Abstract  PURPOSE: To determine the effects of a newly synthesized epalon, ganaxolone (GNX), on primarily generalized seizures in rats of various ages during development. Epalons are classified as neuroactive steroids that interact at unique site of the GABAA receptor-Cl- channel complex in the central nervous system.

METHODS: Sprague-Dawley male rats were used at 9, 15, 30, and 60 postnatal days (PN). GNX dissolved in 2-hydroxypropylbeta-cyclodextrine was administered intraperitoneally in different doses at various time points before flurothyl testing. The incidence and threshold of clonic and tonic-clonic flurothyl seizures were evaluated. Behavioral changes were also assessed.

RESULTS: In all age groups, the effects of GNX were dose dependent and more prominent 10 min after its administration. In PN 60 and PN 30 rats, GNX had dose-dependent anticonvulsant effects; tonic-clonic seizures were more sensitive to GNX treatment than clonic seizures. In PN 15 and PN 9 rats, GNX demonstrated dose-and time-dependent anticonvulsant effects against both types of flurothyl-induced seizures. GNX was more effective in PN 15 rats than in other age groups, but at doses that altered motor behavior.

CONCLUSIONS: GNX has anticonvulsant effects against flurothylinduced seizures in all age groups tested. Its effects are more prominent in the two younger age groups, especially in PN 15 rats, but are associated with motor side effects.

Abstract  Metabolic disorders play an important role in the pathogenesis of many neurological diseases. Recent evidence suggests that leptin levels in peripheral blood and brain are lower in patients with epilepsy. Leptin is an energy-regulating hormone that plays a neuroprotective role in neurodegenerative diseases and brain trauma. However, little is known about the effects and molecular mechanisms of leptin treatment on long-term neurobehavioral impairment caused by developmental seizures. The present study evaluated whether chronic leptin treatment protected against neurobehavioral impairments induced by recurrent seizures in newborns treated with flurothyl. We also examined the effect of leptin on the expression of zinc/cPLA2-related autophagy signaling molecules and CaMKII in the cerebral cortex. Twenty Sprague-Dawley rats (6 days after birth, P6) were randomly divided into two groups, a neonatal seizure group and control group. Rats were subdivided on P13 into control, control + leptin (leptin, 2 mg/kg/day, continuous 10 days), seizure (RS), and seizure + leptin group (RS + leptin, 2 mg/kg/day for 10 consecutive days). Neurological behavioral parameters (negative geotaxis reaction reflex, righting reflex, cliff avoidance reflex, forelimb suspension reflex and open field test) were observed from P23 to P30. mRNA and protein levels in the cerebral cortex were detected using real-time RT-PCR and Western blotting, respectively. Flurothyl-induced seizures (RS group) produced long-term abnormal neurobehavior, which was improved with leptin treatment. Chronic leptin treatment restored several expression parameters affected by neonatal seizures, including seizure-induced up-regulated zinc transporter ZnT1/ZIP7, lipid membrane injury-related cPLA2, autophagy marker beclin-1/bcl2, LC3II/LC3I, and its execution molecule cathepsin-E, and down-regulated memory marker CaMK II alpha. Our results suggest that the early use of leptin after neonatal recurrent seizures may exert neuroprotective effects and antagonize the long-term neurobehavioral impairment caused by seizures. Autophagy-mediated Zn/cPLA2 and CaMK II signaling in the cerebral cortex may be involved in the neuroprotective effect of leptin. Our results provide new clues for anti-epileptogenetic treatment.

Abstract  OBJECTIVE: To investigate the long-term effects of flurothyl-induced neonatal recurrent seizures on GABA A receptor (GABA(A)R) alpha1 and gamma2 subunit expressions in adult rat brain, and discuss the relationship between these alterations of GABA(A)R subunits in mature brain and the changes of spatial memory and seizure susceptibility in adult rats.

METHODS: Thirty-two of 7-day-old (P7) Sprague-Dawley rats were divided randomly into two groups: the control group and the seizure group. Seizures were induced by inhalant flurothyl daily for six consecutive days. Rats were tested for spatial memory by using the Morris water maze task from postnatal 61 d(P61) to P65. Seizure threshold was examined by intraperitoneal injection of pentylenetetrazol, and then the brains were sampled on P75. The expressions of GABA(A)R alpha1 and gamma2 subunits mRNA and protein in cerebral cortex and hippocampus were detected by reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry method.

RESULTS: On P64, the escape latencies in water maze for the rats in the seizure group were significantly longer than those of the control rats [(82 424+/-35 622)ms vs (40 712+/-29 468)ms, P=0.001]. On P65, the frequency of crossing target within 120 s in water maze for the rats in the seizure group were significantly less than those of the control rats [(1.2+/-0.9) vs (3.1+/-1.3), P<0.001]. There was no difference in seizure threshold between the two groups on P75 [(1 487+/-662)s vs (1 841+/-648)s, P=0.137]. Compared to the control rats, the accumulated optical density (AOD) of GABA(A)R alpha1 subunit immunoreactivity in seizure rats decreased significantly in the parietal cortex, CA1-2 and CA4 region (P<0.05). The AOD of alpha1 subunit protein immunoreactivity in the frontal cortex, CA3 and dentate gyrus (DG) were of no significant difference between the two groups on P75 (P>0.05). There were also no differences in the AOD of gamma2 subunit protein immunoreactivity in the parietal lobe, DG, and CA1-3 region between the two groups (P>0.05). The AOD of gamma2 subunit protein immunoreactivity in the frontal lobe and CA4 region decreased significantly in the seizure group than those in the control group (P<0.05). The expressions of alpha1 subunits mRNA in the cerebral cortex and gamma2 subunit mRNA in the hippocampus were similar between the two groups (P>0.05), but the expressions of alpha1 subunits mRNA in the hippocampus and gamma2 subunit mRNA in the cerebral cortex in the seizure group decreased significantly as compared with those in the control group (P< 0.05).

CONCLUSION: Recurrent seizures in neonatal rats modify GABA(A) receptor alpha1 and gamma2 subunits expression in the cerebral cortex and hippocampus of adult rats, and cause long-term cognitive deficit, Which suggests that abnormal GABA(A) receptor expression might play an important role in long-term cognitive deficit induced by early life recurrent seizures.

Abstract  Volatile organic solvents, fuels and anesthetics are subject to abuse. The aim of the present study was to evaluate i.v. self-administration of several of these chemicals in drug- and experiment-naive mice using a commercially available vehicle, intralipid. Two strains of mice (DBA/2 and Swiss) were allowed to self-administer toluene (0.0017-0.17 micromol/infusion), 1,1,1-trichloroethane (0.006-0.19 micromol/infusion), ethanol (0.32-1.6 micromol/infusion), cyclohexane (0.0017-0.052 micromol/infusion), propofol (0.01-0.53 micromol/infusion) and flurothyl (0.00042-0.072 micromol/infusion) or their vehicles during 30-min tests. During the test, each nose-poke of the master mouse resulted in a 1.88-microl i.v. infusion to the master mouse and a yoked control mouse. When the delivery line was loaded with a reinforcing drug solution, the number of nose-pokes of the master mice significantly exceeded that for yoked control mice. In the present experiments, significant differences in rates of nose-poking were observed between mice receiving response-contingent and response-noncontingent deliveries of ethanol and toluene in both strains of mice and of 1,1,1-trichloroethane in Swiss mice. These data suggest that the reinforcing effects of abused inhalants can be studied using i.v. self-administration procedures.

Abstract  1. Effects of two volatile anesthetics [halothane (Hal) and enflurane (Enf)] and a volatile convulsant [hexafluorodiethyl ether (HFE)] on amino acid-induced membrane currents in neurons dissociated from the nucleus tractus solitarius of the rat were examined. The dissociated neurons were voltage clamped in the whole-cell mode of the patch-clamp technique. All drugs were applied with a microperfusion system, termed the "Y-tube" method. 2. The glutamate (Glu)-induced excitatory response was slightly reduced by both the anesthetics. The responses to three agonists at Glu receptor were depressed by Hal (10(-3) M) in the rank order of quisqualate greater than N-methyl-D-aspartate greater than kainate. HFE slightly increased the Glu response at a high concentration of 2 x 10(-3) M. 3. The gamma-aminobutyric acid (GABA)-induced chloride current (ICl) was enhanced by both anesthetics. The dissociation constant (Kd) for the enhancement was 2.3 x 10(-4) M for Hal and 2.1 x 10(-4) M for Enf, and the Hill coefficient was 1.6 for Hal and 1.5 for Enf. HFE depressed the GABA response with a Kd of 8.7 x 10(-5) M and a Hill coefficient of 0.84. 4. Hal (10(-3) M) and Enf (10(-3) M) decreased the Kd of the GABA concentration-response curve from 3.5 x 10(-6) to 10(-6) and 1.9 x 10(-6) M, respectively, without changing the maximum response or the Hill coefficient (1.5). In the presence of HFE (10(-4) M), the Kd was increased to 1.4 x 10(-5) M and the Hill coefficient was slightly changed to 1.2.(ABSTRACT TRUNCATED AT 250 WORDS)

Abstract  The inducible 72-kDa heat shock protein (HSP72) is a highly conserved stress protein that is expressed in CNS cells and may play a role in protection from neural injury. We used a monoclonal antibody to HSP72 and immunocytochemistry to localize HSP72 in the rat brain 24 h following either 30 or 60 min of flurothyl-induced status epilepticus. Sprague-Dawley rats were anesthetized with halothane, paralyzed, and ventilated, and remained normotensive and well oxygenated for the duration of the seizures. Seizure activity was quantified via analysis of the scalp EEG pattern. HSP72-like immunoreactivity (HSP72-LI) was induced in specific brain regions in a graded fashion that correlated, in part, with the duration and degree of seizure activity. Milder seizures produced HSP72-LI limited to layers 2 and 3 of frontoparietal cortex, dentate hilus cells, and CA3 pyramidal neurons. More extensive seizures led to HSP72-LI in layers 2, 3 and 5 of frontoparietal and visual cortex, dentate hilus cells, CA1 and CA3 pyramidal neurons, and certain thalamic and amygdaloid nuclei. These are similar to many, but not all, of the brain regions known to be injured with this model. No HSP72-LI was observed in sham-treated controls or flurothyl-treated animals whose seizures were controlled with pentobarbital. HSP72-LI thus localizes to certain regions of seizure-induced injury, and may provide a sensitive method of detecting neuronal 'stress' or injury relatively soon after status epilepticus. Whether or not HSP72 synthesis plays a protective role in the pathogenesis of seizures, or is only a marker for cell injury, remains to be determined.

Abstract  Earlier work in our and other laboratories suggest that alteration of brain lipids, primarily sterols, could be a precondition for the development of epileptiform activity. The present study further tests this hypothesis by attempting to produce chronic epileptiform activity in the rat by a drug which impairs biosynthesis of cerebral cholesterol. Starting one day after birth, weekly injection of the rat with U18666A, 3-beta(2-diethylaminoethoxy)androst-5-en-17-one hydrochloride (10 mg/kg, s.c.), produced a reduced seizure threshold to flurothyl ether and a recurrent, spontaneous seizure state by the sixth and tenth weeks of life, respectively. These conditions were not seen if treatment was delayed until rats were about 4 weeks old. The seizure pattern, as seen by continuous ECoG and EMG recordings, consisted of a 3--5 sec burst of high voltage spiking and corresponding increases in muscle activity. However, major motor seizures were not produced. The total episode lasted 10--15 sec. Seizure frequency ranged from 5 to 21 per day. U18666A decreases cholesterolsynthesis, presumably by inhibiting enzymatic reduction of desmosterol to cholesterol. After the first two weeks of treatment, cerebral cortical cholesterol levels decreased to about 50% of control cortical levels. However, the concentration of cerebral total sterols did not change because desmosterol levels reciprocally increased. In spite of continued drug dosage, cholesterol and desmosterol levels of treated rats approached those of controls by 8 weeks of age. These observations, plus finding that a seizure-prone state did not develop in rats when the onset of drug treatment was postponed until about 4 weeks of age, suggest that alterations of brain sterols early in development of the mammalian brain can result in development of a chronic, epileptiform condition later in life.

Abstract  Flurothyl kindling initiates a time-dependent process that results in a facilitated propagation from the forebrain to the brainstem seizure system and in an increase in the complexity of behavioral seizure expression. We investigated the involvement of the ventromedial nucleus of the hypothalamus (VMH) in mediating this facilitated propagation between these seizure systems. Bilateral ibotenic acid lesions of the VMH, but not the dorsomedial nucleus of the hypothalamus (DMH), resulted in a disruption in the propagation of seizure activity from the forebrain to the brainstem. Moreover, VMH lesioned mice were able to express brainstem seizures following minimal corneal electroconvulsive shock (mECS). Together, our results indicate that the VMH is a critical substrate involved in propagating seizure activity between the forebrain and brainstem, but is not involved in the expression systems necessary for forebrain or brainstem seizure manifestations.

Abstract  Status epilepticus of greater than 30-min duration in rats gives rise to a conspicuous lesion in the substantia nigra pars reticulata (SNPR) and globus pallidus (GP). The objective of the present study was to explore whether the lesion, which encompasses necrosis of both neurons and glial cells, is related to intra- and extracellular acidosis. Using the flurothyl model previously described to produce seizures, we assessed regional pH values with the autoradiographic 5,5-dimethyl[2-14C]oxazolidine-2,4-dione technique. Regional pH values were assessed in animals with continuous seizures for 20 and 60 min, as well as in those allowed to recover for 30 and 120 min after seizure periods of 20 or 60 min. In additional animals, changes in extracellular fluid pH (pHe) were measured with ion-selective microelectrodes, and extracellular fluid (ECF) volume was calculated from the diffusion profile for electrophoretically administered tetramethylammonium. In structures such as the neocortex and the hippocampus, which show intense metabolic activation during seizures, status epilepticus of 20- and 60-min duration was accompanied by a reduction of the "composite" tissue pH (pHt) of 0.2-0.3 unit. Recovery of pHt was observed upon termination of seizures. In SNPR and in GP, the acidosis was marked to excessive after 20 and 60 min of seizures (delta pHt approximately 0.6 after 60 min).(ABSTRACT TRUNCATED AT 250 WORDS)

Abstract  Folate depletion has often been a problem in chronic antiepileptic drug (AED) therapy. Carbamazepine (CBZ), a commonly used AED, has been implicated in some clinical studies. A rat model was developed to examine the effects of chronic CBZ treatment on folate concentrations in the rat. In the course of developing this model, a common vehicle, propylene glycol, by itself in high doses, was found to exhibit protective properties against induced seizures and inhibited weight gain. Seizures induced by hexafluorodiethyl ether (HFDE) were also found to be a more sensitive measure of protection by CBZ than seizures induced by maximal electroshock (MES). Oral administration of CBZ as an aqueous suspension every 8 h at a dose of 250 mg/kg was continuously protective against HFDE-induced seizures and was minimally toxic as measured by weight gain over 8 weeks of treatment. The CBZ levels measured in plasma and brain of these animals, however, were below those normally considered protective. This treatment with CBZ had no apparent adverse effect on folate concentrations in the rat, and, indeed, the folate concentration increased in liver after 6 weeks of treatment and in plasma at 8 weeks of treatment.

Abstract  Methylazoxymethanol acetate (MAMac) is a potent teratogenic agent which can produce ectopic cell placement in developing rat brains. In the present study, we evaluated (i) whether prenatal exposure to MAMac results in a lowered seizure threshold to flurothyl and (ii) if there is a correlation between the number of ectopic cells in MAMac-exposed hippocampus and flurothyl-induced seizure latency. In 60 day old (P60) rats exposed to MAMac in utero, the latencies to myoclonic jerk (173 +/- 2.3 s) and forelimb clonus (215 +/- 4.6 s) were significantly shorter than those of controls (200 +/- 6.9 s and 238 +/- 8.8 s, respectively). MAMac also increased the proportion of flurothyl-treated rats that progressed from bilateral forelimb clonus to generalized tonic-clonic seizures (control: 33%; MAMac: 91%). Shorter seizure latencies were associated with an increased number of ectopic pyramidal cells in region CA1/CA2. These results suggest seizure susceptibility is enhanced in an animal model (MAMac) characterized by abnormal neuronal migration.

Abstract  Seizure disorders frequently occur early in life. Seizures are classified as reactive, symptomatic, or idiopathic depending on whether their cause can be identified. Reactive seizures are the result of acute environmental perturbations. Early in life, many stressors can produce seizures and the ultimate outcome may depend on the particular precipitating factor and its intensity. Febrile convulsions are the most common reactive seizures, although they must be differentiated from symptomatic seizures precipitated by fever. Symptomatic seizures are often associated with varying degrees of central nervous system (CNS) insults, including congenital malformations and metabolic storage diseases of the gray matter. These seizures may have age-specific characteristics and may at times be difficult to treat with conventional antiepileptic treatments. To develop a better understanding of the pathophysiology of seizures early in life, we have extensively used animal models of epilepsy. In this chapter, we report our findings with a rat model of developmental cortical dysplasias produced by intrauterine injections of methylazoxymethanol acetate. These rats are more susceptible to kainic acid, flurothyl, and hyperthermic seizures than normal rats. Rats with severe cortical dysplasia are most susceptible to seizures. We have also studied the mechanisms involved in the control of seizures during development because status epilepticus is more prevalent in infants than in adults. Our data suggest that the substantia nigra may play a crucial role in status epilepticus as a function of age. In the adult substantia nigra two regions mediate opposing effects on seizures following infusions of gamma-aminobutyric acid type A (GABAA) agents. One region is located in the anterior substantia nigra, and muscimol infusions in this region mediate anticonvulsant effects. The second region is in the posterior substantia nigra, and here muscimol infusions produce proconvulsant effects. In situ hybridization data demonstrate that, at the cellular level, neurons in the two substantia nigra regions differ in the amount of hybridization grains for GABAA receptor alpha 1 and gamma 2L subunit mRNAs. In developing male rats, only the "proconvulsant" region is present up to the age of 21 days. The transition from the immature to mature substantia nigra mediated seizure control occurs between age 25 and 30 days. The identification of age-dependent functional networks involved in the containment of seizures may lead to possible new pharmacologic strategies to control seizures, thus aiding the development of age-appropriate treatments of seizure disorders.

Abstract  Physiological ketosis is a hallmark of metabolism in suckling infants. However, little is known on the impact of physiological ketosis on brain excitability. We addressed this question in suckling rats in vivo. 16-channel extracellular field potential recordings were performed from somatosensory barrel cortex at postnatal days 5-9 non-anaesthetized rat pups. Seizures were induced by the volatile convulsant agent flurothyl. One hour after blockade of physiological ketogenesis using combined administration of beta-oxidation inhibitors mercaptoacetate, insulin and glucose to prevent hypoglycemia, we found no significant change in the flurothyl-induced electrographic seizures. However, build-up of seizures during two repetitive flurothyl applications was strongly aggravated in the animals with blocked ketogenesis. The effect of ketogenesis inhibitors was reversed by exogenous beta-hydroxybutyrate. Diazepam exerted anticonvulsive action both under physiological ketosis and after blockade of ketogenesis, and bumetanide had no significant anticonvulsive effects in both conditions. Thus, physiological ketosis reduces excitability in the immature brain and elimination of physiological ketosis results in elimination of this anticonvulsant effect. Our study raises concern that the changes in diet, and pharmacological manipulations such as glucose infusion, and pathologies such as hyperinsulinism which break natural ketosis, may be a potential risk factor for epileptogenesis in nursing infants.

Abstract  This study reviews the different in vivo experimental models that have been used for the study of epileptogenesis. In this review we will focus on how to replicate the different models that have led to the study of partial seizures, as well as generalized seizures and the status epilepticus. The main characteristics that participate in the processes that generate and modulate the manifestations of different models of epileptogenesis are described. The development of several models of experimental epilepsy in animals has clearly helped the study of specific brain areas capable of causing convulsions. The experimental models of epilepsy also have helped in the study the mechanisms and actions of epilepsy drugs. In order to develop experimental animal models of epilepsy, animals are generally chosen according to the kind of epilepsy that can be developed and studied. It is currently known that animal species can have epileptic seizures similar to those in humans. However, it is important to keep in mind that it has not been possible to entirely evaluate all manifestations of human epilepsy. Notwithstanding, these experimental models of epilepsy have allowed a partial understanding of most of the underlying mechanisms of this disease.

Abstract  Specific binding of [3H]naloxone to rat brain tissue in vitro was inhibited by the excitant organochlorinated insecticides (OCI), by ether (E) and octanol (OCT), and by the convulsant indoklon (IND) and its anesthetic isomer, isoindoklon (ISO). In the presence of 100 mM NaCl the inhibition of naloxone binding by E, OCT and ISO was greatly potentiated, whereas that by OCI and IND was attenuated. KCl (100 mM) was equally effective as NaCl on the action of anesthetics, but the effect of the excitant drugs was, in contrast to NaCl, unaffected by KCl. Specific binding of [3H]ouabain in the absence of Na, was depressed by anesthetics and enhanced by neuroexcitants. In the presence of NaCl, which by itself inhibits ouabain binding to brain, both anesthetics and excitants enhanced ouabain binding. DDE, a non-insecticidal analog of DDT, and the dimethyl derivative of the OCI, lindane, were inactive in the receptor assays. These observations point to a unique isolated system which responds consistently to anesthetic agents as a class and, in a different way, to neuroexcitant compounds.

Abstract  OBJECTIVE: BRD2 is a human gene repeatedly linked to and associated with juvenile myoclonic epilepsy (JME). Here, we define the developmental stage when increased seizure susceptibility first manifests in heterozygous Brd2+/- mice, an animal model of JME. We wanted to determine (1) whether seizure susceptibility correlates with the proven decrease of γ-aminobutyric acidergic (GABAergic) neuron numbers and (2) whether the seizure phenotype can be affected by sex hormones.

METHODS: Heterozygous (Brd2+/-) and wild-type (wt) mice of both sexes were tested for flurothyl-induced seizure susceptibility at postnatal day 15 (P15; wt, n = 13; Brd2+/-, n = 20), at P30 (wt, n = 20; Brd2+/-, n = 20), and in adulthood (5-6 months of age; wt, n = 10; Brd2+/-, n = 12). We measured latency to clonic and tonic-clonic seizure onset (flurothyl threshold). We also compared relative density of parvalbumin-positive (PVA+) and GAD67+ GABA neurons in the striatum and primary motor (M1) neocortex of P15 (n = 6-13 mice per subgroup) and P30 (n = 7-10 mice per subgroup) mice. Additional neonatal Brd2+/- mice were injected with testosterone propionate (females) or formestane (males) and challenged with flurothyl at P30.

RESULTS: P15 Brd2+/- mice showed no difference in seizure susceptibility compared to P15 wt mice. However, even at this early age, Brd2+/- mice showed fewer PVA+ neurons in the striatum and M1 neocortex. Compared to wt, the striatum in Brd2+/- mice showed an increased proportion of immature PVA+ neurons, with smaller cell bodies and limited dendritic arborization. P30 Brd2+/- mice displayed increased susceptibility to flurothyl-induced clonic seizures compared to wt. Both genotype and sex strongly influenced the density of PVA+ neurons in the striatum. Susceptibility to clonic seizures remained increased in adult Brd2+/- mice, and additionally there was increased susceptibility to tonic-clonic seizures. In P30 females, neonatal testosterone reduced the number of flurothyl-induced clonic seizures.

SIGNIFICANCE: A decrease in striatal PVA+ GABAergic neurons developmentally precedes the onset of increased seizure susceptibility and likely contributes to the expression of the syndrome.

Abstract  OBJECTIVE: Early life seizures (ELSs) alter activity-dependent maturation of neuronal circuits underlying learning and memory. The pathophysiological mechanisms underpinning seizure-induced cognitive impairment are not fully understood, and critical variables such as sex and dynamic brain states with regard to cognitive outcomes have not been explored. We hypothesized that in comparison to control (CTL) rats, ELS rats would exhibit deficits in spatial cognition correlating with impaired dynamic neural signal coordination between the hippocampus and medial prefrontal cortex (mPFC).

METHODS: Male and female rat pups were given 50 flurothyl-induced seizures over 10 days starting at postnatal Day 15. As adults, spatial cognition was tested through active avoidance on a rotating arena. Microwire tetrodes were implanted in the mPFC and CA1 subfield. Single cells and local field potentials were recorded and analyzed in each region during active avoidance and sleep.

RESULTS: ELS males exhibited avoidance impairments, whereas female rats were unaffected. During avoidance, hippocampus-mPFC coherence was higher in CTL females than CTL males across bandwidths. In comparison to CTL males, ELS male learners exhibit increased coherence within theta bandwidth as well as altered burst-timing in mPFC cell activity. Hippocampus-mPFC coherence levels are predictive of cognitive outcome in the active avoidance spatial task.

SIGNIFICANCE: Spatial cognitive outcome post-ELS is sex-dependent, as females fare better than males. ELS males that learn the task exhibit increased mPFC coherence levels at low-theta frequency, which may compensate for ELS effects on mPFC cell timing. These results suggest that coherence may serve as a biomarker for spatial cognitive outcome post-ELS and emphasize the significance of analyzing sex and dynamic cognition as variables in understanding seizure effects on the developing brain.

Abstract  OBJECTIVE: PCDH19-related epilepsy is characterized by a distinctive pattern of X-linked inheritance, where heterozygous females exhibit seizures and hemizygous males are asymptomatic. A cellular interference mechanism resulting from the presence of both wild-type and mutant PCDH19 neurons in heterozygous patients or mosaic carriers of PCDH19 variants has been hypothesized. We aim to investigate seizure susceptibility and progression in the Pchd19 mouse model.

METHODS: We assessed seizure susceptibility and progression in the Pcdh19 mouse model using three acute seizure induction paradigms. We first induced focal, clonic seizures using the 6-Hz psychomotor test. Mice were stimulated with increasing current intensities and graded according to a modified Racine scale. We next induced generalized seizures using flurothyl or pentylenetetrazol (PTZ), both γ-aminobutyric acid type A receptor function inhibitors, and recorded latencies to myoclonic and generalized tonic-clonic seizures.

RESULTS: Pcdh19 knockout and heterozygous females displayed increased seizure susceptibility across all current intensities in the 6-Hz psychomotor test, and increased severity overall. They also exhibited shorter latencies to generalized seizures following flurothyl, but not PTZ, seizure induction. Hemizygous males showed comparable seizure incidence and severity to their wild-type male littermates across all paradigms tested.

SIGNIFICANCE: The heightened susceptibility observed in Pcdh19 knockout females suggests additional mechanisms other than cellular interference are at play in PCDH19-related epilepsy. Further experiments are needed to understand the variability in seizure susceptibility so that this model can be best utilized toward development of future therapeutic strategies for PCDH19-related epilepsy.

Abstract  This study aimed to determine the long-term change of seizure susceptibility and the role of nNOS on brain development following recurrent early-life seizures in rats. Video-EEG recordings were conducted between postnatal days 50 and 60. Alterations in seizure susceptibility were assayed on day 22 or 50 using the flurothyl method. Changes in nNOS expression were determined by quantitative immunoblotting on day 50. On average, rats had 8.4+/-2.7 seizures during 10 daily 1 hr behavioral monitoring sessions. As adults (days 50-60), all rats displayed interictal spikes in the hippocampus and/or overlying cortex. Brief electrographic seizures were recorded in only one of five animals. Rats appeared to progress from a period of marked seizure susceptibility (day 22) to one of lessened seizure susceptibility (day 50). Up-regulation of nNOS expression following early-life recurrent seizures was observed on day 50. In conclusion, these data suggested that recurrent early-life seizures had the long-term effects on seizure susceptibility late in life and up-regulatory nNOS expression on the hippocampus during brain development, and nNOS appeared to contribute to the persistent changes in seizure susceptibility, and epileptogenesis.

Abstract  An association has recently been proposed between the incidence of seizures and prolonged consumption of the phenylalanine-containing artificial sweetener, aspartame. Since consumption of aspartame, unlike dietary protein, can elevate phenylalanine in brain, and thereby inhibit the synthesis and release of neurotransmitters known to protect against seizure activity, the effect of oral doses of aspartame on the sensitivity of mice to the proconvulsant agents, pentylenetetrazole and fluorothyl was studied. Doses of aspartame were used which increased phenylalanine more than tyrosine in brain, as occurs in humans after the consumption of any dose of aspartame. Pretreatment with aspartame significantly increased the percentage of animals convulsing after administration of pentylenetetrazole and significantly lowered the CD50 for this convulsant. The average time to onset of seizures induced by fluorothyl in control mice was 510 sec; pretreatment with oral doses of 1000, 1500 and 2000 mg/kg of aspartame 1 hr earlier significantly reduced the time required to elicit seizures (394, 381 and 339 sec, respectively). The seizure-promoting effect of aspartame could be demonstrated 30, 60 or 120 min after the 1000 mg/kg dose. The seizures induced by either convulsant were potentiated by equimolar amounts of phenylalanine, a major endogenous metabolite of aspartame, while the other metabolites, aspartic acid and methanol, were without effect. Administration together with aspartame of the large neutral amino acid valine, which competes with phenylalanine for entry into the brain, completely abolished the seizure-promoting effect of aspartame.(ABSTRACT TRUNCATED AT 250 WORDS)

Abstract  We have recently demonstrated that eight, daily flurothyl-induced generalized clonic seizures, followed by a four week stimulus-free interval, results in a long-lasting reduction in generalized seizure threshold and a change in the type of seizure expressed in response to flurothyl from clonic to tonic. There is a progressive increase in the probability that a mouse will express a tonic seizure during the four week interval, suggesting that prior flurothyl seizures initiate a proepileptogenic process that requires time to develop. In this study, the immunohistochemical detection of the c-fos protein (Fos) was used to evaluate whether seizure-induced epileptogenesis resulted in regional differences in the degree of neuronal activation. Fos immunoreactivity was examined 1.5 h following either a single generalized seizure, the last of eight consecutive daily seizures or a retest seizure evoked two weeks after the last of eight seizures. In each condition, generalized seizure behaviours were elicited in C57BL/6 mice using flurothyl and classified as either "forebrain" (face and forelimb clonus) or "brainstem" (running/bouncing, treading, tonic extension). The spatial distribution of Fos induction was compared on the basis of the seizure phenotype and the seizure history. The predominant differences in Fos distribution were found to be related to the type of seizure expressed regardless of the seizure history. Furthermore, the different motor components that make up a "brainstem" seizure could not be distinguished by the pattern of Fos labelling suggesting that multiple convulsive behaviours are mediated by one anatomical system. Finally, Fos induction in the ventromedial hypothalamic nucleus preceded and predicted the change in seizure type from "forebrain" to "brainstem". These data support the concept that separate anatomical systems mediate the expression of the two generalized seizure phenotypes. In addition, the ventromedial nucleus of the hypothalamus may be a point of interaction between the systems and may play a role in seizure-induced neural reorganization.