![]() The authors of these studies reported that to get an increase in channel open probability, several polyanion molecules needed to bind to the RyR. For example, highly negatively charged polyanions such as pentosan polysulfate, polyvinyl sulfate, and heparin increased the activity of RyRs, whereas they decrease the activity of IP 3 receptors ( Bezprozvanny et al., 1993). RyRs are large tetrameric proteins that show sequence similarity with inositol 1,4,5-trisphosphate (IP 3)-gated calcium channels of the endoplasmic/sarcoplasmic reticulum, but they are distinct in their biophysical and pharmacological properties ( Smith et al., 1986 Ehrlich and Watras, 1988 Supattapone et al., 1988 Mignery et al., 1989 Palade et al., 1989, Ehrlich et al., 1994). They mediate the release of calcium ions from the endoplasmic/sarcoplasmic reticulum into the cytosol and thereby convert a number of extracellular stimuli into intracellular calcium signals. ![]() ![]() Ryanodine receptors (calcium-induced calcium release channels RyR) play a crucial role in most cell types, including muscle cells, neurons, and epithelial cells. The results suggest that protamine acts on the ryanodine receptor in a similar but opposite manner from heparin and that protamine can be used as a potent, reversible inhibitor of ryanodine receptor activity. Microinjection of protamine into differentiated C 2C 12 mouse muscle cells prevented caffeine-induced intracellular calcium release. The block of channel activity by protamine could be reversed either by removal by perfusion with buffer or by the addition of heparin to the cytoplasmic side of the channel. The addition of protamine bound to agarose beads did not change channel activity, implying that the mechanism of action involves an interaction with the ryanodine receptor rather than changes in the bulk calcium concentration of the medium. Recovery of channel activity was not observed while protamine was present. Optimally and suboptimally calcium-activated calcium release channels were inactivated by the application of protamine to the cytoplasmic side of the channel. Single-channel activity was measured after incorporating channels into planar lipid bilayers. Channel activity of the calcium release channel from skeletal muscle, ryanodine receptor type 1, was measured in the presence and absence of protamine sulfate on the cytoplasmic side of the channel.
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