Diagnostic methods and treatment options for focal cortical dysplasia.

Our inability to adequately treat many patients with refractory epilepsy caused by focal cortical dysplasia (FCD), surgical inaccessibility and failures are significant clinical drawbacks. The targeting of physiologic features of epileptogenesis in FCD and colocalizing functionality has enhanced completeness of surgical resection, the main determinant of outcome. Electroencephalography (EEG)-functional magnetic resonance imaging (fMRI) and magnetoencephalography are helpful in guiding electrode implantation and surgical treatment, and high-frequency oscillations help defining the extent of the epileptogenic dysplasia. Ultra high-field MRI has a role in understanding the laminar organization of the cortex, and fluorodeoxyglucose-positron emission tomography (FDG-PET) is highly sensitive for detecting FCD in MRI-negative cases. Multimodal imaging is clinically valuable, either by improving the rate of postoperative seizure freedom or by reducing postoperative deficits. However, there is no level 1 evidence that it improves outcomes. Proof for a specific effect of antiepileptic drugs (AEDs) in FCD is lacking. Pathogenic mutations recently described in mammalian target of rapamycin (mTOR) genes in FCD have yielded important insights into novel treatment options with mTOR inhibitors, which might represent an example of personalized treatment of epilepsy based on the known mechanisms of disease. The ketogenic diet (KD) has been demonstrated to be particularly effective in children with epilepsy caused by structural abnormalities, especially FCD. It attenuates epigenetic chromatin modifications, a master regulator for gene expression and functional adaptation of the cell, thereby modifying disease progression. This could imply lasting benefit of dietary manipulation. Neurostimulation techniques have produced variable clinical outcomes in FCD. In widespread dysplasias, vagus nerve stimulation (VNS) has achieved responder rates >50%; however, the efficacy of noninvasive cranial nerve stimulation modalities such as transcutaneous VNS (tVNS) and noninvasive (nVNS) requires further study. Although review of current strategies underscores the serious shortcomings of treatment-resistant cases, initial evidence from novel approaches suggests that future success is possible.

Multiple sources of striatal inhibition are differentially affected in Huntington's disease mouse models.

In Huntington's disease (HD) mouse models, spontaneous inhibitory synaptic activity is enhanced in a subpopulation of medium-sized spiny neurons (MSNs), which could dampen striatal output. We examined the potential source(s) of increased inhibition using electrophysiological and optogenetic methods to assess feedback and feedforward inhibition in two transgenic mouse models of HD. Single whole-cell patch-clamp recordings demonstrated that increased GABA synaptic activity impinges principally on indirect pathway MSNs. Dual patch recordings between MSNs demonstrated reduced connectivity between MSNs in HD mice. However, while connectivity was strictly unidirectional in controls, in HD mice bidirectional connectivity occurred. Other sources of increased GABA activity in MSNs also were identified. Dual patch recordings from fast spiking (FS) interneuron-MSN pairs demonstrated greater but variable amplitude responses in MSNs. In agreement, selective optogenetic stimulation of parvalbumin-expressing, FS interneurons induced significantly larger amplitude MSN responses in HD compared with control mice. While there were no differences in responses of MSNs evoked by activating single persistent low-threshold spiking (PLTS) interneurons in recorded pairs, these interneurons fired more action potentials in both HD models, providing another source for increased frequency of spontaneous GABA synaptic activity in MSNs. Selective optogenetic stimulation of somatostatin-expressing, PLTS interneurons did not reveal any significant differences in responses of MSNs in HD mice. These findings provide strong evidence that both feedforward and to a lesser extent feedback inhibition to MSNs in HD can potentially be sources for the increased GABA synaptic activity of indirect pathway MSNs.

Identification of ß-Glucocerebrosidase Activators for Glucosylceramide hydrolysis.

Several novel chemical series were identified that modulate glucocerebrosidase (GCase). Compounds from these series are active on glucosylceramide, unlike other known GCase modulators. We obtained GCase crystal structures with two compounds that have distinct chemotypes. Positive allosteric modulators bind to a site on GCase and induce conformational changes, but also induce an equilibrium state between monomer and dimer.

Differential electrophysiological changes in striatal output neurons in Huntington's disease.

There is considerable evidence that alterations in striatal medium-sized spiny neurons (MSSNs) giving rise to the direct (D1 receptor-expressing) and indirect (D2 receptor-expressing) pathways differentially contribute to the phenotype of Huntington's disease (HD). To determine how each subpopulation of MSSN is functionally affected, we examined spontaneous excitatory postsynaptic currents (sEPSCs) and dopamine (DA) modulation in two HD mouse models, the YAC128 and the BACHD (a bacterial-artificial chromosome). These mice also expressed enhanced green fluorescent protein (EGFP) under the control of the promoter for either DA D1 or D2 receptors to identify neurons. In early symptomatic YAC128 and BACHD mice, glutamate transmission was increased in both D1 and D2 MSSNs, but in different ways. D1 cells displayed increased sEPSC frequencies and decreased paired-pulse ratios (PPRs) while D2 cells displayed larger evoked glutamate currents but no change in sEPSC frequencies or PPRs. D1 receptor modulation of sEPSCs was absent in D1-YAC128 cells at the early symptomatic stage but was restored by treating the slices with tetrabenazine. In contrast, in fully symptomatic YAC128 mice, glutamate transmission was decreased specifically in D1 cells, and D1 receptor modulation was normal in D1-YAC128 cells. Behaviorally, early symptomatic mice showed increased stereotypies that were decreased by tetrabenazine treatment. Together, these studies support differential imbalances in glutamate and DA transmission in direct and indirect pathway MSSNs. Stereotypic behavior at an early stage could be explained by increased glutamate activity and DA tone in direct pathway neurons, whereas hypokinesia at later stages could result from reduced input onto these neurons.

A critical window of CAG repeat-length correlates with phenotype severity in the R6/2 mouse model of Huntington's disease.

The R6/2 mouse is the most frequently used model for experimental and preclinical drug trials in Huntington's disease (HD). When the R6/2 mouse was first developed, it carried exon 1 of the huntingtin gene with ~150 cytosine-adenine-guanine (CAG) repeats. The model presented with a rapid and aggressive phenotype that shared many features with the human condition and was particularly similar to juvenile HD. However, instability in the CAG repeat length due to different breeding practices has led to both decreases and increases in average CAG repeat lengths among colonies. Given the inverse relationship in human HD between CAG repeat length and age at onset and to a degree, the direct relationship with severity of disease, we have investigated the effect of altered CAG repeat length. Four lines, carrying ~110, ~160, ~210, and ~310 CAG repeats, were examined using a battery of tests designed to assess the basic R6/2 phenotype. These included electrophysiological properties of striatal medium-sized spiny neurons, motor activity, inclusion formation, and protein expression. The results showed an unpredicted, inverted "U-shaped" relationship between CAG repeat length and phenotype; increasing the CAG repeat length from 110 to 160 exacerbated the R6/2 phenotype, whereas further increases to 210 and 310 CAG repeats greatly ameliorated the phenotype. These findings demonstrate that the expected relationship between CAG repeat length and disease severity observed in humans is lost in the R6/2 mouse model and highlight the importance of CAG repeat-length determination in preclinical drug trials that use this model.

Enhanced GABAergic network and receptor function in pediatric cortical dysplasia Type IIB compared with Tuberous Sclerosis Complex.

Tuberous Sclerosis Complex (TSC) and cortical dysplasia Type IIB (CDIIB) share histopathologic features that suggest similar epileptogenic mechanisms. This study compared the morphological and electrophysiological properties of cortical cells in tissue from pediatric TSC (n=20) and CDIIB (n=20) patients using whole-cell patch clamp recordings and biocytin staining. Cell types were normal-appearing and dysmorphic-cytomegalic pyramidal neurons, interneurons, and giant/balloon cells, including intermediate neuronal-glial cells. In the cortical mantle, giant/balloon cells occurred more frequently in TSC than in CDIIB cases, whereas cytomegalic pyramidal neurons were found more frequently in CDIIB. Cell morphology and membrane properties were similar in TSC and CDIIB cases. Except for giant/balloon and intermediate cells, all neuronal cell types fired action potentials and displayed spontaneous postsynaptic currents. However, the frequency of spontaneous glutamatergic postsynaptic currents in normal pyramidal neurons and interneurons was significantly lower in CDIIB compared with TSC cases and the GABAergic activity was higher in all neuronal cell types in CDIIB. Further, acutely dissociated pyramidal neurons displayed higher sensitivity to exogenous application of GABA in CDIIB compared with TSC cases. These results indicate that, in spite of similar histopathologic features and basic cell membrane properties, TSC and CDIIB display differences in the topography of abnormal cells, excitatory and inhibitory synaptic network properties, and GABA(A) receptor sensitivity. These differences support the notion that the mechanisms of epileptogenesis could differ in patients with TSC and CDIIB. Consequently, pharmacologic therapies should take these findings into consideration.

Alterations in cortical excitation and inhibition in genetic mouse models of Huntington's disease.

Previously, we identified progressive alterations in spontaneous EPSCs and IPSCs in the striatum of the R6/2 mouse model of Huntington's disease (HD). Medium-sized spiny neurons from these mice displayed a lower frequency of EPSCs, and a population of cells exhibited an increased frequency of IPSCs beginning at approximately 40 d, a time point when the overt behavioral phenotype begins. The cortex provides the major excitatory drive to the striatum and is affected during disease progression. We examined spontaneous EPSCs and IPSCs of somatosensory cortical pyramidal neurons in layers II/III in slices from three different mouse models of HD: the R6/2, the YAC128, and the CAG140 knock-in. Results revealed that spontaneous EPSCs occurred at a higher frequency, and evoked EPSCs were larger in behaviorally phenotypic mice whereas spontaneous IPSCs were initially increased in frequency in all models and subsequently decreased in R6/2 mice after they displayed the typical R6/2 overt behavioral phenotype. Changes in miniature IPSCs and evoked IPSC paired-pulse ratios suggested altered probability of GABA release. Also, in R6/2 mice, blockade of GABA(A) receptors induced complex discharges in slices and seizures in vivo at all ages. In conclusion, altered excitatory and inhibitory inputs to pyramidal neurons in the cortex in HD appear to be a prevailing deficit throughout the development of the disease. Furthermore, the differences between synaptic phenotypes in cortex and striatum are important for the development of future therapeutic approaches, which may need to be targeted early in the development of the phenotype.

Age-dependent alterations of corticostriatal activity in the YAC128 mouse model of Huntington disease.

Huntington disease is a genetic neurodegenerative disorder that produces motor, neuropsychiatric, and cognitive deficits and is caused by an abnormal expansion of the CAG tract in the huntingtin (htt) gene. In humans, mutated htt induces a preferential loss of medium spiny neurons in the striatum and, to a lesser extent, a loss of cortical neurons as the disease progresses. The mechanisms causing these degenerative changes remain unclear, but they may involve synaptic dysregulation. We examined the activity of the corticostriatal pathway using a combination of electrophysiological and optical imaging approaches in brain slices and acutely dissociated neurons from the YAC128 mouse model of Huntington disease. The results demonstrated biphasic age-dependent changes in corticostriatal function. At 1 month, before the behavioral phenotype develops, synaptic currents and glutamate release were increased. At 7 and 12 months, after the development of the behavioral phenotype, evoked synaptic currents were reduced. Glutamate release was decreased by 7 months and was markedly reduced by 12 months. These age-dependent alterations in corticostriatal activity were paralleled by a decrease in dopamine D(2) receptor modulation of the presynaptic terminal. Together, these findings point to dynamic alterations at the corticostriatal pathway and emphasize that therapies directed toward preventing or alleviating symptoms need to be specifically designed depending on the stage of disease progression.

Differential susceptibility to excitotoxic stress in YAC128 mouse models of Huntington disease between initiation and progression of disease.

Huntington disease (HD) is a neurodegenerative disorder caused by an expanded CAG tract in the HD gene. Polyglutamine expansion of huntingtin (htt) results in early, progressive loss of medium spiny striatal neurons, as well as cortical neurons that project to the striatum. Excitotoxicity has been postulated to play a key role in the selective vulnerability of striatal neurons in HD. Early excitotoxic neuropathological changes observed in human HD brain include increased quinolinate (QUIN) concurrent with proliferative changes such as increased spine density and dendritic length. In later stages of the disease, degenerative-type changes are apparent, such as loss of dendritic arborization, a reduction in spine density and reduced levels of 3-hydroxykynurenine and QUIN. It is currently unknown whether sensitivity to excitotoxic stress varies between initiation and progression of disease. Here, we have assessed the excitotoxic phenotype in the YAC128 mouse model of HD by examining the response to excitotoxic stress at different stages of disease. Our results demonstrate that YAC128 mice display enhanced sensitivity to NMDA ex vivo and QUIN in vivo before obvious phenotypic changes. In contrast, 10-month-old symptomatic YAC128 mice are resistant to QUIN-induced neurotoxicity. These findings are paralleled by a significant increase in NMDAR-mediated membrane currents in presymptomatic YAC128 dissociated medium spiny neurons progressing to reduced NMDAR-mediated membrane currents with disease progression. These data highlight the dynamic nature of the mutant htt-mediated excitotoxic phenotype and suggests that therapeutic approaches to HD may need to be altered, depending on the stage and development of the disease.

Dopamine modulation of excitatory currents in the striatum is dictated by the expression of D1 or D2 receptors and modified by endocannabinoids.

Striatal medium-sized spiny neurons (MSSNs) receive glutamatergic inputs modulated presynaptically and postsynaptically by dopamine. Mice expressing the gene for enhanced green fluorescent protein as a reporter gene to identify MSSNs containing D1 or D2 receptor subtypes were used to examine dopamine modulation of spontaneous excitatory postsynaptic currents (sEPSCs) in slices and postsynaptic N-methyl-d-aspartate (NMDA) and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) currents in acutely isolated cells. The results demonstrated dopamine receptor-specific modulation of sEPSCs. Dopamine and D1 agonists increased sEPSC frequency in D1 receptor-expressing MSSNs (D1 cells), whereas dopamine and D2 agonists decreased sEPSC frequency in D2 receptor-expressing MSSNs (D2 cells). These effects were fully (D1 cells) or partially (D2 cells) mediated through retrograde signaling via endocannabinoids. A cannabinoid 1 receptor (CB1R) agonist and a blocker of anandamide transporter prevented the D1 receptor-mediated increase in sEPSC frequency in D1 cells, whereas a CB1R antagonist partially blocked the decrease in sEPSC frequency in D2 cells. At the postsynaptic level, low concentrations of a D1 receptor agonist consistently increased NMDA and AMPA currents in acutely isolated D1 cells, whereas a D2 receptor agonist decreased these currents in acutely isolated D2 cells. These results show that both glutamate release and postsynaptic excitatory currents are regulated in opposite directions by activation of D1 or D2 receptors. The direction of this regulation is also specific to D1 and D2 cells. We suggest that activation of postsynaptic dopamine receptors controls endocannabinoid mobilization, acting on presynaptic CB1Rs, thus modulating glutamate release differently in glutamate terminals projecting to D1 and D2 cells.

Interneurons, GABAA currents, and subunit composition of the GABAA receptor in type I and type II cortical dysplasia.

Interneurons, gamma-aminobutyric acid (GABA)(A) receptor density, and subunit composition determine inhibitory function in pyramidal neurons and control excitability in cortex. Abnormalities in GABAergic cells or GABA(A) receptors could contribute to seizures in malformations of cortical development. Herein we review data obtained in resected cortex from pediatric epilepsy surgery patients with type I and type II cortical dysplasia (CD) and non-CD pathologies. Our studies found fewer interneurons immunolabeled for glutamic acid decarboxylase (GAD) in type II CD, whereas there were no changes in tissue from type I CD. GAD-labeled neurons had larger somata, and GABA transporter (VGAT and GAT1) staining showed a dense plexus surrounding cytomegalic neurons in type II CD. Functionally, neurons from type I CD tissue showed GABA currents with increased half maximal effective concentration compared to cells from the other groups. In type II CD, cytomegalic pyramidal neurons showed alterations in GABA currents, decreased sensitivity to zolpidem and zinc, and increased sensitivity to bretazenil. In addition, pyramidal neurons from type II CD displayed higher frequency of spontaneous inhibitory post synaptic currents. The GABAergic system is therefore, altered differently in cortex from type I and type II CD patients. Alterations in zolpidem, zinc, and bretazenil sensitivity and spontaneous inhibitory postsynaptic currents (IPSCs) suggest that type II CD neurons have altered GABA(A) receptor subunit composition and receive dense GABA inputs. These findings support the hypothesis that patients with type I and type II CD will respond differently to GABA receptor-mediated antiepileptic drugs and that cytomegalic neurons have features similar to immature neurons.

Comparative study of cellular and synaptic abnormalities in brain tissue samples from pediatric tuberous sclerosis complex and cortical dysplasia type II.

Tuberous sclerosis complex (TSC) and severe cortical dysplasia (CD), or CD type II according to Palmini classification, share histopathologic similarities, specifically the presence of cytomegalic neurons and balloon cells. In this study we examined the morphologic and electrophysiologic properties of cells in cortical tissue samples from pediatric patients with TSC and CD type II who underwent surgery for pharmacoresistant epilepsy. Normal-appearing pyramidal neurons from TSC and CD type II cases had similar passive membrane properties. However, the frequency of excitatory postsynaptic currents (EPSCs) was higher in neurons from TSC compared to severe CD cases, particularly the frequency of medium- and large-amplitude synaptic events. In addition, EPSCs rise and decay times were slower in normal cells from TSC compared to severe CD cases. Balloon cells were found more frequently in TSC cases, whereas cytomegalic pyramidal neurons occurred more often in CD type II cases. Both cell types were similar morphologically and electrophysiologically in TSC and severe CD. These results suggest that even though the histopathology in TSC and severe CD is similar, there are subtle differences in spontaneous synaptic activity and topographic distribution of abnormal cells. These differences may contribute to variable mechanisms of epileptogenesis in patients with TSC compared with CD type II.

Full-length human mutant huntingtin with a stable polyglutamine repeat can elicit progressive and selective neuropathogenesis in BACHD mice.

To elucidate the pathogenic mechanisms in Huntington's disease (HD) elicited by expression of full-length human mutant huntingtin (fl-mhtt), a bacterial artificial chromosome (BAC)-mediated transgenic mouse model (BACHD) was developed expressing fl-mhtt with 97 glutamine repeats under the control of endogenous htt regulatory machinery on the BAC. BACHD mice exhibit progressive motor deficits, neuronal synaptic dysfunction, and late-onset selective neuropathology, which includes significant cortical and striatal atrophy and striatal dark neuron degeneration. Power analyses reveal the robustness of the behavioral and neuropathological phenotypes, suggesting BACHD as a suitable fl-mhtt mouse model for preclinical studies. Additional analyses of BACHD mice provide novel insights into how mhtt may elicit neuropathogenesis. First, unlike previous fl-mhtt mouse models, BACHD mice reveal that the slowly progressive and selective pathogenic process in HD mouse brains can occur without early and diffuse nuclear accumulation of aggregated mhtt (i.e., as detected by immunostaining with the EM48 antibody). Instead, a relatively steady-state level of predominantly full-length mhtt and a small amount of mhtt N-terminal fragments are sufficient to elicit the disease process. Second, the polyglutamine repeat within fl-mhtt in BACHD mice is encoded by a mixed CAA-CAG repeat, which is stable in both the germline and somatic tissues including the cortex and striatum at the onset of neuropathology. Therefore, our results suggest that somatic repeat instability does not play a necessary role in selective neuropathogenesis in BACHD mice. In summary, the BACHD model constitutes a novel and robust in vivo paradigm for the investigation of HD pathogenesis and treatment.

Assessment and surgical outcomes for mild type I and severe type II cortical dysplasia: a critical review and the UCLA experience.

Recent findings on the clinical, electroencephalography (EEG), neuroimaging, and surgical outcomes are reviewed comparing patients with Palmini type I (mild) and type II (severe) cortical dysplasia. Resources include peer-reviewed studies on surgically treated patients and a subanalysis of the 2004 International League Against Epilepsy (ILAE) Survey of Pediatric Epilepsy Surgery. These sources were supplemented with data from University of California, Los Angeles (UCLA). Cortical dysplasia is the most frequent histopathologic substrate in children, and the second most common etiology in adult epilepsy surgery patients. Cortical dysplasia patients present with seizures at an earlier age than other surgically treated etiologies, and 33-50% have nonlocalized scalp EEG and normal magnetic resonance imaging (MRI) scans. 2-((18)F)Fluoro-2-deoxy-D-glucose positron emission tomography (FDG-PET) is positive in 75-90% of cases. After complete resection, 80% of patients are seizure free compared with 20% with incomplete resections. Compared with type I, patients with type II cortical dysplasia present at younger ages, have higher seizure frequencies, and are extratemporal. Type I dysplasia is found more often in adult patients in the temporal lobe and is often MRI negative. These findings identify characteristics of patients with mild and severe cortical dysplasia that define surgically treated epilepsy syndromes. The authors discuss future challenges to identifying and treating medically refractory epilepsy patients with cortical dysplasia.

The corticostriatal pathway in Huntington's disease.

The corticostriatal pathway provides most of the excitatory glutamatergic input into the striatum and it plays an important role in the development of the phenotype of Huntington's disease (HD). This review summarizes results obtained from genetic HD mouse models concerning various alterations in this pathway. Evidence indicates that dysfunctions of striatal circuits and cortical neurons that make up the corticostriatal pathway occur during the development of the HD phenotype, well before there is significant neuronal cell loss. Morphological changes in the striatum are probably primed initially by alterations in the intrinsic functional properties of striatal medium-sized spiny neurons. Some of these alterations, including increased sensitivity of N-methyl-D-aspartate receptors in subpopulations of neurons, might be constitutively present but ultimately require abnormalities in the corticostriatal inputs for the phenotype to be expressed. Dysfunctions of the corticostriatal pathway are complex and there are multiple changes as demonstrated by significant age-related transient and more chronic interactions with the disease state. There also is growing evidence for changes in cortical microcircuits that interact to induce dysfunctions of the corticostriatal pathway. The conclusions of this review emphasize, first, the general role of neuronal circuits in the expression of the HD phenotype and, second, that both cortical and striatal circuits must be included in attempts to establish a framework for more rational therapeutic strategies in HD. Finally, as changes in cortical and striatal circuitry are complex and in some cases biphasic, therapeutic interventions should be regionally specific and take into account the temporal progression of the phenotype.

Differential electrophysiological properties of dopamine D1 and D2 receptor-containing striatal medium-sized spiny neurons.

The electrophysiological properties of distinct subpopulations of striatal medium-sized spiny neurons (MSSNs) were compared using enhanced green fluorescent protein as a reporter gene for identification of neurons expressing dopamine D1 and D2 receptor subtypes in mice. Whole-cell patch-clamp recordings in slices revealed that passive membrane properties were similar in D1 and D2 cells. All MSSNs displayed hyperpolarized resting membrane potentials but the threshold for firing action potentials was lower in D2 than in D1 neurons. In voltage clamp, the frequency of spontaneous excitatory postsynaptic currents was higher in D2 than in D1 cells and large-amplitude inward currents (> 100 pA) were observed only in D2 cells. After tetrodotoxin this difference was reduced, suggesting that sodium conductances contribute to the increased frequencies in D2 cells. After pharmacological blockade of GABA(A) receptors, a subset of D2 cells also displayed large spontaneous membrane depolarizations and complex responses to stimulation of the corticostriatal pathway. To further characterize ionotropic glutamate receptor function, alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) was applied onto dissociated MSSNs. Application of AMPA alone or in the presence of cyclothiazide (an AMPA receptor desensitization blocker) evoked larger currents in D1 than in D2 cells. Together, these data demonstrate significant differences in electrophysiological properties of subpopulations of MSSNs defined by selective expression of D1 and D2 receptors. D2 cells display increased excitability and reflect ongoing cortical activity more faithfully than D1 cells, an effect that is independent of postsynaptic AMPA receptors and probably results from stronger synaptic coupling. This could help to explain the increased vulnerability of D2 MSSNs in neurodegenerative disorders.

Pyramidal cell responses to gamma-aminobutyric acid differ in type I and type II cortical dysplasia.

Abnormalities in the gamma-aminobutyric acid (GABA)-ergic system could be responsible for seizures in cortical dysplasia (CD). We examined responses of pyramidal neurons to exogenous application of GABA, as well as alterations of GABAergic interneuron number and size in pediatric epilepsy surgery patients with non-CD, type I CD, and type II CD pathologies. We used the dissociated cell preparation for electrophysiology along with immunohistochemistry to identify number and size of GABAergic cells. Pyramidal neurons from type I CD tissue showed increased EC(50) and faster kinetics compared with cells from non-CD and type II CD tissue. Cytomegalic pyramidal neurons showed increased GABA peak currents and decreased peak current densities, longer kinetics, and decreased sensitivity to zolpidem and zinc compared with normal pyramidal cells from non-CD and type I CD. There were fewer but larger glutamic acid decarboxylase (GAD)-containing cells in type II CD tissue with cytomegalic neurons compared with non-CD, type I CD, and type II CD without cytomegalic neurons. In addition, GABA transporters (VGAT and GAT-1) showed increased staining surrounding cytomegalic neurons in type II CD tissue. These results indicate that there are differences in GABA(A) receptor-mediated pyramidal cell responses in type I and type II CD. Alterations in zolpidem and zinc sensitivities also suggest that cytomegalic neurons have altered GABA(A) receptor subunit composition. These findings support the hypothesis that patients with type I and type II CD will respond differently to GABA-mediated antiepileptic drugs and that cytomegalic neurons have features similar to immature neurons with prolonged GABA(A) receptor open channel times.

Cytomegalic interneurons: a new abnormal cell type in severe pediatric cortical dysplasia.

A defining histopathologic feature of Taylor-type cortical dysplasia (CD) is the presence of cytomegalic neurons and balloon cells. Most cytomegalic neurons appear to be pyramidal-shaped and glutamatergic. The present study demonstrates the presence of cytomegalic GABAergic interneurons in a subset of pediatric patients with severe CD. Cortical tissue samples from children with mild, severe, and non-CD pathologies were examined using morphologic and electrophysiologic techniques. By using in vitro slices, cytomegalic cells with characteristics consistent with interneurons were found in 6 of 10 patients with severe CD. Biocytin labeling demonstrated that cytomegalic interneurons had more dendrites than normal-appearing interneurons. Whole-cell patch clamp recordings showed that cytomegalic interneurons had increased membrane capacitance and time constant compared with normal-appearing interneurons. They also displayed signs of cellular hyperexcitability, evidenced by increased firing rates, decreased action potential inactivation, and the occurrence of spontaneous membrane depolarizations. Single-cell reverse transcription-polymerase chain reaction and immunohistochemistry for GABAergic markers provided further evidence that these cells were probably cytomegalic interneurons. The pathophysiologic significance of GABAergic cytomegalic interneurons in severe CD tissue is unknown, but they could inhibit glutamatergic cytomegalic pyramidal neurons, or contribute to the synchronization of neuronal networks and the propagation of ictal activity in a subset of pediatric patients with severe CD.

Altered expression of alpha3-containing GABAA receptors in the neocortex of patients with focal epilepsy.

Impaired transmission in GABAergic circuits is thought to contribute to the pathogenesis of epilepsy. Although it is well established that major reorganization of GABA(A) receptor subtypes occurs in the hippocampus of patients with medically refractory temporal lobe epilepsy (TLE), it is unclear whether this disorder is also associated with alterations in GABA(A) receptor subtypes in the neocortex. Here we have investigated immunohistochemically the subunit composition and neocortical distribution of three major GABA(A) receptor subtypes using antibodies specifically recognizing the subunits alpha1, alpha2, alpha3, beta2/3 and gamma2. Cortical tissue was obtained at surgery from patients with TLE and hippocampal sclerosis (HS; n = 9), TLE associated with neocortical lesions (non-HS; n = 12) and frontal lobe epilepsy (FLE; n = 5), with post-mortem samples serving as controls (n = 4). A distinct laminar and neuronal expression pattern of the alpha-subunit variants was found across the neocortical regions examined in the temporal and frontal lobes in both control and patient tissue samples. In the five patients with FLE, GABA(A) receptor subunit staining was unchanged as compared to controls. In patients with TLE we observed a marked decrease in alpha3-subunit staining in the superficial neocortical layers (I-III), but no change in the deep layers (V and VI) or in the expression pattern of the alpha1 and alpha2-subunits. Reduced expression in alpha3-containing GABA(A) receptors was detected in six out of nine patients of the HS group and four out of twelve patients of the non-HS group. Histopathological changes were present in eight out of the ten patients with decreased alpha3-subunit staining. The selective reduction in alpha3-containing GABA(A) receptors was confirmed using semiquantitative measurements of optical density (OD). The specific changes unique to alpha3-subunit expression in the superficial neocortical layers of patients with TLE suggest that this subtype is of particular significance in the reorganization of cortical GABAergic systems in focal epilepsy.

Contralateral hemimicrencephaly and clinical-pathological correlations in children with hemimegalencephaly.

In paediatric epilepsy surgery patients with hemimegalencephaly (HME; n = 23), this study compared clinical, neuroimaging and pathologic features to discern potential mechanisms for suboptimal post-hemispherectomy developmental outcomes and structural pathogenesis. MRI measured affected and non-affected cerebral hemisphere volumes for HME and non-HME cases, including monozygotic twins where one sibling had HME. Staining against neuronal nuclei (NeuN) determined grey and white matter cell densities and sizes in HME and autopsy cases, including the non-affected side of a HME surgical/autopsy case. By MRI, the affected hemisphere was larger and the non-affected side smaller in HME compared with non-HME children. The affected HME side showed enlarged abnormal deep grey and white matter structures and/or T2-weighted hypointensity in the subcortical white matter in 75% of cases, suggestive of excessive pre-natal neurogenesis and heterotopias. Histopathological examination of the affected HME side revealed immature-appearing neurons in 70%, polymicrogyria (PMG) in 61% and balloon cells in 45% of cases. Compared with autopsy cases, in HME children NeuN cell densities on the affected side were increased in the molecular layer and upper cortex (+244 to +18%), decreased in lower cortical layers (-35%) and increased in the white matter (+139 to +149%). Deep grey matter MRI abnormalities and/or T2-weighted white matter hypointensity correlated with the presence of immature-appearing neurons and PMG on histopathology, decreased NeuN cell densities in lower cortical layers and a positive history of infantile spasms. Post-surgery seizure control was associated with decreased NeuN densities in the molecular layer. In young children with HME and epilepsy, these findings indicate that there are bilateral cerebral hemispheric abnormalities and contralateral hemimicrencephaly is a likely explanation for poorer post-surgery seizure control and cognitive outcomes. In addition, our findings support the hypothesis that HME pathogenesis probably involves somatic mutations that affect each developing cerebral hemisphere differently with more neurons than expected on the HME side.