Marcus D C, Shipley A M
Biophysics Laboratory, Boys Town National Research Hospital, Omaha, NE 68131.
Biophys J. 1994 Jun;66(6):1939-42. doi: 10.1016/S0006-3495(94)80987-7.
Detection of motion and position by the vestibular labyrinth depends on the accumulation of potassium within a central compartment of the inner ear as a source of energy to drive the transduction process. Much circumstantial evidence points to the vestibular dark cell (VDC) epithelium as being responsible for concentrating K+ within the lumen. We have used the vibrating probe technique to directly observe voltage and ion gradients produced by this tissue to put this assumption on a solid experimental footing. Relative current density (Isc,probe) over the apical membrane of VDC epithelium was measured with the vibrating voltage-sensitive probe, and this technique was validated by performing maneuvers known to either stimulate or inhibit the transepithelial equivalent short circuit current. Basolateral bumetanide (5 x 10(-5) M) and ouabain (1 x 10(-3) M) caused a decrease in Isc,probe by 55 +/- 6% and 39 +/- 3%, respectively while raising the basolateral K+ concentration from 4 to 25 mM caused an increase by 35 +/- 8%. A K+ gradient directed toward the apical membrane was detected with the vibrating K(+)-selective electrode, demonstrating that, indeed, the VDC epithelium secretes K+ under control conditions. This secretion was inhibited by bumetanide (by 94 +/- 7%) and ouabain (by 52 +/- 8%). The results substantiate the supposition that dark cells produce a K+ flux and qualitatively support the correlation between this flux and the transepithelial current.
前庭迷路对运动和位置的检测依赖于内耳中央腔室内钾的积累,以此作为驱动转导过程的能量来源。大量间接证据表明,前庭暗细胞(VDC)上皮负责将K⁺浓缩在管腔内。我们使用振动探针技术直接观察该组织产生的电压和离子梯度,以便将这一假设建立在坚实的实验基础上。用振动电压敏感探针测量VDC上皮顶端膜上的相对电流密度(Isc,probe),并通过进行已知能刺激或抑制跨上皮等效短路电流的操作来验证该技术。基底外侧布美他尼(5×10⁻⁵ M)和哇巴因(1×10⁻³ M)分别使Isc,probe降低55±6%和39±3%,而将基底外侧K⁺浓度从4 mM提高到25 mM则使Isc,probe增加35±8%。用振动K⁺选择性电极检测到朝向顶端膜的K⁺梯度,表明在对照条件下,VDC上皮确实分泌K⁺。布美他尼(抑制94±7%)和哇巴因(抑制52±8%)可抑制这种分泌。结果证实了暗细胞产生K⁺通量的假设,并定性地支持了这种通量与跨上皮电流之间的相关性。