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Unitary Ca2+ current through mammalian cardiac and amphibian skeletal muscle ryanodine receptor Channels under near-physiological ionic conditions.

Kettlun C, González A, Ríos E, Fill M - J. Gen. Physiol. (2003)

Bottom Line: Currents measured with 20 mM lumenal Ca2+ as exclusive charge carrier were 4.00 and 4.07 pA, respectively, and not significantly different.Two amplitudes, differing by approximately 35%, were found in amphibian channel studies, probably corresponding to alpha and beta RyR isoforms.Further, symmetric high concentrations of Mg2+ substantially reduced the current carried by 10 mM Ca2+ (approximately 40% at 10 mM Mg2+), suggesting that high Mg2+ may make sparks smaller by both inhibiting RyR gating and reducing unitary current.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, Loyola University Chicago, 2160 S. First Ave., Maywood, IL 60153, USA.

ABSTRACT
Ryanodine receptor (RyR) channels from mammalian cardiac and amphibian skeletal muscle were incorporated into planar lipid bilayers. Unitary Ca2+ currents in the SR lumen-to-cytosol direction were recorded at 0 mV in the presence of caffeine (to minimize gating fluctuations). Currents measured with 20 mM lumenal Ca2+ as exclusive charge carrier were 4.00 and 4.07 pA, respectively, and not significantly different. Currents recorded at 1-30 mM lumenal Ca2+ concentrations were attenuated by physiological [K+] (150 mM) and [Mg2+] (1 mM), in the same proportion (approximately 55%) in mammalian and amphibian channels. Two amplitudes, differing by approximately 35%, were found in amphibian channel studies, probably corresponding to alpha and beta RyR isoforms. In physiological [Mg2+], [K+], and lumenal [Ca2+] (1 mM), the Ca2+ current was just less than 0.5 pA. Comparison of this value with the Ca2+ flux underlying Ca2+ sparks suggests that sparks in mammalian cardiac and amphibian skeletal muscles are generated by opening of multiple RyR channels. Further, symmetric high concentrations of Mg2+ substantially reduced the current carried by 10 mM Ca2+ (approximately 40% at 10 mM Mg2+), suggesting that high Mg2+ may make sparks smaller by both inhibiting RyR gating and reducing unitary current.

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Influence of elevated Mg2+ on Ca2+ current of frog RyR channel. Experiments at 10 mM [Ca2+]L. (A) Current in the presence of symmetric 0, 5 or 10 mM Mg2+ (top to bottom). (B) Marked segments of records in A on an expanded time scale. (C) Squares, average currents (± SEM; n = 3–5) in different symmetrical Mg2+ concentrations. Line, fit with Eq. 1 to predictions of the 4-barrier model. Filled circle, measured current at 1 mM [Mg2+] and 150 mM KCl. Open circle, corresponding prediction of the 4-barrier model.
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fig8: Influence of elevated Mg2+ on Ca2+ current of frog RyR channel. Experiments at 10 mM [Ca2+]L. (A) Current in the presence of symmetric 0, 5 or 10 mM Mg2+ (top to bottom). (B) Marked segments of records in A on an expanded time scale. (C) Squares, average currents (± SEM; n = 3–5) in different symmetrical Mg2+ concentrations. Line, fit with Eq. 1 to predictions of the 4-barrier model. Filled circle, measured current at 1 mM [Mg2+] and 150 mM KCl. Open circle, corresponding prediction of the 4-barrier model.

Mentions: Recent studies have demonstrated that Ca2+ sparks in frog muscle fibers can still be recorded at high Mg2+ concentrations (as high as 7.5 mM), but with diminished amplitude and spatial width (González et al., 2000). Thus, it was of interest to evaluate unitary Ca2+ current at such high Mg2+ concentrations. Three sample channel records of a single amphibian RyR channel are shown in Fig. 8 A. These records were collected in the presence of 0, 5 (top), or 10 mM Mg2+ (added symmetrically). The solutions included 10 mM [Ca2+]L and the usual 10 mM caffeine. An obstacle for the accurate measurement of current attenuation was the clear decrease in Po (expected and observed) at elevated Mg2+ concentrations (even in the presence of caffeine). In Fig. 8 A, highly compressed records are shown to illustrate the decrease in Po. In spite of this complication, a few sufficiently long openings were recorded and adequate measurements of current amplitude were made at high Mg2+ levels.


Unitary Ca2+ current through mammalian cardiac and amphibian skeletal muscle ryanodine receptor Channels under near-physiological ionic conditions.

Kettlun C, González A, Ríos E, Fill M - J. Gen. Physiol. (2003)

Influence of elevated Mg2+ on Ca2+ current of frog RyR channel. Experiments at 10 mM [Ca2+]L. (A) Current in the presence of symmetric 0, 5 or 10 mM Mg2+ (top to bottom). (B) Marked segments of records in A on an expanded time scale. (C) Squares, average currents (± SEM; n = 3–5) in different symmetrical Mg2+ concentrations. Line, fit with Eq. 1 to predictions of the 4-barrier model. Filled circle, measured current at 1 mM [Mg2+] and 150 mM KCl. Open circle, corresponding prediction of the 4-barrier model.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2233776&req=5

fig8: Influence of elevated Mg2+ on Ca2+ current of frog RyR channel. Experiments at 10 mM [Ca2+]L. (A) Current in the presence of symmetric 0, 5 or 10 mM Mg2+ (top to bottom). (B) Marked segments of records in A on an expanded time scale. (C) Squares, average currents (± SEM; n = 3–5) in different symmetrical Mg2+ concentrations. Line, fit with Eq. 1 to predictions of the 4-barrier model. Filled circle, measured current at 1 mM [Mg2+] and 150 mM KCl. Open circle, corresponding prediction of the 4-barrier model.
Mentions: Recent studies have demonstrated that Ca2+ sparks in frog muscle fibers can still be recorded at high Mg2+ concentrations (as high as 7.5 mM), but with diminished amplitude and spatial width (González et al., 2000). Thus, it was of interest to evaluate unitary Ca2+ current at such high Mg2+ concentrations. Three sample channel records of a single amphibian RyR channel are shown in Fig. 8 A. These records were collected in the presence of 0, 5 (top), or 10 mM Mg2+ (added symmetrically). The solutions included 10 mM [Ca2+]L and the usual 10 mM caffeine. An obstacle for the accurate measurement of current attenuation was the clear decrease in Po (expected and observed) at elevated Mg2+ concentrations (even in the presence of caffeine). In Fig. 8 A, highly compressed records are shown to illustrate the decrease in Po. In spite of this complication, a few sufficiently long openings were recorded and adequate measurements of current amplitude were made at high Mg2+ levels.

Bottom Line: Currents measured with 20 mM lumenal Ca2+ as exclusive charge carrier were 4.00 and 4.07 pA, respectively, and not significantly different.Two amplitudes, differing by approximately 35%, were found in amphibian channel studies, probably corresponding to alpha and beta RyR isoforms.Further, symmetric high concentrations of Mg2+ substantially reduced the current carried by 10 mM Ca2+ (approximately 40% at 10 mM Mg2+), suggesting that high Mg2+ may make sparks smaller by both inhibiting RyR gating and reducing unitary current.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, Loyola University Chicago, 2160 S. First Ave., Maywood, IL 60153, USA.

ABSTRACT
Ryanodine receptor (RyR) channels from mammalian cardiac and amphibian skeletal muscle were incorporated into planar lipid bilayers. Unitary Ca2+ currents in the SR lumen-to-cytosol direction were recorded at 0 mV in the presence of caffeine (to minimize gating fluctuations). Currents measured with 20 mM lumenal Ca2+ as exclusive charge carrier were 4.00 and 4.07 pA, respectively, and not significantly different. Currents recorded at 1-30 mM lumenal Ca2+ concentrations were attenuated by physiological [K+] (150 mM) and [Mg2+] (1 mM), in the same proportion (approximately 55%) in mammalian and amphibian channels. Two amplitudes, differing by approximately 35%, were found in amphibian channel studies, probably corresponding to alpha and beta RyR isoforms. In physiological [Mg2+], [K+], and lumenal [Ca2+] (1 mM), the Ca2+ current was just less than 0.5 pA. Comparison of this value with the Ca2+ flux underlying Ca2+ sparks suggests that sparks in mammalian cardiac and amphibian skeletal muscles are generated by opening of multiple RyR channels. Further, symmetric high concentrations of Mg2+ substantially reduced the current carried by 10 mM Ca2+ (approximately 40% at 10 mM Mg2+), suggesting that high Mg2+ may make sparks smaller by both inhibiting RyR gating and reducing unitary current.

Show MeSH
Related in: MedlinePlus