Developmental maturation of presynaptic ribbon numbers in chicken basilar-papilla hair cells and its perturbation by long-term overexpression of Wnt9a

The avian basilar papilla is a valuable model system for exploring the developmental determination and differentiation of sensory hair cells and their innervation. In the mature basilar papilla, hair cells form a well-known continuum between two extreme types—tall and short hair cells—that differ strikingly in their innervation. Previous work identified Wnt9a as a crucial factor in this differentiation. Here, we quantified the number and volume of immunolabelled presynaptic ribbons in tall and short hair cells of chickens, from developmental stages shortly after ribbons first appear to the mature posthatching condition. Two longitudinal locations were sampled, responding to best frequencies of approximately 1 kHz and approximately 5.5 kHz when mature. We found significant reductions of ribbon number during normal development in the tall-hair-cell domains, but stable, low numbers in the short-hair-cell domains. Exposing developing hair cells to continuous, excessive Wnt9a levels (through virus-mediated overexpression) led to transiently abnormal high numbers of ribbons and a delayed reduction of ribbon numbers at all sampled locations. Thus, (normally) short-hair-cell domains also showed tall-hair-cell like behaviour, confirming previous findings (Munnamalai et al., 2017). However, at 3 weeks posthatching, ribbon numbers had decreased to the location-specific typical values of control hair cells at all sampled locations. Furthermore, as shown previously, mature hair cells at the basal, high-frequency location harboured larger ribbons than more apically located hair cells. This was true for both normal and Wnt9a-overexposed basilar papillae.