Analysis of connexin subunits required for the survival of vestibular hair cells

Mutations in connexin (Cx) genes are responsible for a large proportion of human inherited prelingual deafness cases. The most commonly found human Cx mutations are either Cx26 or Cx30 deletions. Histological observations made in the organ of Corti of homozygous Cx26 and Cx30 gene knockout mice show that cochlear hair cells degenerate after the onset of hearing. However, it is unclear whether vestibular hair cells undergo similar degeneration in connexin knockout mice. To address this question, we first examined expression patterns of Cx26 and Cx30 in the saccule, utricle, and ampulla by immunolabeling. In wild-type mice, Cx26 and Cx30 immunoreactivity was found extensively in vestibular supporting cells and connective tissue cells, and the two Cxs were co-localized in most gap junction (GJ) plaques. Targeted deletion of the Cx30 gene, which caused little change in Cx26 expression pattern, resulted in a significant and age-related loss of vestibular hair cells only in the saccule. dUTP nick end labeling (TUNEL) staining also revealed on-going apoptosis specifically in saccular hair cells of Cx30^-/- mice. These results indicated that hair cell survival in the utricle and ampulae does not require Cx30. Importantly, over-expressing the Cx26 gene from a modified bacterial artificial chromosome in the Cx30^-/- background rescued the saccular hair cells. These results suggest that it is the reduction in the total amount of GJs rather than the specific loss of Cx30 that underlies saccular hair cell death in Cx30^-/- mice. Hybrid GJs co-assembled from Cx26 and Cx30 were not essential for the survival of saccular hair cells.