Neuromast number and size varies with killifish age. (a) Representative images of DAPI-labeled superficial neuromasts from killifish opercula. Images show variation in neuromast size, rather than neuromasts from fish of different ages, to demonstrate the size variability seen in all age classes. Scale bar in the left image applies to all images. (b) Quantification of superficial neuromasts on the opercula of 2-, 3-, and 4–6-month-old killifish, data combined from both sexes. There is a significant age-dependent decrease in neuromast number (one-way ANOVA, F2,45 = 4.651, p = 0.019). (c) Neuromast quantification separated by sex. There is a significant decrease in neuromast number in 4–6-month-old male killifish (p = 0.017) but not female killifish (p = 0.249) (Mann–Whitney U tests). (d) Quantification of the number of hair cells (HCs) per superficial neuromast (sexes combined). We quantified hair cells in up to 10 opercular neuromasts per fish (or all visible neuromasts, for fish with fewer than 10), then calculated the average hair cell number for each fish. There was an age-dependent increase in hair cell number per neuromast (one-way ANOVA, F2,28 = 6.674, p = 0.043). Tukey’s multiple comparison posthoc tests show a significant difference between the 2- and 4–6-month-old age classes. (e) HC counts per neuromast separated by sex. There is a significant increase in average HC numbers in old females as compared to young females (p = 0.008) but no age difference in males (p = 0.065) (Mann–Whitney U tests). N = 12–13 fish per group for 2 and 4–6-month-old groups, even split between males and females. N = 5–6 for the 3-month-old group, with only male animals represented. *p < 0.05, **p < 0.01.
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