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BACKGROUND: Chronobiology is the scientific field focused on studying periodicity in biological processes. In mammals, most physiological variables exhibit circadian rhythmicity, such as metabolism, body temperature, locomotor activity, and sleep. The biological rhythmicity can be statistically evaluated by examining the time series and extracting parameters that correlate to the period of oscillation, its amplitude, phase displacement, and overall variability. NEW METHOD: We have developed a library called CircadiPy, which encapsulates methods for chronobiological analysis and data inspection, serving as an open-access toolkit for the analysis and interpretation of chronobiological data. The package was designed to be flexible, comprehensive and scalable in order to assist research dealing with processes affected or influenced by rhythmicity. RESULTS: The results demonstrate the toolkit's capability to guide users in analyzing chronobiological data collected from various recording sources, while also providing precise parameters related to the circadian rhythmicity. COMPARISON WITH EXISTING METHODS: The analysis methodology from this proposed library offers an opportunity to inspect and obtain chronobiological parameters in a straightforward and cost-free manner, in contrast to commercial tools. CONCLUSIONS: Moreover, being an open-source tool, it empowers the community with the opportunity to contribute with new functions, analysis methods, and graphical visualizations given the simplified computational method of time series data analysis using an easy and comprehensive pipeline within a single Python object.

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BACKGROUND: The phytocannabinoid cannabidiol (CBD) has previously shown to have anticonvulsant effects in preclinical and clinical studies. Recently, CBD has been approved to treat certain types of drug-resistant epileptic syndromes. However, the underlying mechanism of action remains unclear. The phosphatidylinositol 3-kinase (PI3K) signaling pathway has been proposed to modulate seizures and might be recruited by CBD. Thus, we tested the hypothesis that the anticonvulsant effect of CBD involves PI3K in a seizure model induced by pentylenetetrazole (PTZ). METHODS: We employed pharmacological and genetic approaches to inhibit PI3K and quantified its effects on seizure duration, latency, and number. RESULTS: PI3K genetic ablation increased the duration and number of seizures. CBD inhibited PTZ-induced seizures in mice. Genetic deletion of PI3K or pretreatment with the selective inhibitor LY294002 prevented CBD effects. CONCLUSION: Our data strengthen the hypothesis that the CBD anticonvulsant effect requires the PI3K signaling pathway.

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Based on the rationale that neural hypersynchronization underlies epileptic phenomena, nonperiodic stimulation (NPS) was designed and successfully tested as an electrical stimulus with robust anticonvulsant action. Considering the scale-free temporal structure of NPS mimics natural-like activity, here we hypothesized its application to the amygdala would induce minor to none impairment of neural function in treated animals. Wistar rats underwent gold-standard behavioral tests such as open field (OF), elevated plus-maze (EPM), novel object recognition, and social interaction test in order to evaluate the functions of base-level anxiety, motor function, episodic memory, and sociability. We also performed daily (8 days, 6 h per day) electrophysiological recordings (local field potential/LFP and electromyography) to assess global forebrain dynamics and the sleep-wake cycle architecture and integrity. All animals displayed an increased proportion of time exploring new objects, spent more time in the closed arms of the EPM and in the periphery of the OF arena, with similar numbers of crossing between quadrants and no significant changes of social behaviors. In the sleep-wake cycle electrophysiology experiments, we found no differences regarding duration and proportion of sleep stages and the number of transitions between stages. Finally, the power spectrum of LFP recordings and neurodynamics were also unaltered. We concluded that NPS did not impair neural functions evaluated and thus, it may be safe for clinical studies. Additionally, results corroborate the notion that NPS may exert an on-demand only desynchronization effect by efficiently competing with epileptiform activity for the physiological and healthy recruitment of neural circuitry. Considering the very dynamical nature of circuit activation and functional activity underlying neural function in general (including cognition, processing of emotion, memory acquisition, and sensorimotor integration) and its corruption leading to disorder, such mechanism of action may have important implications in the investigation of neuropsychological phenomena and also in the development of rehabilitation neurotechnology.

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

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The extraction of electrophysiological features that reliably forecast the occurrence of seizures is one of the most challenging goals in epilepsy research. Among possible approaches to tackle this problem is the use of active probing paradigms in which responses to stimuli are used to detect underlying system changes leading up to seizures. This work evaluates the theoretical and mechanistic underpinnings of this strategy using two coupled populations of the well-studied Wendling neural mass model. Different model settings are evaluated, shifting parameters (excitability, slow inhibition, or inter-population coupling gains) from normal towards ictal states while probing stimuli are applied every 2 seconds to the input of either one or both populations. The correlation between the extracted features and the ictogenic parameter shifting indicates if the impending transition to the ictal state may be identified in advance. Results show that not only can the response to the probing stimuli forecast seizures but this is true regardless of the altered ictogenic parameter. That is, similar feature changes are highlighted by probing stimuli responses in advance of the seizure including: increased response variance and lag-1 autocorrelation, decreased skewness, and increased mutual information between the outputs of both model subsets. These changes were mostly restricted to the stimulated population, showing a local effect of this perturbational approach. The transition latencies from normal activity to sustained discharges of spikes were not affected, suggesting that stimuli had no pro-ictal effects. However, stimuli were found to elicit interictal-like spikes just before the transition to the ictal state. Furthermore, the observed feature changes highlighted by probing the neuronal populations may reflect the phenomenon of critical slowing down, where increased recovery times from perturbations may signal the loss of a systems' resilience and are common hallmarks of an impending critical transition. These results provide more evidence that active probing approaches highlight information about underlying system changes involved in ictogenesis and may be able to play a role in assisting seizure forecasting methods which can be incorporated into early-warning systems that ultimately enable closing the loop for targeted seizure-controlling interventions.

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Memory impairment is the most common cognitive deficit in patients with temporal lobe epilepsy (TLE). This type of epilepsy is currently regarded as a network disease because of its brain-wide alterations in functional connectivity between temporal and extra-temporal regions. In patients with TLE, network dysfunctions can be observed during ictal states, but are also described interictally during rest or sleep. Here, we examined the available literature supporting the hypothesis that hippocampal-cortical coupling during sleep is hijacked in TLE. First, we look at studies showing that the coordination between hippocampal sharp-wave ripples (100-200 Hz), corticothalamic spindles (9-16 Hz), and cortical delta waves (1-4 Hz) during nonrapid eye movement (NREM) sleep is critical for spatial memory consolidation. Then, we reviewed studies showing that animal models of TLE display precise coordination between hippocampal interictal epileptiform discharges (IEDs) and spindle oscillations in the prefrontal cortex. This aberrant oscillatory coupling seems to surpass the physiological ripple-delta-spindle coordination, which could underlie memory consolidation impairments. We also discuss the role of rapid eye movement (REM) sleep for local synaptic plasticity and memory. Sleep episodes of REM provide windows of opportunity for reactivation of expression of immediate early genes (i.e., zif-268 and Arc). Besides, hippocampal theta oscillations during REM sleep seem to be critical for memory consolidation of novel object place recognition task. However, it is still unclear which extend this particular phase of sleep is affected in TLE. In this context, we show some preliminary results from our group, suggesting that hippocampal theta-gamma phase-amplitude coupling is exacerbated during REM in a model of basolateral amygdala fast kindling. In conclusion, there is an increasing body of evidence suggesting that circuits responsible for memory consolidation during sleep seem to be gradually coopted and degraded in TLE. This article is part of the Special Issue "NEWroscience 2018".

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Electrophysiological recordings lead amongst the techniques that aim to investigate the dynamics of neural activity sampled from large neural ensembles. However, the financial costs associated with the state-of-the-art technology used to manufacture probes and multi-channel recording systems make these experiments virtually inaccessible to small laboratories, especially if located in developing countries. Here, we describe a new method for implanting several tungsten electrode arrays, widely distributed over the brain. Moreover, we designed a headstage system, using the Intan® RHD2000 chipset, associated with a connector (replacing the expensive commercial Omnetics connector), that allows the usage of disposable and inexpensive cranial implants. Our results showed high-quality multichannel recording in freely moving animals (detecting local field, evoked responses and unit activities) and robust mechanical connections ensuring long-term continuous recordings. Our project represents an open source and inexpensive alternative to develop customized extracellular records from multiple brain regions.

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The understanding that hyper-excitability and hyper-synchronism in epilepsy are indissociably bound by a cause-consequence relation has only recently been challenged. Thus, therapeutic strategies for seizure suppression have often aimed at inhibiting excitatory circuits and/or activating inhibitory ones. However, new approaches that aim to desynchronize networks or compromise abnormal coupling between adjacent neural circuitry have been proven effective, even at the cost of enhancing local neuronal activation. Although most of these novel perspectives targeting circuitry desynchronization and network coupling have been implemented by non-pharmacological devices, we argue that there may be endogenous neurochemical systems that act primarily in the desynchronization component of network behavior rather than dampening excitability of individual neurons. This review explores the endocannabinoid system as one such possible pharmacological landmark for mimicking a form of "on-demand" desynchronization analogous to those proposed by deep brain stimulation in the treatment of epilepsy. This essay discusses the evidence supporting the role of the endocannabinoid system in modulating the synchronization and/or coupling of distinct local neural circuitry; which presents obvious implications on the physiological setting of proper sensory-motor integration. Accordingly, the process of ictogenesis involves pathological circuit coupling that could be avoided, or at least have its spread throughout the containment of other areas, if such endogenous mechanisms of control could be activated or potentiated by pharmacological intervention. In addition, we will discuss evidence that supports not only a weaker role played on neuronal excitability but the potential of the endocannabinoid system strengthening its modulatory effect, only when circuitry coupling surpasses a level of activation.

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The phosphatidylinositol 3-kinase (PI3K)/protein kinase B (PKB/Akt)/mechanistic target of rapamycin (mTOR) signaling pathway has been associated with several pathologies in the central nervous system (CNS), including epilepsy. There is evidence supporting the hypothesis that the PI3Kγ signaling pathway may mediate the powerful anticonvulsant properties associated with the cannabinoidergic system. This work aims to investigate if the anticonvulsant and neuroprotective effects of cannabidiol (CBD) are mediated by PI3Kγ. In vitro and in vivo experiments were performed on C57Bl/6 wild-type (WT) and PI3Kγ-/- mice. Behavioral seizures were induced by bilateral intra-hippocampal pilocarpine microinjection. Twenty-four hours after the first behavioral seizure, animals were perfused and their brains removed and processed, for histological analysis of neurodegeneration, microgliosis and astrocytosis. Primary cultures of hippocampal neurons were used for glutamate-induced cell death assay. CDB increased latency and reduced the severity of pilocarpine-induced behavioral seizures, as well as prevented postictal changes, such as neurodegeneration, microgliosis and astrocytosis, in WT animals, but not in PI3Kγ-/-. CBD in vivo effects were abolished by pharmacological inhibition of cannabinoid receptor or mTOR. In vitro, PI3Kγ inhibition or deficiency also changed CBD protection observed in glutamate-induced cell death assay. Thus, we suggest that the modulation of PI3K/mTOR signaling pathway is involved in the anticonvulsant and neuroprotective effects of CBD. These findings are important not only for the elucidation of the mechanisms of action of CBD, which are currently poorly understood, but also to allow the prediction of therapeutic and side effects, ensuring efficacy and safety in the treatment of patients with epilepsy.

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Animal behavioral paradigms, such as classical conditioning and operant conditioning, are an important tool to study the neural basis of cognition and behavior. These paradigms involve manipulating sensory stimuli in a way that learning processes are induced under controlled experimental conditions. However, the majority of the commercially available equipment did not offer flexibility to manipulate stimuli. Therefore, the development of most versatile devices would allow the study of more complex cognitive functions. The purpose of this work is to present a low-cost, customized and wireless-operated chamber for animal behavior conditioning, based on the joint operation of two microcontroller modules: Arduino Due and ESP8266-12E. Our results showed that the auditory stimulation system allows setting the carrier frequency in the range of 1 Hz up to more than 100 kHz and the sound stimulus can be modulated in amplitude, also over a wide range of frequencies. Likewise, foot-shock could be precisely manipulated regarding its amplitude (from ∼200 µA to ∼1500 µA) and frequency (up to 20 pulses per second). Finally, adult rats exposed to a protocol of cued fear conditioning in our device showed consistent behavioral response and electrophysiological evoked responses in the midbrain auditory pathway. Furthermore, the device developed in the current study represents an open source alternative to develop customized protocols to study fear memory under conditions of varied sensory stimuli.

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Evidence suggests that the pathophysiology associated with epileptic susceptibility may disturb the functional connectivity of neural circuits and compromise the brain functions, even when seizures are absent. Although memory impairment is a common comorbidity found in patients with epilepsy, it is still unclear whether more caudal structures may play a role in cognitive deficits, particularly in those cases where there is no evidence of hippocampal sclerosis. This work used a genetically selected rat strain for seizure susceptibility (Wistar audiogenic rat, WAR) and distinct behavioral (motor and memory-related tasks) and electrophysiological (inferior colliculus, IC) approaches to access acoustic primary integrative network properties. The IC neural assemblies' response was evaluated by auditory transient (focusing on bottom-up processing) and steady-state evoked response (ASSR, centering on feedforward and feedback forces over neural circuitry). The results show that WAR displayed no disturbance in motor performance or hippocampus-dependent memory tasks. Nonetheless, WAR animals exhibited significative impairment for auditory fear conditioning (AFC) along with no indicative of IC plastic changes between the pre-conditioning and test phases (ASSR coherence analysis). Furthermore, WAR's IC response to transient stimuli presented shorter latency and higher amplitude compared with Wistar; and the ASSR analysis showed similar results for WAR and Wistar animals under subthreshold dose of pentylenetetrazol (pro-convulsive drug) for seizure-induction. Our work demonstrated alterations at WAR IC neural network processing, which may explain the associated disturbance on AFC memory.

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Neuroimmune interactions underlying the development of pain sensitization in models of neuropathic pain have been widely studied. In this study, we evaluated the development of allodynia and its reduction associated with peripheral antineuroinflammatory effects induced by a dexamethasone-loaded biodegradable implant. Chronic constriction injury (CCI) of the sciatic nerve was performed in Wistar rats. The electronic von Frey test was applied to assess mechanical allodynia. The dexamethasone-loaded implant was placed perineurally at the moment of CCI or 12 days after surgery. Dorsal root ganglia (DRG; L4-L5) were harvested and nuclear extracts were assayed by Western blot for detection of nuclear factor (NF)-κB p65/RelA translocation. Dexamethasone delivered from the implant delayed the development of allodynia for approximately three weeks in CCI rats when the implantation was performed at day 0, but allodynia was not reversed when the implantation was performed at day 12. NF-κB was activated in CCI rat DRG compared with naïve or sham animals (day 15), and dexamethasone implant inhibited p65/RelA translocation in CCI rats compared with control. This study demonstrated that the dexamethasone-loaded implant suppresses allodynia development and peripheral neuroinflammation. This device can reduce the potential side effects associated with oral anti-inflammatory drugs.

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The Wistar Audiogenic Rat (WAR) is a model whose rats are predisposed to develop seizures following acoustic stimulation. We aimed to establish the transcriptional profile of the WAR model, searching for genes that help in understanding the molecular mechanisms involved in the predisposition and seizures expression of this strain. RNA-Seq of the corpora quadrigemina of WAR and Wistar rats subjected to acoustic stimulation revealed 64 genes differentially regulated in WAR. We validated twelve of these genes by qPCR in stimulated and naive (non-stimulated) WAR and Wistar rats. Among these, Acsm3 was upregulated in WAR in comparison with both control groups. In contrast, Gpr126 and Rtel1 were downregulated in naive and stimulated WAR rats in comparison with the Wistar controls. Qdpr was upregulated only in stimulated WAR rats that exhibited audiogenic seizures. Our data show that there are genes with differential intrinsic regulation in the WAR model and that seizures can alter gene regulation. We identified new genes that might be involved in the epileptic phenotype and comorbidities of the WAR model.

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PURPOSE: To combine the technique of respiratory gating and compressed sensing (CS) with the objective of accelerating mouse abdominal magnetic resonance imaging (MRI). MATERIALS AND METHODS: To obtain the maximum acceleration, phase-encoding data from a phantom and mouse were obtained on a 4.7 Tesla scanner using the respiratory gating technique. The fully sampled data (FSD) were used to construct reference images and to provide samples to simulate retrospective undersampled data (UD) acquisition using respiratory gating. The UD and 95% of the UD on acceleration 2-5 rates were acquired and used for image reconstruction by CS. Quantitative assessment of reconstructed images was performed by structural similarity index (SSIM), peak signal-to-noise ratio (PSNR) and root mean square error (RMSE). RESULTS: The proposed method can accelerate phantom and mouse abdominal MRI acquisition between 2 and 4 rates by reducing the amount of FSD. For phantom UD acquisition, the mean time was reduced in 45.9% and for the acquisition of 95% of UD in 67.8%. For mouse abdominal image UD acquisition, the mean time was reduced in 44.6% and for the acquisition of 95% of UD in 62.5%. The metrics results show that the reconstructed image from UD and 95% of UD by using CS maintains an optimal agreement with their reference images (similarity above 0.88 for phantom and 0.93 for mouse). CONCLUSION: This study presents a novel approach to accelerate mouse abdominal MRI combining respiratory gating technique and CS without the use of expensive hardware and capable of achieving up to 4 acceleration rate without image degradation.

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The brain oscillations may play a critical role in synchronizing neuronal assemblies in order to establish appropriate sensory-motor integration. In fact, studies have demonstrated phase-amplitude coupling of distinct oscillatory rhythms during cognitive processes. Here we investigated whether olfacto-hippocampal coupling occurs when mice are detecting familiar odors located in a spatially restricted area of a new context. The spatial olfactory task (SOT) was designed to expose mice to a new environment in which only one quadrant (target) contains odors provided by its own home-cage bedding. As predicted, mice showed a significant higher exploration preference to the target quadrant; which was impaired by olfactory epithelium lesion (ZnSO4). Furthermore, mice were able to discriminate odors from a different cage and avoided the quadrant with predator odor 2,4,5-trimethylthiazoline (TMT), reinforcing the specificity of the SOT. The local field potential (LFP) analysis of non-lesioned mice revealed higher gamma activity (35-100Hz) in the main olfactory bulb (MOB) and a significant theta phase/gamma amplitude coupling between MOB and dorsal hippocampus, only during exploration of home-cage odors (i.e. in the target quadrant). Our results suggest that exploration of familiar odors in a new context involves dynamic coupling between the olfactory bulb and dorsal hippocampus.

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The rat inferior colliculus (IC) is a major midbrain relay for ascending inputs from the auditory brain stem and has been suggested to play a key role in the processing of aversive sounds. Previous studies have demonstrated that auditory fear conditioning (AFC) potentiates transient responses to brief tones in the IC, but it remains unexplored whether AFC modifies responses to sustained periodic acoustic stimulation-a type of response called the steady-state evoked potential (SSEP). Here we used an amplitude-modulated tone-a 10-kHz tone with a sinusoidal amplitude modulation of 53.7 Hz-as the conditioning stimulus (CS) in an AFC protocol (5 CSs per day in 3 consecutive days) while recording local field potentials (LFPs) from the IC. In the preconditioning session (day 1), the CS elicited prominent 53.7-Hz SSEPs. In the training session (day 2), foot shocks occurred at the end of each CS (paired group) or randomized in the inter-CS interval (unpaired group). In the test session (day 3), SSEPs markedly differed from preconditioning in the paired group: in the first two trials the phase to which the SSEP coupled to the CS amplitude envelope shifted ~90°; in the last two trials the SSEP power and the coherence of SSEP with the CS amplitude envelope increased. LFP power decreased in frequency bands other than 53.7 Hz. In the unpaired group, SSEPs did not change in the test compared with preconditioning. Our results show that AFC causes dissociated changes in the phase and power of SSEP in the IC.NEW & NOTEWORTHY Local field potential oscillations in the inferior colliculus follow the amplitude envelope of an amplitude-modulated tone, originating a neural response called the steady-state evoked potential. We show that auditory fear conditioning of an amplitude-modulated tone modifies two parameters of the steady-state evoked potentials in the inferior colliculus: first the phase to which the evoked oscillation couples to the amplitude-modulated tone shifts; subsequently, the evoked oscillation power increases along with its coherence with the amplitude-modulated tone.

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Epilepsy is a neurological disease related to the occurrence of pathological oscillatory activity, but the basic physiological mechanisms of seizure remain to be understood. Our working hypothesis is that specific sensory processing circuits may present abnormally enhanced predisposition for coordinated firing in the dysfunctional brain. Such facilitated entrainment could share a similar mechanistic process as those expediting the propagation of epileptiform activity throughout the brain. To test this hypothesis, we employed the Wistar audiogenic rat (WAR) reflex animal model, which is characterized by having seizures triggered reliably by sound. Sound stimulation was modulated in amplitude to produce an auditory steady-state-evoked response (ASSR; -53.71Hz) that covers bottom-up and top-down processing in a time scale compatible with the dynamics of the epileptic condition. Data from inferior colliculus (IC) c-Fos immunohistochemistry and electrographic recordings were gathered for both the control Wistar group and WARs. Under 85-dB SLP auditory stimulation, compared to controls, the WARs presented higher number of Fos-positive cells (at IC and auditory temporal lobe) and a significant increase in ASSR-normalized energy. Similarly, the 110-dB SLP sound stimulation also statistically increased ASSR-normalized energy during ictal and post-ictal periods. However, at the transition from the physiological to pathological state (pre-ictal period), the WAR ASSR analysis demonstrated a decline in normalized energy and a significant increase in circular variance values compared to that of controls. These results indicate an enhanced coordinated firing state for WARs, except immediately before seizure onset (suggesting pre-ictal neuronal desynchronization with external sensory drive). These results suggest a competing myriad of interferences among different networks that after seizure onset converge to a massive oscillatory circuit.

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Accumulating evidence from different animal models has contributed to the understanding of the bidirectional comorbidity associations between the epileptic condition and behavioral abnormalities. A strain of animals inbred to enhance seizure predisposition to high-intensity sound stimulation, the Wistar audiogenic rat (WAR), underwent several behavioral tests: forced swim test (FST), open-field test (OFT), sucrose preference test (SPT), elevated plus maze (EPM), social preference (SP), marble burying test (MBT), inhibitory avoidance (IAT), and two-way active avoidance (TWAA). The choice of tests aimed to investigate the correlation between underlying circuits believed to be participating in both WAR's innate susceptibility to sound-triggered seizures and the neurobiological substrates associated with test performance. Comparing WAR with its Wistar counterpart (i.e., resistant to audiogenic seizures) showed that WARs present behavioral despair traits (e.g., increased FST immobility) but no evidence of anhedonic behavior (e.g., increased sucrose consumption in SPT) or social impairment (e.g., no difference regarding juvenile exploration in SP). In addition, tests suggested that WARs are unable to properly evaluate degrees of aversiveness (e.g., performance on OFT, EPM, MBT, IAT, and TWAA). The particularities of the WAR model opens new venues to further untangle the neurobiology underlying the co-morbidity of behavioral disorders and epilepsy. This article is part of a Special Issue entitled "Genetic and Reflex Epilepsies, Audiogenic Seizures and Strains: From Experimental Models to the Clinic".

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Temporal lobe epilepsy (TLE) is the most common form of epilepsy in adults. The pilocarpine (PILO) experimental model of TLE portrays behavioral and pathophysiological changes in rodents that are very similar to those found in humans with TLE. However, this model is associated with an unfortunate high mortality rate. Studies have shown that intrahippocampal injection of PILO, while having a much smaller mortality rate, induces status epilepticus (SE) that secondarily leads to TLE. To the best of our knowledge, the present study was the first to evaluate the cognitive and histological alterations 72h after intrahippocampal microinjection of PILO in C57BL/6 mice. Seventy percent of mice developed status epilepticus (SE) after PILO administration, and all animals survived after SE. Seventy-two hours after SE, mice presented memory impairment in both Novel Object Recognition (recognition index - vehicle: 67.57±4.46% vs PILO: 52.33±3.29%) and Contextual Fear Conditioning (freezing time - vehicle: 203±20.43 vs PILO: 107.80±25.17s) tasks. Moreover, using Nissl and NeuN staining, we observed in PILO-treated mice a significant decrease in cell viability and an increase in neuronal loss in all three hippocampal regions analyzed, cornus ammonis (CA) 1, CA3, and dentate gyrus (DG), in comparison with the control group. Additionally, using Iba-1 staining, we observed in PILO-treated mice a significant increase in microglial proliferation in CA1, CA3, and DG of the hippocampus. Therefore, intrahippocampal PILO microinjection is an efficient route to induce SE and acute postictal epileptogenic-like alterations in C57BL/6 mice.

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Many patients with epilepsy do not obtain proper control of their seizures through conventional treatment. We review aspects of the pathophysiology underlying epileptic phenomena, with a special interest in the role of the amygdala, stressing the importance of hypersynchronism in both ictogenesis and epileptogenesis. We then review experimental studies on electrical stimulation of mesiotemporal epileptogenic areas, the amygdala included, as a means to treat medically refractory epilepsy. Regular high-frequency stimulation (HFS) commonly has anticonvulsant effects and sparse antiepileptogenic properties. On the other hand, HFS is related to acute and long-term increases in excitability related to direct neuronal activation, long-term potentiation, and kindling, raising concerns regarding its safety and jeopardizing in-depth understanding of its mechanisms. In turn, the safer regular low-frequency stimulation (LFS) has a robust antiepileptogenic effect, but its pro- or anticonvulsant effect seems to vary at random among studies. As an alternative, studies by our group on the development and investigation of temporally unstructured electrical stimulation applied to the amygdala have shown that nonperiodic stimulation (NPS), which is a nonstandard form of LFS, is capable of suppressing both acute and chronic spontaneous seizures. We hypothesize two noncompetitive mechanisms for the therapeutic role of amygdala in NPS, 1) a direct desynchronization of epileptic circuitry in the forebrain and brainstem and 2) an indirect desynchronization/inhibition through nucleus accumbens activation. We conclude by reintroducing the idea that hypersynchronism, rather than hyperexcitability, may be the key for epileptic phenomena and epilepsy treatment.

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