Quantifying calretinin-labeled afferent terminals in vestibular sensory epithelia of young and older gerbils

Vestibular dysfunction becomes increasingly prevalent with aging and it is estimated that more than 80 % of people over 80 years old experience balance problems linked to vestibular dysfunction. Within the peripheral vestibular organs, sensory hair cells transform hair bundle motion into receptor potentials and information is next relayed to the brain by electrical activity in vestibular afferent nerves. There are three types of vestibular afferents: calyx-only afferents innervate one or more type I hair cells; bouton dendrites innervate type II hair cells and dimorphic afferents contact both hair cell types. Calyx-only afferents are found solely in central areas of vestibular neuroepithelia and have distinct physiological characteristics. Previous studies have shown changes in vestibular-mediated responses with aging in addition to age-related degeneration of afferent synapses in rodent inner ear epithelia. However, whether calyx-only afferent synapses are lost with aging in vestibular epithelia remains unresolved. Here we used an antibody to the Ca^2 + -binding protein calretinin as a marker of calyx-only afferent terminals in gerbil vestibular epithelia at different ages. We used fluorescent immunohistochemistry and confocal imaging to identify afferent neurons in the utricle and cristae of young (1-2 months), adult (1-2 years), and older adult (≥3 years) gerbils. Counts were made of single, double and triple calretinin-positive calyx terminals in central regions of vestibular epithelia. Overall, a mild decrease in numbers occurred with aging between adult and older adult animals suggesting that aging-related decline in vestibular function can be linked to partial loss of calretinin-positive calyx-only afferent terminals.