Long-term effect of postnatal hypoxia on the seizure susceptibility in rats

The effects of postnatal hypoxia at ten days of age on the pentylenetetrazol (PTZ)-induced seizure and amygdaloid kindling were investigated in male adult rats. The rats with postnatal hypoxia were significantly more susceptible to PTZ and had a significantly more easily induced amygdaloid kindling with a rapid propagation of afterdischarges to the contralateral amygdala than the control group. Light microscopic examination in one adult rat brain with postnatal hypoxia revealed no abnormal histopathological changes. The present study suggests that the consequences of postnatal hypoxia in rats remain for a long time as enhancement in seizure susceptibility.     

The piriform cortex and the cortical nucleus of the amygdala in epileptogenesis--the role of the rostrocaudal gradient

The seizure susceptibility of amygdaloid complex in rat was investigated. In piriform cortex and cortical nucleus of amygdaloid complex the structural and electrophysiological rostro-caudal differences were found (using relative spectral densities EEG, seizure thresholds, electrical kindling rate). The fundamental dependence of severity of motor seizures from structural (nuclear or cortical) organization of stimulating area was shown. There were more of limbic stages while stimulating anterior and posterior cortical nuclei, and there were more generalized stages while stimulating piriform and periamygdaloid cortex. Using the model of electrical kindling anticonvulsant effects of Sacricin were demonstrated. Sacricin is one of the compounds of polycarbonic acid. Sacricin has fully coped the process of secondary generalization of epileptic seizures.     

Involvement of Putrescine in the Development of Kindled Seizure in Rats

During the development of kindling by daily electrical stimulations applied to the left amygdala of rats, concentrations of the polyamines putrescine, spermidine, and spermine were measured in the left amygdala and the remainder of the cerebrum. A significant increase of putrescine concentration appeared first at the left amygdala in prekindled rats and then propagated to the remainder of the cerebrum with the development of kindling. This increase in putrescine concentration in the left amygdala was higher in prekindled rats than in fully kindled rats and lasted for at least 24 h after the final kindling stimulation. The concentrations of spermidine and spermine were slightly increased in a fully kindled state. To clarify the role of putrescine in kindling, the development of amygdaloid kindling was examined in rats after microinjections of alpha-difluoromethylornithine, a specific inhibitor of polyamine synthesis, and putrescine into the ipsilateral amygdala. Pretreatment with alpha-difluoromethylornithine (50 nmol) for 10 days accelerated both the development of behavioral kindling and the propagation of the afterdischarge from the left amygdala to the frontal cortex. In contrast, pretreatment with putrescine (200 nmol) for 10 days retarded the development of kindling. These results suggest that the increase in putrescine concentration in the kindled brain has an inhibitory effect on the development of kindling.     

Opioid peptides, particularly dynorphin, after amygdaloid-kindled seizures

The influence of amygdaloid kindling on brain and pituitary content of immunoreactive dynorphin (IR-DYN) and other opioid peptides was studied in rabbits. The kindling was very effective in increasing the hippocampal levels of IR-DYN, alpha-neoendorphin and Leu-enkephalin, but remained without any significant effect on the levels of IR-DYN and beta-endorphin in the majority of brain structures studied. The concentration of IR-DYN in the hippocampus remained at the control level throughout the development but was increased dramatically after completion of kindling. Biochemical alterations persisted for at least one month following the completion of kindling. The obtained results suggest that the hippocampal IR-DYN and related peptides may play some role in the maintenance of amygdaloid-kindled seizures.     

Changes of immunoreactive somatostatin and beta-endorphin content in rat brain after amygdaloid kindling

A possible contribution of brain beta-endorphin and somatostatin to the epileptogenicity established by amygdaloid kindling was investigated in rats. Fourteen male rats were chronically implanted with electrodes placed bilaterally into the amygdala. The rats received 1 sec of electrical stimulation to the left amygdala each day. Generalized seizures were observed on average 10 days after initiation of kindling and the electrical stimulation was continued up to twenty-one days. Two months after the completion of the kindling procedure, each kindled and control rat was killed by microwave irradiation and the brains were dissected on ice into thirteen subregions. Each region was homogenized and centrifuged twice in 0.1 N acetic acid. The supernatant extracts were decanted and stored at - 20 degrees C until assay. Immunoreactive beta-endorphin and somatostatin were measured by radioimmunoassays. There were no significant differences in brain beta-endorphin contents between the two groups. In kindled rats, immunoreactive somatostatin was increased significantly in amygdala, sensorimotor, piriform, and entorhinal cortex. The results suggest that changes in somatostatin may be associated with epileptic susceptibility induced by the electrical kindling procedure.     

Changes in Polyamine Concentrations in Amygdaloid-Kindled Rats

Concentrations of the polyamines putrescine, spermidine, and spermine were investigated in the left and right amygdala and in the remaining cerebrum, in which kindling was induced by repeated application of electrical stimulation of the left amygdala of rats. In kindled rats, the concentrations of spermidine and spermine increased slightly, but elevations did not reach significant levels in any brain regions. The most profound increase was detected in the putrescine concentration in all parts of the cerebrum 1-8 h after the final stimulation. These results suggest that the increases in concentrations of polyamines, particularly of putrescine, are involved in the pathogenesis of amygdaloid kindling.     

Anticonvulsive properties of peptide ACTH4-7 pro-gly-pro detected in amygdaloid kindling and audiogenic epilepsy in rats

Effects of the ACTH4-7 pro-gly-pro, calcium valproate ("Germed", DDR) and nembutal on kindling preparation and audiogenic epilepsy were investigated. Development of after-discharges in response to repeated amygdaloid electrical stimulation was assessed in normal rats and in rats susceptible to audiogenic epilepsy (KM line of rats). ACTH4-7 pro-gly-pro had an anticonvulsant profile. ACTH4-7 pro-gly-pro decreased seizure threshold in the audiogenic epilepsy test, but did not prevent the motor convulsions.     

Increased levels of messenger RNAs for neurotrophic factors in the brain during kindling epileptogenesis.

Kindling, induced by repeated subconvulsive electrical or chemical stimulations leads to progressive and permanent amplification of seizure activity, culminating in generalized seizures. We report that kindling induced by electrical stimulation in the ventral hippocampus leads to a marked and transient increase in mRNA for NGF and BDNF in the dentate gyrus, the parietal cortex, and the piriform cortex. BDNF mRNA increased also in the pyramidal layer of hippocampus and in the amygdaloid complex. No change was seen in the level of HDNF/NT-3 mRNA. The increased expression of NGF and BDNF mRNAs was not influenced by pretreatment with the NMDA receptor antagonist MK801, but was partially blocked by the quisqualate, AMPA receptor antagonist NBQX. The presumed subsequent increase of the trophic factors themselves may be important for kindling-associated plasticity in specific neuronal systems in the hippocampus, which could promote hyperexcitability and contribute to the development of epileptic syndromes.     

Amygdala kindled seizure stage is related to altered benzodiazepine binding site density

Benzodiazepine receptor binding was examined in rats at 3 stages of amygdaloid kindling (i.e., initial afterdischarge, Stage 3 and Stage 5) immediately or 24 hr after seizure. 3H-diazepam binding site density (Bmax) was significantly increased 24 hr after Stage 3 and Stage 5 kindled seizures in the hippocampus but not in the amygdala. There were no significant differences in the dissociation constants (KD) between kindled and control rats at any time point examined for either brain region. These results demonstrate that changes in benzodiazepine binding are observed with partial kindled seizures (i.e., Stage 3), indicating that generalized seizures are not prerequisite to increased benzodiazepine receptor site density.     

Naloxone and the kindling of seizures.

In view of evidence indicating that endogenous opiate-like peptides have epileptiform effects, we examined the effect of the opiate antagonist naloxone on the kindling of seizures produced by repeated electrical stimulation of the amygdala or the caudate nucleus in rats. Naloxone had no effect on the threshold for local after-discharge in the two areas and failed to retard the rate of kindling of clinical seizures. These results suggest that an interaction of opiate-like peptides with central opiate receptors does not play any critical role in the kindling of seizures.     

Identification of QTLs Involved in the Development of Amygdala Kindling in the Rat

Amygdala kindling is useful for modeling human epilepsy development. It has been known that genetic factors are involved in the development of amygdala kindling. The purpose of this study was to identify the loci that are responsible for the development of amygdala kindling. To achieve this, rat strains from a LEXF/FXLE recombinant inbred (RI) strain panel were used. The phenotypes of amygdala kindling-related parameters for seven RI strains and parental LE/Stm and F344/Stm strains were determined. They included the afterdischarge threshold (ADT), the afterdischarge duration (ADD), and the kindling rate, an incidence of development of kindling. Quantitative trait loci (QTL) analysis was performed to identify linkage relationships between these phenotypes and 1,033 SNP markers. Although no significant differences in pre-kindling ADT and ADD were observed, a significant difference in the kindling rate was found for the LEXF/FXLE RI strain. Two QTLs for the amygdala kindling rate (Agkr1 and Agkr2) were identified on rat chromosome 2. These findings clearly prove the existence of genetic influences that are involved in kindling development and suggest that substantial genetic components contribute to the progression of partial seizures into generalized seizures.     

Induction of c-fos mRNA Expression by Afterdischarge in the Hippocampus of Naive and Kindled Rats

Periodic induction of focal electrical seizure [afterdischarge (AD)] is an absolute prerequisite for the development of kindling, an animal model of complex partial epilepsy. Once established, it is a permanent condition. The mechanism(s) that translate ADs, which last tens of seconds, into life-long alterations in the CNS is unclear. Cellular immediate-early genes have been implicated in the conversion of short-term stimuli to long-term alterations in cellular phenotypes by regulating target gene expression. We have investigated the contribution of one such early gene, c-fos, to this process. The relationship between ADs and expression of c-fos gene in the rat hippocampus, a key structure in kindling development, was studied by analysis of mRNA levels. The low constitutive expression of c-fos mRNA in the hippocampus was not altered by kindling. There was an "all-or-none" relationship between induction of c-fos and the duration of AD. The threshold for induction was approximately 30 s of AD. Above-threshold ADs induced c-fos in both naive and kindled animals to the same extent and with identical temporal profiles. Although the expression of c-fos is unchanged with kindling, c-fos may nonetheless contribute to many long-term changes of kindling, both adaptive and epileptogenic.     

Accumulation of 7S SNARE complexes in hippocampal synaptosomes from chronically kindled rats

Kindling is a model of complex partial epilepsy wherein periodic application of an initially subconvulsive stimulus leads to first limbic and then generalized tonic-clonic seizures. Several laboratories have reported that augmented neurotransmitter release of l-glutamate is associated with the chronically kindled state. Neurotransmitter release requires membrane proteins called SNAREs, which form transmembrane complexes that participate in vesicle docking and are required for membrane fusion. We show here that kindling by entorhinal stimulation is associated with an accumulation of 7S SNARE complexes in the ipsilateral hippocampus. This increase of 7S SNARE complexes appears to begin early in the kindling process, achieves a peak with full kindling, and remains at this level for at least a month following cessation of further kindling stimuli. The increase is focal and permanently limited to the ipsilateral hippocampus despite progression to generalized electrographic and behavioral seizures. It is not seen in animals that receive electroconvulsive seizures, suggesting it is related to the kindling process itself. The duration and focality of increased 7S SNARE complexes with entorhinal kindling suggest that this is an altered molecular process associated with epileptogenesis.     

Organic calcium antagonists verapamil and ryodipine prevent an increase of free calcium in synaptosomes of the rat brain during corazol kindling]

In experiments on Wistar male rats it was shown that i.p. administration of verapamil or ryodipine (1,4-dihydropyridine) 15 min before each daily injection of pentylenetetrazol (PTZ) in a subconvulsive dose 30 mg/kg during 28 days significantly delayed the development of PTZ-induced kindling and attenuated kindled seizure reactions during 21-23 days as compared with control. One week after kindling an increase in free Ca2+ concentration in control rat brain synaptosomes by 60% was revealed; the treatment by verapamil and ryodipine prevented this increase. Nevertheless, Ca2+ antagonists studied at least under our experimental conditions inspite of a significant decrease in free Ca2+ concentration in synaptosomes would not completely prevent the development of kindling. Thus, an alteration in calcium homeostasis will not be the only mechanism of plasticity and long-term changes of neurons during kindling.     

Synaptic degeneration and remodelling after fast kindling of the olfactory bulb

Kindling of the olfactory bulb using a novel fast protocol (within 24 h) was studied in rats. In target brain regions, the effects of kindling were measured on the concentration of glial fibrillary acidic protein (GFAP) by dot-blot and on the concentrations of neural cell adhesion molecule (NCAM) and the 25 kDa synaptosomal associated protein of the D3 immunoprecipitate (D3(SNAP-25)) by crossed immunoelectrophoresis. Bilateral increases in the levels of GFAP, indicating activation of astrocytes, were detected in primary olfactory cortical projection areas, including the piriform cortex, and also in the basolateral amygdala and dentate gyrus, suggesting that these regions may be functionally altered during the kindling process. In the piriform cortex and dentate gyrus increased NCAM/D3(SNAP-25) ratios found ipsilaterally at seven days after kindling probably reflect an elevated rate of synaptic remodelling. At this time, however, an overall pattern of ipsilateral decreases in the synaptic marker proteins NCAM and D3(SNAP-25) indicated that this remodelling occurred on a background of synaptic degeneration. These results confirm previous studies showing that kindling is associated with synaptic remodelling and neuronal degeneration in the hippocampal formation and extends the area of plasticity to include the piriform cortex which is believed to be central to the kindling process.     

Epileptogenesis caused by the kindling effect in diving at 3 ATA of air

The kindling effect is an experimental model of epilepsy which results from an intermittent application of focal electrical stimulations to certain regions of the brain. Generalized seizures are usually obtained after about fifteen stimulations of amygdala in the rat. We studied the kindling effect in hyperbaric-hyperoxic conditions. Eleven Wistar rats were stimulated daily in a caisson and all stimulations were delivered after 30 min of diving at 3 ATA of air. Diving inhibited kindling in half animals. In the others, which all presented generalised convulsions, diving produced a reduction in seizure duration, especially during the generalisation phase.     

The development of kindled seizures is accelerated in the genetically epilepsy-prone rat

The kindling phenomenon was examined in genetically epilepsy-prone (GEPR) and non-epileptic control Sprague-Dawley rats. Kindling stimulations were administered three times a day until each rat had exhibited three Class 5 kindled motor seizures. The mean total number of kindling stimulations required for each experimental group to exhibit three motor seizures of each motor seizure class was determined. The results indicated that the early stage of kindling development was accelerated significantly in both the GEPR-3 and GEPR-9 rats, compared to non-epileptic control rats. Later stages of kindling development were accelerated in GEPR-9 but not GEPR-3 rats. Thus a differential acceleration of kindling development was exhibited by GEPR-3 and GEPR-9 rats. The results suggest the possibility that some brain region(s) involved in the early stages of kindling development may be hyperexcitable in both GEPR-3 and GEPR-9 rats. Other brain region(s) involved with the later stages of kindling development may be more excitable in GEPR-9 rats. These putative alterations may, in part, contribute to the seizure prone state of GEPR rats and the differential seizure responses of GEPR-3 and GEPR-9 rats.     

Effect of the destruction and activation of the caudate nuclei on the convulsive action of corazol kindling

Chronic experiments on the rats have shown that the pharmacological destruction of caudate nuclei significantly elevates the general brain excitation and induces rapid development of the corazol kindling. The hippocampal destruction exerts an opposite effect. Regression analysis of this processes has shown that mechanism of the general brain excitation and those of epileptogenesis are different on the stage of the developing kindling. Caudate nucleus activation induces powerful inhibition of kindling behavioral convulsive reactions and its electrographic epileptic activity. These data suggest that the caudate nucleus is a significant structure of the antiepileptic brain system and confirm G. N. Kryzhanovsky's concept about the system-antisystem interrelationship in case of neuropathologic syndromes as a result of the system hyperactivity.     

Modifications induced on the amygdaloid paroxysmal activity by entopeduncolar or nigral injection of kainic acid, in the cat

Entopeduncolar or nigral injection of kainic acid determines an early decrease of the evoked amygdaloid paroxysmal activity and a later increase of the after discharge duration. This biphasic effect is likely due to the structural analogy of the drug with monosodium glutamate, a neuroexcitant amino acid. The data suggest that the basal ganglia exert a tonic control on the amygdaloid activity.