Studies on the ion channels in the plasma membrane of isolated guinea pig outer hair cells.

Abstract

Membrane potential controls the length of cochlear outer hair cells (OHCs). Ion channels in the plasma membrane of OHCs may play a role in auditory transduction by affecting membrane potential changes, thus modulating the mechanical properties of OHCs. In this study, the whole cell voltage-clamp technique was used to study kinetic and pharmacological properties of ionic currents in response to depolarization and to extracellularly applied adenosine triphosphate (ATP) in guinea-pig isolated OHCs. Depolarization of OHCs evoked voltage-dependent Ca$\sp{++}$ currents and K$\sp+$ currents. The Ca$\sp{++}$ current was a small inward component in the whole cell current. Kinetic and ionic properties of the voltage-dependent Ca$\sp{++}$ current suggested that L-type Ca$\sp{++}$ channels existed in OHCs. The dominant current in OHCs was K$\sp+$ current, which was subdivided into delayed rectifier and Ca$\sp{++}$ dependent K$\sp+$ currents. The OHC delayed rectifier K$\sp+$ current had novel pharmacological properties. Agents usually considered to be Ca$\sp{++}$ channel ligands (dihydropyridines and verapamil) were found to directly inhibit this K$\sp+$ current. The presence of ATP receptors in OHCs suggests that ATP may be associated with signaling or regulatory mechanisms in the cochlea. The studies of this dissertation showed that a well known ototoxic aminoglycoside-neomycin-reversibly blocked ATP-activated channels of OHCs in a voltage-dependent fashion. The kinetics and the voltage dependence of the neomycin block of the ATP-induced OHC whole cell current suggested an open channel block mechanism. Variance analysis and spectral density analysis were used to estimate the single channel current and the total number of ATP-activated channels on OHCs. The two methods gave similar estimates of the single channel parameters. The voltage-dependence of the neomycin steady state inhibitory effect suggested that the neomycin binding site was about 26% of the electrical distance into the OHC membrane electrical field from the outside. The results of this dissertation extended our understanding of the properties of OHC ion channels and provide new findings on the pharmacological properties of these channels. These results suggest a way to test the role of specific classes of ion channels in the function of the cochlea.