Balance and Hearing Deficits in Mice with a Null Mutation in the Gene Encoding Plasma Membrane Ca2+-ATPase Isoform 2

Abstract

Plasma membrane Ca 2؉-ATPase isoform 2 (PMCA2) exhibits a highly restricted tissue distribution, suggesting that it serves more specialized physiological functions than some of the other isoforms. A unique role in hearing is indicated by the high levels of PMCA2 expression in cochlear outer hair cells and spiral ganglion cells. To analyze the physiological role of PMCA2 we used gene targeting to produce PMCA2-deficient mice. Breeding of heterozygous mice yielded live homozygous mutant offspring. PMCA2-null mice grow more slowly than heterozygous and wild-type mice and exhibit an unsteady gait and difficulties in maintaining balance. Histological analysis of the cerebellum and inner ear of mutant and wild-type mice revealed that null mutants had slightly increased numbers of Purkinje neurons (in which PMCA2 is highly expressed), a decreased thickness of the molecular layer, an absence of otoconia in the vestibular system, and a range of abnormalities of the organ of Corti. Analysis of auditory evoked brainstem responses revealed that homozygous mutants were deaf and that heterozygous mice had a significant hearing loss. These data demonstrate that PMCA2 is required for both balance and hearing and suggest that it may be a major source of the calcium used in the formation and maintenance of otoconia.Calmodulin-dependent plasma membrane Ca 2ϩ -ATPases (PMCAs) 1 are highly regulated enzymes that maintain the appropriate concentrations of intracellular free Ca 2ϩ by extruding Ca 2ϩ from the cell (1, 2). There are four mammalian PMCA isoforms (PMCA1-4), each encoded by a distinct gene (3-8), and additional diversity is generated by alternative splicing of exons encoding the regulatory domains (7, 9 -11). Variants of PMCA1 and PMCA4 are expressed in many different tissues and cell types, whereas variants of PMCA2 and PMCA3 exhibit a highly restricted distribution (5,8,12). This suggests that specific isoforms and splice variants serve different physiological functions. However, despite extensive information about PMCA structural diversity, expression patterns, and biochemical and regulatory characteristics, little is known about the functions of individual isoforms in vivo.Its unique biochemical characteristics (13, 14) and tissue specificity (5, 10 -12) suggest that PMCA2 might serve specialized physiological functions. In situ hybridization studies revealed that expression of PMCA2 is particularly high in Purkinje neurons of the cerebellum (15) and in the spiral ganglion nerves of the inner ear and outer hair cells of the cochlea (16). The observation that PMCA2 is the predominant isoform in outer hair cells (16) suggests that it might be the isoform that is expressed at high levels in stereocilia (17), which comprise hair bundles, the sensory organelles that mediate mechanoelectrical transduction by hair cells of both the vestibular and auditory systems (18,19). A recent study demonstrated that PMCA activity in the stereocilia of vestibular hair cells regulates hair bundle Ca 2ϩ concentrations and indicated that it ...

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