Understanding the dynamics of cellular responsiveness to modifications of metabolic substrates in perifusion.
A novel microperifusion system with capabilities for continuous, real-time, potentiometric monitoring of extracellular hydrogen ion concentration has been used to define the response of HeLa cells to abrupt changes in extracellular energy sources or introduction of an inhibitor of glycolysis. Glycolytic inhibition, induced by removal of glucose or introduction of iodoacetate, each led to a rapid, continuous decrease in acid release. The response to iodoacetate took longer than removal of glucose, perhaps due to the time required for binding and activation. Once inhibition began, however, the rate of change was greater than following glucose removal. Conversely, recovery time following iodoacetate inhibition was much slower than with glucose removal. Unlike the response to short-term glucose depletion, a second pulse of iodoacetate resulted in a faster response followed by an even longer recovery time. The response to switching between glucose and glutamine began almost without evident delay. The response patterns revealed that HeLa cells prefer glutamine to glucose, but, in the presence of both energy sources, some glucose continues to be used. In summary, these results indicate that continuous, real-time monitoring of the kinetics of hydrogen-ion release can be used to gain new insights into the dynamics of cellular response to perturbations of extracellular energy sources.
Reproductive Sciences Program, University of Michigan, Ann Arbor 48109-0404.
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Brand
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Lyons
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Midgley
D004734:Energy Metabolism; D005947:Glucose / Q000378:metabolism; D005973:Glutamine / Q000378:metabolism; D006019:Glycolysis / Q000502:physiology; D006367:HeLa Cells; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007461:Iodoacetates / Q000494:pharmacology; D019807:Iodoacetic Acid; D013379:Substrate Specificity; D013997:Time Factors
D007461:Iodoacetates; D005973:Glutamine; D005947:Glucose; D019807:Iodoacetic Acid