Differences in mitochondrial function in homogenated samples from healthy and epileptic specific brain tissues revealed by high-resolution respirometry

Abstract

Mitochondrial dysfunction and oxidative stress are strongly implicated in neurodegenerative diseases and epilepsy. Strikingly, neurodegenerative diseases show regional specificity in vulnerability and follow distinct patterns of neuronal loss. A challenge is to understand, why mitochondria fail in particular brain regions under specific pathological conditions. A potential explanation could be provided by regional or cellular specificity of mitochondrial function. We applied high-resolution respirometry to analyze the integrated Complex I- and II (CI and CII)-linked respiration, the activity of Complex IV, and the combined CI&II-linked oxidative phosphorylation (OXPHOS)- and electron-transfer system (ETS)-capacity in microsamples obtained from distinct regions of the mouse brain. We compared different approaches to assess mitochondrial density and suggest flux control ratios as a valid method to normalize respiration to mitochondrial density. This approach revealed significant differences of CI- and CII-linked OXPHOS capacity and coupling control between motor cortex, striatum, hippocampus and pons of naïve mice. CI-linked respiration was highest in motor cortex, while CII-linked respiration predominated in the striatum. To investigate if this method could also determine differences in normal and disease states within the same brain region, we compared hippocampal homogenates in a chronic epilepsy model. Three weeks after stereotaxic injection of kainate, there was a down-regulation of CI- and upregulation of CII-linked respiration in the resulting epileptic ipsilateral hippocampus compared to the contralateral one. In summary, respirometric OXPHOS analysis provides a very sensitive diagnostic approach using small amounts of distinct brain tissues. In a single assay, information is obtained on numerous OXPHOS parameters as indicators of tissue-specific mitochondrial performance.

Distinct differences in rates of oxygen consumption and ATP synthesis of regionally isolated non‐synaptic mouse brain mitochondria

“…Furthermore, the absolute OCR of striatal mitochondria, when provided succinate, was larger than mitochondria of the cerebral cortex, which was not the case when using pyruvate and malate as respiratory substrates. Several recent studies have reported elevations in both coupled and uncoupled OCRs of striatal mitochondria when provided succinate as substrate, which is in line with our results (Burtscher et al, ; Pandya et al, ; Subramaniam, Vergnes, Franich, Reue, & Chesselet, ). A general hypothesis on the pathogenesis of Huntington’s disease, a disease primarily affecting the striatum, is that the mitochondria of the striatum are more sensitive to metabolic insults and therefore more susceptible to neurodegeneration (Browne & Beal, ; Damiano, Galvan, Deglon, & Brouillet, ; Mochel & Haller, ).…”

“…Compared to the literature, these values signify well-functioning mitochondria, validating the quality of our preparation (Burtscher, Zangrandi, Schwarzer, & Gnaiger, 2015;Cardoso, Santos, Seica, & Moreira, 2010;Czerniczyniec et al, 2015;Dagani et al, 1988;Lai et al, 1977;Sauerbeck et al, 2011;Sims, 1990;Varela, Schwartz, & Horvath, 2016). We observed that hippocampal mitochondria exhibited a larger proton leak, with both substrate conditions, resulting in reduced RCR values for this region.…”

High-resolution respirometry of fine-needle muscle biopsies in pre-manifest Huntington’s disease expansion mutation carriers shows normal mitochondrial respiratory function

“…No significant changes between HDEMCs and controls were detected in these parameters (Fig 2A). In addition complex I and complex II activity were examined in the limited amount of tissue sample, by accepting the restriction that complex II activity could be determined in an uncoupled state only, as it was described before [22, 29]. Neither complex I activity nor complex II activity showed significant changes between the HDEMCs and the control group.…”

Tert-buthylhydroquinone pre-conditioning exerts dual effects in old female rats exposed to 3-nitropropionic acid

“…Briefly, 3-NP inhibits the succinate dehydrogenase (SDH, or mitochondrial complex II) catalytic site; blocking the electron transport chain and the Krebs cycle, decreasing ATP synthesis and promoting reactive oxygen species (ROS) formation, thus fomenting mitochondrial dysfunction [7] and neuronal death [8], [9].…”