Brain Mitochondrial Metabolic Dysfunction and Glutamate Level Reduction in the Pilocarpine Model of Temporal Lobe Epilepsy in Mice

Abstract

Although certain metabolic characteristics such as interictal glucose hypometabolism are well established for temporal lobe epilepsy (TLE), its pathogenesis still remains unclear. Here, we performed a comprehensive study of brain metabolism in a mouse model of TLE, induced by pilocarpine-status epilepticus (SE). To investigate glucose metabolism, we injected mice 3.5-4 weeks after SE with [1,2-13 C]glucose before microwave fixation of the head. Using 1 H and 13 C nuclear magnetic resonance spectroscopy, gas chromatography-mass spectrometry and high-pressure liquid chromatography, we quantified metabolites and 13 C labeling in extracts of cortex and hippocampal formation (HF). Hippocampal levels of glutamate, glutathione and alanine were decreased in pilocarpine-SE mice compared with controls. Moreover, the contents of N-acetyl aspartate, succinate and reduced nicotinamide adenine dinucleotide (phosphate) NAD(P)H were decreased in HF indicating impairment of mitochondrial function. In addition, the reduction in 13 C enrichment of hippocampal citrate and malate suggests decreased tricarboxylic acid (TCA) cycle turnover in this region. In cortex, we found reduced 13 C labeling of glutamate, glutamine and aspartate via the pyruvate carboxylation and pyruvate dehydrogenation pathways, suggesting slower turnover of these amino acids and/or the TCA cycle. In conclusion, mitochondrial metabolic dysfunction and altered amino-acid metabolism is found in both cortex and HF in this epilepsy model. Keywords: 13 C isotope; glutamate; mitochondria; neurometabolism; NMR spectroscopy; temporal lobe epilepsy Journal of Cerebral Blood INTRODUCTIONTemporal lobe epilepsy (TLE) is one of the most common forms of human epilepsy and is associated with a high level of drug resistance among patients. The mechanisms underlying the pathogenesis of TLE still remain unclear, although increasing evidence points to a disturbance in amino-acid neurotransmitter homeostasis and energy metabolism, as well as neuronal injury. Metabolic characteristics of human TLE include interictal glucose hypometabolism and a decrease in the levels of the neuronal marker N-acetyl aspartate (NAA) in the epileptogenic region. 1,2 Increased levels of extracellular glutamate in epileptogenic hippocampi have been reported before and during seizures 3 and interictally. 4 Moreover, it has been suggested that the uptake of glutamate from the extracellular space by astrocytes is slower in the epileptic brain as the expression of glutamine synthetase, the glial enzyme that converts glutamate to glutamine, was decreased in the epileptogenic hippocampi from patients with TLE. 5,6 Changes in brain metabolism in rat models of TLE resemble those reported in human TLE such as interictal glucose hypometabolism demonstrated in lithium-pilocarpine rats, 7 as well as alterations in NAA and glutamate reported in post-status epilepticus (SE) models induced by kainic acid and lithiumpilocarpine. [8][9][10] There are, however, few comprehensive studies on brain metabolism in mouse model...

Alterations of Hippocampal Glucose Metabolism by Even versus Uneven Medium Chain Triglycerides

“…17 Samples were lyophilized and liquid chromatography tandem mass spectrometry analysis was performed by investigators masked to the treatment. Metabolite concentrations are within reported ranges 18,19 and the average coefficient of variation of metabolite levels was 21%, which is expected for biologic samples, despite no internal standard being used to monitor extraction efficiency.…”

“…All data are represented as mean ± s.e.m. We chose to use 9 to 11 animals in each group for metabolite analyses and 15 mice per group in the 6 Hz model, based on previous experience of variations in metabolite levels 4,19 and the 6 Hz model.…”

“…3,4,13,14 Previously, it has been hypothesized that anaplerosis is an advantage, specifically to the 'epileptic' brain in which TCA cycle intermediates levels are reduced. 4,19 So far, to our knowledge trioctanoin has not been tested in chronic epilepsy models. From this study, one could expect that trioctanoin may be less effective than triheptanoin in chronic epilepsy models because trioctanoin reduces the levels of TCA cycle intermediates and is not anaplerotic.…”

“…26 Conversely [1,2-13 C] glucose that was derived from the labeled heptanoate was primarily metabolized in neurons, indicated by the higher levels of label found in glutamate. 19 The lack of triheptanoin's effect on these major metabolic pathways would appear to be beneficial, as fewer metabolic side effects would be expected to result from the therapeutic administration of triheptanoin.…”

The Relationship between Glucose Metabolism, Resting-State fMRI BOLD Signal, and GABAA-Binding Potential: A Preliminary Study in Healthy Subjects and Those with Temporal Lobe Epilepsy

“…If we assume a relatively intact neurovascular coupling, we can infer that the blood oxygen leveldependent activity measured by ALFFs accurately reflects underlying neural activity, and that the mechanisms underlying the glutamate-glutamine cycle in neurons and astrocytes are disturbed. Some evidence from animal models suggests that impaired glutamate cycling is widespread throughout the cortex, particularly implicating neuronal mitochondrial dysfunction, 41,42 though inadequate evidence is available in humans to firmly support this hypothesis. An alternate explanation for our finding is that neurovascular coupling is impaired in TLE.…”