and quantified using an in vitro AChE assay. The seizures correlated with the induction of IEG mRNA's with all agents used. The pre-treatment with 150 mg/kg of PBN prevented DFP- and PHY-induced seizures and the related expression of IEG mRNA's, but had no effect on PIL- or KA-induced seizures and associated IEG mRNA's changes. PBN prevented seizures and significantly protected AChE activity against DFP inhibition when given before, but not when given after DFP. This study shows that PBN specifically protects against anticholinesterase-induced seizures by reversible protection of AChE activity and not by the blockade of muscarinic or glutamate receptors, reactivation of AChE or scavenging of ROS. The anticholinesterase properties should be considered when using PBN in studies of cholinergic dysfunction.     

Effect of free radical spin trap N-tert-butyl-alpha-phenylnitrone (PBN) on seizures induced in immature rats by homocysteic acid

The present study has examined the effect of free radical spin trap N-tert-butyl-alpha-phenylnitrone (PBN) in the model of seizures induced in immature 12-day-old rats by bilateral intracerebroventricular infusion of dl-homocysteic acid (dl-HCA, 600 nmol/side). PBN was given i.p. in two doses (100 mg/kg each), 30 min prior and 30 min after dl-HCA infusion. PBN did not significantly influence the severity of seizures, evident both from the behavioral symptoms and EEG recordings. PBN normalized decreased ATP levels in the hippocampus, occurring during the acute phase of seizures ( approximately 45-50 min after infusion) and persisting until the end of the 24-h recovery period. PBN also led to normalization of decreased glucose levels and to a significant reduction of lactate accumulation in the cerebral cortex and hippocampus. The neuroprotective effect of PBN was evaluated after 24 h and 6 days of survival following dl-HCA-induced seizures (Nissl and Fluoro-Jade B staining). The administration of PBN resulted in a partial amelioration of severe damage observed in many brain regions following infusion of dl-HCA alone. The data suggest that increased free radical production is apparently occurring during seizures induced in immature rats by homocysteic acid. Free radical scavenger PBN had a clear-cut protective effect, evident as the improved recovery of brain energy status and as a partial, but significant, attenuation of neuronal degeneration associated with this model of seizures.     

Glutamatergic and morphological alterations associated with early life seizure-induced preconditioning in young rats

Postnatal day (P)20 rats are sensitive to CA1 injury following a single injection of kainic acid (KA) but are resistant to this injury when animals have a history of two neonatal seizures. We hypothesized that the two earlier seizures led to neuroprotection by a preconditioning mechanism. Therefore, morphology, [Ca(2+)](i) and NMDA subunit proteins of the hippocampus were examined after KA was administered once (1 × KA, on P6, P9, P13 or P20), twice (2 × KA, on P6 and P9) or three times (3 × KA, on P6, P9, P13 or P20). After 1 × KA on P20, the Golgi method revealed marked decreases in spine densities and aborization of CA1 and CA3 apical dendrites. After 3 × KA, morphological alterations were attenuated in CA1 neurons and were similar to pruning observed after 1 × KA on P6 or 2 × KA. After 1 × KA at P13, baseline [Ca(2+)](i) was elevated within pyramidal and dentate granule cells. N-methyl-D-aspartate (NMDA) responses were simultaneously enhanced. After 3 × KA, Ca(2+) elevations were attenuated. Immunohistochemistry revealed selective depletion of the NR2A/B subunit modulator in the same areas. NR1 subunit expression was downregulated in the subiculum and increased in the CA3, causing a significant shift in the NR1:NR2A/B ratio throughout the hippocampus. After 1 × KA or 3 × KA at P20, reduced expression was only observed in areas of cell injury. Results indicate that different changes in morphology and excitatory responses occur depending upon when seizures begin. Partial pruning and persistent shift in the NR1:NR2A/B ratio among excitatory synapses of the hippocampus early in life may produce epileptic tolerance and protect against subsequent insults.     

Epileptogenesis in immature rats following recurrent status epilepticus

Strong evidences link status epilepticus (SE) in childhood with the later development of epilepsy. Pilocarpine-induced SE in developing rats leads to late appearance of spontaneous epileptic seizures only when SE is induced after the 18th day of life. We examined the possibility that 3 consecutive episodes of pilocarpine-induced SE on postnatal days 7, 8 and 9 could induce behavioral, electrographic and histological epileptic changes in adult life. The animals also underwent behavioral tests (inhibitory step-down avoidance, skinner box, rota-rod, open field and elevated plus-maze). EEG recordings made at the age of 30, 60 and 90 days showed the occurrence of several episodes of spikes and/or polyspikes appearing simultaneously in hippocampus and cortex. Only three isolated spontaneous seizures were observed during the whole period of observation (120 days). The long-term effects of three consecutive episodes of SE include increased spontaneous exploratory activity, learning impairment, and reduced anxiety when tested on P60. Our findings provide evidence for EEG changes and cognitive deficits in adult life following recurrent SE on postnatal days 7–9.     

Effects of chronic morphine and N(6)-cyclopentyl-adenosine administration on kainic acid-induced status epilepticus

The aim of the present study was to investigate if the upregulation of mu or A(1) receptors modifies the expression of the kainic acid (KA)-induced status epilepticus (SE). Male Wistar rats received one of the following treatments: saline solution (SS) (1 ml/kg, i.p. for 7 days); morphine (M) (20 mg/kg, i.p. for 7 days) or N(6)-cyclopentyl-adenosine (CPA) (1 mg/kg, i.p. for 9 days). Twenty-four hours after the last administration rats were sacrificed. Membranes were obtained mu and and A(1) receptor binding experiments were carried out. Furthermore, an injection of SS (1 ml/kg, i.p.) or KA (10 mg/kg, i.p.) was applied in rats pretreated chronically with M, CPA or SS, 48 h after the last administration. Seizure activity, death rate and a postictal explosive motor behavior were evaluated after KA administration. Chronic M administration increased mu receptor number in hippocampus (115%) and cortex (265%), whereas chronic CPA treatment enhanced A(1) receptor number in hippocampus (55%), amygdala (39%) and cortex (51%). The pretreatment with M facilitated the KA-induced SE and reduced the death rate as well as the postictal explosive motor behavior. The pretreatment with CPA delayed the SE presentation, increased the death rate and decreased the postictal explosive motor behavior. These findings suggest that upregulation of mu receptors enhances the KA seizures, whereas upregulation of A(1) receptors depresses these seizures.     

Comparison of neuroprotective effects induced by alpha-phenyl-N-tert-butyl nitrone (PBN) and N-tert-butyl-alpha-(2 sulfophenyl) nitrone (S-PBN) in lithium-pilocarpine status epilepticus

The status epilepticus (SE) induced in rats by lithium-pilocarpine (Li-pilo) shares many common features with soman-induced SE including extensive limbic neuropathology. Reactive oxygen species are hypothesized to play a role in the SE induced neuropathology and we propose that the free radical scavengers alpha-phenyl-N-tert-butyl nitrone (PBN) and N-tert-butyl-alpha-(2 sulfophenyl) nitrone (S-PBN) may be neuroprotective. PBN or S-PBN were administered either immediately following pilocarpine (exposure treatment) or 5 min after the onset of SE as determined by ECoG activity. SE was allowed to continue for 3 h before termination with propofol. The rats were sacrified 24 h following pilocarpine administration. S-PBN induced minor effects to reduce SE duration and improve neurological deficit 24 h following pilocarpine administration. One hundred and fifty milligrams per kilograms PBN administered 5 min after SE onset produced significant neuroprotection in the parietal, occipital, perirhinal and piriform cortices as well as the lateral amygdala. One hundred and fifty milligrams per kilograms S-PBN was neuroprotective only in the occipital and perirhinal cortex while 300 mg/kg S-PBN exacerbated cortical neuropathology. S-PBN administered 5 min after SE onset exacerbated neuropathology in thalamic regions. In contrast, PBN and S-PBN administered as exposure treatment exacerbated neuropathology in thalamic and CA3 regions. The differential neuroprotective effects of PBN and S-PBN may be the result of the poor brain penetration by S-PBN. The results suggest that free radical scavenger activity is neuroprotective in cortical regions during cholinergic convulsions. Regional variations in drug-induced neuroprotectant activity in Li-pilo SE are common and suggest multiple mechanisms of neuropathology.     

[31P]/[1H] Nuclear Magnetic Resonance Study of Mitigating Effects of GYKI 52466 on Kainate-Induced Metabolic Impairment in Perfused Rat Cerebrocortical Slices

Summary: Purpose: Kainic acid (KA) has long been used in experimental animals to induce status epilepticus (SE). A mechanistic implication of this is the association between ex-citotoxicity and brain damage during or after SE. We evaluated KA-induced metabolic impairment and the potential mitigating effects of GYKI 52466 [1-(4-aminophenyl)-4-methyl-7,8-methylenedioxy-5H-2,3-benzodiazepine] in superfused rat cerebral cortical slices. Methods: Interleaved [³¹P]/[¹H] magnetic resonance spectroscopy (MRS) was used to assess energy metabolism, intra-cellular pH (pH_i), N-acetyl-L-aspartate (NAA) level, and lactate (Lac) formation before, during, and after a 56-min exposure to 4 mM KA in freshly oxygenated artificial cerebrospinal fluid (OXY-ACSF). Results: In the absence of GYKI 52466 and during the KA exposure, NAA, PCr, and ATP levels were decreased to 91.1 ± 0.8, 62.4 ± 3.9, and 59.1 ± 4.3% of the control, respectively; Lac was increased to 118.2 ± 2.1%, and pH, was reduced from 7.27 ± 0.02 to 7.13 2 0.02. During 4-h recovery with KA-free ACSF, pH_i rapidly and Lac gradually recovered, NAA decreased further to 85.5 ± 0.3%, and PCr and AW showed little recovery. Removal of Mg²⁺ from ACSF during KA exposure caused a more profound Lac increase (to 147.1 ± 4.0%) during KA exposure and a further NAA decrease (to 80.4 ± 0.5%) during reperfusion, but did not exacerbate PCr, ATP, and pH, changes. Inclusion of 100 μM GYKI 52466 during KA exposure significantly improved energy metabolism: the PCr and ATP levels were above 76.6 ± 2.1 and 82.0 & 2.9% of the control, respectively, during KA exposure and recovered to 101.4 ± 2.4 and 95.0 ± 2.4%, respectively, during reperfusion. NAA level remained at 99.8 ± 0.6% during exposure and decreased only slightly at a later stage of reperfusion. Conclusions: Our finding supports the notion that KA-induced SE causes metabolic disturbance and neuronal injury mainly by overexcitation through non-N-methy1-D-aspartate (NMDA) receptor functions.     

Progression of brain damage after status epilepticus and its association with epileptogenesis: a quantitative MRI study in a rat model of temporal lobe epilepsy

PURPOSE: This study examined the hypothesis that neurodegeneration continues after status epilepticus (SE) ends and that the severity of damage at the early phase of the epileptogenic process predicts the outcome of epilepsy in a long-term follow-up. METHODS: SE was induced in rats by electrical stimulation of the amygdala, and the progression of structural alterations was monitored with multiparametric magnetic resonance imaging (MRI). Absolute T2, T1rho, and diffusion (Dav) images were acquired from amygdala, piriform cortex, thalamus, and hippocampus for < or = 4.5 months after SE. Frequency and type of spontaneous seizures were monitored with video-electroencephalography recordings. Histologic damage was assessed from Nissl, Timm, and Fluoro-Jade B preparations at 8 months. RESULTS: At the acute phase (2 days after SE induction), quantitative MRI revealed increased T2, T1rho, and Dav values in the primary focal area (amygdala), reflecting disturbed water homeostasis and possible early structural damage. Pathologic T2 and T1rho were observed in mono- or polysynaptically connected regions, including the piriform cortex, midline thalamus, and hippocampus. The majority of acute MRI abnormalities were reversed by 9 days after SE. In later time points (> 20 days after induction), both the T1rho and diffusion MRI revealed secondarily affected areas, most predominantly in the amygdala and hippocampus. At this time, animals began to have spontaneous seizures. The initial pathology revealed by MRI had a low predictive value for the subsequent severity of epilepsy and tissue damage. CONCLUSIONS: The results demonstrate progressive neurodegeneration after SE in the amygdala and the hippocampus and stress the need for continued administration of neuroprotectants in the treatment of SE even after electrographic seizure activity has ceased.     

Prolonged neonatal seizures exacerbate hypoxic-ischemic brain damage: correlation with cerebral energy metabolism and excitatory amino acid release

BACKGROUND: Perinatal hypoxia-ischemia (HI) is the most common precipitant of seizures in the first 24-48 h of a newborn's life. In a previous study, our laboratory developed a model of prolonged, continuous electrographic seizures in 10-day-old rat pups using kainic acid (KA) as a proconvulsant. Groups of animals included those receiving only KA, or HI for 15 or 30 min, followed by KA infusion. Our results showed that prolonged electrographic seizures following 30 min of HI resulted in a marked exacerbation of brain damage. We have undertaken studies to determine alterations in hippocampal high-energy phosphate reserves and the extracellular release of hippocampal amino acids in an attempt to ascertain the underlying mechanisms responsible for the damage promoted by the combination of HI and KA seizures. METHODS: All studies were performed on 10-day-old rats. Five groups were identified: (1) group I--KA alone, (2) group II--15 min of HI plus KA, (3) group III--15 min of HI alone, (4) group IV--30 min of HI plus KA, and (5) group VI--30 min of HI alone. HI was induced by right common carotid artery ligation and exposure to 8% oxygen/balance nitrogen. Glycolytic intermediates and high-energy phosphates were measured. Prior to treatment, at the end of HI (both 15 and 30 min), prior to KA injection, and at 1 (onset of seizures), 3, 5 (end of seizures), 7, 24 and 48 h, blood samples were taken for glucose, lactate and beta-hydroxybutyrate. At the same time points, animals were sacrificed by decapitation and brains were rapidly frozen for subsequent dissection of the hippocampus and measurement of glucose, lactate, beta-hydroxybutyrate, adenosine triphosphate (ATP) and phosphocreatine (PCr). In separate groups of rats as defined above, microdialysis probes (CMA) were stereotactically implanted into the CA2-3 region of the ipsilateral hippocampus for measurement of extracellular amino acid release. Dialysate was collected prior to any treatment, at the end of HI (15 and 30 min), prior to KA injection, and at 1 (onset of seizures), 3, 5 (end of seizures), 7 and 9 h. Determination of glutamate, serine, glutamine, glycine, taurine, alanine, and GABA was accomplished using high-performance liquid chromatography with EC detection. RESULTS: Blood and hippocampal glucose concentrations in all groups receiving KA were significantly lower than control during seizures (p < 0.05). beta-Hydroxybutyrate values displayed the inverse, in that values were significantly higher (p < 0.01) in all KA groups compared with pretreatment controls during seizure activity. Values returned to control by 2 h following the cessation of seizures. Lactate concentrations in brain and blood mimicked those of beta-hydroxybutyrate. ATP values declined to 0.36 mmol/l in both the 15 and 30 min hypoxia groups compared with 1.85 mmol/l for controls (p < 0.01). During seizures, ATP and PCr values declined significantly below their homologous controls. Following seizures, ATP values only for those animals receiving KA plus HI for 30 min remained below their homologous controls for at least 24 h. Determination of amino acid release revealed elevations of glutamate, glycine, taurine, alanine and GABA above pretreatment control during HI, with a return to normal prior to KA injections. During seizures and for the 4 h of recovery monitored, only glutamate in the combined HI and KA group rose significantly above both the 15 min of HI plus KA and the KA alone group (p < 0.05). CONCLUSION: Under circumstances in which there is a protracted depletion of high-energy phosphate reserves, as occurs with a combination of HI- and KA-induced seizures, excess amounts of glutamate become toxic to the brain. The latter may account for the exacerbation of damage to the newborn hippocampus, and serve as a target for future therapeutic intervention.     

Kainic acid seizure syndrome and binding sites in developing rats

In rats up to 16 days after birth, parenteral kainic acid (KA) produced tonico-clonic convulsions, metabolic activation limited to the hippocampus, and no brain damage. Starting with the 19th day after birth, KA produced limbic seizures associated with metabolic activation and subsequent damage in the hippocampus, the amygdala, and other limbic structures. Membranes prepared from hippocampi 10 days after birth bound [3H]KA with a high affinity component, which was localized in the mossy fiber region by slice autoradiography. In contrast, on amygdaloid membranes this component appeared only 17-19 days after birth. Our results further stress the crucial role of the amygdala in the KA seizure syndrome.     

Complex partial status epilepticus induced by a microinjection of kainic acid into unilateral amygdala in dogs and its brain damage

OBJECTIVE: In order to investigate kainic acid (KA)-induced amygdaloid seizure and seizure-induced brain damage in dogs, and to compare these findings with that in other species, a KA-induced seizure model in dogs was produced. MATERIAL AND METHODS: Normal beagle dogs were used. A Teflon cannula for KA injection was inserted into the left amygdala, and cortical or depth electrodes were positioned. One week after surgery, 1.5 microg of KA was microinjected into the left amygdala. EEGs and the behavior of the animals were monitored for 2 months after KA injection. In addition, neuron-specific enolase levels in the cerebrospinal fluid (CSF-NSE) were measured intermittently. At 2 months after the injection, histopathological studies were performed. RESULTS: KA-treated dogs showed limbic seizures that started from the left amygdala within 30 min after injection. The seizures developed into complex partial status epilepticus (CPSE), and started independently from the bilateral amygdala during the CPSE. The CPSE lasted for 1-3 days, and the animals showed no spontaneous seizures during the 2-month observation period. A significant increase in CSF-NSE was observed immediately after CPSE. Histopathologically, extensive necrosis, which formed large cavity lesions, was observed around the bilateral amygdala. SUMMARY: A microinjection of KA into unilateral amygdala in dogs induced CPSE. The seizures elicited independently from bilateral amygdala, and bilateral limbic structures suffered extensive injury. In addition, CSF-NSE was demonstrated as a useful marker of acute neuronal damage.     

Effect of 8-OH-DPAT on electrographic activity during the kainic acid-induced status epilepticus in rats.

The effect of 8-OH-DPAT, a 5-HT1A receptor agonist, on electrographic activity during the kainic acid (KA)-induced status epilepticus (SE) was evaluated in male Wistar rats. Electrographic (EEG) recordings from the ventral hippocampus and the frontal cortex along with behavioral changes were evaluated in animals that received KA administration (10mg/kg, i.p.) 20 min after saline solution (control group) or 8-OH-DPAT (1mg/kg, s.c.) injection. Rats pretreated with 8-OH-DPAT presented augmented latency for wet dog shakes (71%), generalized seizures (54%) and behavioral SE (31%). 8-OH-DPAT delayed occurrence of the first KA-induced paroxystic spikes (70%), increased latency to the EEG SE (39%) and decreased spike frequency (35-43%) recorded from the frontal cortex, and increased the time necessary for the high voltage EEG activity synchronization of the hippocampus and the frontal cortex (125%). However, EEG ictal activity recorded in hippocampus was not modified after 8-OH-DPAT pretreatment. These results indicate that 8-OH-DPAT reduces the EEG activity associated with the KA-induced SE in the frontal cortex, but not the hippocampus, and suggest an inhibitory effect in the propagation of epileptic seizures during the KA-induced SE.     

Regional expression of Bcl-2 mRNA and mitochondrial cytochrome c release after experimental brain injury in the rat

Regional levels of anti-apoptotic Bcl-2 mRNA and the cytosolic cytochrome c protein were measured after lateral fluid percussion (FP) brain injury in rats. Levels of Bcl-2 mRNA were significantly decreased in the injured left cortex (IC) and ipsilateral hippocampus (IH), but not in the contralateral right cortex (CC) and hippocampus (CH) after brain injury. Levels of Bcl-2 mRNA were significantly decreased as early as 2 h and stayed decreased as long as 48 h in the IC and IH after injury. Levels of the cytosolic cytochrome c protein were significantly increased in the IC and IH, but not in the CC and CH after brain injury. Levels of cytosolic cytochrome c were significantly increased in the IC at 30 min, 48 and 72 h, and in the IH at 2 h and as long as 72 h after injury. The increase of cytosolic cytochrome c suggests that the mitochondrial release of cytochrome is increased in the IC and IH after lateral FP brain injury. These data show that the reduction of anti-apoptotic Bcl-2 and increases of mitochondrial release of cytochrome c protein occur only in the IC and IH, regions which have been observed to undergo apoptosis and neuronal cell loss after lateral FP brain injury. Therefore, it is likely that the reduction of Bcl-2 and the increased cytochrome c protein in the cytosol contribute to the observed apoptosis and neuronal cell death in the IC and IH after lateral FP brain injury in rats.     

Long-term cosequences of intrahippocampal kainate injection in the Proechimys guyannensis rodent

The Proechimys guyannensis (PG), a spiny rodent specie living in the Amazonian region has been recently studied as an animal model of anti-convulsant mechanisms. The PG was found to be resistant to the administration of the muscarinic cholinergic agonist pilocarpine or the amygdala kindling development. This study examined the susceptibility of this animal species to the intrahippocampal kainic acid (KA) injection. Electrographic, behavioral and neuropathological changes induced by intrahippocampal KA injections were analyzed. PG showed to be extremely sensitive to the acute effects of the KA injection. Although the EEG findings in PG rodents were similar to those typically obtained in Wistar rats the pattern of electrographic activity in PG animals was longer than in Wistar rats. Neuropathological examinations of PG brains that survived KA-induced SE revealed severe cell loss in CA1/CA3 areas of the hippocampus, an extensive cell dispersion in the hilus of DG at the injected site with mossy fiber sprouting in the dentate gyrus supragranular layer. None of PG animals presented spontaneous seizures during the 120 days of observation. These findings confirm our previous observation on the resistance of this animal specie to experimental models of limbic epilepsy.