Audiogenic Seizures in the Streptozotocin-Induced Rat Alzheimer's Disease Model.

BACKGROUND: Alzheimer's disease (AD) is a neurodegenerative and progressive disorder with no cure and constant failures in clinical trials. The main AD hallmarks are amyloid-ß (Aß) plaques, neurofibrillary tangles, and neurodegeneration. However, many other events have been implicated in AD pathogenesis. Epilepsy is a common comorbidity of AD and there is important evidence indicating a bidirectional link between these two disorders. Some studies suggest that disturbed insulin signaling might play an important role in this connection. OBJECTIVE: To understand the effects of neuronal insulin resistance in the AD-epilepsy link. METHODS: We submitted the streptozotocin (STZ) induced rat AD Model (icv-STZ AD) to an acute acoustic stimulus (AS), a known trigger of seizures. We also assessed animals' performance in the memory test, the Morris water maze and the neuronal activity (c-Fos protein) induced by a single audiogenic seizure in regions that express high levels of insulin receptors. RESULTS: We identified significant memory impairment and seizures in 71.43% of all icv-STZ/AS rats, in contrast to 22.22% of the vehicle group. After seizures, icv-STZ/AS rats presented higher number of c-Fos immunopositive cells in hippocampal, cortical, and hypothalamic regions. CONCLUSION: STZ may facilitate seizure generation and propagation by impairment of neuronal function, especially in regions that express high levels of insulin receptors. The data presented here indicate that the icv-STZ AD model might have implications not only for AD, but also for epilepsy. Finally, impaired insulin signaling might be one of the mechanisms by which AD presents a bidirectional connection to epilepsy.

A Genetic Model of Epilepsy with a Partial Alzheimer's Disease-Like Phenotype and Central Insulin Resistance.

Studies have suggested an important connection between epilepsy and Alzheimer's disease (AD), mostly due to the high number of patients diagnosed with AD who develop epileptic seizures later on. However, this link is not well understood. Previous studies from our group have identified memory impairment and metabolic abnormalities in the Wistar audiogenic rat (WAR) strain, a genetic model of epilepsy. Our goal was to investigate AD behavioral and molecular alterations, including brain insulin resistance, in naïve (seizure-free) animals of the WAR strain. We used the Morris water maze (MWM) test to evaluate spatial learning and memory performance and hippocampal tissue to verify possible molecular and immunohistochemical alterations. WARs presented worse performance in the MWM test (p < 0.0001), higher levels of hyperphosphorylated tau (S396) (p < 0.0001) and phosphorylated glycogen synthase kinase 3 (S21/9) (p < 0.05), and lower insulin receptor levels (p < 0.05). Conversely, WARs and Wistar controls present progressive increase in amyloid fibrils (p < 0.0001) and low levels of soluble amyloid-ß. Interestingly, the detected alterations were age-dependent, reaching larger differences in aged than in young adult animals. In summary, the present study provides evidence of a partial AD-like phenotype, including altered regulation of insulin signaling, in a genetic model of epilepsy. Together, these data contribute to the understanding of the connection between epilepsy and AD as comorbidities. Moreover, since both tau hyperphosphorylation and altered insulin signaling have already been reported in epilepsy and AD, these two events should be considered as important components in the interconnection between epilepsy and AD pathogenesis and, therefore, potential therapeutic targets in this field.

Putative Causal Variant on Vlgr1 for the Epileptic Phenotype in the Model Wistar Audiogenic Rat.

Wistar Audiogenic Rat is an epilepsy model whose animals are predisposed to develop seizures induced by acoustic stimulation. This model was developed by selective reproduction and presents a consistent genetic profile due to the several generations of inbreeding. In this study, we performed an analysis of WAR RNA-Seq data, aiming identified at genetic variants that may be involved in the epileptic phenotype. Seventeen thousand eighty-five predicted variants were identified as unique to the WAR model, of which 15,915 variants are SNPs and 1,170 INDELs. We filter the predicted variants by pre-established criteria and selected five for validation by Sanger sequencing. The genetic variant c.14198T>C in the Vlgr1 gene was confirmed in the WAR model. Vlgr1 encodes an adhesion receptor that is involved in the myelination process, in the development of stereocilia of the inner ear, and was already associated with the audiogenic seizures presented by the mice Frings. The transcriptional quantification of Vlgr1 revealed the downregulation this gene in the corpus quadrigeminum of WAR, and the protein modeling predicted that the mutated residue alters the structure of a domain of the VLGR1 receptor. We believe that Vlgr1 gene may be related to the predisposition of WAR to seizures and suggest the mutation Vlgr1/Q4695R as putative causal variant, and the first molecular marker of the WAR strain.

Neuroendocrine changes in the hypothalamic-neurohypophysial system in the Wistar audiogenic rat (WAR) strain submitted to audiogenic kindling.

The Wistar audiogenic rat (WAR) strain is used as an animal model of epilepsy, which when submitted to acute acoustic stimulus presents tonic-clonic seizures, mainly dependent on brainstem (mesencephalic) structures. However, when WARs are exposed to chronic acoustic stimuli (audiogenic kindling-AK), they usually present tonic-clonic seizures, followed by limbic seizures, after recruitment of forebrain structures such as the cortex, hippocampus and amygdala. Although some studies have reported that hypothalamic-hypophysis function is also altered in WAR through modulating vasopressin (AVP) and oxytocin (OXT) secretion, the role of these neuropeptides in epilepsy still is controversial. We analyzed the impact of AK and consequent activation of mesencephalic neurocircuits and the recruitment of forebrain limbic (LiR) sites on the hypothalamic-neurohypophysial system and expression of Avpr1a and Oxtr in these structures. At the end of the AK protocol, nine out of 18 WARs presented LiR. Increases in both plasma vasopressin and oxytocin levels were observed in WAR when compared to Wistar rats. These results were correlated with an increase in the expressions of heteronuclear (hn) and messenger (m) RNA for Oxt in the paraventricular nucleus (PVN) in WARs submitted to AK that presented LiR. In the paraventricular nucleus, the hnAvp and mAvp expressions increased in WARs with and without LiR, respectively. There were no significant differences in Avp and Oxt expression in supraoptic nuclei (SON). Also, there was a reduction in the Avpr1a expression in the central nucleus of the amygdala and frontal lobe in the WAR strain. In the inferior colliculus, Avpr1a expression was lower in WARs after AK, especially those without LiR. Our results indicate that both AK and LiR in WARs lead to changes in the hypothalamic-neurohypophysial system and its receptors, providing a new molecular basis to better understaind epilepsy.

Inflammatory markers in the hippocampus after audiogenic kindling.

Here, we investigated the participation of pro and anti-inflammatory cytokines in the spread of repeated audiogenic seizures from brainstem auditory structures to limbic areas, including the hippocampus. We used Wistar Audiogenic Rats (WARs) and Wistars submitted to the audiogenic kindling protocol with a loud broad-band noise. We measured pro and anti-inflammatory cytokines and nitrate levels in the hippocampus of stimulated animals. Our results show that all WARs developed audiogenic seizures that evolved to limbic seizures whereas seizure-resistant controls did not present any seizures. However, regardless of seizure severity, we did not observe differences in the pro inflammatory cytokines IL-1ß, IL-6, TNF-α and IFN-α or in the anti-inflammatory IL-10 in the hippocampi of audiogenic and resistant animals. We also did not find any differences in nitrate content. Our data indicate that the spread of seizures during the audiogenic kindling is not dependent on hippocampal release of cytokines or oxidative stress, but the severity of brainstem seizures will be higher in animals with higher levels of cytokines and the oxidative stress marker, nitrate.

Alterations in brainstem auditory processing, the acoustic startle response and sensorimotor gating of startle in Wistar audiogenic rats (WAR), an animal model of reflex epilepsies.

While acute audiogenic seizures in response to acoustic stimulus appear as an alteration in sensory-motor processing in the brainstem, the repetition of the stimulus leads to the spread of epileptic activity to limbic structures. Here, we investigated whether animals of the Wistar Audiogenic Rat (WAR) strain, genetically selected by inbreeding for seizure susceptibility, would have alterations in their auditory response, assessed by the auditory brainstem responses (ABR) and sensory-motor gating, measured as pre-pulse inhibition (PPI), which could be related to their audiogenic seizures susceptibility or severity. We did not find differences between the amplitudes and latencies of ABR waves in response to clicks for WARs when compared to Wistars. Auditory gain and symmetry between ears were also similar. However, hearing thresholds in response to some tones were lower and amplitudes of wave II were larger in WARs. WARs had smaller acoustic startle reflex amplitudes and the percentages of startle inhibited by an acoustic prepulse were higher for WARs than for Wistars. However, no correlation was found between these alterations and brainstem-dependent seizure severity or limbic seizure frequency during audiogenic kindling. Our data show that while WARs present moderate alterations in primary auditory processing, the sensory motor gating measured in startle/PPI tests appears to be more drastically altered. The observed changes might be correlated with audiogenic seizure susceptibility but not seizures severity.

Energy Metabolism and Redox State in Brains of Wistar Audiogenic Rats, a Genetic Model of Epilepsy.

The Wistar Audiogenic Rat (WAR) strain is a genetic model of epilepsy, specifically brainstem-dependent tonic-clonic seizures, triggered by acute auditory stimulation. Chronic audiogenic seizures (audiogenic kindling) mimic temporal lobe epilepsy, with significant participation of the hippocampus, amygdala, and cortex. The objective of the present study was to characterize the mitochondrial energy metabolism in hippocampus and cortex of WAR and verify its relationship with seizure severity. Hippocampus of WAR naïve (no seizures) presented higher oxygen consumption in respiratory states related to the maximum capacities of phosphorylation and electron transfer system, elevated mitochondrial density, lower GSH/GSSG and catalase activity, and higher protein carbonyl and lactate contents, compared with their Wistar counterparts. Audiogenic kindling had no adding functional effect in WAR, but in Wistar, it induced the same alterations observed in the audiogenic strain. In the cortex, WAR naïve presented elevated mitochondrial density, lower GSH/GSSG and catalase activity, and higher protein carbonyl levels. Chronic acoustic stimulation in Wistar induced the same alterations in cortex and hippocampus. Mainly in the hippocampus, WAR naïve presented elevated mRNA expression of glucose, lactate and excitatory amino acids transporters, several glycolytic enzymes, lactate dehydrogenase, and Na+/K+ ATPase in neurons and in astrocytes. In vivo treatment with mitochondrial uncoupler 2,4-dinitrophenol (DNP) or N-acetylcysteine (NAC) in WAR had no effect on mitochondrial metabolism, but lowered oxidative stress. Unlike DNP, NAC downregulated all enzyme genes involved in glucose and lactate uptake, and metabolism in neurons and astrocytes. Additionally, it was able to reduce brainstem seizure severity in WAR. In conclusion, in WAR naïve animals, both cerebral cortex and hippocampus display elevated mitochondrial density and/or activity associated with oxidative damage, glucose and lactate metabolism pathways upregulation, and increased Na+/K+ ATPase mRNA expression. Only in vivo treatment with NAC was able to reduce seizure severity of kindled WARs, possibly via down regulation of glucose/lactate metabolism. Taken together, our results are a clear contribution to the field of mitochondrial metabolism associated to epileptic seizures.

Intrinsic and synaptic properties of hippocampal CA1 pyramidal neurons of the Wistar Audiogenic Rat (WAR) strain, a genetic model of epilepsy.

Despite the many studies focusing on epilepsy, a lot of the basic mechanisms underlying seizure susceptibility are mainly unclear. Here, we studied cellular electrical excitability, as well as excitatory and inhibitory synaptic neurotransmission of CA1 pyramidal neurons from the dorsal hippocampus of a genetic model of epilepsy, the Wistar Audiogenic Rat (WARs) in which limbic seizures appear after repeated audiogenic stimulation. We examined intrinsic properties of neurons, as well as EPSCs evoked by Schaffer-collateral stimulation in slices from WARs and Wistar parental strain. We also analyzed spontaneous IPSCs and quantal miniature inhibitory events. Our data show that even in the absence of previous seizures, GABAergic neurotransmission is reduced in the dorsal hippocampus of WARs. We observed a decrease in the frequency of IPSCs and mIPSCs. Moreover, mIPSCs of WARs had faster rise times, indicating that they probably arise from more proximal synapses. Finally, intrinsic membrane properties, firing and excitatory neurotransmission mediated by both NMDA and non-NMDA receptors are similar to the parental strain. Since GABAergic inhibition towards CA1 pyramidal neurons is reduced in WARs, the inhibitory network could be ineffective to prevent the seizure-dependent spread of hyperexcitation. These functional changes could make these animals more susceptible to the limbic seizures observed during the audiogenic kindling.

Intense olfactory stimulation blocks seizures in an experimental model of epilepsy.

There are reports of patients whose epileptic seizures are prevented by means of olfactory stimulation. Similar findings were described in animal models of epilepsy, such as the electrical kindling of amygdala, where olfactory stimulation with toluene (TOL) suppressed seizures in most rats, even when the stimuli were 20% above the threshold to evoke seizures in already kindled animals. The Wistar Audiogenic Rat (WAR) strain is a model of tonic-clonic seizures induced by acute acoustic stimulation, although it also expresses limbic seizures when repeated acoustic stimulation occurs - a process known as audiogenic kindling (AK). The aim of this study was to evaluate whether or not the olfactory stimulation with TOL would interfere on the behavioral expression of brainstem (acute) and limbic (chronic) seizures in the WAR strain. For this, animals were exposed to TOL or saline (SAL) and subsequently exposed to acoustic stimulation in two conditions that generated: I) acute audiogenic seizures (only one acoustic stimulus, without previous seizure experience before of the odor test) and II) after AK (20 acoustic stimuli [2 daily] before of the protocol test). We observed a decrease in the seizure severity index of animals exposed only to TOL in both conditions, with TOL presented 20s before the acoustic stimulation in both protocols. These findings were confirmed by behavioral sequential analysis (neuroethology), which clearly indicated an exacerbation of clusters of specific behaviors such as exploration and grooming (self-cleaning), as well as significant decrease in the expression of brainstem and limbic seizures in response to TOL. Thus, these data demonstrate that TOL, a strong olfactory stimulus, has anticonvulsant properties, detected by the decrease of acute and AK seizures in WARs.

Morphofunctional alterations in the olivocochlear efferent system of the genetic audiogenic seizure-prone hamster GASH:Sal.

The genetic audiogenic seizure hamster (GASH:Sal) is a model of a form of reflex epilepsy that is manifested as generalized tonic-clonic seizures induced by external acoustic stimulation. The morphofunctional alterations in the auditory system of the GASH:Sal that may contribute to seizure susceptibility have not been thoroughly determined. In this study, we analyzed the olivocochlear efferent system of the GASH:Sal from the organ of Corti, including outer and inner hair cells, to the olivocochlear neurons, including shell, lateral, and medial olivocochlear (LOC and MOC) neurons that innervate the cochlear receptor. To achieve this, we carried out a multi-technical approach that combined auditory hearing screenings, scanning electron microscopy, morphometric analysis of labeled LOC and MOC neurons after unilateral Fluoro-Gold injections into the cochlea, and 3D reconstruction of the lateral superior olive (LSO). Our results showed that the GASH:Sal exhibited higher auditory brain response (ABR) thresholds than their controls, as well as absence of distortion-product of otoacoustic emissions (DPOAEs) in a wide range of frequencies. The ABR and DPOAE results also showed differences between the left and right ears, indicating asymmetrical hearing alterations in the GASH:Sal. These alterations in the peripheral auditory activity correlated with morphological alterations. At the cochlear level, the scanning electron microscopy analysis showed marked distortions of the stereocilia from basal to apical cochlear turns in the GASH:Sal, which were not observed in the control hamsters. At the brainstem level, MOC, LOC, and shell neurons had reduced soma areas compared with control animals. This LOC neuron shrinkage contributed to reduction in the LSO volume of the GASH:Sal as shown in the 3D reconstruction analysis. Our study demonstrated that the morphofunctional alterations of the olivocochlear efferent system are innate components of the GASH:Sal, which might contribute to their susceptibility to audiogenic seizures. This article is part of a Special Issue entitled "Genetic and Reflex Epilepsies, Audiogenic Seizures and Strains: From Experimental Models to the Clinic".

Evaluation of Cardiovascular Risk Factors in the Wistar Audiogenic Rat (WAR) Strain.

INTRODUCTION: Risk factors for life-threatening cardiovascular events were evaluated in an experimental model of epilepsy, the Wistar Audiogenic Rat (WAR) strain. METHODS: We used long-term ECG recordings in conscious, one year old, WAR and Wistar control counterparts to evaluate spontaneous arrhythmias and heart rate variability, a tool to assess autonomic cardiac control. Ventricular function was also evaluated using the pressure-volume conductance system in anesthetized rats. RESULTS: Basal RR interval (RRi) was similar between WAR and Wistar rats (188 ± 5 vs 199 ± 6 ms). RRi variability strongly suggests that WAR present an autonomic imbalance with sympathetic overactivity, which is an isolated risk factor for cardiovascular events. Anesthetized WAR showed lower arterial pressure (92 ± 3 vs 115 ± 5 mmHg) and exhibited indices of systolic dysfunction, such as higher ventricle end-diastolic pressure (9.2 ± 0.6 vs 5.6 ± 1 mmHg) and volume (137 ± 9 vs 68 ± 9 µL) as well as lower rate of increase in ventricular pressure (5266 ± 602 vs 7320 ± 538 mmHg.s-1). Indices of diastolic cardiac function, such as lower rate of decrease in ventricular pressure (-5014 ± 780 vs -7766 ± 998 mmHg.s-1) and a higher slope of the linear relationship between end-diastolic pressure and volume (0.078 ± 0.011 vs 0.036 ± 0.011 mmHg.µL), were also found in WAR as compared to Wistar control rats. Moreover, Wistar rats had 3 to 6 ventricular ectopic beats, whereas WAR showed 15 to 30 ectopic beats out of the 20,000 beats analyzed in each rat. CONCLUSIONS: The autonomic imbalance observed previously at younger age is also present in aged WAR and, additionally, a cardiac dysfunction was also observed in the rats. These findings make this experimental model of epilepsy a valuable tool to study risk factors for cardiovascular events in epilepsy.

Looking for complexity in quantitative semiology of frontal and temporal lobe seizures using neuroethology and graph theory.

Epileptic syndromes and seizures are the expression of complex brain systems. Because no analysis of complexity has been applied to epileptic seizure semiology, our goal was to apply neuroethology and graph analysis to the study of the complexity of behavioral manifestations of epileptic seizures in human frontal lobe epilepsy (FLE) and temporal lobe epilepsy (TLE). We analyzed the video recordings of 120 seizures of 18 patients with FLE and 28 seizures of 28 patients with TLE. All patients were seizure-free >1 year after surgery (Engel Class I). All patients' behavioral sequences were analyzed by means of a glossary containing all behaviors and analyzed for neuroethology (Ethomatic software). The same series were used for graph analysis (CYTOSCAPE). Behaviors, displayed as nodes, were connected by edges to other nodes according to their temporal sequence of appearance. Using neuroethology analysis, we confirmed data in the literature such as in FLE: brief/frequent seizures, complex motor behaviors, head and eye version, unilateral/bilateral tonic posturing, speech arrest, vocalization, and rapid postictal recovery and in the case of TLE: presence of epigastric aura, lateralized dystonias, impairment of consciousness/speech during ictal and postictal periods, and development of secondary generalization. Using graph analysis metrics of FLE and TLE confirmed data from flowcharts. However, because of the algorithms we used, they highlighted more powerfully the connectivity and complex associations among behaviors in a quite selective manner, depending on the origin of the seizures. The algorithms we used are commonly employed to track brain connectivity from EEG and MRI sources, which makes our study very promising for future studies of complexity in this field.

Behavioral and cardiorespiratory responses to bilateral microinjections of oxytocin into the central nucleus of amygdala of Wistar rats, an experimental model of compulsion.

INTRODUCTION: The central nucleus of amygdala plays an important role mediating fear and anxiety responses. It is known that oxytocin microinjections into the central nucleus of amygdala induce hypergrooming, an experimental model of compulsive behavior. We evaluated the behavioral and cardiorespiratory responses of conscious rats microinjected with oxytocin into the central nucleus of amygdala. METHODS: Male Wistar rats were implanted with guide cannulae into the central nucleus of amygdala and microinjected with oxytocin (0.5 µg, 1 µg) or saline. After 24 h, rats had a catheter implanted into the femoral artery for pulsatile arterial pressure measurement. The pulsatile arterial pressure was recorded at baseline conditions and data used for cardiovascular variability and baroreflex sensitivity analysis. Respiratory and behavioral parameters were assessed during this data collection session. RESULTS: Microinjections of oxytocin (0.5 µg) into the central nucleus of amygdala produced hypergrooming behavior but did not change cardiorespiratory parameters. However, hypergrooming evoked by microinjections of oxytocin (1 µg) into the central nucleus of amygdala was accompanied by increase in arterial pressure, heart rate and ventilation and augmented the power of low and high (respiratory-related) frequency bands of the systolic arterial pressure spectrum. No changes were observed in power of the low and high frequency bands of the pulse interval spectrum. Baroreflex sensitivity was found lower after oxytocin microinjections, demonstrating that the oxytocin-induced pressor response may involve an inhibition of baroreflex pathways and a consequent facilitation of sympathetic outflow to the cardiovascular system. CONCLUSIONS: The microinjection of oxytocin (1 µg) into the central nucleus of amygdala not only induces hypergrooming but also changes cardiorespiratory parameters. Moreover, specific oxytocin receptor antagonism attenuated hypergrooming but did not affect pressor, tachycardic and ventilatory responses to oxytocin, suggesting the involvement of distinct neural pathways.

Behavioral and EEG effects of GABAergic manipulation of the nigrotectal pathway in the Wistar audiogenic rat strain.

The superior colliculus (SC), substantia nigra pars reticulata (SNPr), and striatum have been characterized as important structures involved in the modulation of seizure activity. In the current study, bicuculline (GABA(A) antagonist) and muscimol (GABA(A) agonist) were microinjected into the deep layers of either the anterior SC (aSC) or posterior SC (pSC) of genetically developed Wistar audiogenic rats. Behavior and EEG activity were studied simultaneously. Only muscimol microinjected into the pSC had behavioral and EEG anticonvulsant effects in Wistar audiogenic rats, eliciting EEG oscillation changes in both SNPr and pSC, primarily during tonic seizures. The SC of Wistar audiogenic rats thus comprises two functionally different subregions, pSC and aSC, defined by distinct behavioral and EEG features. The pSC has proconvulsant audiogenic seizure activity in Wistar audiogenic rats. Our data suggest that this phenomenon may be a consequence of the genetic selection of the Wistar audiogenic rat strain.

The neurobiological substrates of behavioral manifestations during temporal lobe seizures: a neuroethological and ictal SPECT correlation study.

Ictal behavior coupled with SPECT findings during 28 seizures in patients with temporal lobe epilepsy (TLE) with unilateral hippocampal sclerosis (13 left; 15 right) was displayed as flowcharts from right-sided (RTLE) plus left-sided (LTLE) seizures. Ictal SPECT was classified blind to neuroethology. Behaviors were categorized as ipsilateral to the epileptogenic zone (IL), contralateral to the epileptogenic zone (CL), or bilateral. SPECT intensity and region were categorized as IL or CL to the epileptogenic zone. All patients developed automatisms and had hyperperfusion in their temporal lobes. Patients' verbal responses to questions had statistical interactions in RTLE but not in LTLE sum. Most CL dystonic posturing was correlated to IL basal ganglia hyperperfusion. Basal ganglia activation occurred in seizures without dystonic posturing and CL manual automatisms, and lack of IL dystonic posturing and the presence of CL cerebellar hemispheric hyperperfusion were also observed. Coupling of neuroethology and SPECT findings reliably evaluates ictal behavior and functionality of associated brain areas.

Increased expression of GluR2-flip in the hippocampus of the Wistar audiogenic rat strain after acute and kindled seizures.

The Wistar Audiogenic Rat (WAR) is an epileptic-prone strain developed by genetic selection from a Wistar progenitor based on the pattern of behavioral response to sound stimulation. Chronic acoustic stimulation protocols of WARs (audiogenic kindling) generate limbic epileptogenesis, confirmed by ictal semiology, amygdale, and hippocampal EEG, accompanied by hippocampal and amygdala cell loss, as well as neurogenesis in the dentate gyrus (DG). In an effort to identify genes involved in molecular mechanisms underlying epileptic process, we used suppression-subtractive hybridization to construct normalized cDNA library enriched for transcripts expressed in the hippocampus of WARs. The most represented gene among the 133 clones sequenced was the ionotropic glutamate receptor subunit II (GluR2), a member of the alpha-amino-3-hydroxy-5-methyl-4-isoxazoleopropionic acid (AMPA) receptor. Although semiquantitative RT-PCR analysis shows that the hippocampal levels of the GluR2 subunits do not differ between naïve WARs and their Wistar counterparts, we observed that the expression of the transcript encoding the splice-variant GluR2-flip is increased in the hippocampus of WARs submitted to both acute and kindled audiogenic seizures. Moreover, using in situ hybridization, we verified upregulation of GluR2-flip mainly in the CA1 region, among the hippocampal subfields of audiogenic kindled WARs. Our findings on differential upregulation of GluR2-flip isoform in the hippocampus of WARs displaying audiogenic seizures is original and agree with and extend previous immunohistochemical for GluR2 data obtained in the Chinese P77PMC audiogenic rat strain, reinforcing the association of limbic AMPA alterations with epileptic seizures.

Sexual differentiation of cortical spreading depression propagation after acute and kindled audiogenic seizures in the Wistar audiogenic rat (WAR).

SUMMARY: Brain excitability diseases like epilepsy constitute one factor that influences brain electrophysiological features. Cortical spreading depression (CSD) is a phenomenon that can be altered by changes in brain excitability. CSD propagation was presently characterized in adult male and female rats from a normal Wistar strain and from a genetically audiogenic seizure-prone strain, the Wistar audiogenic rat (WAR), both previously submitted (RAS(+)), or not (RAS(-)), to repetitive acoustic stimulation, to provoke audiogenic kindling in the WAR-strain. A gender-specific change in CSD-propagation was found. Compared to seizure-resistant animals, in the RAS(-) condition, male and female WARs, respectively, presented CSD-propagation impairment and facilitation, characterized, respectively, by lower and higher propagation velocities (P<0.05). In contraposition, in the RAS(+) condition, male and female WARs displayed, respectively, higher and lower CSD-propagation rates, as compared to the corresponding controls. In some Wistar and WAR females, we determined estrous cycle status on the day of the CSD-recording as being either estrous or diestrous; no cycle-phase-related differences in CSD-propagation velocities were detected. In contrast to other epilepsy models, such as Status Epilepticus induced by pilocarpine, despite the CSD-velocity reduction, in no case was CSD propagation blocked in WARs. The results suggest a gender-related, estrous cycle-phase-independent modification in the CSD-susceptibility of WAR rats, both in the RAS(+) and RAS(-) situation.

EEG wavelet analyses of the striatum-substantia nigra pars reticulata-superior colliculus circuitry: audiogenic seizures and anticonvulsant drug administration in Wistar audiogenic rats (War strain).

The importance of the substantia nigra pars reticulata (SNPr), striatum (STR) and superior colicullus (SC) in the blockade of experimental seizures is well known. But, in audiogenic seizures (brainstem tonic-clonic seizures), the anticonvulsant activity of these nuclei is still controversial. In the present study we aimed to analyze the STR-SNPr-CS circuitry in the audiogenic seizures of Wistar audiogenic rat (WAR). Behavioral and electroencephalographic (EEG) data were collected from WARs under no treatment or injection with systemic (phenobarbital) or intracerebral (intranigral) drugs (muscimol and phenobarbital). The main EEG frequency oscillation of STR, SNPr and SC seen before, during and after audiogenic seizures or during seizure protection, was determinated with wavelet spectral analyses. This method allows the association between behavior and EEG (video-EEG). Audiogenic seizures last only for half a minute in average, suggesting that the interruptions of seizures are probably not due to exhaustion. Systemic phenobarbital caused an acute and dose-dependent behavioral and EEGraphic anticonvulsant effect both in WARs. The dose of phenobarbital 15mg/kg protected animals almost completely, without side effects such as ataxia and sedation. In our data, this endogenous "natural" seizure blockade (or termination) seems to be similar to the "forced" seizure abolition, like the one caused by a systemic non-ataxic phenobarbital dose, because in both cases an intense decrease in the EEG main frequency oscillation can be seen in SNPr and SC. Intranigral phenobarbital or muscimol did not protect animals, and actually induced an increase in the main EEG frequency oscillation in SC. The main finding of the present study is that, in contrast to what is well believed about the incapacity to control audiogenic seizures by the striato-nigro-tectal circuitry, we collected here evidences that these nuclei are involved in the ability to block these seizures. However, the striato-nigro-tectal circuitry in WARs, a genetically developed strain, seems to have different functional mechanisms when compared with normal rats.