Schuhmeier R P, Tewes S, Szentesi P, Melzer W
Department of Applied Physiology, University of Ulm, Germany.
Pflugers Arch. 2000 Jan;439(3):385-93. doi: 10.1007/s004249900102.
An experimental set-up is described that allows the combination of rapid transmembrane voltage changes and photometric calcium recording with the fast photochemical turnover of substances applied externally or intracellularly to cut skeletal muscle fibres. It consists of a double-vaseline-gap system, designed for use with a xenon-flash-lamp device and a dual-wavelength microscope photometer. The pools of the vaseline gap chamber that contain the solutions surrounding the cut ends and the voltage-clamped segment of the muscle fibre are closed and have volumes of 20-50 microl. Thin tubes allow rapid solution change or continuous perfusion in the chamber compartments. Accessory tools were constructed to simplify focussing and measuring the flash-light intensity. A pilot light delivered from a red laser diode is used as a guide beam to target the ultraviolet (UV) flash to the preparation. The light distribution in the focal region and the relative changes in flash intensity with increasing numbers of flashes were quantified with an instrument that integrates the photo-current of a UV-sensitive silicon diode. The function of the set-up was demonstrated by measuring the efficiency of Ca2+ release from DM-nitrophen in quartz capillaries using the Ca(2+)-sensitive dye antipyrylazo III and by recording the flash-induced recovery of L-type calcium currents in muscle fibres blocked by the light-sensitive dihydropyridine drug nifedipine.
本文描述了一种实验装置，该装置能够将快速跨膜电压变化、光度钙记录与快速光化学周转相结合，可将物质从外部或细胞内施加到离体骨骼肌纤维上。它由一个双凡士林间隙系统组成，设计用于与氙闪光灯装置和双波长显微镜光度计配合使用。凡士林间隙腔室的池包含围绕肌肉纤维切断端和电压钳制段的溶液，这些池是封闭的，体积为20 - 50微升。细管允许在腔室隔室中快速更换溶液或持续灌注。构建了辅助工具以简化聚焦和测量闪光强度。从红色激光二极管发出的引导光用作引导光束，将紫外（UV）闪光对准标本。使用集成了紫外敏感硅二极管光电流的仪器，对焦点区域的光分布以及随着闪光次数增加闪光强度的相对变化进行了量化。通过使用钙敏感染料安替比拉宗III测量石英毛细管中DM - 硝基苯酚的Ca2+释放效率，以及记录被光敏二氢吡啶药物硝苯地平阻断的肌肉纤维中闪光诱导的L型钙电流恢复，证明了该装置的功能。
