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Simultaneous recording of calcium transients in skeletal muscle using high- and low-affinity calcium indicators
- Source :
- Biophysical Journal. 53:971-988
- Publication Year :
- 1988
- Publisher :
- Elsevier BV, 1988.
-
Abstract
- To monitor cytosolic [Ca2+] over a wide range of concentrations in functioning skeletal muscle cells, we have used simultaneously the rapid but relatively low affinity calcium indicator antipyrylazo III (AP III) and the slower but higher affinity indicator fura-2 in single frog twitch fibers cut at both ends and voltage clamped with a double vaseline gap system. When both dyes were added to the end pool solution the cytosolic fura-2 concentration reached a steady level equal to the end pool concentration within approximately 2.5 h, a time when the AP III concentration was still increasing. For depolarizing pulses of increasing amplitude, the fura-2 fluorescence signal approached saturation when the simultaneously recorded AP III absorbance change was far from saturation. Comparison of simultaneously recorded fura-2 and AP III signals indicated that the mean values of the on and off rate constants for calcium binding to fura-2 in 18 muscle fibers were 1.49 x 10(8) M-1 s-1 and 11.9 s-1, respectively (mean KD = 89 nM), if all AP III in the fiber is assumed to behave as in calibrating solution and to be in instantaneous equilibrium with [Ca2+]. [Ca2+] transients calculated from the fura-2 signals using these rate constants were consistent with the [Ca2+] transients calculated from the AP III signals. Resting [Ca2+] or small changes in [Ca2+] which could not be reliably monitored with AP III could be monitored with fura-2 with little or no interference from changes in [Mg2+] or from intrinsic signals. The fura-2 signal was also less sensitive to movement artifacts than the AP III signal. After a [Ca2+] transient the fura-2 signal demonstrated a relatively small elevation of [Ca2+] that was maintained for many seconds.
- Subjects :
- Ranidae
Fura-2
Biophysics
Analytical chemistry
chemistry.chemical_element
Calcium
Signal
Absorbance
03 medical and health sciences
chemistry.chemical_compound
Cytosol
0302 clinical medicine
Naphthalenesulfonates
medicine
Animals
Benzofurans
Fluorescent Dyes
030304 developmental biology
0303 health sciences
Chemistry
Muscles
Skeletal muscle
Depolarization
Fluorescence
Spectrometry, Fluorescence
medicine.anatomical_structure
Saturation (chemistry)
030217 neurology & neurosurgery
Research Article
Subjects
Details
- ISSN :
- 00063495
- Volume :
- 53
- Database :
- OpenAIRE
- Journal :
- Biophysical Journal
- Accession number :
- edsair.doi.dedup.....3d05e9648e946c9e8e9846d0b57516b7