Protective potential of dimethyl fumarate in a mouse model of thalamocortical demyelination.

Alterations in cortical cellular organization, network functionality, as well as cognitive and locomotor deficits were recently suggested to be pathological hallmarks in multiple sclerosis and corresponding animal models as they might occur following demyelination. To investigate functional changes following demyelination in a well-defined, topographically organized neuronal network, in vitro and in vivo, we focused on the primary auditory cortex (A1) of mice in the cuprizone model of general de- and remyelination. Following myelin loss in this model system, the spatiotemporal propagation of incoming stimuli in A1 was altered and the hierarchical activation of supra- and infragranular cortical layers was lost suggesting a profound effect exerted on neuronal network level. In addition, the response latency in field potential recordings and voltage-sensitive dye imaging was increased following demyelination. These alterations were accompanied by a loss of auditory discrimination abilities in freely behaving animals, a reduction of the nuclear factor-erythroid 2-related factor-2 (Nrf-2) protein in the nucleus in histological staining and persisted during remyelination. To find new strategies to restore demyelination-induced network alteration in addition to the ongoing remyelination, we tested the cytoprotective potential of dimethyl fumarate (DMF). Therapeutic treatment with DMF during remyelination significantly modified spatiotemporal stimulus propagation in the cortex, reduced the cognitive impairment, and prevented the demyelination-induced decrease in nuclear Nrf-2. These results indicate the involvement of anti-oxidative mechanisms in regulating spatiotemporal cortical response pattern following changes in myelination and point to DMF as therapeutic compound for intervention.

Directional spread of activity in synaptic networks of the human lateral amygdala.

Spontaneous epileptiform activity has previously been observed in lateral amygdala (LA) slices derived from patients with intractable-temporal lobe epilepsy. The present study aimed to characterize intranuclear LA synaptic connectivity and to test the hypothesis that differences in the spread of flow of neuronal activity may relate to spontaneous epileptiform activity occurrence. Electrical activity was evoked through electrical microstimulation in acute human brain slices containing the LA, signals were recorded as local field potentials combined with fast optical imaging of voltage-sensitive dye fluorescence. Sites of stimulation and recording were systematically varied. Following recordings, slices were anatomically reconstructed using two-dimensional unitary slices as a reference for coronal and parasagittal planes. Local spatial patterns and spread of activity were assessed by incorporating the coordinates of electrical and optical recording sites into the respective unitary slice. A preferential directional spread of evoked electrical signals was observed from ventral to dorsal, rostral to caudal and medial to lateral regions in the LA. No differences in spread of evoked activity were observed between spontaneously and non-spontaneously active LA slices, i.e. basic properties of evoked synaptic responses were similar in the two functional types of LA slices, including input-output relationship, and paired-pulse depression. These results indicate a directed propagation of synaptic signals within the human LA in spontaneously active epileptic slices. We suggest that the lack of differences in local and in systemic information processing has to be found in confined epileptiform circuits within the amygdala likely involving well-known "epileptic neurons".

Interictal-like network activity and receptor expression in the epileptic human lateral amygdala.

While the amygdala is considered to play a critical role in temporal lobe epilepsy, conclusions on underlying pathophysiological mechanisms have been derived largely from experimental animal studies. Therefore, the present study aimed to characterize synaptic network interactions, focusing on spontaneous interictal-like activity, and the expression profile of transmitter receptors in the human lateral amygdala in relation to temporal lobe epilepsy. Electrophysiological recordings, obtained intra-operatively in vivo in patients with medically intractable temporal lobe epilepsy, revealed the existence of interictal activity in amygdala and hippocampus. For in vitro analyses, slices were prepared from surgically resected specimens, and sections from individual specimens were used for electrophysiological recordings, receptor autoradiographic analyses and histological visualization of major amygdaloid nuclei for verification of recording sites. In the lateral amygdala, interictal-like activity appeared as spontaneous slow rhythmic field potentials at an average frequency of 0.39 Hz, which occurred at different sites with various degrees of synchronization in 33.3% of the tested slices. Pharmacological blockade of glutamate α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors, but not N-methyl-D-aspartate receptors, abolished interictal-like activity, while the γ-aminobutyric acid A-type receptor antagonist bicuculline resulted in a dampening of activity, followed by highly synchronous patterns of slow rhythmic activity during washout. Receptor autoradiographic analysis revealed significantly higher α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid, kainate, metabotropic glutamate type 2/3, muscarinic type 2 and adrenoceptor α(1) densities, whereas muscarinergic type 3 and serotonergic type 1A receptor densities were lower in the lateral amygdala from epileptic patients in comparison to autopsy controls. Concerning γ-aminobutyric acid A-type receptors, agonist binding was unaltered whereas antagonist binding sites were downregulated in the epileptic lateral amygdala, suggesting an altered high/low-affinity state ratio and concomitant reduced pool of total γ-aminobutyric acid A-type receptors. Together these data indicate an abnormal pattern of receptor densities and synaptic function in the lateral nucleus of the amygdala in epileptic patients, involving critical alterations in glutamate and γ-aminobutyric acid receptors, which may give rise to domains of spontaneous interictal discharges contributing to seizure activity in the amygdala.

GABAergic interneurons in the mouse lateral amygdala: a classification study.

Whole cell patch-clamp recordings were performed in GABAergic interneurons labeled by green fluorescent protein (GFP) in the lateral amygdala (LA) in vitro from glutamic acid decarboxylase 67 (GAD67)-GFP mice. Neurons were characterized by electrotonic and electrogenic parameters. Cytoplasm was collected from individual neurons, and single-cell RT-PCR was used for detection of molecular markers typifying LA interneurons. Hierarchical cluster and multiple discriminant analysis demonstrated the existence of five types of GABAergic interneurons, which can be reliably identified through electrophysiological criteria. Action potentials were of a short duration followed by pronounced fast afterhyperpolarization (AHP) in interneurons of all types, except for type V, which generated broad action potentials and displayed typical spike bursts at the beginning of depolarizing stimuli and prominent anomalous inward rectification. Interneurons of type I and II generated series of action potentials with frequency adaptation on maintained depolarizing current stimulation with overall frequencies at high levels and presented delayed firing, stuttering or fast-spiking behavior. Further distinguishing features of type II interneurons were a medium AHP following spike trains and pronounced anomalous inward rectification. Types III and IV of neurons fired regularly, whereas type IV displayed no prominent spike frequency adaptation. Additionally, interneurons of all five types contained mRNA of glutamic acid decarboxylase 65 and cholecystokinin, whereas only type I interneurons were somatostatin-positive. Overall, these data represent a detailed and reliable classification scheme of LA GABAergic interneurons and will provide a feasible basis for subsequent functional studies.

Alteration of NMDA receptor-mediated synaptic interactions in the lateral amygdala associated with seizure activity in a mouse model of chronic temporal lobe epilepsy.

PURPOSE: Because results from both animal models and human temporal lobe epilepsy (TLE) have pointed to synaptic network alterations in the amygdala, we have tested the hypothesis that glutamatergic transmission in the lateral amygdala (LA) is critically involved. METHODS: Using the pilocarpine mouse model, LA slices were prepared ex vivo in the recurrent phase of TLE (Pilo group), and LA projection neurons (PNs) were recorded using patch-clamp techniques. Intrinsic and synaptic properties of LA PNs were analyzed and compared with those in age-matched saline-injected controls. RESULTS: Only mild changes were observed in intrinsic properties of LA PNs, whereas both spontaneous excitatory postsynaptic currents (sEPSCs) and miniature EPSCs (mEPSCs) were significantly increased in Pilo as compared to saline controls. This difference was sensitive to AP5, but persisted during action of NBQX, indicating mediation by N-methyl-d-aspartate (NMDA) receptors. Moreover, these changes were associated with an increase in frequency but not amplitude of mEPSCs, indicative of a contribution of presynaptic mechanisms. DISCUSSION: In conclusion, dynamic changes seem to occur in glutamatergic transmission within the amygdala during TLE, to which a functional upregulation of presynaptic NMDA receptors in LA PNs makes a significant contribution.