Stimulation Of Amygdala Research Articles

Deep brain stimulation of the Tbr1-deficient mouse model of autism spectrum disorder at the basolateral amygdala alters amygdalar connectivity, whole-brain synchronization, and social behaviors.

Autism spectrum disorders (ASDs) are considered neural dysconnectivity syndromes. To better understand ASD and uncover potential treatments, it is imperative to know and dissect the connectivity deficits under conditions of autism. Here, we apply a whole-brain immunostaining and quantification platform to demonstrate impaired structural and functional connectivity and aberrant whole-brain synchronization in a Tbr1+/- autism mouse model. We express a channelrhodopsin variant oChIEF fused with Citrine at the basolateral amygdala (BLA) to outline the axonal projections of BLA neurons. By activating the BLA under blue light theta-burst stimulation (TBS), we then evaluate the effect of BLA activation on C-FOS expression at a whole brain level to represent neural activity. We show that Tbr1 haploinsufficiency almost completely disrupts contralateral BLA axonal projections and results in mistargeting in both ipsilateral and contralateral hemispheres, thereby globally altering BLA functional connectivity. Based on correlated C-FOS expression among brain regions, we further show that Tbr1 deficiency severely disrupts whole-brain synchronization in the absence of salient stimulation. Tbr1+/- and wild-type (WT) mice exhibit opposing responses to TBS-induced amygdalar activation, reducing synchronization in WT mice but enhancing it in Tbr1+/- mice. Whole-brain modular organization and intermodule connectivity are also affected by Tbr1 deficiency and amygdalar activation. Following BLA activation by TBS, the synchronizations of the whole brain and the default mode network, a specific subnetwork highly relevant to ASD, are enhanced in Tbr1+/- mice, implying a potential ameliorating effect of amygdalar stimulation on brain function. Indeed, TBS-mediated BLA activation increases nose-to-nose social interactions of Tbr1+/- mice, strengthening evidence for the role of amygdalar connectivity in social behaviors. Our high-resolution analytical platform reveals the inter- and intrahemispheric connectopathies arising from ASD. Our study emphasizes the defective synchronization at a whole-brain scale caused by Tbr1 deficiency and implies a potential beneficial effect of deep brain stimulation at the amygdala for TBR1-linked autism.

Quantitative cellular pathology of the amygdala in temporal lobe epilepsy and correlation with magnetic resonance imaging volumetry, tissue microstructure, and sudden unexpected death in epilepsy risk factors.

Amygdala enlargement can occur in temporal lobe epilepsy, and increased amygdala volume is also reported in sudden unexpected death in epilepsy (SUDEP). Apnea can be induced by amygdala stimulation, and postconvulsive central apnea (PCCA) and generalized seizures are both known SUDEP risk factors. Neurite orientation dispersion and density imaging (NODDI) has recently provided additional information on altered amygdala microstructure in SUDEP. In a series of 24 surgical temporal lobe epilepsy cases, our aim was to quantify amygdala cellular pathology parameters that could predict enlargement, NODDI changes, and ictal respiratory dysfunction. Using whole slide scanning automated quantitative image analysis methods, parallel evaluation of myelin, axons, dendrites, oligodendroglia, microglia, astroglia, neurons, serotonergic networks, mTOR-pathway activation (pS6) and phosphorylated tau (pTau; AT8, AT100, PHF) in amygdala, periamygdala cortex, and white matter regions of interest were compared with preoperative magnetic resonance imaging data on amygdala size, and in 13 cases with NODDI and evidence of ictal-associated apnea. We observed significantly higher glial labeling (Iba1, glial fibrillary acidic protein, Olig2) in amygdala regions compared to cortex and a strong positive correlation between Olig2 and Iba1 in the amygdala. Larger amygdala volumes correlated with lower microtubule-associated protein (MAP2), whereas higher NODDI orientation dispersion index correlated with lower Olig2 cell densities. In the three cases with recorded PCCA, higher MAP2 and pS6-235 expression was noted than in those without. pTau did not correlate with SUDEP risk factors, including seizure frequency. Histological quantitation of amygdala microstructure can shed light on enlargement and diffusion imaging alterations in epilepsy to explore possible mechanisms of amygdala dysfunction, including mTOR pathway activation, that in turn may increase the risk for SUDEP.

Single-pulse electrical stimulation artifact removal using the novel matching pursuit-based artifact reconstruction and removal method (MPARRM)

Objective. Single-pulse electrical stimulation (SPES) has been widely used to probe effective connectivity. However, analysis of the neural response is often confounded by stimulation artifacts. We developed a novel matching pursuit-based artifact reconstruction and removal method (MPARRM) capable of removing artifacts from stimulation-artifact-affected electrophysiological signals. Approach. To validate MPARRM across a wide range of potential stimulation artifact types, we performed a bench-top experiment in which we suspended electrodes in a saline solution to generate 110 types of real-world stimulation artifacts. We then added the generated stimulation artifacts to ground truth signals (stereoelectroencephalography signals from nine human subjects recorded during a receptive speech task), applied MPARRM to the combined signal, and compared the resultant denoised signal with the ground truth signal. We further applied MPARRM to artifact-affected neural signals recorded from the hippocampus while performing SPES on the ipsilateral basolateral amygdala in nine human subjects. Main results. MPARRM could remove stimulation artifacts without introducing spectral leakage or temporal spread. It accommodated variable stimulation parameters and recovered the early response to SPES within a wide range of frequency bands. Specifically, in the early response period (5–10 ms following stimulation onset), we found that the broadband gamma power (70–170 Hz) of the denoised signal was highly correlated with the ground truth signal ( R=0.98±0.02 , Pearson), and the broadband gamma activity of the denoised signal faithfully revealed the responses to the auditory stimuli within the ground truth signal with 94%±1.47% sensitivity and 99%±1.01% specificity. We further found that MPARRM could reveal the expected temporal progression of broadband gamma activity along the anterior-posterior axis of the hippocampus in response to the ipsilateral amygdala stimulation. Significance. MPARRM could faithfully remove SPES artifacts without confounding the electrophysiological signal components, especially during the early-response period. This method can facilitate the understanding of the neural response mechanisms of SPES.

Failure to breathe persists without air hunger or alarm following amygdala seizures.

Postictal apnea is thought to be a major cause of sudden unexpected death in epilepsy (SUDEP). However, the mechanisms underlying postictal apnea are unknown. To understand causes of postictal apnea, we used a multimodal approach to study brain mechanisms of breathing control in 20 patients (ranging from pediatric to adult) undergoing intracranial electroencephalography for intractable epilepsy. Our results indicate that amygdala seizures can cause postictal apnea. Moreover, we identified a distinct region within the amygdala where electrical stimulation was sufficient to reproduce prolonged breathing loss persisting well beyond the end of stimulation. The persistent apnea was resistant to rising CO2 levels, and air hunger failed to occur, suggesting impaired CO2 chemosensitivity. Using es-fMRI, a potentially novel approach combining electrical stimulation with functional MRI, we found that amygdala stimulation altered blood oxygen level-dependent (BOLD) activity in the pons/medulla and ventral insula. Together, these findings suggest that seizure activity in a focal subregion of the amygdala is sufficient to suppress breathing and air hunger for prolonged periods of time in the postictal period, likely via brainstem and insula sites involved in chemosensation and interoception. They further provide insights into SUDEP, may help identify those at greatest risk, and may lead to treatments to prevent SUDEP.

Discovering how the amygdala shapes human behavior: From lesion studies to neuromodulation

Case studies of patients with amygdala damage or those receiving direct amygdala stimulation have informed our understanding of the amygdala's role in emotion and cognition. These foundational studies illustrate how the human amygdala influences our present behavior and prioritizes memories of our past in service of future experiences. This broad influence makes the amygdala a novel target for clinical neuromodulation.

Signatures of Electrical Stimulation Driven Network Interactions in the Human Limbic System.

Stimulation-evoked signals are starting to be used as biomarkers to indicate the state and health of brain networks. The human limbic network, often targeted for brain stimulation therapy, is involved in emotion and memory processing. Previous anatomic, neurophysiological, and functional studies suggest distinct subsystems within the limbic network (Rolls, 2015). Studies using intracranial electrical stimulation, however, have emphasized the similarities of the evoked waveforms across the limbic network. We test whether these subsystems have distinct stimulation-driven signatures. In eight patients (four male, four female) with drug-resistant epilepsy, we stimulated the limbic system with single-pulse electrical stimulation. Reliable corticocortical evoked potentials (CCEPs) were measured between hippocampus and the posterior cingulate cortex (PCC) and between the amygdala and the anterior cingulate cortex (ACC). However, the CCEP waveform in the PCC after hippocampal stimulation showed a unique and reliable morphology, which we term the "limbic Hippocampus-Anterior nucleus of the thalamus-Posterior cingulate, HAP-wave." This limbic HAP-wave was visually distinct and separately decoded from the CCEP waveform in ACC after amygdala stimulation. Diffusion MRI data show that the measured end points in the PCC overlap with the end points of the parolfactory cingulum bundle rather than the parahippocampal cingulum, suggesting that the limbic HAP-wave may travel through fornix, mammillary bodies, and the anterior nucleus of the thalamus (ANT). This was further confirmed by stimulating the ANT, which evoked the same limbic HAP-wave but with an earlier latency. Limbic subsystems have unique stimulation-evoked signatures that may be used in the future to help network pathology diagnosis.SIGNIFICANCE STATEMENT The limbic system is often compromised in diverse clinical conditions, such as epilepsy or Alzheimer's disease, and characterizing its typical circuit responses may provide diagnostic insight. Stimulation-evoked waveforms have been used in the motor system to diagnose circuit pathology. We translate this framework to limbic subsystems using human intracranial stereo EEG (sEEG) recordings that measure deeper brain areas. Our sEEG recordings describe a stimulation-evoked waveform characteristic to the memory and spatial subsystem of the limbic network that we term the "limbic HAP-wave." The limbic HAP-wave follows anatomic white matter pathways from hippocampus to thalamus to the posterior cingulum and shows promise as a distinct biomarker of signaling in the human brain memory and spatial limbic network.

Stimulation-induced respiratory enhancement in corticothalamic regions.

We aimed to identify corticothalamic areas and electrical stimulation paradigms that optimally enhance breathing. Twenty-nine patients with medically intractable epilepsy were prospectively recruited in an epilepsy monitoring unit while undergoing stereoelectroencephalographic evaluation. Direct electrical stimulation in cortical and thalamic regions was carried out using low (<1 Hz) and high (≥10 Hz) frequencies, and low (<5 mA) and high (≥5 mA) current intensities, with pulse width of .1 ms. Electrocardiography, arterial oxygen saturation (SpO2 ), end-tidal carbon dioxide (ETCO2 ), oronasal airflow, and abdominal and thoracic plethysmography were monitored continuously during stimulations. Airflow signal was used to estimate breathing rate, tidal volume, and minute ventilation (MV) changes during stimulation, compared to baseline. Electrical stimulation increased MV in the amygdala, anterior cingulate, anterior insula, temporal pole, and thalamus, with an average increase in MV of 20.8% ± 28.9% (range = 0.2%-165.6%) in 19 patients. MV changes were associated with SpO2 and ETCO2 changes (p < .001). Effects on respiration were parameter and site dependent. Within amygdala, low-frequency stimulation of the medial region produced 78.49% greater MV change (p < .001) compared to high-frequency stimulation. Longer stimulation produced greater MV changes (an increase of 4.47% in MV for every additional 10 s, p = .04). Stimulation of amygdala, anterior cingulate gyrus, anterior insula, temporal pole, and thalamus, using certain stimulation paradigms, enhances respiration. Among tested paradigms, low-frequency, low-intensity, long-duration stimulation of the medial amygdala is the most effective breathing enhancement stimulation strategy. Such approaches may pave the way for the future development of neuromodulatory techniques that aid rescue from seizure-related apnea, potentially as a targeted sudden unexpected death in epilepsy prevention method.

Integrative roles of human amygdala subdivisions: Insight from direct intracerebral stimulations via stereotactic EEG.

Substantial studies of human amygdala function have revealed its importance in processing emotional experience, autonomic regulation, and sensory information; however, the neural substrates and circuitry subserving functions have not been directly mapped at the level of the subnuclei in humans. We provide a useful overview of amygdala functional characterization by using direct electrical stimulation to various amygdala regions in 48 patients with drug-resistant epilepsy undergoing stereoelectroencephalography recordings. This stimulation extends beyond the anticipated emotional, neurovegetative, olfactory, and somatosensory responses to include visual, auditory, and vestibular sensations, which may be explained by the functional connectivity with cortical and subcortical regions due to evoked amygdala-cortical potentials. Among the physiological symptom categories for each subnucleus, the most frequently evoked neurovegetative symptoms were distributed in almost every subnucleus. Laterobasal subnuclei are mainly associated with emotional responses, somatosensory responses, and vestibular sensations. Superficial subnuclei are mainly associated with emotional responses and olfactory and visual hallucinations. Our findings contribute to a better understanding of the functional architecture of the human amygdala at the subnuclei level and as a mechanistic basis for the clinical practice of amygdala stimulation in treating patients with neuropsychiatric disorders.

Focal seizures producing loss of wakefulness without ictal asystole: Does temporal lobe syncope exist?

Focal seizures are frequently associated with alteration of consciousness, mainly of awareness, rather than with complete loss of wakefulness. We aimed to explore whether episodes of complete loss of wakefulness (LOW) could be attributed to focal seizures alone, out of the context of ictal asystole or secondary generalization. From a database of adult patients with refractory, focal epilepsy, evaluated for presurgical work-up we searched for patients having the following criteria: (1) focal epilepsy, and (2) transient loss of consciousness, documented in video EEG or/and video SEEG, characterized by an alteration in the level of wakefulness ("syncope like", LOW), with eye closure, hypotonia and non-reactivity state. Patients with motor signs of secondary generalization and patients with non-epileptic psychogenic seizures were excluded. Fifteen patients with transient ictal LOW during focal seizures were found. Among them, 12 patients had ictal asystole. We found 3 patients who had complete loss of wakefulness during focal seizures, without asystole or documented hypotension. In two patients the episodes were provoked by high frequency stimulation of hippocampus and amygdala. The third patient had LOW appearing during a spontaneous temporal lobe seizure. Syncope semiology without ictal asystole can be attributed to temporal lobe seizures but remains an exceptional phenomenon. A crucial clinical requirement is the exclusion of cardiac arrhythmias, especially asystole.

Using a Bipolar Electrode to Create a Temporal Lobe Epilepsy Mouse Model by Electrical Kindling of the Amygdala

The amygdala is one of the most common origins of seizures, and the amygdala mouse model is essential for the illustration of epilepsy. However, few studies have described the experimental protocol in detail. This paper illustrates the whole process of amygdala electrical kindling epilepsy model making, with the introduction of a method of bipolar electrode fabrication. This electrode can both stimulate and record, reducing brain injury caused by implanting separate electrodes for stimulation and recording. For long-term electroencephalogram (EEG) recording purposes, slip rings were used to eliminate the record interruption caused by cable tangles and falling off. After periodic stimulation (60 Hz, 1 s every 15 min) of the basolateral amygdala (AP: 1.67 mm, L: 2.7 mm, V: 4.9 mm) for 19.83 ± 5.742 times, full kindling was observed in six mice (defined as induction of three continuous grade V episodes classified by Racine's scale). An intracranial EEG was recorded throughout the entire kindling process, and an epileptic discharge in the amygdala lasting 20-70 s was observed after kindling. Therefore, this is a robust protocol for modeling epilepsy originating from the amygdala, and the method is suitable for revealing the role of the amygdala in temporal lobe epilepsy. This research contributes to future studies on the mechanisms of mesial temporal lobe epilepsy and novel antiepileptogenic drugs.

Effects of electrical stimulation and temporary inactivation of basolateral amygdala on morphine-induced conditioned place preference in rats

In the present study, to evaluate the role of the basolateral amygdala (BLA) in morphine addiction, the BLA was stimulated electrically, or inactivated temporarily using lidocaine. The electrical stimulation (ES) was delivered to BLA with low or high intensities (LI or HI: 25 or 150 µA, respectively), and five minutes before morphine administration with effective or ineffective doses, lidocaine was microinjected into the BLA. Using a 5-day conditioned place preference (CPP) paradigm, the dependence on morphine was evaluated. The results showed that LI-ES of BLA induced CPP in both the acquisition and expression phases, in the control and the ineffective dose of morphine groups. HI-ES had no effect on CPP acquisition but induced aversion in the expression, with both effective and ineffective doses of morphine. Inactivation of BLA using lidocaine, inhibited morphine-induced CPP in both acquisition and expression phases. The results of the present study indicate the prominent role of BLA in morphine addiction and dependence. Considering the contradictory results of different intensities of ES, it can be inferred that there are different neural circuits in this area of the brain, in relation to the reward responses.

Modulation of Emotion Perception via Amygdala Stimulation in Humans.

BackgroundMultiple lines of evidence show that the human amygdala is part of a neural network important for perception of emotion from environmental stimuli, including for processing of intrinsic attractiveness/“goodness” or averseness/“badness,” i.e., affective valence.Objective/HypothesisWith this in mind, we investigated the effect of electrical brain stimulation of the human amygdala on perception of affective valence of images taken from the International Affective Picture Set (IAPS).MethodsUsing intracranial electrodes in patients with epilepsy, we first obtained event-related potentials (ERPs) in eight patients as they viewed IAPS images of varying affective valence. Next, in a further cohort of 10 patients (five female and five male), we measured the effect of 50 Hz electrical stimulation of the left amygdala on perception of affective valence from IAPS images.ResultsWe recorded distinct ERPs from the left amygdala and found significant differences in the responses between positively and negatively valenced stimuli (p = 0.002), and between neutral and negatively valenced stimuli (p = 0.017) 300–500 ms after stimulus onset. Next, we found that amygdala stimulation did not significantly affect how patients perceived valence for neutral images (p = 0.58), whereas stimulation induced patients to report both positively (p = 0.05) and negatively (< 0.01) valenced images as more neutral.ConclusionThese results render further evidence that the left amygdala participates in a neural network for perception of emotion from environmental stimuli. These findings support the idea that electrical stimulation disrupts this network and leads to partial disruption of perception of emotion. Harnessing this effect may have clinical implications in treatment of certain neuropsychiatric disorders using deep brain stimulation (DBS) and neuromodulation.

The Dostoyevsky effect: epileptogenesis and memory enhancement after kindling stimulation in the primate basolateral amygdala.

Kindling is an electrical stimulation technique used to lower the threshold for epileptogenic activity in the brain. It can also be used as a tool to investigate electrophysiologic alterations that occur as a result of seizures. Epileptiform activity, like seizures and after-discharges (AD; evoked epileptiform activity), commonly cause memory impairment but rarely, can elicit vivid memory retrieval. We kindled the basolateral amygdala of a non-human primate (NHP) once weekly and had him perform a spatial memory task in a 3D virtual environment before, during and after kindling. AD were associated with an initial average performance increase of 46.6%. The enhancement which followed AD persisted up to 2 days. Memory task performance enhancement was accompanied by significant resetting of hippocampal theta oscillations and robust hippocampal potentiation as measured by field evoked potentials. However, neither lasted throughout the duration of performance enhancement. Sharp-wave ripples (SWR), a local field event that supports episodic memory, were generated more often throughout the period of enhancement. SWR rate increased from 14.38 SWR per min before kindling to 24.22 SWR per min after kindling on average. Our results show that kindling can be associated with improved memory. Memory function appears to depend on the particular induction circuit and the resultant excitation/inhibition ratio of the mesial temporal lobe network. Investigating the electrophysiologic underpinnings of this observed memory enhancement is an important step towards understanding the network alterations that occur after seizures and stimulation.Clinical Relevance- Our findings provide new insight into the effects of kindling stimulation in the primate brain. Kindling can cause increase MTL synchrony and the frequency of spontaneous seizures in a primate. This work highlights important considerations for therapeutic deep brain stimulation.

Identifying the neurophysiological effects of memory-enhancing amygdala stimulation using interpretable machine learning

BackgroundDirect electrical stimulation of the amygdala can enhance declarative memory for specific events. An unanswered question is what underlying neurophysiological changes are induced by amygdala stimulation. ObjectiveTo leverage interpretable machine learning to identify the neurophysiological processes underlying amygdala-mediated memory, and to develop more efficient neuromodulation technologies. MethodPatients with treatment-resistant epilepsy and depth electrodes placed in the hippocampus and amygdala performed a recognition memory task for neutral images of objects. During the encoding phase, 160 images were shown to patients. Half of the images were followed by brief low-amplitude amygdala stimulation. For local field potentials (LFPs) recorded from key medial temporal lobe structures, feature vectors were calculated by taking the average spectral power in canonical frequency bands, before and after stimulation, to train a logistic regression classification model with elastic net regularization to differentiate brain states. ResultsClassifying the neural states at the time of encoding based on images subsequently remembered versus not-remembered showed that theta and slow-gamma power in the hippocampus were the most important features predicting subsequent memory performance. Classifying the post-image neural states at the time of encoding based on stimulated versus unstimulated trials showed that amygdala stimulation led to increased gamma power in the hippocampus. ConclusionAmygdala stimulation induced pro-memory states in the hippocampus to enhance subsequent memory performance. Interpretable machine learning provides an effective tool for investigating the neurophysiological effects of brain stimulation.

Basolateral amygdala stimulation plus water maze training restore dentate gyrus LTP and improve spatial learning and memory

Synaptic plasticity is a key mechanism of neural plasticity involved in learning and memory. A reduced or impaired synaptic plasticity could lead to a deficient learning and memory. On the other hand, besides reducing hipocampal dependent learning and memory, fimbria-fornix lesion affects LTP. However, we have consistently shown that stimulation of the basolateral amygdala (BLA) 15 min after water maze training is able to improve spatial learning and memory in fimbria fornix lesioned rats while also inducing changes in the expression of plasticity-related genes expression in memory associated brain regions like the hippocampus and prefrontal cortex. In this study we test that hypothesis: whether BLA stimulation 15 min after water maze training can improve LTP in the hippocampus of fimbria-fornix lesioned rats. To address this question, we trained fimbria-fornix lesioned rats in water maze for four consecutive days, and the BLA was bilaterally stimulated 15 min after each training session.Our data show that trained fimbria-fornix lesioned rats develop a partially improved LTP in dentated gyrus compared with the non-trained fimbria-fornix lesioned rats. In contrast, dentated gyrus LTP in trained and BLA stimulated fimbria-fornix lesioned rats improved significantly compared to the trained fimbria-fornix lesioned rats, but was not different from that shown by healthy animals. BLA stimulation in non-trained FF lesioned rats did not improve LTP; instead produces a transient synaptic depression. Restoration of the ability to develop LTP by the combination of training and BLA stimulation would be one of the mechanisms involved in ameliorating memory deficits in lesioned animals.

Prefrontal neuronal morphology in kindling-prone (FAST) and kindling-resistant (SLOW) rats.

The epileptogenic-prone (FAST) and epileptogenic-resistant (SLOW) rat strains have become a valuable tool for investigating neural plasticity. The strains were generated by breeding the rats that required the fewest amygdala stimulations to elicit a stage-5 convulsive seizure (FAST) and rats requiring the most stimulations (SLOW). Previous studies have shown differences in behavior and amygdala physiology in the two strains. This study examined the dendritic morphology of pyramidal neurons in the brains of adult male and female rats of the two strains. The brains were stained with the Golgi-Cox method and the length and branching from layer III pyramidal cells were measured in parietal cortex (Zilles Par1), medial frontal cortex (Zilles Cg3), and orbitofrontal cortex (Zilles AID) in these two strains of rats. We observed significantly longer dendrites in Cg3 in the FAST group but longer dendrites in the SLOW group in AID and Par1. There was also a sex difference (M>F) in Par1 in both strains. These morphological differences can provide insights into the neurobiological basis of the behavioral differences and suggest that localized changes in the amygdala do not occur independently of changes in other brain regions, and especially prefrontal cortex.

Optogenetic stimulation of the basolateral amygdala accelerates acquisition of object-context associations.

The basolateral complex of the amygdala (BLA) is capable of modulating memory and is thought to do so via projections to regions such as the hippocampus. The present study used optogenetic stimulation of glutamatergic projection neurons in the BLA as rats learned object-context associations during a well-studied hippocampus-dependent memory task. Relative to a control condition, optogenetic BLA stimulation resulted in the accelerated acquisition of when stimulation was delivered following correct choices but not when it was delivered during the intertrial interval. These results extend prior examples of amygdala-mediated memory enhancement to a canonical example of hippocampus-dependent memory and provide an opportunity for future dissection of amygdalar modulation of object-context associative memory. (PsycInfo Database Record (c) 2021 APA, all rights reserved).

Limbic system involvement in modulation of breathing during seizures and arousal

Automatic control of breathing is mediated by a neural network in the brainstem, modulated by central and peripheral inputs. A major source of this modulation comes from chemoreceptors from peripheral and central level. In addition, automatic control of breathing can be overridden by voluntary or conscious control. Pathways involved in this conscious control may be usurped by seizures, resulting in ictal apnea, at the same time that chemoreceptor drive is impaired. Here we studied mechanisms by which the limbic system might modulate breathing, and how this may lead to fatal apnea induced by a seizure. We used Scn1aR1407X/+ mice, a genetic mouse model of Dravet Syndrome (DS) and SUDEP, to study how activation of the Central Amygdala (CeA) can modify breathing and induce fatal apneas. Methods In DS mice (n=4), the CeA was stimulated under light anesthesia by injecting current with monopolar electrodes guided by a stereotaxic manipulator while measuring breathing using head-out plethysmography. The locations of the electrode were verified post hoc by electrolytic lesion and histology. A 3D map was generated of the portions of the CeA where breathing was modulated. In a separate group of DS (n=7) mice, whole-body plethysmography was used to assess baseline breathing, the hypercapnic ventilatory response (HCVR) and the hypoxic ventilatory response (HVR) before and after lesioning the CeA. In a third group of DS mice (n=11), electrolytic lesions were induced in the CeA. Mice were then instrumented with headmounts with EEG, ECG and EMG electrodes. After 5 days of recovery, headmounts were connected to a preamplifier and a commutator, and mice were placed in a plethysmography chamber to monitor breathing as well as EEG, ECG and EMG activity. Body temperature was increased with a heat lamp by 0.5°C per minute until a seizure was induced, or a maximum of 42.5°C was reached. Results CeA lesions did not affect baseline breathing, the HCVR or HVR, and did not prevent seizures in DS mice. Respiratory arrest and death occurred in control (sham-lesioned) DS mice following heat-induced seizures in 54.4% of cases (n=11). In contrast, only 9.1% of DS mice with CeA lesions showed ventilatory arrest and death after heat-induced seizures (n=11; p=0.016). Unilateral rostral amygdala lesions increased the survival rate from spontaneous seizures in DS mice (26.10% vs 49.12.5%, n=14 control, n=20 lesioned p=0.022). Stimulation of the rostral CeA produced apnea without causing seizures. Complete apnea could be induced for 1 minute without causing death in three animals, and for four minutes in one animal. Breathing returned immediately upon termination of amygdala stimulation in each case. There was also no reversal of anesthetic immobility, suggesting lack of an arousal response. Conclusions Our data suggest that the CeA modulates breathing and can induce complete apnea but is not directly involved in baseline respiratory rhythm generation or the ventilatory response to hypercapnia or hypoxia. Lesions of the CeA reduce the risk of apnea during seizures. We conclude that the CeA is a critical component of the neural pathway responsible for spread of seizures in the forebrain to key respiratory centers in the brainstem that elicits ventilatory arrest.

Comparison of the Effects of Deep Brain Stimulation of the Prelimbic Cortex and Basolateral Amygdala for Facilitation of Extinction Process of Conditioned Fear

Background: The study of the biological basis of fear in animal models has progressed considerably because of the energy and space that the brain devotes to this basic emotion. Electrical stimulation targets several structures of the brain to examine its behavioral effects and to understand the role of different regions in underlying mechanisms of fear processing and anxiety in preclinical models. Objectives: In this study, the effects of high-frequency deep brain stimulation (DBS) of the basolateral amygdala (BLA) and prelimbic (PL) sub-region of the prefrontal cortex were evaluated on the extinction process of conditioned fear. Methods: This study was performed on 35 male Wistar rats in the weight range of 220 – 250 g. After selecting the animals, they were separated into five groups. Then, we did stereotactic surgery on rats for electrode implantation. After recovery, some rats were conditioned, followed by a 10-day treatment schedule via high-frequency DBS in the BLA or PL. Next, freezing behavior was measured as a predicted response dedicated to extinction, without shock (re-exposure). In addition, we used ELISA and Western blot to estimate blood serum corticosterone levels and c-Fos protein expression. Results: The mean freezing time recorded for the PL group was significantly lower than that of both the BLA group and the PC group (P &lt; 0.01). The BLA group and PC group were also significantly different (P &lt; 0.001). Corticosterone results indicated that the PL group had significantly higher serum corticosterone levels compared with both the BLA group and the PC group (P &lt; 0.01). In addition, the BLA group revealed a significant reduction in c-Fos expression compared with the PC (P &lt; 0.001). Conclusions: This study provides further evidence for the contribution of the prelimbic cortex and amygdala both in acquisition and extinction processes during contextual fear conditioning. However, the PL stimulation by high-frequency DBS might be more involved in the extinction process and play a more important role as an enhancer.

Amygdala stimulation ameliorates memory impairments and promotes c-Fos activity in fimbria-fornix-lesioned rats.

Recently we provided data showing that amygdala stimulation can ameliorate spatial memory impairments in rats with lesion in the fimbria-fornix (FF). The mechanisms for this improvement involve early gene expression and synthesis of BDNF, MAP-2, and GAP43 in the hippocampus and prefrontal cortex. Now we have studied which brain structures are activated by the amygdala using c-Fos as a marker of neural activation. First, we studied neuronal activation after tetanic stimulation to the amygdala in intact rats. We then carried out a second study in FF-lesioned rats in which the amygdala was stimulated 15min after daily spatial memory training in the water maze. Our results showed that amygdala stimulation produces widespread brain activation, that includes cortical, thalamic, and brain stem structures. Activation was particularly intense in the dentate gyrus and the prefrontal cortex. Training in the water maze increased c-Fos positive nuclei in the dentate gyrus of the hippocampus and in medial prefrontal cortex. Amygdala stimulation to trained FF-lesioned rats induced an increase of neural activity in the dentate gyrus and medial prefrontal cortex relative to the FF-lesioned, but not stimulated group, like the c-Fos activity seen in trained control rats. Based on these and previous results we explain the mechanisms of amygdala reinforcement of neural plasticity and the partial recovery of spatial memory deficits.