A neuronal disruption in redox homeostasis elicited by ammonia alters the glycine/glutamate (GABA) cycle and contributes to MMA-induced excitability

Abstract

Hyperammonemia is a common finding in children with methylmalonic acidemia. However, its contribution to methylmalonate-induced excitotoxicty is poorly understood. The aim of this study was to evaluate the mechanisms by which ammonia influences in the neurotoxicity induced by methylmalonate (MMA) in mice. The effects of ammonium chloride (NH4Cl 3, 6, and 12 mmol/kg; s.c.) on electroencephalographic (EEG) and behavioral convulsions induced by MMA (0.3, 0.66, and 1 µmol/2 µL, i.c.v.) were observed in mice. After, ammonia, TNF-α, IL1β, IL-6, nitrite/nitrate (NOx) levels, mitochondrial potential (ΔΨ), reactive oxygen species (ROS) generation, Methyl-Tetrazolium (MTT) reduction, succinate dehydrogenase (SDH), and Na(+), K(+)-ATPase activity levels were measured in the cerebral cortex. The binding of [(3)H]flunitrazepam, release of glutamate-GABA; glutamate decarboxylase (GAD) and glutamine synthetase (GS) activity and neuronal damage [opening of blood brain barrier (BBB) permeability and cellular death volume] were also measured. EEG recordings showed that an intermediate dose of NH4Cl (6 mmol/kg) increased the duration of convulsive episodes induced by MMA (0.66 μmol/2 μL i.c.v). NH4Cl (6 mmol/kg) administration also induced neuronal ammonia and NOx increase, as well as mitochondrial ROS generation throughout oxidation of 2,7-dichlorofluorescein diacetate (DCFH-DA) to DCF-RS, followed by GS and GAD inhibition. The NH4Cl plus MMA administration did not alter cytokine levels, plasma fluorescein extravasation, or neuronal damage. However, it potentiated DCF-RS levels, decreased the ΔΨ potential, reduced MTT, inhibited SDH activity, and increased Na(+), K(+)-ATPase activity. NH4Cl also altered the GABA cycle characterized by GS and GAD activity inhibition, [(3)H]flunitrazepam binding, and GABA release after MMA injection. On the basis of our findings, the changes in ROS and reactive nitrogen species (RNS) levels elicited by ammonia alter the glycine/glutamate (GABA) cycle and contribute to MMA-induced excitability.

Altered Redox Homeostasis in Branched‐Chain Amino Acid Disorders, Organic Acidurias, and Homocystinuria

“…Intracellular ROS generation throughout oxidation of H 2 DCF has been detected in PA, cobalamin disorders (cblB, cblC, cblE, and cblG), MTHFR deficiency, and MSUD patient-derived fibroblasts [ 26 , 42 , 98 ]. Recently, mitochondrial ROS generation has been measured in the cortex of a mouse model of MMA (induced by MMA and ammonia) [ 147 ]. DCF measurements should be carefully considered, and extra controls for the assessment of ROS production are required.…”

Methylmalonic Aciduria may be an Innegligible Etiology of Cerebral Venous Sinus Thrombosis

“…In this case, similar to the infarcted brain lesions distributed at left frontal and right parietal lobes, bilateral cerebellar infarctions were likely to be associated with MMA-related CVST. The pathogenesis of cerebral infarctions in MMA is currently understood to be induced by the disruption of mitochondrial aerobic glucose oxidation as a result of excessive accumulation of organic acids, leading to a decrease in cellular energy generation and subsequent excitotoxicity [9]. Nevertheless, it cannot rule out another possibility, that is, cerebellar infarctions may be derived from PFO related arterial emboli [10], considering the fact that blood flow of PICA was interrupted in the right and absent in the left on CTA, even though the infarctions did not seem to be located within the typical PICA territory.…”

Pathophysiology of propionic and methylmalonic acidemias. Part 1: Complications

“…Additionally, propionic acid, methylmalonic acid and ammonia all induce inhibition of glutamate decarboxylase. Less glutamate is converted into GABA, leading to NMDA‐receptor activation and promoting excitotoxicity . In addition to these two routes, two more processes contribute to excitotoxicity.…”

“…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.…”

“…Less glutamate is converted into GABA, leading to NMDA-receptor activation and promoting excitotoxicity. 52,53 In addition to these two routes, two more processes contribute to excitotoxicity. First, glutamate dehydrogenase activity is inhibited by 2-methylcitric acid.…”