The Role of Glycolysis and Gluconeogenesis in the Cytoprotection of Neuroblastoma Cells against 1-Methyl 4-Phenylpyridinium Ion Toxicity

doi:10.1016/S0161-813X(02)00110-9

NeuroToxicology

Volume 24, Issue 1, January 2003, Pages 137-147

https://doi.org/10.1016/S0161-813X(02)00110-9 Get rights and content

Abstract

1-Methyl-4-phenylpyridinium (MPP+) is a mitochondrial Complex I inhibitor and is frequently used to investigate the pathological degeneration of neurons associated with Parkinson’s disease (PD). In vitro, extracellular concentration of glucose is one of the most critical factors in establishing the vulnerability of neurons to MPP+ toxicity. While glucose is the primary energy fuel for the brain, central nervous system (CNS) neurons can also take up and utilize other metabolic intermediates for energy. In this study, we compared various monosaccharides, disaccharides, nutritive/non-nutritive sugar alcohols, glycolytic and gluconeogenic metabolic intermediates for their cytoprotection against MPP+ in murine brain neuroblastoma cells. Several monosaccharides were effective against MPP+ (500 μM) including glucose, fructose and mannose, which restored cell viability to 109 ± 5%, 70 ± 5%, 99 ± 3% of live controls, respectively. Slight protective effects were observed in the presence of 3-phosphoglyceric acid and glucose-6-phosphate; however, no protective effects were exhibited by galactose, sucrose, sorbitol, mannitol, glycerol or various gluconeogenic and ketogenic amino acids. On the other hand, fructose 1,6 bisphosphate and gluconeogenic energy intermediates [pyruvic acid, malic acid and phospho(enol)pyruvate (PEP)] were neuroprotective against MPP+. The gluconeogenic intermediates elevated intracellular levels of ATP and reduced propidium iodide (PI) nucleic acid staining to live controls, but did not alter the MPP+-induced loss of mitochondrial O₂ consumption. These data indicate that malic acid, pyruvic acid and PEP contribute to anaerobic substrate level phosphorylation. The use of hydrazine sulfate to impede gluconeogenesis through PEP carboxykinase (PEPCK) inhibition heightened the protective effects of energy substrates possibly due to attenuated ATP demands from pyruvate carboxylase (PC) activity and pyruvate mitochondrial transport. It was concluded from these studies that several metabolic intermediates are effective in fueling anaerobic glycolysis during mitochondrial inhibition by MPP+.

Section snippets

INTRODUCTION

Administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) can lead to selective degeneration of dopaminergic neurons in the substantia nigra, resembling pathological features of Parkinson’s disease (PD) (Langston et al., 1983). 1-Methyl-4-phenylpyridinium (MPP+), the neurotoxic metabolite of MPTP, can reduce energy metabolism by impeding oxidative phosphorylation (OXPHOS) through inhibiting NADH dehydrogenase in Complex I of the electron transport chain (ETC) (Suzuki et al., 1990).

Materials

Mouse brain neuro-2A neuroblastoma cells (N2A) were obtained from American Type Culture Collection (Manassas, VA, USA). Dulbecco’s Modified Eagle Medium (DMEM), l-glutamine, fetal bovine serum—heat inactivated (FBS), Hank’s balanced salt solution (HBSS), phosphate buffered saline (PBS) and penicillin/streptomycin were supplied by Fischer Scientific, Mediatech (Pittsburgh, PA, USA). All other chemicals and supplies were purchased from Sigma Chemical (St. Louis, MO, USA).

Cell Culture Growth and Experimental Conditions

N2A cells were chosen for

RESULTS

A concentration–time curve for MPP+ toxicity was established and it was determined that 500 μM of MPP+ was adequate to achieve significant, but not total cell death at 24–36 h (data not shown). N2A cells were pretreated with experimental compounds for 2 h prior to treatment with MPP+ and samples were returned to the incubator for 24–36 h at 37 °C in 5% CO₂/atmosphere. Each experimental batch was monitored with a timed control, to ensure a desired level of toxicity had been achieved. Moreover,

DISCUSSION

In the present study, we examined the propensity of alternative energy substrates to support anaerobic glycolysis and provide neuroprotection against MPP+. The data indicate that in addition to glucose, several monosaccharides and metabolic intermediates are capable of attenuating MPP+-induced toxicity. Monosaccharides are the smallest molecular form of carbohydrates and are the building blocks of all complex carbohydrates. The most common monosaccharides are glucose, fructose, galactose and

Acknowledgements

This work was supported by grants received from the National Institutes of Health (NCRR 03020 and GM 08111).

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The Role of Glycolysis and Gluconeogenesis in the Cytoprotection of Neuroblastoma Cells against 1-Methyl 4-Phenylpyridinium Ion Toxicity

Abstract

Section snippets

INTRODUCTION

Materials

Cell Culture Growth and Experimental Conditions

RESULTS

DISCUSSION

Acknowledgements

Neuropharmacology

Brain Res. Brain Res. Rev.

Biochem. Biophys. Res. Commun.

Toxicol. In Vitro

Brain Res. Brain Res. Protoc.

Neurochem. Int.

Brain Res.

Neuropharmacology

J. Biol. Chem.

Brain Res.

Immunopharmacology

Intestinal digestion and absorption of sucrose in experimental diabetes

Acta Diabetol. Lat.

Effects of 1-methyl-4-phenylpyridinium on isolated rat brain mitochondria: evidence for a primary involvement of energy depletion

J. Neurochem.

Resting regional cerebral glucose metabolism in advanced Parkinson’s disease studied in the off and on conditions with [(18) F] FDG-PET

Mov. Disord.

Use of fructose, xylitol, or sorbitol as a sweetener in diabetes mellitus

Diabetes Care

Interaction among mitochondria, mitogen-activated protein kinases, and nuclear factor-kappa B in cellular models of Parkinson’s disease

J. Neurochem.