Metabolic dysfunction has been implicated in the pathogenesis of temporal lobe epilepsy (TLE), but its manifestation during neuronal activation in the ex vivo hippocampus from TLE patients has not been shown. We characterized metabolic and mitochondrial functions in acute hippocampal slices from pilocarpine-treated, chronic epileptic rats and from pharmaco-resistant TLE patients. Recordings of NAD(P)H fluorescence indicated the status of cellular energy metabolism, and simultaneous monitoring of extracellular potassium concentration ([K+]o) allowed us to control the induction of neuronal activation. In control rats, electrical stimulation elicited biphasic NAD(P)H fluorescence transients that were characterized by a brief initial 'drop' and a subsequent prolonged 'overshoot' correlating to enhanced NAD(P)+ reduction. In chronic epileptic rats, overshoots were significantly smaller in area CA1, but not in the subiculum as compared to controls. In TLE patients, who were histopathologically classified in groups with and without Ammon's horn sclerosis (AHS, non-AHS), large drops and very small overshoots of NAD(P)H transients were observed in dentate gyrus, CA3, CA1 and subiculum. Nevertheless, monitoring mitochondrial membrane potential (DeltaPsi(m)) by mitochondria-specific, voltage-sensitive dye (rhodamine-123) revealed similar mitochondrial responses during neuronal activation with glutamate and protonophore application in area CA1 of control and chronic-epileptic rats. Applying confocal laser scanning microscopy, these findings were confirmed in individual neurons of AHS tissue, indicating a negative DeltaPsi(m) and activation-dependent mitochondrial depolarization. Our data demonstrate severe metabolic dysfunction during neuronal activation in the hippocampus from chronic epileptic rats and humans, although mitochondria maintain negative DeltaPsi(m). Thus, our findings provide a cellular correlate for 'hypometabolism' as described for epilepsy patients and suggest mitochondrial enzyme defects in TLE.
In the hippocampus of patients with therapy-refractory temporal lobe epilepsy, glial cells of area CA1 might be less able to take up potassium ions via barium-sensitive inwardly rectifying and voltage-independent potassium channels. Using ion-selective microelectrodes we investigated the effects of barium on rises in [K+]o induced by repetitive alvear stimulation in slices from surgically removed hippocampi with and without Ammon's horn sclerosis (AHS and non-AHS). In non-AHS tissue, barium augmented rises in [K+]o by 147% and prolonged the half time of recovery by 90%. The barium effect was reversible, concentration dependent, and persisted in the presence of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA), N-methyl-D-aspartate (NMDA) and gamma-aminobutyric acid [GABA(A)] receptor antagonists. In AHS tissue, barium caused a decrease in the baseline level of [K+]o. In contrast to non-AHS slices, in AHS slices with intact synaptic transmission, barium had no effect on the stimulus-induced rises of [K+]o, and the half time of recovery from the rise was less prolonged (by 57%). Under conditions of blocked synaptic transmission, barium augmented stimulus-induced rises in [K+]o, but only by 40%. In both tissues, barium significantly reduced negative slow-field potentials following repetitive stimulation but did not alter the mean population spike amplitude. The findings suggest a significant contribution of glial barium-sensitive K+-channels to K+-buffering in non-AHS tissue and an impairment of glial barium-sensitive K+-uptake in AHS tissue.
Summary:Purpose: Comparison of extracellular K' regulation in sclerotic and nonsclerotic epileptic hippocampus.Methods: Measurements of K' signals with double-barreled K+-selective reference microelectrodes in area CAI of slices from human and rat hippocampus, induction of increases in extracellular potassium concentration by repetitive alvear stimulation or iontophoresis, and block of inward-rectifying and background K' channels in astrocytes by barium.Results: In the CA1 pyramidal layer from normal rat hippocampus, barium augmented extracellular K+ accumulation induced by iontophoresis or antidromic stimulation in a dosedependent manner. Similarly, barium augmented stimulusinduced K+ signals from nonsclerotic hippocampi (human mesial temporal lobe epilepsy). In contrast, barium failed to do so in sclerotic hippocampi (human mesial temporal lobe epilepsy, rat pilocarpine model).Conclusions: Our findings suggest that in areas of reduced neuronal density (hippocampal sclerosis), glial cells adapt to permit rather large increases in extracellular potassium accumulation. Such increases might be involved in the transmission of activity through the sclerotic area. Key Words: Temporal lobe epilepsy-Human-Pilocarpine model-Hippocampus-Pharmacoresistent mesial temporal lobe epilepsy is often accompanied by structural alterations of hippocampal glial cells ("gliosis"). These are most pronounced in the hippocampus of patients with Ammon' s horn sclerosis (AHS), whereas they are less pronounced in patients who have their main lesions outside the hippocampus (non-AHS). The role of gliosis in epilepsy is not clear. It was first suggested by Pollen and Trachtenberg (1) that epileptogenic foci are characterized by a disturbance of K+ homeostasis and that this disturbance leads to the generation of seizures. Now, it is well known that increases in neuronal activity lead to increases in extracellular potassium concentration ([K+],), which can reach levels of up to 12 mmol/L during repetitive activation and seizure activity and even higher levels during anoxia and spreading depression. Otherwise, it has been shown that such increases in [K+], can augment neuronal excitability and thereby contribute to the initiation and spread of seizure activity (2,3). In adult tissue, glial cells are thought to limit increases in [K+], (4). Therefore, disturbances of this glial function could facilitate extracellular accumulation of K+ and thereby promote the generation of sei- Glial cells contribute to K+ regulation by uptake of KC1 (6) and spatial K+ buffering (7). Both spatial K' buffering and KC1 uptake depend on K+ fluxes through barium-sensitive background and inward-rectifying K+ channels. Recently, loss of inward-rectifying K' currents and the low density of background K+ currents associated with high membrane resistance have been observed in astrocytes of human epileptogenic foci (8). High membrane resistance and a reduced density of inwardrectifying K+ currents have also been found in astrocytes of area CAI from sclerotic human hippoc...
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