Determination of Glucose Utilization Rates in Cultured Astrocytes and Neurons with [14C]deoxyglucose: Progress, Pitfalls, and Discovery of Intracellular Glucose Compartmentation
Autor: | Louis Sokoloff, Nancy F. Cruz, Gerald A. Dienel, Bernard F. Driscoll |
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Rok vydání: | 2015 |
Předmět: |
0301 basic medicine
medicine.medical_specialty Hypoglycemia Biology Biochemistry 03 medical and health sciences Cellular and Molecular Neuroscience chemistry.chemical_compound 0302 clinical medicine Internal medicine medicine Extracellular Hexokinase Deoxyglucose General Medicine Metabolism medicine.disease 030104 developmental biology Endocrinology medicine.anatomical_structure chemistry Neuron 030217 neurology & neurosurgery Intracellular Astrocyte |
Zdroj: | Neurochemical Research. 42:50-63 |
ISSN: | 1573-6903 0364-3190 |
Popis: | 2-Deoxy-d-[14C]glucose ([14C]DG) is commonly used to determine local glucose utilization rates (CMRglc) in living brain and to estimate CMRglc in cultured brain cells as rates of [14C]DG phosphorylation. Phosphorylation rates of [14C]DG and its metabolizable fluorescent analog, 2-(N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)-2-deoxyglucose (2-NBDG), however, do not take into account differences in the kinetics of transport and metabolism of [14C]DG or 2-NBDG and glucose in neuronal and astrocytic cells in cultures or in single cells in brain tissue, and conclusions drawn from these data may, therefore, not be correct. As a first step toward the goal of quantitative determination of CMRglc in astrocytes and neurons in cultures, the steady-state intracellular-to-extracellular concentration ratios (distribution spaces) for glucose and [14C]DG were determined in cultured striatal neurons and astrocytes as functions of extracellular glucose concentration. Unexpectedly, the glucose distribution spaces rose during extreme hypoglycemia, exceeding 1.0 in astrocytes, whereas the [14C]DG distribution space fell at the lowest glucose levels. Calculated CMRglc was greatly overestimated in hypoglycemic and normoglycemic cells because the intracellular glucose concentrations were too high. Determination of the distribution space for [14C]glucose revealed compartmentation of intracellular glucose in astrocytes, and probably, also in neurons. A smaller metabolic pool is readily accessible to hexokinase and communicates with extracellular glucose, whereas the larger pool is sequestered from hexokinase activity. A new experimental approach using double-labeled assays with DG and glucose is suggested to avoid the limitations imposed by glucose compartmentation on metabolic assays. |
Databáze: | OpenAIRE |
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