Methods for calibration of fluorescent calcium indicators in skeletal muscle fibers

Calibration of fluorescent calcium indicators.

During the past decades, many different fluorescent indicators have been developed for measuring intracellular ion concentrations. Of particular interest are fluorescent calcium indicators because of the fundamental role of Ca in various cellular processes such as contraction, secretion, and gene activation. For a quantitative understanding of the physiological roles of Ca, fluorescence signals...

Calibration protocols for fluorescent calcium indicators.

Myoplasmic calcium transients in intact frog skeletal muscle fibers monitored with the fluorescent indicator furaptra

Furaptra (Raju, B., E. Murphy, L. A. Levy, R. D. Hall, and R. E. London. 1989. Am. J. Physiol. 256:C540-C548) is a "tri-carboxylate" fluorescent indicator with a chromophore group similar to that of fura-2 (Grynkiewicz, G., M. Poenie, and R. Y. Tsien. 1985. J. Biol. Chem. 260:3440-3450). In vitro calibrations indicate that furaptra reacts with Ca2+ and Mg2+ with 1:1 stoichiometry, with dissocia...

Dimensions of calcium release domains in frog skeletal muscle fibers

The spatiotemporal properties of the Ca release process in skeletal muscle fibers were determined using an improved confocal spot detection system. Muscle fibers were loaded with the low affinity fluorescent Ca indicator OGB-5N and localized action potential-induced fluorescence signals were recorded from consecutive locations separated by 200 nm within a single sarcomere. Three-dimensional rec...

Calcium Sparks in Intact Skeletal Muscle Fibers of the Frog

Calcium sparks were studied in frog intact skeletal muscle fibers using a home-built confocal scanner whose point-spread function was estimated to be approximately 0.21 microm in x and y and approximately 0.51 microm in z. Observations were made at 17-20 degrees C on fibers from Rana pipiens and Rana temporaria. Fibers were studied in two external solutions: normal Ringer's ([K(+)] = 2.5 mM; es...