An Examination of Plasma Membrane Calcium ATPase 2 localization in the Medial Nucleus of the Trapezoid Body Reveals a Cell Size Gradient

Thesis (Ph.D.)--University of Washington, 2014An Examination of Plasma Membrane Calcium ATPase 2 localization in the Medial Nucleus of the Trapezoid Body Reveals a Cell Size Gradient Jessica H. Weatherstone Chair of Supervisory Committee: Professor Bruce L Tempel Otolaryngology-Head and Neck Surgery, Pharmacology The plasma membrane calcium ATPase 2 (PMCA2) pumps calcium from the cytoplasm to the extracellular space. We show PMCA2 is highly expressed in the medial nucleus of the trapezoid body (MNTB). <italic>dfw<super>2J</super></italic> mutant mice have a null mutation in the gene that codes for PMCA2 causing deafness in homozygotes. Because MNTB neurons highly express PMCA2, we postulated that MNTB neurons lacking PMCA2 may have abnormal morphology. We found no significant difference in the gross morphology of MNTBs from<italic> dfw<super>2J</super></italic> mutants, but did find a decrease in cell size between wildtype and <italic>dfw<super>2J</super>/dfw<super>2J</super></italic> mutants. The MNTB exhibits a tonotopic gradient with medial cells responding best to high frequencies and lateral cells responding best to low frequencies. In wildtype, the medial cells were significantly smaller than the lateral cells suggesting the presence of a cell size gradient. This size gradient was absent in <italic>dfw<super>2J</super>/dfw<super>2J</super></italic>. To determine if the absence of the neuronal size gradient was caused by abnormal inputs from the cochlea, we used diphtheria toxin receptor (DTR) mice in which hair cells can be selectively ablated (Tong et al., 2011; Golub et al., 2012). These mice show no cell size gradient after hair cells were ablated suggesting auditory activity is necessary to maintain the gradient. We also show the gradient can return after a period of auditory deprivation suggesting changes in cell size are plastic and reversible. In addition to a cell size gradient, there are gradients of ion channels along the tonotopic axis in the MNTB (von Hehn et al., 2004; Leao et al,. 2006; Gazula et al, 2010). We used ICC to determine if PMCA2 is homogenously expressed throughout the MNTB. While total PMCA2 is greater in lateral cells than medial cells, this is due to the cell size gradient. The density of PMCA2 is homogenous along the tonotopic axis. Additionally, we show auditory activity does not affect the density of PMCA2 in the MNTB. In summary, PMCA2 is highly expressed in the MNTB, there is a cell size gradient in the MNTB that requires auditory activity, and homogenous expression of PMCA2 in the MNTB is not affected by auditory inputs