Figure 3. Mito-Ca²⁺ is important for ribbon size and synapse integrity in mature hair cells.

(A-D) Representative images of mature neuromasts (5 dpf) immunostained with Ribeye b (magenta, ribbons) and MAGUK (green, postsynapses) after a 1 hr 0.1% DMSO (A), a 1 hr 2 μM Ru360 (B), a 30 min 10 μM Ru360 (C), or a 1 hr 10 μM Ru360 (D) treatment. Insets show three example synapses (white squares). E-F, Scatter plots show synapse counts (E), and ribbon area (F) in controls and in treatment groups. Ribbon areas, synapse numbers, and hair-cell counts are unaffected after a 1 hr 2 µM Ru360 treatment. Ribbon areas are larger and there are fewer synapses without significant loss of hair cells after a 30 min treatment with 10 µM Ru360 (F). After a 1 hr 10 µM Ru360 treatment there is an increase in ribbon area and a decrease in synapse (E) and hair-cell counts. N ≥ 9 neuromasts per treatment. Error bars in E-F represent SEM. An unpaired t-test was used in E and a Welch’s unequal variance t-test was used in F. ****p<0.0001. Scale bar = 5 µm in A, and 2 µm in inset.

Figure 3—figure supplement 1. Ribbon and postsynapse size in mature ALL neuromasts.

(A-B) Scatter plots show that ribbon areas (A) and postsynaptic density areas (B) within the same fish are similar between mature anterior lateral-line (ALL) and posterior lateral-line (PLL) neuromasts. (C) Scatter plots show ribbon areas in controls and after a 1 hr treatment with 2 µM Ru360 are similar in mature hair cells within the ALL. Ribbon sizes of untreated anterior lateral-line hair cells are from the same data as A and (C) n ≥ 10 neuromasts per treatment; error bars represent SEM; and a Welch’s unequal variance t-test was used for comparisons.

Figure 3—figure supplement 2. MCU block does not impact postsynapse size in mature hair cells.

(A) Quantification of postsynapse size assayed by MAGUK immunolabel in mature neuromasts indicate the treatments with 2 µM Ru360 and 10 µM Ru360 do not significantly alter postsynapse size compared to controls, n ≥ 9 neuromasts per treatment. Error bars represent SEM. A Welch’s unequal variance t-test was used for comparisons.