MEASUREMENT OF CELLULAR RESPONSES TO TOXIC AGENTS USING A SILICON MICROPHYSIOMETER

H.G. WADAI1, J.C. OWICKI1, L.H. BRUNER2, K.R. MILLERS3, K.M. RALEY-SUSMAN4, P.R. PANFILI1, G.M.K. HUMPHRIES1 & J.W. PARCE1

1Molecular Devices Corp, Menlo Park, CA 94025; 2Procter & Gamble Co., Miami Valley Lab., Cincinnati, OH 45239-8707; 3Microbiological Associates, Rockville, MD 20850; 4Vassar College, Poughkeepsie, NY 12601

AATEX 1(3):154-164

Abstract
The silicon microphysiometer monitors cellular metabolism in vitro and has been used to detect and study biological responses to xenobiotics. This instrument uses a light addressable potentiometric sensor to measure millipH changes in micro-flow chambers maintained at 37C. Cells are immobilized in the flow chambers. Cell proton excretion can be measured as the acidification of the medium when medium flow through the chamber is stopped. The rate of acidification is a measure of catabolism, which produces lactic and carbonic acids. Through continual cycling of on and off periods of flow, nondestructive metabolic measurements may be made every few minutes. When a cell affecting agent is introduced into the fluid stream a change in acidification rate indicates either the stimulatory or toxic effect of the agent on the cells. Thus, rapid cellular responses to chemical agents can be detected within minutes and quantitated by the magnitude of acidification rate change.
Receptor-mediated cell activation by specific ligands, such as hormones, neurotransmitters, and growth factors, causes increases in acidification rate within minutes of receptor-ligand binding. This provides a receptor-specific response that can be monitored during exposure to materials as a possible indicator of cell-type specific toxicity. Receptor specific toxicity in hippocampal neurons has also been tested using extracellular acidification as the measurement of the toxicity of glutamate receptor overstimulation.
In tests for chemotherapeutic efficacy, antiviral drug activity and toxicity, acidification rates have been used as the cells activity and viability indicator. Agents, such as detergents, that produce irritancy or other non-specific toxicity have been evaluated for their effects on extracellular acidification rate and found to decrease acidification rate. Concentrations at which acidification rates are reduced by 50% were determined for test substances and correlated with animal ocular irritation test results. Recovery after insult is readily measured and may be an important index of irritancy or toxicity. These examples illustrate the broad range of cell affecting agents that cause cellular responses detectable in the microphysiometer using extracellular acidification rate.


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