Article date: January 1995
By: Riccardo Zucchi, Simonetta Ronca‐Testoni, Gongyuan Yu, Paola Galbani, Giovanni Ronca, Mario Mariani, in Volume 114, Issue 1, pages 85-92
In a sarcoplasmic reticulum fraction obtained from rat hearts, the analysis of equilibrium [3H]‐ryanodine binding showed high and low affinity sites (KD = 1.3 nm and 2.8 μm, Bmax = 2.2 pmol mg−1 and 27.8 pmol mg−1). The dissociation rate constant increased at 1 μm VS 4 nm [3H]‐ryanodine concentration, and micromolar ryanodine slowed the dissociation of nanomolar ryanodine.
The binding of 4 nm [3H]‐ryanodine was not affected by gallopamil, while the binding of 100 nm to 18 μm [3H]‐ryanodine was partly displaced. Data analysis suggested that gallopamil inhibited low affinity [3H]‐ryanodine binding, with IC50 in the micromolar range.
Gallopamil decreased the dissociation rate constant of 1 μm [3H]‐ryanodine. While gallopamil alone did not affect the dissociation of 4 nm [3H]‐ryanodine, gallopamil and micromolar ryanodine slowed it to a greater extent than micromolar ryanodine alone.
Our results are consistent with the hypothesis that the ryanodine receptor is a negatively cooperative oligomer, which undergoes a sequential alteration after ryanodine binding. Gallopamil has complex actions: it inhibits ryanodine binding to its low affinity site(s), and probably modulates the cooperativity of ryanodine binding and/or the transition to a receptor state characterized by slow ryanodine dissociation. These molecular actions could account for the previously reported effect of gallopamil on the sarcoplasmic reticulum calcium release channel.
In a sarcoplasmic reticulum fraction obtained from rat hearts, the analysis of equilibrium [3H]‐ryanodine binding showed high and low affinity sites (KD = 1.3 nm and 2.8 μm, Bmax = 2.2 pmol mg−1 and 27.8 pmol mg−1). The dissociation rate constant increased at 1 μm VS 4 nm [3H]‐ryanodine concentration, and micromolar ryanodine slowed the dissociation of nanomolar ryanodine.
The binding of 4 nm [3H]‐ryanodine was not affected by gallopamil, while the binding of 100 nm to 18 μm [3H]‐ryanodine was partly displaced. Data analysis suggested that gallopamil inhibited low affinity [3H]‐ryanodine binding, with IC50 in the micromolar range.
Gallopamil decreased the dissociation rate constant of 1 μm [3H]‐ryanodine. While gallopamil alone did not affect the dissociation of 4 nm [3H]‐ryanodine, gallopamil and micromolar ryanodine slowed it to a greater extent than micromolar ryanodine alone.
Our results are consistent with the hypothesis that the ryanodine receptor is a negatively cooperative oligomer, which undergoes a sequential alteration after ryanodine binding. Gallopamil has complex actions: it inhibits ryanodine binding to its low affinity site(s), and probably modulates the cooperativity of ryanodine binding and/or the transition to a receptor state characterized by slow ryanodine dissociation. These molecular actions could account for the previously reported effect of gallopamil on the sarcoplasmic reticulum calcium release channel.
DOI: 10.1111/j.1476-5381.1995.tb14909.x
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