Use of microphysiometry for analysis of heterologous ion channels expressed in yeast.
Measurement of extracellular acidification rates by microphysiometry provides a means to analyze the function of ion channels expressed in yeast cells. These measurements depend on the proton pumping action of the H(+)-ATPase, a central component of the yeast plasma membrane. We used microphysiometry to analyze the activity of two ion channels expressed in yeast. In one example, an inwardly rectifying K+ channel, gpIRK1, provides a potassium uptake function when expressed in a potassium transporter-defective yeast strain. Rates of acidification in gpIRK1-expressing cells directly reflect channel function. Addition of cesium, an inhibitor of gpIRK1 activity, results in an immediate reduction in acidification rates. In a second example, expression of a nonselective cation channel, the influenza virus M2 protein, is believed to interfere with the maintenance of the electrochemical proton gradient by the H(+)-ATPase. In cells expressing the M2 channel, addition of inhibitors increases the rate of proton extrusion. Moreover, functional differences between two M2 inhibitors, amantadine and BL-1743, are distinguished by the microphysiometer. This application demonstrates the utility of the microphysiometer for functional studies of ion channels; it is adaptable to a screening process for compounds that modulate ion channel activity.
Hahnenberger
K M
KM
Bristol-Myers Squibb Pharmaceutical Research Institute, Seattle, WA 98121, USA.
Krystal
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M
Esposito
K
K
Tang
W
W
Kurtz
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D015374:Biosensing Techniques; D018408:Patch-Clamp Techniques; D015221:Potassium Channels / Q000235:genetics / Q000378:metabolism*; D024661:Potassium Channels, Inwardly Rectifying; D011199:Potentiometry / Q000379:methods*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics*
D015221:Potassium Channels; D024661:Potassium Channels, Inwardly Rectifying; D011994:Recombinant Proteins