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Ryanoid modification of the cardiac muscle ryanodine receptor channel results in relocation of the tetraethylammonium binding site.

Tanna B, Welch W, Ruest L, Sutko JL, Williams AJ - J. Gen. Physiol. (2001)

Bottom Line: In all cases, channel open probability increases dramatically and single-channel current amplitude is reduced.It has been proposed that these alterations result from a reorganization of channel structure induced by the binding of the ryanoid.The degree of change of these parameters correlates broadly with the change in conductance of permeant cations induced by the ryanoids, indicating that modification of RyR channel structure by ryanoids is likely to underlie both phenomena.

View Article: PubMed Central - PubMed

Affiliation: Department of Cardiac Medicine, National Heart and Lung Institute, Imperial College of Science, Technology and Medicine, London SW3 6LY, United Kingdom.

ABSTRACT
The interaction of ryanodine and derivatives of ryanodine with the high affinity binding site on the ryanodine receptor (RyR) channel brings about a characteristic modification of channel function. In all cases, channel open probability increases dramatically and single-channel current amplitude is reduced. The amplitude of the ryanoid-modified conductance state is determined by structural features of the ligand. An investigation of ion handling in the ryanodine-modified conductance state has established that reduced conductance results from changes in both the affinity of the channel for permeant ions and the relative permeability of ions within the channel (Lindsay, A.R.G., A. Tinker, and A.J. Williams. 1994. J. Gen. Physiol. 104:425-447). It has been proposed that these alterations result from a reorganization of channel structure induced by the binding of the ryanoid. The experiments reported here provide direct evidence for ryanoid-induced restructuring of RyR. TEA+ is a concentration- and voltage-dependent blocker of RyR in the absence of ryanoids. We have investigated block of K+ current by TEA+ in the unmodified open state and modified conductance states of RyR induced by 21-amino-9alpha-hydroxyryanodine, 21-azido-9alpha-hydroxyryanodine, ryanodol, and 21-p-nitrobenzoylamino-9alpha-hydroxyryanodine. Analysis of the voltage dependence of block indicates that the interaction of ryanoids with RyR leads to an alteration in this parameter with an apparent relocation of the TEA+ blocking site within the voltage drop across the channel and an alteration in the affinity of the channel for the blocker. The degree of change of these parameters correlates broadly with the change in conductance of permeant cations induced by the ryanoids, indicating that modification of RyR channel structure by ryanoids is likely to underlie both phenomena.

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(A) Single-channel current-voltage relationships for unmodified channels in the absence (•; mean ± SEM of 22–51 experiments) and presence (▴; mean ± SEM of 4–25 experiments) of 20 mM TEA+. (B) Single-channel current-voltage relationships for 21-amino-9α-hydroxyryanodine–modified channels in the absence (•; ± SEM of 4–15 experiments) and presence (▴; ± SEM of 4–10 experiments) of 20 mM TEA+. Where no error bars are visible they are contained within the symbol.
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Figure 3: (A) Single-channel current-voltage relationships for unmodified channels in the absence (•; mean ± SEM of 22–51 experiments) and presence (▴; mean ± SEM of 4–25 experiments) of 20 mM TEA+. (B) Single-channel current-voltage relationships for 21-amino-9α-hydroxyryanodine–modified channels in the absence (•; ± SEM of 4–15 experiments) and presence (▴; ± SEM of 4–10 experiments) of 20 mM TEA+. Where no error bars are visible they are contained within the symbol.

Mentions: The right panel of Fig. 2 shows current fluctuations of the same channel after the addition of 20 mM TEA+ to the solutions at both sides of the channel. In the absence of ryanoid, TEA+ is a concentration- and voltage-dependent blocker of K+ conductance in the RyR channel. Dwell times in the TEA+ blocked state are too short to be resolved and, as a consequence, RyR channel block by this cation appears as a time-averaged reduction in single-channel current amplitude (Lindsay et al. 1991; Tinker et al. 1992). Block of this form can be seen in Fig. 2 in the periods of unmodified gating when the ryanoid is not bound to the channel. In agreement with previous observations (Lindsay et al. 1991), the degree of block of this unmodified state of the channel by TEA+ varies as transmembrane potential is changed; block increases as the holding potential is made more positive. Current-voltage relationships for unmodified conductance states of several RyR channels in the absence and presence of 20 mM TEA+ are shown in Fig. 3 A. TEA+ has no effect on K+ translocation at negative holding potentials but blocks K+ current with increasing effectiveness as the holding potential is shifted to more positive values.


Ryanoid modification of the cardiac muscle ryanodine receptor channel results in relocation of the tetraethylammonium binding site.

Tanna B, Welch W, Ruest L, Sutko JL, Williams AJ - J. Gen. Physiol. (2001)

(A) Single-channel current-voltage relationships for unmodified channels in the absence (•; mean ± SEM of 22–51 experiments) and presence (▴; mean ± SEM of 4–25 experiments) of 20 mM TEA+. (B) Single-channel current-voltage relationships for 21-amino-9α-hydroxyryanodine–modified channels in the absence (•; ± SEM of 4–15 experiments) and presence (▴; ± SEM of 4–10 experiments) of 20 mM TEA+. Where no error bars are visible they are contained within the symbol.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2233661&req=5

Figure 3: (A) Single-channel current-voltage relationships for unmodified channels in the absence (•; mean ± SEM of 22–51 experiments) and presence (▴; mean ± SEM of 4–25 experiments) of 20 mM TEA+. (B) Single-channel current-voltage relationships for 21-amino-9α-hydroxyryanodine–modified channels in the absence (•; ± SEM of 4–15 experiments) and presence (▴; ± SEM of 4–10 experiments) of 20 mM TEA+. Where no error bars are visible they are contained within the symbol.
Mentions: The right panel of Fig. 2 shows current fluctuations of the same channel after the addition of 20 mM TEA+ to the solutions at both sides of the channel. In the absence of ryanoid, TEA+ is a concentration- and voltage-dependent blocker of K+ conductance in the RyR channel. Dwell times in the TEA+ blocked state are too short to be resolved and, as a consequence, RyR channel block by this cation appears as a time-averaged reduction in single-channel current amplitude (Lindsay et al. 1991; Tinker et al. 1992). Block of this form can be seen in Fig. 2 in the periods of unmodified gating when the ryanoid is not bound to the channel. In agreement with previous observations (Lindsay et al. 1991), the degree of block of this unmodified state of the channel by TEA+ varies as transmembrane potential is changed; block increases as the holding potential is made more positive. Current-voltage relationships for unmodified conductance states of several RyR channels in the absence and presence of 20 mM TEA+ are shown in Fig. 3 A. TEA+ has no effect on K+ translocation at negative holding potentials but blocks K+ current with increasing effectiveness as the holding potential is shifted to more positive values.

Bottom Line: In all cases, channel open probability increases dramatically and single-channel current amplitude is reduced.It has been proposed that these alterations result from a reorganization of channel structure induced by the binding of the ryanoid.The degree of change of these parameters correlates broadly with the change in conductance of permeant cations induced by the ryanoids, indicating that modification of RyR channel structure by ryanoids is likely to underlie both phenomena.

View Article: PubMed Central - PubMed

Affiliation: Department of Cardiac Medicine, National Heart and Lung Institute, Imperial College of Science, Technology and Medicine, London SW3 6LY, United Kingdom.

ABSTRACT
The interaction of ryanodine and derivatives of ryanodine with the high affinity binding site on the ryanodine receptor (RyR) channel brings about a characteristic modification of channel function. In all cases, channel open probability increases dramatically and single-channel current amplitude is reduced. The amplitude of the ryanoid-modified conductance state is determined by structural features of the ligand. An investigation of ion handling in the ryanodine-modified conductance state has established that reduced conductance results from changes in both the affinity of the channel for permeant ions and the relative permeability of ions within the channel (Lindsay, A.R.G., A. Tinker, and A.J. Williams. 1994. J. Gen. Physiol. 104:425-447). It has been proposed that these alterations result from a reorganization of channel structure induced by the binding of the ryanoid. The experiments reported here provide direct evidence for ryanoid-induced restructuring of RyR. TEA+ is a concentration- and voltage-dependent blocker of RyR in the absence of ryanoids. We have investigated block of K+ current by TEA+ in the unmodified open state and modified conductance states of RyR induced by 21-amino-9alpha-hydroxyryanodine, 21-azido-9alpha-hydroxyryanodine, ryanodol, and 21-p-nitrobenzoylamino-9alpha-hydroxyryanodine. Analysis of the voltage dependence of block indicates that the interaction of ryanoids with RyR leads to an alteration in this parameter with an apparent relocation of the TEA+ blocking site within the voltage drop across the channel and an alteration in the affinity of the channel for the blocker. The degree of change of these parameters correlates broadly with the change in conductance of permeant cations induced by the ryanoids, indicating that modification of RyR channel structure by ryanoids is likely to underlie both phenomena.

Show MeSH
Related in: MedlinePlus