Tools for flash-photolysis experiments on voltage-clamped muscle fibre segments

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...

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Bibliographic Details
Published in:Pflügers Archiv 2000-01, Vol.439 (3), p.385-393
Main Authors: Schuhmeier, R P, Tewes, S, Szentesi, P, Melzer, W
Format: Article
Language:English
Subjects:
Acetates - pharmacology
Algorithms
Animals
Calcium Channel Blockers - pharmacology
Calcium Channels, L-Type - drug effects
Calcium Channels, L-Type - metabolism
Calcium Signaling - drug effects
Chelating Agents - pharmacology
Ethylenediamines - pharmacology
In Vitro Techniques
Membrane Potentials - physiology
Muscle Fibers, Skeletal - metabolism
Muscle Fibers, Skeletal - physiology
Nifedipine - pharmacology
Patch-Clamp Techniques
Photolysis
Photometry
Rana pipiens
Studies
Ultraviolet Rays
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Summary: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.
ISSN:0031-6768
1432-2013
DOI:10.1007/s004249900102