The spatial and temporal distribution changes of [Ca]i were studied in hair cells mechanically isolated from the semicircular canals of the frog. Cells were stimulated by depolarizing voltage commands during whole cell recordings and concurrently imaged at 247 Hz frame rate using single wavelength fluorescent dyes selective for Ca. Voltage-gated Ca entry occurred at selected sites (hotspots) located at the celi synaptic pole. During brief depolarizations, Ca fluorescence at individuai hotspots rose exponentially with a time constant of 70 ms and decayed with a bi-exponential time course (time constants: 0.16 and 2.5s). In cells exhibiting run-up or run-down of the Ca current, the fluorescence variation at distinct hotspots underwent independent amplitude changes, suggesting that distinct Ca channel clusters may be regulated differently by intracellular messengers. Results of applying the selective blocker nifedipine suggest that L-type Ca channels are not segregated within particular hotspots, but may be present at different densities within distinct channel clusters. The study of diffusion away from the hotspot indicates that Ca does not penetrate the nuclear region, no major diffusional barriers exists in the cytoplasm, and the Ca clearance machinery is also likely clustered at selected locations.
CALCIUM MICRODOMAIN DYNAMICS IN HAIR CELLS OF THE FROG SEMICIRCULAR CANAL
RISPOLI, Giorgio;MARTINI, Marta;ROSSI, Marialisa;
2001
Abstract
The spatial and temporal distribution changes of [Ca]i were studied in hair cells mechanically isolated from the semicircular canals of the frog. Cells were stimulated by depolarizing voltage commands during whole cell recordings and concurrently imaged at 247 Hz frame rate using single wavelength fluorescent dyes selective for Ca. Voltage-gated Ca entry occurred at selected sites (hotspots) located at the celi synaptic pole. During brief depolarizations, Ca fluorescence at individuai hotspots rose exponentially with a time constant of 70 ms and decayed with a bi-exponential time course (time constants: 0.16 and 2.5s). In cells exhibiting run-up or run-down of the Ca current, the fluorescence variation at distinct hotspots underwent independent amplitude changes, suggesting that distinct Ca channel clusters may be regulated differently by intracellular messengers. Results of applying the selective blocker nifedipine suggest that L-type Ca channels are not segregated within particular hotspots, but may be present at different densities within distinct channel clusters. The study of diffusion away from the hotspot indicates that Ca does not penetrate the nuclear region, no major diffusional barriers exists in the cytoplasm, and the Ca clearance machinery is also likely clustered at selected locations.I documenti in SFERA sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
