Clinical Correlation of Altered Molecular Signatures in Epileptic Human Hippocampus and Amygdala.

Widespread alterations in the expression of various genes could contribute to the pathogenesis of epilepsy. The expression levels of various genes, including major inhibitory and excitatory receptors, ion channels, cell type-specific markers, and excitatory amino acid transporters, were assessed and compared between the human epileptic hippocampus and amygdala, and findings from autopsy controls. Moreover, the potential correlation between molecular alterations in epileptic brain tissues and the clinical characteristics of patients undergoing epilepsy surgery was evaluated. Our findings revealed significant and complex changes in the expression of several key regulatory genes in both the hippocampus and amygdala of patients with intractable epilepsy. The expression changes in various genes differed considerably between the epileptic hippocampus and amygdala. Different correlation patterns were observed between changes in gene expression and clinical characteristics, depending on whether the patients were considered as a whole or were subdivided. Altered molecular signatures in different groups of epileptic patients, defined within a given category, could be viewed as diagnostic biomarkers. Distinct patterns of molecular changes that distinguish these groups from each other appear to be associated with epilepsy-specific functional consequences.

Antiepileptic effects of cobalt, manganese and magnesium on bicuculline-induced epileptiform activity in hippocampal neurons.

BACKGROUND: Calcium signaling is described as a relevant factor in synchronization of neurons and increased excitability in epileptogenesis. Aim of the present investigations was to test the antiepileptic effect of the classical inorganic calcium channel blockers cobalt (Co2+), manganese (Mn2+) and magnesium (Mg2+). METHODS: Experiments were carried out on hippocampal slices of guinea pigs. Epileptiform field potentials (EFP) were elicited by adding bicuculline (10 µmol/l) to the artificial cerebrospinal fluid (CSF). Kalium was elevated from normal (4 mmol/l) to 8 mmol/l. Co2+ (CoCl2; 2, 1, 0.5 and 0.1 mmol/l), Mn2+ (MnCl2; 2, 1, 0.5 and 0.1 mmol/l) and Mg2+ (MgCl2; 8, 6, 5, 4 and 2 mmol/l) were added to the superfusate. RESULTS: Concentrations of 2, 1 and 0.5 mmol/l Co2+, 2 and 1 mmol/l Mn2+ and 8 respectively 6 mmol/l Mg2+ were able to suppress EFP sufficient in a dose dependent manner. In concentrations of 0.1 mmol/l Co2+, 0.5 mmol/l and 0.1 mmol/l Mn2+ and 5 respectively 4 and 2 mmol/l Mg2+ suppression was incomplete. With washout of the inorganic calcium channel blockers the EFP reappeared. DISCUSSION: All tree inorganic calcium channel blockers were able to suppress EFP in a dosage dependent and reversible manner. Weak reappearance of EFP after washout of Co2+ might be due to additional cytotoxic effects. The following mechanisms may contribute: i) blockade of voltage-activated calcium channels in the postsynaptic membrane, ii) changes in the activation of voltage-dependent sodium channels, iii) blockade of synaptic transmission.

The next-generation sphingosine-1 receptor modulator BAF312 (siponimod) improves cortical network functionality in focal autoimmune encephalomyelitis.

Autoimmune diseases of the central nervous system (CNS) like multiple sclerosis (MS) are characterized by inflammation and demyelinated lesions in white and grey matter regions. While inflammation is present at all stages of MS, it is more pronounced in the relapsing forms of the disease, whereas progressive MS (PMS) shows significant neuroaxonal damage and grey and white matter atrophy. Hence, disease-modifying treatments beneficial in patients with relapsing MS have limited success in PMS. BAF312 (siponimod) is a novel sphingosine-1-phosphate receptor modulator shown to delay progression in PMS. Besides reducing inflammation by sequestering lymphocytes in lymphoid tissues, BAF312 crosses the blood-brain barrier and binds its receptors on neurons, astrocytes and oligodendrocytes. To evaluate potential direct neuroprotective effects, BAF312 was systemically or locally administered in the CNS of experimental autoimmune encephalomyelitis mice with distinct grey- and white-matter lesions (focal experimental autoimmune encephalomyelitis using an osmotic mini-pump). Ex-vivo flow cytometry revealed that systemic but not local BAF312 administration lowered immune cell infiltration in animals with both grey and white matter lesions. Ex-vivo voltage-sensitive dye imaging of acute brain slices revealed an altered spatio-temporal pattern of activation in the lesioned cortex compared to controls in response to electrical stimulation of incoming white-matter fiber tracts. Here, BAF312 administration showed partial restore of cortical neuronal circuit function. The data suggest that BAF312 exerts a neuroprotective effect after crossing the blood-brain barrier independently of peripheral effects on immune cells. Experiments were carried out in accordance with German and EU animal protection law and approved by local authorities (Landesamt für Natur, Umwelt und Verbraucherschutz Nordrhein-Westfalen; 87-51.04.2010.A331) on December 28, 2010.

Cell injury and receptor expression in the epileptic human amygdala.

Neuropathological findings in the amygdala obtained from patients with mesial temporal lobe epilepsy (MTLE) indicate varying degrees of histopathological alterations, such as neuronal loss and gliosis. The mechanisms underlying cellular damage in the amygdala of patients with MTLE have not been fully elucidated. In the present study, we assess cellular damage, determine the receptor expression of major inhibitory and excitatory neurotransmitters, and evaluate the correlation between the expression of various receptors and cell damage in the basolateral complex and the centromedial areas in the amygdala specimens resected during brain surgery on 30 patients with medically intractable MTLE. Our data reveal an increased rate of cell damage and apoptosis as well as decreased expression levels of several GABAergic receptor subunits (GABAARα1, GABAARß3, and GABABR1) and GAD65 in the amygdalae obtained during epilepsy surgery compared to autopsy specimens. Analyses of the expression of glutamate excitatory receptor subunits (NR1, NR2B, mGluR1α, GluR1, and GluR2) reveal no significant differences between the epileptic amygdalae and autopsy control tissues. Furthermore, the increased occurrence of apoptotic cells in the amygdala is negatively correlated with the reduced expression of the studied GABAergic receptor subunits and GAD65 but is not correlated with the expression of excitatory receptors. The present data point to the importance of GABAergic neurotransmission in seizure-induced cell injury in the amygdala of patients with MTLE and suggest several GABA receptor subunits as potential druggable target structures to control epilepsy and its comorbid disorders, such as anxiety.

Levetiracetam mediates subtle pH-shifts in adult human neocortical pyramidal cells via an inhibition of the bicarbonate-driven neuronal pH-regulation - Implications for excitability and plasticity modulation.

The intracellular pH (pHi) of mammalian central neurons is tightly regulated and small pHi-fluctuations can fine-tune inter-/intracellular signaling, excitability, and synaptic plasticity. The research-gap about the pHi-regulation of human brain neurons is addressed here by testing possible influences of the anticonvulsant levetiracetam (LEV). BCECF-AM-loaded neocortical pyramidal cells were fluorometrically investigated in slice-preparations of tissue resected from the middle temporal gyrus of five adults with intractable temporal-lobe epilepsy. Recovery-slope from intracellular acidification following an ammonium prepulse (APP) was used to measure the pHi-regulation. Among twenty pyramidal cells exposed to 50 µM LEV, the resting pHi (7.09 ±â€¯0.14) was lowered in eight (40%) neurons, on average by 0.02 ±â€¯0.011 pH-units. In three (15%) and nine (45%) neurons, a minimal alkaline shift (0.017 ±â€¯0.004 pH-units) and no pHi-shift occurred, respectively. The LEV-induced pHi-shifts were positively correlated with the resting pHi (r = 0.6, p = 0.006, n = 20). In five neurons, which all had responded on LEV with an acidification before, the recovery from APP-acidification was significantly delayed during LEV (p < 0.001). This inhibitory LEV-effect on pHi-regulation i) was similar to that of 200 µM 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (n = 2) and ii) did not occur under nominal bicarbonate-free conditions (n = 2). Thus, LEV lowered the pHi of human neocortical pyramidal cells most likely by a weakening of the transmembrane HCO3(-)-mediated acid-extrusion. This might contribute to LEV's anticonvulsive potency. Neurons with more acidic resting pHi-values showed a minimal alkalization upon LEV providing a mechanism for paradoxical proconvulsive LEV-effects rarely observed in epilepsy patients. The significance of these subtle pHi-shifts for cortical excitability and plasticity is discussed.

Aging is associated with a mild acidification in neocortical human neurons in vitro.

The intracellular pH (pHi) in the cytosol of mammalian central neurons is tightly regulated and small pHi-fluctuations are deemed to modulate inter-/intracellular signaling, excitability, and synaptic plasticity. The resting pHi of young rodent hippocampal pyramidal neurons is known to decrease alongside aging for about 0.1 pH-units. There is no information about the relationship between age and pHi of human central neurons. We addressed this knowledge gap using 26 neocortical slices from 12 patients (1-56-years-old) who had undergone epilepsy surgery. For fluorometric recordings, the slice-neurons were loaded with the pHi-sensitive dye BCECF-AM. We found that the pyramidal cells' resting pHi (n = 26) descended linearly alongside aging (r = - 0.71, p < 0.001). This negative relationship persisted, when the sample was confined to specific brain regions (i.e., middle temporal gyrus, 23 neurons, r = - 0.68, p < 0.001) or pathologies (i.e., hippocampus sclerosis, 8 neurons, r = - 0.78, p = 0.02). Specifically, neurons (n = 9, pHi 7.25 ± 0.12) from young children (1.5 ± 0.46-years-old) were significantly more alkaline than neurons from adults (n = 17, 38.53 ± 12.38 years old, pHi 7.08 ± 0.07, p < 0.001). Although the samples were from patients with different pathologies the results were in line with those from the rodent hippocampal pyramidal neurons. Like a hormetin, the age-related mild pHi-decrease might contribute to neuroprotection, e.g., via limiting excitotoxicity. On the other hand, aging cortical neurons could become more vulnerable to metabolic overstress by a successive pHi-decrease. Certainly, its impact for the dynamics in short and long-term synaptic plasticity and, ultimately, learning and memory provides a challenge for further research.

Small intraneuronal acidification via short-chain monocarboxylates: First evidence of an inhibitory action on over-excited human neocortical neurons.

AIMS: In cortical mammalian neurons, small fluctuations of intracellular pH (pHi) play a crucial role for inter- and intracellular signaling as well as for cellular and synaptic plasticity. Yet, there have been no respective data about humans. Thus, we investigated the interrelation of pHi and excitability of human cortical neurons. MATERIALS AND METHODS: Intracellular electrophysiological and pH-recordings were made in neurons in slices taken from brain tissue resected from the middle temporal gyrus of two male children (26 months and 35 months old) who suffered from pharmacotherapy-resistant temporal lobe epilepsy. To excite the tissue (n = 13), we used the 0-Mg2+/high-K+-in vitro epilepsy model producing robust epileptiform discharges (ED). To evoke an intracellular acidification (n = 12), we used the well-established propionate-model and applied 10 mM propionate to the bath solutions. In addition, we recorded the effects of other strongly related short-chain monocarboxylates (l-lactate (10 mM) and the ketone body DL-ß-hydroxybutyrate (10 mM)) on ED and pHi. KEY FINDINGS: The ED-frequency was reversibly reduced by propionate (n = 5), l-lactate (n = 5), or DL-ß-hydroxybutyrate (n = 3), while the durations of EDs and their after-depolarizations increased. In parallel experiments, all three short-chain monocarboxylates (each n = 4) lowered the pHi of the neurons (n = 12) by 0.05-0.07 pH units which was temporally related to the reported changes in bioelectric activity. SIGNIFICANCE: A mild drop of the intraneuronal pH was associated with the control of even over-excited human neocortical tissue. This is identical with prior observations in non-human mammalian cortical neurons. Possible implications for neuroplasticity and the treatment of neuropsychiatric disorders are discussed.

Uncensored EEG: The role of DC potentials in neurobiology of the brain.

Brain direct current (DC) potentials denote sustained shifts and slow deflections of cerebral potentials superimposed with conventional electroencephalography (EEG) waves and reflect alterations in the excitation level of the cerebral cortex and subcortical structures. Using galvanometers, such sustained displacement of the EEG baseline was recorded in the early days of EEG recordings. To stabilize the EEG baseline and eliminate artefacts, EEG was performed later by voltage amplifiers with high-pass filters that dismiss slow DC potentials. This left slow DC potential recordings as a neglected diagnostic source in the routine clinical setting over the last few decades. Brain DC waves may arise from physiological processes or pathological phenomena. Recordings of DC potentials are fundamental electro-clinical signatures of some neurological and psychological disorders and may serve as diagnostic, prognostic, and treatment monitoring tools. We here review the utility of both physiological and pathological brain DC potentials in different aspects of neurological and psychological disorders. This may enhance our understanding of the role of brain DC potentials and improve our fundamental clinical and research strategies for brain disorders.

ATPase N-ethylmaleimide-sensitive Fusion Protein: A Novel Key Player for Causing Spontaneous Network Excitation in Human Temporal Lobe Epilepsy.

The molecular basis for onset, maintenance and propagation of excitation along neuronal networks in epilepsy is still poorly understood. Besides different neurotransmitter receptors that control signal transfer at the synapse, one key regulator involved in all of these processes is the ATPase N-ethylmaleimide-sensitive fusion protein (NSF). Therefore, we analyzed receptor subunits and NSF levels in tissues from the medial temporal gyrus (MTG) of patients with pharmaco-resistant focal temporal lobe epilepsy resected during epilepsy surgery and autopsy controls. The resected tissues were further characterized by field potential recordings into tissues with and without spontaneous sharp wave activity. We detected increased levels of NSF, NMDA 1.1, 2A and GABAAγ2 receptor subunits associated with spontaneous sharp wave spiking activity. We further identified correlations between NSF, AMPA receptor subunit, metabotropic glutamate receptor and adenosine 1 receptor levels in the spontaneous sharp wave spiking tissues. Our findings suggest that NSF plays a key role in controlling spontaneous network excitation in epilepsy by two mechanisms of action: (1) directly via controlling transmitter release at the presynaptic side, and (2) indirectly via altering the function of possible receptor crosstalk and directing/integrating specific receptor compounds through/into the postsynaptic membrane.

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".

Effects of garlic extract on spreading depression: In vitro and in vivo investigations.

OBJECTIVES: The potential use of garlic for prevention and treatment of different types of headaches has been suggested by several medieval literatures. Cortical spreading depression (CSD), a propagating wave of neuroglial depolarization, was established as a target for anti-migraine drugs. This study was designed to investigate the effect of garlic extract on CSD in adult rats. METHODS: CSD was induced by KCl microinjection in the somatosensory cortex. The effects of five different concentrations of garlic oil (1-500 µl/l) were tested on different characteristic features of CSD in necocortical slices. In in vivo experiments, the effects of garlic oil on electrophysiological and morphological changes induced by CSD were investigated. RESULTS: Garlic oil in a dose-dependent manner decreased the amplitude of CSD but not its duration and velocity in neocortical brain slices. Garlic oil at concentration of 500 µl/l reversibly reduced the amplitude of the field excitatory post-synaptic potentials and inhibited induction of long-term potentiation in the third layer of neocortical slices. In in vivo studies, systemic application of garlic oil (1 ml/l) for three consecutive days reduced the amplitude and repetition rate of CSD. Garlic oil also prevented of CSD-induced reactive astrocytosis in the neocortex. DISCUSSION: Garlic oil suppresses CSD, likely via inhibition of synaptic plasticity, and prevents its harmful effects on astrocyte. Further studies are required to identify the exact active ingredient(s) of garlic oil that inhibit CSD and may have the potential to use in treatment of CSD-related disorders.

Non-invasive monitoring of spreading depression.

Spreading depression (SD), a slow propagating depolarization wave, plays an important role in pathophysiology of different neurological disorders. Yet, research into SD-related disorders has been hampered by the lack of non-invasive recording techniques of SD. Here we compared the manifestations of SD in continuous non-invasive electroencephalogram (EEG) recordings to invasive electrocorticographic (ECoG) recordings in order to obtain further insights into generator structures and electrogenic mechanisms of surface recording of SD. SD was induced by KCl application and simultaneous SD recordings were performed by scalp EEG as well as ECoG electrodes of somatosensory neocortex of rats using a novel homemade EEG amplifier, AgCl recording electrodes, and high chloride conductive gel. Different methods were used to analyze the data; including the spectrogram, bi-spectrogram, pattern distribution, relative spectrum power, and multivariable Gaussian fit analysis. The negative direct current (DC) shifts recorded by scalp electrodes exhibited a high homogeneity to those recorded by ECoG electrodes. Furthermore, this novel method of recording and analysis was able to separate SD recorded by scalp electrodes from non-neuronal DC shifts induced by other potential generators, such as the skin, muscles, arteries, dura, etc. These data suggest a novel application for continuous non-invasive monitoring of DC potential changes, such as SD. Non-invasive monitoring of SD would allow early intervention and improve outcome in SD-related neurological disorders.

Thalamocortical-auditory network alterations following cuprizone-induced demyelination.

BACKGROUND: Demyelination and remyelination are common pathological processes in many neurological disorders, including multiple sclerosis (MS). Clinical evidence suggests extensive involvement of the thalamocortical (TC) system in patients suffering from MS. METHODS: Using murine brain slices of the primary auditory cortex, we investigated the functional consequences of cuprizone-induced de- and remyelination on neuronal activity and auditory TC synaptic transmission in vitro. RESULTS: Our results revealed an impact of myelin loss and restoration on intrinsic cellular firing patterns, synaptic transmission, and neuronal plasticity in layer 3 and 4 neurons of the auditory TC network. While there was a complex hyper- and depolarizing shift of the resting membrane potential, spontaneous and induced action potential firing was reduced during demyelination and early remyelination. In addition, excitatory postsynaptic potential amplitudes were decreased and induction of LTP was reduced during demyelination. CONCLUSIONS: These data indicate that demyelination-induced impairment of neurons and network activity within the TC system may underlie clinical symptoms observed in demyelinating diseases, corroborating human findings that disease progression is significantly correlated with microstructural tissue damage of the TC system. Further investigation into focal inflammation-induced demyelination models ex vivo and in vivo are needed to understand the functional implication of local and remote lesion formation on TC network activity in MS.

Anticonvulsant effect of neural regeneration peptide 2945 on pentylenetetrazol-induced seizures in rats.

Neuron regeneration peptides (NRPs) are small synthetic peptides that stimulate neural proliferation, migration, and differentiation with no apparent toxicity and high target specificity in CNS. The aim of this study was to investigate the effect of NRP2945 on seizure activity induced by pentylenetetrazol (PTZ) in rats. Using behavioural assessment and electrocorticographical recordings, the effects of different doses of NRP2945 (5-20 µg/kg) were tested on seizure attacks induced by PTZ injection. In addition, the effect of NRP2945 was evaluated on the production of dark neurons and expression of GABAA receptor α and ß subunits and GAD-65 in the hippocampus and somatosensory cortex of the rat brain. Intraperitoneal injection of NRP2945 at 20 µg/kg prevented seizure attacks after PTZ injection. NRP2945 at doses of 5 and 10 µg/kg significantly decreased the total duration of seizure attacks and reduced the amplitude, duration and latency of epileptiform burst discharges induced by PTZ. In addition, the peptide significantly inhibited the production of dark neurons in the hippocampus and somatosensory cortex of epileptic rats. NRP2945 also significantly increased the expression of GABAA receptor α and ß subunits and GAD-65 in the hippocampus and somatosensory cortex compared with PTZ treated rats. This study indicates that NRP2945 is able to prevent the seizure attacks and neuronal injuries induced by PTZ, likely by stimulating GABAA and GAD-65 protein expression and/or protecting these components of GABAergic signalling from PTZ-induced alteration. Further studies are needed to elucidate the potential role of NRP2945 as an antiepileptic drug.

Familial hemiplegic migraine and spreading depression.

OBJECTIVE: Familial hemiplegic migraine (FHM) is an autosomal dominantly inherited subtype of migraine with aura, characterized by transient neurological signs and symptoms. Typical hemiplegic migraine attacks start in the first or second decade of life. Some patients with FHM suffer from daily recurrent attacks since childhood. Results from extensive studies of cellular and animal models have indicated that gene mutations in FHM increase neuronal excitability and reduce the threshold for spreading depression (SD). SD is a transient wave of profound neuronal and glial depolarization that slowly propagates throughout the brain tissue and is characterized by a high amplitude negative DC shift. After induction of SD, S218L mutant mice exhibited neurological signs highly reminiscent of clinical attacks in FHM type 1 patients carrying this mutation. FHM1 with ataxia is attributable to specific mutations that differ from mutations that cause pure FHM1 and have peculiar consequences on cerebellar Cav2.1 currents that lead to profound Purkinje cell dysfunction and neuronal loss with atrophy. SD in juvenile rats produced neuronal injury and death. Hormonal factors involved in FHM affect SD initiation and propagation. The data identify SD as a possible target of treatment of FHM. In addition, FHM is a useful model to explore the mechanisms of more common types of migraine.

Propagation of cortical spreading depression into the hippocampus: The role of the entorhinal cortex.

Propagation of cortical spreading depression (CSD) to the subcortical structures could be the underlying mechanism of some neurological deficits in migraine with aura. The entorhinal cortex (EC) as a gray matter bridge between the neocortex and subcortical regions plays an important role in this propagation. In vitro combined neocortex-hippocampus brain slices were used to study the propagation pattern of CSD between the neocortex and the hippocampus. The effects of different compounds as well as tetanic electrical stimulations in the EC on propagation of CSD to the hippocampus were investigated. Repetitive induction of CSD by KCl injection in the somatosensory cortex enhanced the probability of CSD entrance to the hippocampus via EC. Local application of AMPA receptor blocker CNQX and cannabinoid receptor agonist WIN 55212-2 in EC facilitated the propagation of CSD to the hippocampus, whereas application of NMDA receptor blocker APV and GABAA receptor blocker bicuculline in this region reduced the probability of CSD penetration to the hippocampus. Application of tetanic stimulation in EC also facilitated the propagation of CSD entrance to the hippocampus. Our data suggest the importance of synaptic plasticity of EC in filtering the propagation of CSD into subcortical structures and possibly the occurrence of concomitant neurological deficits. Synapse 68:574-584, 2014. © 2014 Wiley Periodicals, Inc.

Interstitial amino acid concentrations in rodent brain tissue during chemical ischemia.

The significance of electrophysiological phenomena is well validated in brain ischemia research. A close link with interstitial amino acid levels has not been proved convincingly but is generally assumed. This has given widespread rise to the clinical method of amino acid, especially glutamate, microdialysis. We combined microdialytic and electrophysiological techniques in an in vitro ischemia model to test for such a correlation. Rodent hippocampal brain slices were subjected to various patterns of ischemic simulation by depletion of glucose and oxygen and to K+ superfusion, which is often used as an alternative stressor. Our data do not strengthen the significance of clinically standardized glutamate measurements, insofar as ischemia-induced damage was demonstrated by electrophysiology and histology before being clearly mirrored by interstitial glutamate levels. Taurine would be a more promising candidate. K+ is not an adequate substitute for ischemic simulation, because biochemical and electrophysiological reactions of the tissue are clearly different. In vitro microdialysis during ischemic simulation is feasible and might provide a tool to inquire into glial functions during ischemic stress. It is probably not able to elucidate processes within the synaptic cleft.

Spreading depression triggers ictaform activity in partially disinhibited neuronal tissues.

There is unequivocal electrophysiological evidence that spreading depression (SD) can trigger epileptiform field potentials. In vitro experiments on human brain tissues indicated that γ-aminobutyric acid (GABA)-mediated inhibition prevented this process. Intra- and extracellular recordings of bioelectrical activities were performed in the rodent neocortex, hippocampus and amygdala after perfusion of low concentrations of the GABAA antagonist bicuculline and induction of SD by KCl application. Induction of SD in combined amygdala-hippocampus-cortex slices pre-treated with low concentration of bicuculline triggered epileptiform burst discharges in cortical as well as subcortical brain structures. Propagation of SD significantly depolarized the membrane, decreased the amplitude and duration of action potentials (APs) and after-hyperpolarization as well as the neuronal membrane input resistance and the amplitude of threshold potentials. Ten to twenty minutes after induction of SD, the pattern of APs changed from regular firing to a series of APs riding on an underlying paroxysmal depolarization shift before the appearance of typical ictaform activities. Changes of characteristic features of APs occurred after SD persisted during the appearance of epileptiform activities. These results indicate that SD increases neuronal excitability and facilitates synchronization of neuronal discharges in the presence of partial disinhibition of neuronal tissues. Our findings might explain the occurrence of seizures in neurological disorders with partial impairment of inhibitory tone, such as brain ischemia and epilepsy.

The effects of tetanic stimulation on plasticity of remote synapses in the hippocampus-perirhinal cortex-amygdala network.

In the temporal lobe, multiple synaptic pathways reciprocally link different structures. These multiple pathways play an important role in the integrity of the function of the temporal lobe and malfunction in this network has been suggested to underlie some neurological disorders such as epilepsy. To test whether the induction of long-term potentiation (LTP) in one temporal lobe structure would modulate functional synaptic plasticity in other structures of this network, tetanic stimulation was applied to the white matter of the perirhinal cortex, Schaffer collaterals of the hippocampus, or the external capsule in combined rat amygdala-hippocampus-cortex slices. This tetanic stimulation was accompanied by enhancement of the evoked field potential slope in the third layer of perirhinal cortex, hippocampal CA1 area, and the lateral amygdala. Induction of LTP in each of these structures was concomitant with increased evoked field potentials in the neighboring structures. Surgical disconnection of anatomical pathways between these structures inhibited this concomitant enhancement of the evoked field potential slope. Both NMDA and AMPA glutamate sub-receptors were involved in changes of synaptic plasticity elicited by induction of LTP in the neighboring structures. The present data indicate a reciprocal control among the perirhinal cortex, the amygdala, and the hippocampus plasticity. This could be important for the formation and retention of the medial temporal lobe-dependent memory and may play a role in the involvement of all different regions of the temporal lobe in pathological conditions such as epilepsy that affect this brain structure.

Effect of cannabinoid receptor activation on spreading depression.

OBJECTIVE(S): The objective of this study was to evaluate the effect of cannabinoid on cortical spreading depression (CSD) in rat brain. Cannabis has been used for centuries for both symptomatic and prophylactic treatment of different types of headaches including migraine. CSD is believed to be a putative neuronal mechanism underlying migraine aura and subsequent pain. MATERIALS AND METHODS: The effects of Delta9-tetrahydrocannabinol (THC), as well as, cannabinoid CB1 and CB2 receptor agonists on CSD in rat neocortical slices were investigated. Furthermore, the effect of cannabinoid CB1 agonist was tested on field excitatory postsynaptic potentials (fEPSP) and long-term potentiation (LTP). RESULTS: HC (1-20 microM) dose dependently suppressed CSD amplitude, duration, and propagation velocity. Cannabinoid CB1 agonist, WIN 55,212-2 mesylate (1-10 microM), also significantly suppressed all characteristic features of CSD. However, cannabinoid CB2 agonist, JWH-133 (1-20 microM), did not affect CSD. FEPSP and induction of LTP were suppressed by application of WIN55212-2. CONCLUSION: Suppression of CSD by activation of CB1 receptors points to the potential therapeutic effects of cannabinoids in migraine with aura. More research is needed before we know whether cannabinoids may be helpful in treating migraine pain.