Nanophysiology approach reveals diversity in calcium microdomains across Zebrafish retinal bipolar ribbon synapses

Rapid and high local calcium (Ca2+) signals are essential for triggering neurotransmitter release from presynaptic terminals. In specialized bipolar ribbon synapses of the retina, these local Ca2+ signals control multiple processes, including the priming, docking, and translocation of vesicles on the ribbon before exocytosis, endocytosis, and the replenishment of release-ready vesicles to the fusion sites for sustained neurotransmission. However, our knowledge about Ca2+ signals along the axis of the ribbon active zone is limited. Here, we used fast confocal quantitative dual-color ratiometric line-scan imaging of a fluorescently labeled ribbon binding peptide and Ca2+ indicators to monitor the spatial and temporal aspects of Ca2+ transients of individual ribbon active zones in zebrafish retinal rod bipolar cells (RBCs). We observed that a Ca2+ transient elicited a much greater fluorescence amplitude when the Ca2+ indicator was conjugated to a ribeye-binding peptide than when using a so..., Rearing of zebrafish. Male and female zebrafish (Danio rerio; 16 ~20 months) were raised under a 14 h light/10 h dark cycle and housed according to NIH guidelines and the University of Tennessee Health Science Center (UTHSC) Guidelines for Animals in Research. All procedures were approved by the UTHSC Institutional Animal Care and Use Committee (IACUC; protocol # 23-0459).  
Isolation of zebrafish retinal RBCs. Dissociation of RBCs was performed using established procedures 95. Briefly, retinas were dissected from zebrafish eyes and incubated in hyaluronidase (1100 units/ml) for 20 minutes. The tissue was washed with a saline solution containing 120 mM NaCl, 2.5 mM KCl, 0.5 mM CaCl2, 1 mM MgCl2, 10 mM glucose, and 10 mM HEPES, pH = 7.4] before being cut into quadrants. Each quadrant was incubated at room temperature for 25 to 40 min in the same saline solution, to which was added DL-cysteine and papain (20-30 units/ml; Sigma Millipore, St. Louis, MO) and triturated using ..., # Data from: Nanophysiology approach reveals diversity in calcium microdomains across Zebrafish retinal bipolar ribbon synapses

Dataset DOI: [10.5061/dryad.qfttdz0vh](https://doi.org/10.5061/dryad.qfttdz0vh)

## Description of the data and file structure

The present dataset compares calcium domains in retinal rod bipolar cells using free and ribeye-bound calcium indicators. Computational models were established to estimate the spatial distribution of the Ca2+ signals accurately. The ultrastructure of the zebrafish retina, obtained using Serial Block-Face Scanning Electron Microscopy, was used to count and measure the rod bipolar cell synaptic ribbons and active zones. Experiment data and quantities contained in the spreadsheets are described below under `Description` and `Glossary` sections. For a detailed description of the experiments outlined here, see the associated article at [https://elifesciences.org/reviewed-preprints/105875](https://elifesciences.org/reviewed-preprints/105875...,