2‐Methylcitric acid impairs glutamate metabolism and induces permeability transition in brain mitochondria

Abstract

Accumulation of 2-methylcitric acid (2MCA) is observed in methylmalonic and propionic acidemias, which are clinically characterized by severe neurological symptoms. The exact pathogenetic mechanisms of brain abnormalities in these diseases are poorly established and very little has been reported on the role of 2MCA. In the present work we found that 2MCA markedly inhibited ADP-stimulated and uncoupled respiration in mitochondria supported by glutamate, with a less significant inhibition in pyruvate plus malate respiring mitochondria. However, no alterations occurred when a-ketoglutarate or succinate was used as respiratory substrates, suggesting a defect on glutamate oxidative metabolism. It was also observed that 2MCA decreased ATP formation in glutamate plus malate or pyruvate plus malate-supported mitochondria. Furthermore, 2MCA inhibited glutamate dehydrogenase activity at concentrations as low as 0.5 mM. Kinetic studies revealed that this inhibitory effect was competitive in relation to glutamate. In contrast, assays of osmotic swelling in non-respiring mitochondria suggested that 2MCA did not significantly impair mitochondrial glutamate transport. Finally, 2MCA provoked a significant decrease in mitochondrial membrane potential and induced swelling in Ca 2+ -loaded mitochondria supported by different substrates. These effects were totally prevented by cyclosporine A plus ADP or ruthenium red, indicating induction of mitochondrial permeability transition. Taken together, our data strongly indicate that 2MCA behaves as a potent inhibitor of glutamate oxidation by inhibiting glutamate dehydrogenase activity and as a permeability transition inducer, disturbing mitochondrial energy homeostasis. We presume that 2MCA-induced mitochondrial deleterious effects may contribute to the pathogenesis of brain damage in patients affected by methylmalonic and propionic acidemias.

Pathophysiology of propionic and methylmalonic acidemias. Part 2: Treatment strategies

“…Whereas Amaral et al could not confirm an altered glutamine synthetase activity; they demonstrated that 2‐methylcitric acid inhibited glutamate dehydrogenase activity towards the formation of α‐ketoglutarate in rat brain mitochondria, through competition with glutamate. This affects the availability of α‐ketoglutarate for proper functioning of the citric acid cycle.…”

“…This affects the availability of α‐ketoglutarate for proper functioning of the citric acid cycle. In addition, Amaral et al demonstrated that 2‐methylcitric acid induced mitochondrial permeability transition pore opening, that ultimately led to mitochondrial energy dysfunction, as illustrated by decreased ATP synthesis.…”

Rhabdomyolysis in organic acidemia patients manifesting with metabolic decompensation

“…Some researchers reported on the dysfunction of mitochondrial energy metabolism in patients with MMA and PA . Patients with MMA had mitochondrial abnormalities, reduced oxygen uptake, significantly lowered enzyme activities including cytochrome C oxidase, and respiratory chain complexes I‐IV in various tissues.…”

Pathophysiology of propionic and methylmalonic acidemias. Part 1: Complications

“…In addition to these two routes, two more processes contribute to excitotoxicity. First, glutamate dehydrogenase activity is inhibited by 2‐methylcitric acid . Glutamate oxidation is reduced, ATP production is reduced and the glutamine/glutamate ratio is altered, also inducing excitotoxicity .…”

“…First, glutamate dehydrogenase activity is inhibited by 2‐methylcitric acid . Glutamate oxidation is reduced, ATP production is reduced and the glutamine/glutamate ratio is altered, also inducing excitotoxicity . Second, oxidative stress (both reactive oxygen species [ROS] and reactive nitrogen species) leads to decreased Na + /K + /ATPase activity, mitochondrial depolarization and decreased ATP synthesis, again promoting excitotoxicity.…”