Figure 6.

Spontaneous and ACh evoked [Ca²⁺]_i transients at P14. (A) Resting G5 fluorescence (blue) and peak G5 ΔF/F₀ (green) showing superposition of spontaneous (“s” and ACh evoked events (“a”). (B) Spontaneous transients in 23 example 5 μm diameter ROIs, with transients in 3 ACh sensitive ROIs highlighted (1,2,3), at higher magnification of in (C). (D) ACh evoked transients were observed in a subset of cells (e.g., 1,2,3), with morphology consistent with calyx terminals contacting hair cells. (E) G5 ΔF/F₀ was blocked by 5μM ATR in a subset of cells (closed arrows) but persisted in other cells (open arrows). (F) ATR blocked long-lasting responses in ACh sensitive cells but not in other cells. (G) Confocal image of P14 fixed tissue showing the morphology of tdT positive hair cells (red) and synaptic contacts (anti-Tubulin) in an imaging slice similar to (A), suggesting ATR-sensitive cells are calyx endings. (H) ACh evoked transients could be distinguished from spontaneous transients by their kinetics, with ACh evoked events rising more slowly and lasting longer. (I) ACh evoked transients were largely blocked by ATR, while short-duration transients were not. *p = 0.05.