Mechanisms contributing to synaptic Ca2+ signals and their heterogeneity in hair cells.

Sound coding at hair cell ribbon synapses is tightly regulated by Ca(2+). Here, we used patch-clamp, fast confocal Ca(2+) imaging and modeling to characterize synaptic Ca(2+) signaling in cochlear inner hair cells (IHCs) of hearing mice. Submicrometer fluorescence hotspots built up and collapsed at the base of IHCs within a few milliseconds of stimulus onset and cessation. They most likely represented Ca(2+) microdomains arising from synaptic Ca(2+) influx through Ca(V)1.3 channels. Synaptic Ca(2+) microdomains varied substantially in amplitude and voltage dependence even within single IHCs. Testing putative mechanisms for the heterogeneity of Ca(2+) signaling, we found the amplitude variability unchanged when blocking mitochondrial Ca(2+) uptake or Ca(2+)-induced Ca(2+) release, buffering cytosolic Ca(2+) by millimolar concentrations of EGTA, or elevating the Ca(2+) channel open probability by the dihydropyridine agonist BayK8644. However, we observed substantial variability also for the fluorescence of immunolabeled Ca(V)1.3 Ca(2+) channel clusters. Moreover, the Ca(2+) microdomain amplitude correlated positively with the size of the corresponding synaptic ribbon. Ribbon size, previously suggested to scale with the number of synaptic Ca(2+) channels, was approximated by using fluorescent peptide labeling. We propose that IHCs adjust the number and the gating of Ca(V)1.3 channels at their active zones to diversify their transmitter release rates.