EF-hand protein Ca 2+ buffers regulate Ca 2+ influx and exocytosis in sensory hair cells

• Published in: Proceedings of the National Academy of Sciences - PNAS 2015-03, Vol.112 (9)
• Main Authors: Pangršič, Tina, Gabrielaitis, Mantas, Michanski, Susann, Schwaller, Beat, Wolf, Fred, Strenzke, Nicola, Moser, Tobias
• Format: Article
• Language: English
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.1416424112

Ca 2+ ions serve as a key cellular signal and are tightly controlled. One mechanism to limit free Ca 2+ ions is buffering by Ca 2+ -binding proteins, which are strongly expressed in sensory hair cells of the ear. Here we studied how genetic disruption of the Ca 2+ -binding proteins parvalbumin-α, calbindin-D28k, and calretinin affects exocytosis and sound encoding at the synapses of mouse inner hair cells (IHCs) and spiral ganglion neurons (SGNs). Mutant IHCs showed increased exocytosis, but the sound-evoked spiking activity in SGNs was unaltered. Together with mathematical modeling, this finding indicates that a large fraction of exocytosis in mutant IHCs occurred outside synapses. We conclude that Ca 2+ -binding proteins shape presynaptic Ca 2+ signals to restrict exocytosis to active zones, thus enabling metabolically efficient sound encoding. EF-hand Ca 2+ -binding proteins are thought to shape the spatiotemporal properties of cellular Ca 2+ signaling and are prominently expressed in sensory hair cells in the ear. Here, we combined genetic disruption of parvalbumin-α, calbindin-D28k, and calretinin in mice with patch-clamp recording, in vivo physiology, and mathematical modeling to study their role in Ca 2+ signaling, exocytosis, and sound encoding at the synapses of inner hair cells (IHCs). IHCs lacking all three proteins showed excessive exocytosis during prolonged depolarizations, despite enhanced Ca 2+ -dependent inactivation of their Ca 2+ current. Exocytosis of readily releasable vesicles remained unchanged, in accordance with the estimated tight spatial coupling of Ca 2+ channels and release sites (effective “coupling distance” of 17 nm). Substitution experiments with synthetic Ca 2+ chelators indicated the presence of endogenous Ca 2+ buffers equivalent to 1 mM synthetic Ca 2+ -binding sites, approximately half of them with kinetics as fast as 1,2-Bis(2-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid (BAPTA). Synaptic sound encoding was largely unaltered, suggesting that excess exocytosis occurs extrasynaptically. We conclude that EF-hand Ca 2+ buffers regulate presynaptic IHC function for metabolically efficient sound coding.