USE OF MICROPHYSIOMETRY FOR ANALYSIS OF HETEROLOGOUS ION CHANNELS EXPRESSED IN YEAST

Measurement of extracellular acidification rates by microphysiometry p rovides a means to analyze the function of ion channels expressed in y east cells. These measurements depend on the proton pumping action of the H+-ATPase, a central component of the yeast plasma membrane. We us ed microphysiometry to analyze the activity of two ion channels expres sed in yeast. In one example, an inwardly rectifying K+ channel, gpIRK 1, provides a potassium uptake function when expressed in a potassium transporter-defective yeast strain. Rates of acidification in gpIRK1-e xpressing 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 nonselectiv e cation channel, the influenza virus M2 protein, is believed to inter fere with the maintenance of the electrochemical proton gradient by th e H+-ATPase. In cells expressing the M2 channel, addition of inhibitor s increases the rate of proton extrusion. Moreover, functional differe nces between two M2 inhibitors, amantadine and BL-1743, are distinguis hed 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 chan nel activity.