Absence of CNTNAP2 leads to epilepsy, neuronal migration abnormalities, and core autism-related deficits.

Although many genes predisposing to autism spectrum disorders (ASD) have been identified, the biological mechanism(s) remain unclear. Mouse models based on human disease-causing mutations provide the potential for understanding gene function and novel treatment development. Here, we characterize a mouse knockout of the Cntnap2 gene, which is strongly associated with ASD and allied neurodevelopmental disorders. Cntnap2(-/-) mice show deficits in the three core ASD behavioral domains, as well as hyperactivity and epileptic seizures, as have been reported in humans with CNTNAP2 mutations. Neuropathological and physiological analyses of these mice before the onset of seizures reveal neuronal migration abnormalities, reduced number of interneurons, and abnormal neuronal network activity. In addition, treatment with the FDA-approved drug risperidone ameliorates the targeted repetitive behaviors in the mutant mice. These data demonstrate a functional role for CNTNAP2 in brain development and provide a new tool for mechanistic and therapeutic research in ASD.

Pathological neurons generate ripples at the UP-DOWN transition disrupting information transfer.

OBJECTIVE: To confirm and investigate why pathological high-frequency oscillations (pHFOs), including ripples (80-200 Hz) and fast ripples (200-600 Hz), are generated during the UP-DOWN transition of the slow wave and if information transmission mediated by ripple temporal coupling is disrupted in the seizure-onset zone (SOZ). METHODS: We isolated 217 total units from 175.95 intracranial electroencephalography (iEEG) contact-hours of synchronized macro- and microelectrode recordings from 6 patients. Sleep slow oscillation (.1-2 Hz) epochs were identified in the iEEG recording. iEEG HFOs that occurred superimposed on the slow wave were transformed to phasors and adjusted by the phase of maximum firing in nearby units (i.e., maximum UP). We tested whether, in the SOZ, HFOs and associated action potentials (APs) occur more often at the UP-DOWN transition. We also examined ripple temporal correlations using cross-correlograms. RESULTS: At the group level in the SOZ, HFO and HFO-associated AP probability was highest during the UP-DOWN transition of slow wave excitability (p < < .001). In the non-SOZ, HFO and HFO-associated AP was highest during the DOWN-UP transition (p < < .001). At the unit level in the SOZ, 15.6% and 20% of units exhibited more robust firing during ripples (Cohen's d = .11-.83) and fast ripples (d = .36-.90) at the UP-DOWN transition (p < .05 f.d.r. corrected), respectively. By comparison, also in the SOZ, 6.6% (d = .14-.30) and 8.5% (d = .33-.41) of units had significantly less firing during ripples and fast ripples at the UP-DOWN transition, respectively. Additional data shows that ripple and fast ripple temporal correlations, involving global slow waves, between the hippocampus, entorhinal cortex, and parahippocampal gyrus were reduced by >50% in the SOZ compared to the non-SOZ (N = 3). SIGNIFICANCE: The UP-DOWN transition of slow wave excitability facilitates the activation of pathological neurons to generate pHFOs. Ripple temporal correlations across brain regions may be important in memory consolidation and are disrupted in the SOZ, perhaps by pHFO generation.

Spatial and temporal profile of high-frequency oscillations in posttraumatic epileptogenesis.

We studied the role of temporal and spatial changes in high-frequency oscillation (HFO, 80-500 Hz) generation in epileptogenesis following traumatic brain injury (TBI). Experiments were conducted on adult male Sprague Dawley rats. For the TBI group, fluid percussion injury (FPI) on the left sensorimotor area was performed to induce posttraumatic epileptogenesis. For the sham control group, only the craniotomy was performed. After TBI, 8 bipolar micro-electrodes were implanted bilaterally in the prefrontal cortex, perilesional area and homotopic contralateral site, striatum, and hippocampus. Long-term video/local field potential (LFP) recordings were performed for up to 21 weeks to identify and characterize seizures and capture HFOs. The electrode tip locations and the volume of post TBI brain lesions were further estimated by ex-vivo MRI scans. HFOs were detected during slow-wave sleep and categorized as ripple (80-200 Hz) and fast ripple (FR, 250-500 Hz) events. HFO rates and the HFO peak frequencies were compared in the 8 recording locations and across 8-weeks following TBI. Data from 48 rats (8 sham controls and 40 TBI rats) were analyzed. Within the TBI group, 22 rats (55%) developed recurrent spontaneous seizures (E+ group), at an average of 62.2 (+17.1) days, while 18 rats (45%) did not (E- group). We observed that the HFOs in the E+ group had a higher mean peak frequency than the E- group and the sham group (P < 0.05). Furthermore, the FR rate of the E+ group showed a significant increase compared to the E-group (P < 0.01) and sham control group (P < 0.01), specifically in the perilesional area, homotopic contralateral site, bilateral hippocampus, and to a lesser degree bilateral striatum. When compared across time, the increased FR rate in the E+ group occurred immediately after the insult and remained stable across the duration of the experiment. In addition, lesion size was not statistically different in the E+ and E- group and was not correlated with HFO rates. Our results suggest that TBI results in the formation of a widespread epileptogenic network. FR rates serve as a biomarker of network formation and predict the future development of epilepsy, however FR are not a temporally specific biomarker of TBI sequelae responsible for epileptogenesis. These results suggest that in patients, future risk of post-TBI epilepsy can be predicted early using FR.

Spike and wave discharges and fast ripples during posttraumatic epileptogenesis.

OBJECTIVE: The goal of the present study was to determine whether spike and wave discharges (SWDs) and SWDs with superimposed fast ripples (SWDFRs) could be biomarkers of posttraumatic epileptogenesis. METHODS: Fluid percussion injury was conducted on 13-14-week old male Sprague Dawley rats. Immediately after traumatic brain injury (TBI), they were implanted with microelectrodes in the neocortex, hippocampus, and striatum bilaterally. Age-matched sham rats with the same electrode implantation montage acted as controls. Wideband brain electrical activity was recorded intermittently from Day 1 of TBI, and continued from 2 to 21 weeks after TBI. SWD and SWDFR analysis was performed during the first 2 weeks to investigate whether the occurrence of this pattern predicted development of epilepsy. The remaining 3-21 weeks were used for identifying which rats became epileptic (E+ group) and which did not (E- group). RESULTS: The E+ group (n = 9) showed a significant increase in SWD rate in prefrontal cortex during Weeks 1 and 2 after TBI. The E- group showed a significant increase in SWD rate only in the second week. One hundred percent of rats in the E+ group displayed SWDFRs beginning from the first week after TBI. The SWDFR pattern was observed in all recorded brain areas: prefrontal and perilesional cortices, hippocampus, and striatum. None of rats in the E- group showed coexistence of fast ripples with SWDs. SIGNIFICANCE: Occurrence of SWDFRs after TBI, but not an increase in the rate of SWDs, could be a noninvasive electroencephalographic biomarker of posttraumatic epileptogenesis.

Topographical reorganization of brain functional connectivity during an early period of epileptogenesis.

OBJECTIVE: The current study aims to investigate functional brain network representations during the early period of epileptogenesis. METHODS: Eighteen rats with the intrahippocampal kainate model of mesial temporal lobe epilepsy were used for this experiment. Functional magnetic resonance imaging (fMRI) measurements were made 1 week after status epilepticus, followed by 2-4-month electrophysiological and video monitoring. Animals were identified as having (1) developed epilepsy (E+, n = 9) or (2) not developed epilepsy (E-, n = 6). Nine additional animals served as controls. Graph theory analysis was performed on the fMRI data to quantify the functional brain networks in all animals prior to the development of epilepsy. Spectrum clustering with the network features was performed to estimate their predictability in epileptogenesis. RESULTS: Our data indicated that E+ animals showed an overall increase in functional connectivity strength compared to E- and control animals. Global network features and small-worldness of E- rats were similar to controls, whereas E+ rats demonstrated increased small-worldness, including increased reorganization degree, clustering coefficient, and global efficiency, with reduced shortest pathlength. A notable classification of the combined brain network parameters was found in E+ and E- animals. For the local network parameters, the E- rats showed increased hubs in sensorimotor cortex, and decreased hubness in hippocampus. The E+ rats showed a complete loss of hippocampal hubs, and the appearance of new hubs in the prefrontal cortex. We also observed that lesion severity was not related to epileptogenesis. SIGNIFICANCE: Our data provide a view of the reorganization of topographical functional brain networks in the early period of epileptogenesis and how it can significantly predict the development of epilepsy. The differences from E- animals offer a potential means for applying noninvasive neuroimaging tools for the early prediction of epilepsy.

Electrophysiological biomarkers of epileptogenicity after traumatic brain injury.

Post-traumatic epilepsy is the architype of acquired epilepsies, wherein a brain insult initiates an epileptogenic process culminating in an unprovoked seizure after weeks, months or years. Identifying biomarkers of such process is a prerequisite for developing and implementing targeted therapies aimed at preventing the development of epilepsy. Currently, there are no validated electrophysiological biomarkers of post-traumatic epileptogenesis. Experimental EEG studies using the lateral fluid percussion injury model have identified three candidate biomarkers of post-traumatic epileptogenesis: pathological high-frequency oscillations (HFOs, 80-300 Hz); repetitive HFOs and spikes (rHFOSs); and reduction in sleep spindle duration and dominant frequency at the transition from stage III to rapid eye movement sleep. EEG studies in humans have yielded conflicting data; recent evidence suggests that epileptiform abnormalities detected acutely after traumatic brain injury carry a significantly increased risk of subsequent epilepsy. Well-designed studies are required to validate these promising findings, and ultimately establish whether there are post-traumatic electrophysiological features which can guide the development of 'antiepileptogenic' therapies.

Unit firing and oscillations at seizure onset in epileptic rodents.

Epileptic seizures result from a variety of pathophysiological processes, evidenced by different electrographic ictal onset patterns, as seen on direct brain recordings. The two most common electrographic patterns of focal ictal onset in patients are hypersynchronous (HYP) and low-voltage fast (LVF). Whereas LVF ictal onsets were believed to result from disinhibition; based on similarities with absence seizures, focal HYP ictal onsets were believed to result from increased synchronizing inhibition. Recent findings, however, suggest the differences between these seizure onset types are more complicated and, in some cases, the opposite of these concepts are true. The following review presents evidence that a reduction of tonic inhibition on small pathologically interconnected neuron (PIN) clusters generating pathological high-frequency oscillations (pHFOs), which reflect abnormal synchronously bursting neurons may be the cause of HYP ictal onsets. Increased inhibition preceding LVF ictal onsets are discussed in other reviews in this issue. We postulate that neuronal cell loss following epileptogenic insults can result in structural reorganization, giving rise to small PIN clusters, which generate pHFOs. These clusters have a heterogeneous distribution and are spatially stable over time. Studies have demonstrated that a transient reduction in tonic inhibition causes these clusters to increase in size. This could result in consolidation and synchronization of pHFOs until a critical mass leads to propagation of HYP ictal discharges. Viewed within a network neuroscience framework, local disturbances such as PIN clusters are likely to contribute to large-scale brain network alterations: a better understanding of these epileptogenic networks promises to elucidate mechanisms of ictogenesis, epileptogenesis, and certain comorbidities of epilepsy.

Low-voltage fast seizures in humans begin with increased interneuron firing.

OBJECTIVE: Intracellular recordings from cells in entorhinal cortex tissue slices show that low-voltage fast (LVF) onset seizures are generated by inhibitory events. Here, we determined whether increased firing of interneurons occurs at the onset of spontaneous mesial-temporal LVF seizures recorded in patients. METHODS: The seizure onset zone (SOZ) was identified using visual inspection of the intracranial electroencephalogram. We used wavelet clustering and temporal autocorrelations to characterize changes in single-unit activity during the onset of LVF seizures recorded from microelectrodes in mesial-temporal structures. Action potentials generated by principal neurons and interneurons (ie, putative excitatory and inhibitory neurons) were distinguished using waveform morphology and K-means clustering. RESULTS: From a total of 200 implanted microelectrodes in 9 patients during 13 seizures, we isolated 202 single units; 140 (69.3%) of these units were located in the SOZ, and 40 (28.57%) of them were classified as inhibitory. The waveforms of both excitatory and inhibitory units remained stable during the LVF epoch (p > > 0.05). In the mesial-temporal SOZ, inhibitory interneurons increased their firing rate during LVF seizure onset (p < 0.01). Excitatory neuron firing rates peaked 10 seconds after the inhibitory neurons (p < 0.01). During LVF spread to the contralateral mesial temporal lobe, an increase in inhibitory neuron firing rate was also observed (p < 0.01). INTERPRETATION: Our results suggest that seizure generation and spread during spontaneous mesial-temporal LVF onset events in humans may result from increased inhibitory neuron firing that spawns a subsequent increase in excitatory neuron firing and seizure evolution. Ann Neurol 2018;84:588-600.

Extrahippocampal high-frequency oscillations during epileptogenesis.

The current study aimed to investigate the spatial and temporal patterns of high-frequency oscillations (HFOs) in the intra-/extrahippocampal areas during epileptogenesis. Local field potentials were bilaterally recorded from hippocampus (CA1), thalamus, motor cortex, and prefrontal cortex in 13 rats before and after intrahippocampal kainic acid (KA) lesions. HFOs in the ripple (100-200 Hz) and fast ripple (250-500 Hz) ranges were detected and their rates were computed during different time periods (1-5 weeks) after KA-induced status epilepticus (SE). Recurrent spontaneous seizures were observed in 7 rats after SE, and the other 6 rats did not develop epilepsy. During the latent period, the rate of hippocampal HFOs increased at the ipsilateral site of the KA lesion in both groups, and the HFO rate was significantly higher in the animals that later developed epilepsy. Animals that later developed epilepsy also demonstrated widespread appearance of HFOs, in both the ripple and the fast ripple range, whereas animals that did not develop epilepsy only exhibited changes in the ipsilateral intrahippocampal HFO rate. This study demonstrates an association between an increased rate of widespread HFOs and the later development of epilepsy, suggesting the formation of large-scale distributed pathological networks during epileptogenesis.

Bimodal coupling of ripples and slower oscillations during sleep in patients with focal epilepsy.

OBJECTIVE: Differentiating pathologic and physiologic high-frequency oscillations (HFOs) is challenging. In patients with focal epilepsy, HFOs occur during the transitional periods between the up and down state of slow waves. The preferred phase angles of this form of phase-event amplitude coupling are bimodally distributed, and the ripples (80-150 Hz) that occur during the up-down transition more often occur in the seizure-onset zone (SOZ). We investigated if bimodal ripple coupling was also evident for faster sleep oscillations, and could identify the SOZ. METHODS: Using an automated ripple detector, we identified ripple events in 40-60 min intracranial electroencephalography (iEEG) recordings from 23 patients with medically refractory mesial temporal lobe or neocortical epilepsy. The detector quantified epochs of sleep oscillations and computed instantaneous phase. We utilized a ripple phasor transform, ripple-triggered averaging, and circular statistics to investigate phase event-amplitude coupling. RESULTS: We found that at some individual recording sites, ripple event amplitude was coupled with the sleep oscillatory phase and the preferred phase angles exhibited two distinct clusters (p < 0.05). The distribution of the pooled mean preferred phase angle, defined by combining the means from each cluster at each individual recording site, also exhibited two distinct clusters (p < 0.05). Based on the range of preferred phase angles defined by these two clusters, we partitioned each ripple event at each recording site into two groups: depth iEEG peak-trough and trough-peak. The mean ripple rates of the two groups in the SOZ and non-SOZ (NSOZ) were compared. We found that in the frontal (spindle, p = 0.009; theta, p = 0.006, slow, p = 0.004) and parietal lobe (theta, p = 0.007, delta, p = 0.002, slow, p = 0.001) the SOZ incidence rate for the ripples occurring during the trough-peak transition was significantly increased. SIGNIFICANCE: Phase-event amplitude coupling between ripples and sleep oscillations may be useful to distinguish pathologic and physiologic events in patients with frontal and parietal SOZ.

The progression of electrophysiologic abnormalities during epileptogenesis after experimental traumatic brain injury.

OBJECTIVE: Posttraumatic epilepsy (PTE) accounts for 20% of acquired epilepsies. Experimental models are important for studying epileptogenesis. We previously reported that repetitive high-frequency oscillations with spikes (rHFOSs) occur early after lateral fluid percussion injury (FPI) and may be a biomarker for PTE. The objective of this study was to use multiple electrodes in rat hippocampal and neocortical regions to describe the long-term electroencephalographic and behavioral evolution of rHFOSs and epileptic seizures after traumatic brain injury (TBI). METHODS: Adult male rats underwent mild, moderate, or severe FPI or sham injury followed by video-electroencephalography (EEG) recordings with a combination of 16 neocortical and hippocampal electrodes at an early, intermediate, or late time-point after injury, up to 52 weeks. Recordings were analyzed for the presence of rHFOSs and seizures. RESULTS: Analysis was done on 28 rats with FPI and 7 shams. Perilesional rHFOSs were recorded in significantly more rats after severe (70.3%) than mild (20%) injury or shams (14.3%). Frequency of occurrence was significantly highest in the early (10.8/h) versus late group (3.2/h). Late focal seizures originating from the same electrodes were recorded in significantly more rats in the late (87.5%) versus early period (22.2%), occurring almost exclusively in injured rats. Seizure duration increased significantly over time, averaging 19 s at the beginning of the early period and 27 s at the end of the late period. Seizure frequency also increased significantly over time, from 4.4 per week in the early group to 26.4 per week in the late group. Rarely, rats displayed early seizures or generalized seizures. SIGNIFICANCE: FPI results in early rHFOSs and later spontaneous focal seizures arising from peri-lesional neocortex, supporting its use as a model for PTE. Epilepsy severity increased over time and was related to injury severity. The association between early rHFOSs and later focal seizures suggests that rHFOSs may be a potential noninvasive biomarker of PTE.

Pathologic electrographic changes after experimental traumatic brain injury.

OBJECTIVE: To investigate possible electroencephalography (EEG) correlates of epileptogenesis after traumatic brain injury (TBI) using the fluid percussion model. METHODS: Experiments were conducted on adult 2- to 4-month-old male Sprague-Dawley rats. Two groups of animals were studied: (1) the TBI group with depth and screw electrodes implanted immediately after the fluid percussion injury (FPI) procedure, and (2) a naive age-matched control group with the same electrode implantation montage. Pairs of tungsten microelectrodes (50 µm outer diameter) and screw electrodes were implanted in neocortex inside the TBI core, areas adjacent to TBI, and remote areas. EEG activity, recorded on the day of FPI, and continuously for 2 weeks, was analyzed for possible electrographic biomarkers of epileptogenesis. Video-EEG monitoring was also performed continuously in the TBI group to capture electrographic and behavioral seizures until the caps came off (28-189 days), and for 1 week, at 2, 3, and 6 months of age, in the control group. RESULTS: Pathologic high-frequency oscillations (pHFOs) with a central frequency between 100 and 600 Hz, were recorded from microelectrodes, beginning during the first two post-FPI weeks, in 7 of 12 animals in the TBI group (58%) and never in the controls. pHFOs only occurred in cortical areas within or adjacent to the TBI core. These were associated with synchronous multiunit discharges and popSpikes, duration 15-40 msec. Repetitive pHFOs and EEG spikes (rHFOSs) formed paroxysmal activity, with a unique arcuate pattern, in the frequency band 10-16 Hz in the same areas as isolated pHFOs, and these events were also recorded by screw electrodes. Although loss of caps prevented long-term recordings from all rats, pHFOs and rHFOSs occurred during the first 2 weeks in all four animals that later developed seizures, and none of the rats without these events developed late seizures. SIGNIFICANCE: pHFOs, similar to those associated with epileptogenesis in the status rat model of epilepsy, may also reflect epileptogenesis after FPI. rHFOSs could be noninvasive biomarkers of epileptogenesis.

Ictal onset patterns of local field potentials, high frequency oscillations, and unit activity in human mesial temporal lobe epilepsy.

OBJECTIVE: To characterize local field potentials, high frequency oscillations, and single unit firing patterns in microelectrode recordings of human limbic onset seizures. METHODS: Wide bandwidth local field potential recordings were acquired from microelectrodes implanted in mesial temporal structures during spontaneous seizures from six patients with mesial temporal lobe epilepsy. RESULTS: In the seizure onset zone, distinct epileptiform discharges were evident in the local field potential prior to the time of seizure onset in the intracranial EEG. In all three seizures with hypersynchronous (HYP) seizure onset, fast ripples with incrementally increasing power accompanied epileptiform discharges during the transition to the ictal state (p < 0.01). In a single low voltage fast (LVF) onset seizure a triad of evolving HYP LFP discharges, increased single unit activity, and fast ripples of incrementally increasing power were identified ~20 s prior to seizure onset (p < 0.01). In addition, incrementally increasing fast ripples occurred after seizure onset just prior to the transition to LVF activity (p < 0.01). HYP onset was associated with an increase in fast ripple and ripple rate (p < 0.05) and commonly each HYP discharge had a superimposed ripple followed by a fast ripple. Putative excitatory and inhibitory single units could be distinguished during limbic seizure onset, and heterogeneous shifts in firing rate were observed during LVF activity. SIGNIFICANCE: Epileptiform activity is detected by microelectrodes before it is detected by depth macroelectrodes, and the one clinically identified LVF ictal onset was a HYP onset at the local level. Patterns of incrementally increasing fast ripple power are consistent with observations in rats with experimental hippocampal epilepsy, suggesting that limbic seizures arise when small clusters of synchronously bursting neurons increase in size, coalesce, and reach a critical mass for propagation.

Ripples on spikes show increased phase-amplitude coupling in mesial temporal lobe epilepsy seizure-onset zones.

OBJECTIVE: Ripples (80-150 Hz) recorded from clinical macroelectrodes have been shown to be an accurate biomarker of epileptogenic brain tissue. We investigated coupling between epileptiform spike phase and ripple amplitude to better understand the mechanisms that generate this type of pathologic ripple (pRipple) event. METHODS: We quantified phase amplitude coupling (PAC) between epileptiform electroencephalography (EEG) spike phase and ripple amplitude recorded from intracranial depth macroelectrodes during episodes of sleep in 12 patients with mesial temporal lobe epilepsy. PAC was determined by (1) a phasor transform that corresponds to the strength and rate of ripples coupled with spikes, and a (2) ripple-triggered average to measure the strength, morphology, and spectral frequency of the modulating and modulated signals. Coupling strength was evaluated in relation to recording sites within and outside the seizure-onset zone (SOZ). RESULTS: Both the phasor transform and ripple-triggered averaging methods showed that ripple amplitude was often robustly coupled with epileptiform EEG spike phase. Coupling was found more regularly inside than outside the SOZ, and coupling strength correlated with the likelihood a macroelectrode's location was within the SOZ (p < 0.01). The ratio of the rate of ripples coupled with EEG spikes inside the SOZ to rates of coupled ripples in non-SOZ was greater than the ratio of rates of ripples on spikes detected irrespective of coupling (p < 0.05). Coupling strength correlated with an increase in mean normalized ripple amplitude (p < 0.01), and a decrease in mean ripple spectral frequency (p < 0.05). SIGNIFICANCE: Generation of low-frequency (80-150 Hz) pRipples in the SOZ involves coupling between epileptiform spike phase and ripple amplitude. The changes in excitability reflected as epileptiform spikes may also cause clusters of pathologically interconnected bursting neurons to grow and synchronize into aberrantly large neuronal assemblies.

Molecular alterations in areas generating fast ripples in an animal model of temporal lobe epilepsy.

The molecular basis of epileptogenesis is poorly characterized. Studies in humans and animal models have identified an electrophysiological signature that precedes the onset of epilepsy, which has been termed fast ripples (FRs) based on its frequency. Multiple lines of evidence implicate regions generating FRs in epileptogenesis, and FRs appear to demarcate the seizure onset zone, suggesting a role in ictogenesis as well. We performed gene expression analysis comparing areas of the dentate gyrus that generate FRs to those that do not generate FRs in a well-characterized rat model of epilepsy. We identified a small cohort of genes that are differentially expressed in FR versus non-FR brain tissue and used quantitative PCR to validate some of those that modulate neuronal excitability. Gene expression network analysis demonstrated conservation of gene co-expression between non-FR and FR samples, but examination of gene connectivity revealed changes that were most pronounced in the cm-40 module, which contains several genes associated with synaptic function and the differentially expressed genes Kcna4, Kcnv1, and Npy1r that are down-regulated in FRs. We then demonstrate that the genes within the cm-40 module are regulated by seizure activity and enriched for the targets of the RNA binding protein Elavl4. Our data suggest that seizure activity induces co-expression of genes associated with synaptic transmission and that this pattern is attenuated in areas displaying FRs, implicating the failure of this mechanism in the generation of FRs.

Simulated resections and RNS placement can optimize post-operative seizure outcomes when guided by fast ripple networks.

In medication-resistant epilepsy, the goal of epilepsy surgery is to make a patient seizure free with a resection/ablation that is as small as possible to minimize morbidity. The standard of care in planning the margins of epilepsy surgery involves electroclinical delineation of the seizure onset zone (SOZ) and incorporation of neuroimaging findings from MRI, PET, SPECT, and MEG modalities. Resecting cortical tissue generating high-frequency oscillations (HFOs) has been investigated as a more efficacious alternative to targeting the SOZ. In this study, we used a support vector machine (SVM), with four distinct fast ripple (FR: 350-600 Hz on oscillations, 200-600 Hz on spikes) metrics as factors. These metrics included the FR resection ratio (RR), a spatial FR network measure, and two temporal FR network measures. The SVM was trained by the value of these four factors with respect to the actual resection boundaries and actual seizure free labels of 18 patients with medically refractory focal epilepsy. Leave one out cross-validation of the trained SVM in this training set had an accuracy of 0.78. We next used a simulated iterative virtual resection targeting the FR sites that were highest rate and showed most temporal autonomy. The trained SVM utilized the four virtual FR metrics to predict virtual seizure freedom. In all but one of the nine patients seizure free after surgery, we found that the virtual resections sufficient for virtual seizure freedom were larger in volume (p<0.05). In nine patients who were not seizure free, a larger virtual resection made five virtually seizure free. We also examined 10 medically refractory focal epilepsy patients implanted with the responsive neurostimulator system (RNS) and virtually targeted the RNS stimulation contacts proximal to sites generating FR at highest rates to determine if the simulated value of the stimulated SOZ and stimulated FR metrics would trend toward those patients with a better seizure outcome. Our results suggest: 1) FR measures can accurately predict whether a resection, defined by the standard of care, will result in seizure freedom; 2) utilizing FR alone for planning an efficacious surgery can be associated with larger resections; 3) when FR metrics predict the standard of care resection will fail, amending the boundaries of the planned resection with certain FR generating sites may improve outcome; and 4) more work is required to determine if targeting RNS stimulation contact proximal to FR generating sites will improve seizure outcome.

Connectomics and epilepsy.

PURPOSE OF REVIEW: Tremendous advances have occurred in recent years in elucidating basic mechanisms of epilepsy at the level of ion channels and neurotransmitters. Epilepsy, however, is ultimately a disease of functionally and/or structurally aberrant connections between neurons and groups of neurons at the systems level. Recent advances in neuroimaging and electrophysiology now make it possible to investigate structural and functional connectivity of the entire brain, and these techniques are currently being used to investigate diseases that manifest as global disturbances of brain function. Epilepsy is such a disease, and our understanding of the mechanisms underlying the development of epilepsy and the generation of epileptic seizures will undoubtedly benefit from research utilizing these connectomic approaches. RECENT FINDINGS: MRI using diffusion tensor imaging provides structural information, whereas functional MRI and electroencephalography provide functional information about connectivity at the whole brain level. Optogenetics, tracers, electrophysiological approaches, and calcium imaging provide connectivity information at the level of local circuits. These approaches are revealing important neuronal network disturbances underlying epileptic abnormalities. SUMMARY: An understanding of the fundamental mechanisms underlying the development of epilepsy and the generation of epileptic seizures will require delineation of the aberrant functional and structural connections of the whole brain. The field of connectomics now provides approaches to accomplish this.

Intracranial electrophysiological recordings on a swine model of mesial temporal lobe epilepsy.

Objective: To test the feasibility and reliability of intracranial electrophysiological recordings in an acute status epilepticus model on laboratory swine. Method: Intrahippocampal injection of kainic acid (KA) was performed on 17 male Bama pigs (Sus scrofa domestica) weighing between 25 and 35 kg. Two stereoelectroencephalography (SEEG) electrodes with a total of 16 channels were implanted bilaterally along the sensorimotor cortex to the hippocampus. Brain electrical activity was recorded 2 h daily for 9-28 days. Three KA dosages were tested to evaluate the quantities capable of evoking status epilepticus. Local field potentials (LFPs) were recorded and compared before and after the KA injection. We quantified the epileptic patterns, including the interictal spikes, seizures, and high-frequency oscillations (HFOs), up to 4 weeks after the KA injection. Test-retest reliability using intraclass correlation coefficients (ICCs) were performed on interictal HFO rates to evaluate the recording stability of this model. Results: The KA dosage test suggested that a 10 µl (1.0 µg/µl) intrahippocampal injection could successfully evoke status epilepticus lasting from 4 to 12 h. At this dosage, eight pigs (50% of total) had prolonged epileptic events (tonic-chronic seizures + interictal spikes n = 5, interictal spikes alone n = 3) in the later 4 weeks of the video-SEEG recording period. Four pigs (25% of total) had no epileptic activities, and another four (25%) had lost the cap or did not complete the experiments. Animals that showed epileptiform events were grouped as E + (n = 8) and the four animals showing no signs of epileptic events were grouped as E- (n = 4). A total of 46 electrophysiological seizures were captured in the 4-week post-KA period from 4 E + animals, with the earliest onset on day 9. The seizure durations ranged from 12 to 45 s. A significant increase of hippocampal HFOs rate (num/min) was observed in the E+ group during the post-KA period (weeks 1, 2,4, p < 0.05) compared to the baseline. But the E-showed no change or a decrease (in week 2, p = 0.43) compared to their baseline rate. The between-group comparison showed much higher HFO rates in E + vs. E - (F = 35, p < 0.01). The high ICC value [ICC (1, k) = 0.81, p < 0.05] quantified from the HFO rate suggested that this model had a stable measurement of HFOs during the four-week post-KA periods. Significance: This study measured intracranial electrophysiological activity in a swine model of KA-induced mesial temporal lobe epilepsy (mTLE). Using the clinical SEEG electrode, we distinguished abnormal EEG patterns in the swine brain. The high test-retest reliability of HFO rates in the post-KA period suggests the utility of this model for studying mechanisms of epileptogenesis. The use of swine may provide satisfactory translational value for clinical epilepsy research.

Pathological neurons generate ripples at the UP-DOWN transition disrupting information transfer.

Objective: To confirm and investigate why pathological HFOs (pHFOs), including Ripples [80-200 Hz] and fast ripples [200-600 Hz], are generated during the UP-DOWN transition of the slow wave and if pHFOs interfere with information transmission. Methods: We isolated 217 total units from 175.95 iEEG contact-hours of synchronized macro- and microelectrode recordings from 6 patients. Sleep slow oscillation (0.1-2 Hz) epochs were identified in the iEEG recording. iEEG HFOs that occurred superimposed on the slow wave were transformed to phasors and adjusted by the phase of maximum firing in nearby units (i.e., maximum UP). We tested whether, in the seizure onset zone (SOZ), HFOs and associated action potentials (AP) occur more often at the UP-DOWN transition. We also examined ripple temporal correlations using cross correlograms. Results: At the group level in the SOZ, HFO and HFO-associated AP probability was highest during the UP-DOWN transition of slow wave excitability (p<<0.001). In the non-SOZ, HFO and HFO-associated AP was highest during the DOWN-UP transition (p<<0.001). At the unit level in the SOZ, 15.6% and 20% of units exhibited more robust firing during ripples (Cohen's d=0.11-0.83) and fast ripples (d=0.36-0.90) at the UP-DOWN transition (p<0.05 f.d.r corrected), respectively. By comparison, also in the SOZ, 6.6% (d=0.14-0.30) and 8.5% (d=0.33-0.41) of units had significantly less firing during ripples and fast ripples at the UP-DOWN transition, respectively. Additional data shows ripple temporal correlations, involving global slow waves, between the hippocampus, entorhinal cortex, and parahippocampal gyrus were reduced by ~50-80% in the SOZ compared to the non-SOZ (N=3). Significance: The UP-DOWN transition of slow wave excitability facilitates the activation of pathological neurons to generate pHFOs. The pathological neurons and pHFOs disrupt ripple temporal correlations across brain regions that transfer information and may be important in memory consolidation.