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

ABSTRACT: 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.

SUBMITTER: Frank T

PROVIDER: S-EPMC2657422 | biostudies-literature | 2009 Mar

REPOSITORIES: biostudies-literature

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

Frank Thomas T Khimich Darina D Neef Andreas A Moser Tobias T

Proceedings of the National Academy of Sciences of the United States of America 20090225 11

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( ...[more]

PMID: 19246382

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Project description:Merkel cell-neurite complexes are highly sensitive touch receptors comprising epidermal Merkel cells and sensory afferents. Based on morphological and molecular studies, Merkel cells are proposed to be mechanosensory cells that signal afferents via neurotransmission; however, functional studies testing this hypothesis in intact skin have produced conflicting results. To test this model in a simplified system, we asked whether purified Merkel cells are directly activated by mechanical stimulation. Cell shape was manipulated with anisotonic solution changes and responses were monitored by Ca2+ imaging with fura-2. We found that hypotonic-induced cell swelling, but not hypertonic solutions, triggered cytoplasmic Ca2+ transients. Several lines of evidence indicate that these signals arise from swelling-activated Ca2+-permeable ion channels. First, transients were reversibly abolished by chelating extracellular Ca2+, demonstrating a requirement for Ca2+ influx across the plasma membrane. Second, Ca2+ transients were initially observed near the plasma membrane in cytoplasmic processes. Third, voltage-activated Ca2+ channel (VACC) antagonists reduced transients by half, suggesting that swelling-activated channels depolarize plasma membranes to activate VACCs. Finally, emptying internal Ca2+ stores attenuated transients by 80%, suggesting Ca2+ release from stores augments swelling-activated Ca2+ signals. To identify candidate mechanotransduction channels, we used RT-PCR to amplify ion-channel transcripts whose pharmacological profiles matched those of hypotonic-evoked Ca2+ signals in Merkel cells. We found 11 amplicons, including PKD1, PKD2, and TRPC1, channels previously implicated in mechanotransduction in other cells. Collectively, these results directly demonstrate that Merkel cells are activated by hypotonic-evoked swelling, identify cellular signaling mechanisms that mediate these responses, and support the hypothesis that Merkel cells contribute to touch reception in the Merkel cell-neurite complex.

Dataset Information

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

Publications

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

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