CLUSTERING OF CA2-ACTIVATED K+ CHANNELS AT FLUORESCENTLY LABELED PRESYNAPTIC ACTIVE ZONES OF HAIR-CELLS( CHANNELS AND CA2+)

Electrical resonance, which in some hair cells provides a mechanism fo r frequency tuning, is mediated by clusters of Ca2+ channels and Ca2+- activated K+ channels that have been proposed to occur at presynaptic active zones. To localize Ca2+ channels on the cellular surface, we lo aded hair cells from the frog's sacculus with the Ca2+ indicator fluo- 3 and imaged them by fluorescence confocal microscopy. When a cell was depolarized, we observed on its basolateral surface several foci of t ransiently enhanced fluorescence due to local Ca2+ influx. After protr acted recording, each cell displayed on average 18 brightly and perman ently fluorescent spots at the same positions. We mapped these spots i n four hair cells and compared their locations with those of presynapt ic active zones, as determined from transmission electron micrographs of serial sections through the same cells. The results demonstrated th at enhanced fluo-3 fluorescence marks active zones. Measurement of cur rents through membrane patches at fluorescently labeled active zones d emonstrated that both voltage-activated Ca2+ channels and Ca2+-activat ed K+ channels occur there. These results confirm that the ion channel s involved in electrical tuning and synaptic transmission by hair cell s cluster together at presynaptic active zones.