ATP-Evoked Intracellular Ca2+ Signaling of Different Supporting Cells in the Hearing Mouse Hemicochlea
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Hearing and its protection is regulated by ATP-evoked Ca2+ signaling in the supporting cells of the organ of Corti, however, the unique anatomy of the cochlea hampers observing these mechanisms. For the first time, we have performed functional ratiometric Ca2+ imaging (fura-2) in three different supporting cell types in the hemicochlea preparation of hearing mice to measure purinergic receptor-mediated Ca2+ signaling in pillar, Deiters’ and Hensen’s cells. Their resting [Ca2+]i was determined and compared in the same type of preparation. ATP evoked reversible, repeatable and dose-dependent Ca2+ transients in all three cell types, showing desensitization. Inhibiting the Ca2+ signaling of the ionotropic P2X (omission of extracellular Ca2+) and metabotropic P2Y purinergic receptors (depletion of intracellular Ca2+ stores) revealed the involvement of both receptor types. Detection of P2X2,3,4,6,7 and P2Y1,2,6,12,14 receptor mRNAs by RT-PCR supported this finding and antagonism by PPADS suggested different functional purinergic receptor population in pillar versus Deiters’ and Hensen’s cells. The sum of the extra- and intracellular Ca2+-dependent components of the response was about equal with the control ATP response (linear additivity) in pillar cells, and showed supralinearity in Deiters’ and Hensen’s cells. Calcium-induced calcium release might explain this synergistic interaction. The more pronounced Ca2+ leak from the endoplasmic reticulum in Deiters’ and Hensen’s cells, unmasked by cyclopiazonic acid, may also suggests the higher activity of the internal stores in Ca2+ signaling in these cells. Differences in Ca2+ homeostasis and ATP-induced Ca2+ signaling might reflect the distinct roles these cells play in cochlear function and pathophysiology.
KeywordsHemicochlea Ca2+ imaging ATP Pillar cells Deiters’ cells Hensen’s cells
Intracellular Ca2+ concentration
Calcium-induced calcium release
Half maximal effective concentration
Ethylene glycol-bis(2-aminoethylether)-N,N,N′,N′-tetraacetic acid
Real-time polymerase chain reaction
Sarco/endoplasmic reticulum Ca2+-ATPase
This work was supported by the Hungarian-French Collaborative R&I Programme on Biotechnologies (TÉT_10-1-2011-0421) and the Hungarian Research and Development Fund (NN107234 and K116654). We thank Peter Dallos and Claus-Peter Richter for teaching us the preparation of the hemicochlea and László Köles for his advices concerning purinergic receptor pharmacology.
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