In vivo assessment of hair cell damage and developmental toxicity caused by gestational caffeine exposure using zebrafish (Danio rerio) models.

The aim of the present study was to evaluate hair cell damage and associated developmental toxicity caused by gestational caffeine exposure. We exposed embryos to various caffeine concentrations (25μM, 125μM, 250μM, and 500μM) and evaluated developmental toxicity of the embryos at 72 and 120h and hair cell damage at 120h after fertilization. The average number of total hair cells within four neuromasts exposed to various concentrations of caffeine was compared with that of the control group. To seek the underlying mechanisms, TUNEL and DASPEI assay were carried out to evaluate hair cell apoptosis and mitochondrial damage, respectively. Morphologic abnormality, mortality, hatching rate, and heart rate were also evaluated. Caffeine induced significant hair cell damage compared with control group (p<0.01, control; 35.64±10.48 cells, 500μM caffeine; 23.32±12.14 cells, n=25-30). Significant increase in the hair cell apoptosis was confirmed in a dose-dependent manner (p<0.01, TUNEL assay) and the mitochondrial damage in high caffeine concentrations (250, 500μM) (p<0.01, DASPEI assay).Morphologic abnormalities were significantly increased in high caffeine concentrations (250 or 500μM) for body shape, notochord, and heart at both 3-, and 5-dpf. The control group exhibited 3.3% mortality which increased up to 11.6% at 500μM caffeine. Rapid hatching was present at 48h (control; 46.6%, 500μM caffeine; 100%). In conclusion, gestational caffeine exposure caused significant hair cell damage and developmental toxicities in zebrafish at early developmental stages.