Journal
EXPERIMENTAL EYE RESEARCH
Volume 76, Issue 6, Pages 715-723Publisher
ACADEMIC PRESS LTD- ELSEVIER SCIENCE LTD
DOI: 10.1016/S0014-4835(03)00052-6
Keywords
retinal Muller cells; retinal pigment epithelial cells; glucose metabolism; mitochondria; aerobic and anaerobic glycolysis; Pasteur effect; metabolic stress
Categories
Funding
- NEI NIH HHS [R01 EY010015-10, EY 10015, EY 05230] Funding Source: Medline
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The purpose of the present experiments was to enhance understanding of the factors that are critical for the survival of retinal cells exposed to mitochondrial inhibition. Confluent cultures of Muller cells (rMC-1) and human retinal pigment epithelial cells (hRPE) were incubated in Dulbecco's minimal essential medium in the presence and absence of 1 X 10(-5) m Antimycin A, an inhibitor of mitochondrial electron transport. To modulate the rates of aerobic and anaerobic glycolysis, cells were incubated in media containing varying concentrations of glucose and 1-100 muM of iodoacetic acid (IAA), an inhibitor of glyceraldehdye-3-phosphate dehydrogenase (G3PDH). Measurements were made of G3PDH, lactic acid production, and cellular ATP levels, along with an examination of cellular morphology, the latter providing an index of cellular viability. Control rMC-1 and hRPE produced lactate aerobically, respectively, at 0.48 and 1.50 mumol hr(-1)/10(6) cells. Anaerobically, lactate production increased 2-fold in rMC-1 and 3-fold in hRPE. Anaerobic ATP levels in both types of cells were maintained at control levels over 8 hr. Experimental conditions were sought that would modulate only the capacity of rMC-1 and hRPE to increase glycolysis following mitochondrial inhibition, i.e. alter their Pasteur effect. We used low concentrations of LA-A to partially inhibit G3PDH. Incubation of rMC-1 with IAA for 6 hr caused a graded inhibition of G3PDH: 70% inhibition with 1 muM, 90% with 5 muM, 97% with 10 muM, and 100% with 100 muM. While the aerobic and anaerobic rates of lactic acid production were not altered by 1 muM IAA, both were suppressed completely by 100 muM IAA. However, incubation of rMC-I with 5 muM IAA caused a decrease of 30% in the rate of anaerobic lactic acid production but no change in the rate of aerobic glycolysis. Moreover, with 5 muM IAA, rMC-1 incubated aerobically maintained ATP levels, but anaerobic ATP content decreased to a low level and cell morphology and viability were compromised. Essentially similar results were observed with hRPE. Both rMC-I and hRPE are remarkably resistant to mitochondrial inhibition. This resistance is linked directly to the magnitude of the increase in the Pasteur effect. When the capacity of rMC-1 and hRPE to generate a Pasteur effect is selectively curtailed, these cells no longer are resistant to mitochondrial inhibition. It is suggested that in an intact tissue the ability of a cell to withstand a metabolic challenge will depend very much on the adequacy of the supply of glucose. Even a small limitation in the availability of this utilizable substrate and in the rate of the compensatory increase in the rate of anaerobic glycolysis could put the cell at greater risk during the challenge. (C) 2003 Elsevier Science Ltd. All rights reserved.
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