Yamoah E N, Lumpkin E A, Dumont R A, Smith P J, Hudspeth A J, Gillespie P G
Department of Physiology, The Johns Hopkins University, Baltimore, Maryland 21205, USA.
J Neurosci. 1998 Jan 15;18(2):610-24. doi: 10.1523/JNEUROSCI.18-02-00610.1998.
Mechanically sensitive hair cells of the auditory and vestibular systems use Ca2+ to control adaptation of mechanical transduction, to effect frequency tuning, to trigger neurotransmitter release, and to mediate efferent synaptic signaling. To determine the role that pumps play in regulation of Ca2+ in the hair bundle, the organelle responsible for mechanoelectrical transduction, we localized and quantified the plasma membrane Ca2+-ATPase (PMCA) of the bundle. We found that each hair bundle contains approximately 10(6) PMCA molecules or approximately 2000 per square micrometer of bundle membrane and that PMCA is the principal calmodulin binding protein of the bundle. Consistent with biochemical estimates of PMCA density, we measured with extracellular Ca2+-selective electrodes a substantial Ca2+ efflux from bundles. The number of bundle Ca2+ pumps and magnitude of resting Ca2+ efflux suggested that PMCA should generate a substantial membrane current as bundles expel Ca2+. Measurement of whole-cell currents revealed a transduction-dependent outward current that was consistent with the activity of PMCA. Finally, dialysis of hair cells with PMCA inhibitors led to a large increase in the concentration of Ca2+ in bundles, which suggests that PMCA plays a major role in regulating bundle Ca2+ concentration. Our data further indicate that PMCA could elevate the extracellular Ca2+ concentration close to hair bundles above the low level found in bulk endolymph.
听觉和前庭系统中对机械刺激敏感的毛细胞利用Ca2+来控制机械转导的适应性、实现频率调谐、触发神经递质释放以及介导传出突触信号传递。为了确定泵在毛束（负责机械电转导的细胞器）中Ca2+调节过程中所起的作用，我们对毛束中的质膜Ca2+ -ATP酶（PMCA）进行了定位和定量分析。我们发现，每个毛束含有约10^6个PMCA分子，即每平方微米束膜约含2000个，并且PMCA是毛束中主要的钙调蛋白结合蛋白。与PMCA密度的生化估计结果一致，我们使用细胞外Ca2+选择性电极测量到大量Ca2+从毛束流出。毛束Ca2+泵的数量和静息Ca2+流出量表明，随着毛束排出Ca2+，PMCA应该会产生大量的膜电流。全细胞电流测量显示出一种与转导相关的外向电流，这与PMCA的活性一致。最后，用PMCA抑制剂对毛细胞进行透析导致毛束中Ca2+浓度大幅增加，这表明PMCA在调节毛束Ca2+浓度方面起主要作用。我们的数据进一步表明，PMCA可以将靠近毛束的细胞外Ca2+浓度提高到高于内淋巴总体中发现的低水平。
