VOLTAGE-DEPENDENT BLOCK BY NEOMYCIN OF THE ATP-INDUCED WHOLE-CELL CURRENT OF GUINEA-PIG OUTER HAIR-CELLS

1. The effects of externally applied ATP and neomycin on whole cell cu rrents of isolated guinea pig cochlear outer hair cells (OHCs) were st udied using the whole cell voltage-clamp technique. In OHCs held at -7 0 mV, ATP activated a large inward current. In the presence of neomyci n, the ATP-induced whole cell current activated along a relatively una ltered time course, but the current then decreased to a reduced steady level. The neomycin inhibition of the ATP-induced current was dose de pendent. The half-inhibitory concentration (IC50) of neomycin measured at steady state was estimated to be 90 muM. 2. Neomycin inhibition of the ATP response could not be reversed by increasing the concentratio n of ATP, indicating that the effect was noncompetitive. The inhibitio n was voltage dependent and was greatly reduced when OHCs were held at positive potentials. 3. Cells treated with 100 muM ATP gave maximal c urrent responses. Addition of neomycin substantially increased membran e current noise of the 100 muM ATP responses. When neomycin concentrat ion was varied from 10 to 500 muM, the current noise level peaked betw een 50 and 100 muM. The noise increase was observed at negative holdin g potentials but not at positive potentials. 4. The neomycin-induced w hole cell current noise was used to estimate the size of the underlyin g elementary current. The ATP-induced single channel current of OHCs a t -70 mV was estimated to be approximately 0.3 pA. The number of ATP-a ctivated channels in a single OHC was estimated to be in the range of a few thousand. 5. The characteristics of the neomycin inhibition of A TP-induced currents were consistent with an open channel blocking mech anism. Analysis of the voltage dependence of the steady state neomycin inhibition suggested a neomycin binding site at an electrical distanc e of 0.3 from the extracellular side.