Voltage-Dependent Modulation of Cardiac Ryanodine Receptors (RyR2) by Protamine

Abstract

It has been reported that protamine (>10 µg/ml) blocks single skeletal RyR1 channels and inhibits RyR1-mediated Ca2+ release from sarcoplasmic reticulum microsomes. We extended these studies to cardiac RyR2 reconstituted into planar lipid bilayers. We found that protamine (0.02–20 µg/ml) added to the cytosolic surface of fully activated RyR2 affected channel activity in a voltage-dependent manner. At membrane voltage (Vm; SR lumen - cytosol) = 0 mV, protamine induced conductance transitions to several intermediate states (substates) as well as full block of RyR2. At Vm>10 mV, the substate with the highest level of conductance was predominant. Increasing Vm from 0 to +80 mV, decreased the number of transitions and residence of the channel in this substate. The drop in current amplitude (full opening to substate) had the same magnitude at 0 and +80 mV despite the ∼3-fold increase in amplitude of the full opening. This is more similar to rectification of channel conductance induced by other polycations than to the action of selective conductance modifiers (ryanoids, imperatoxin). A distinctive effect of protamine (which might be shared with polylysines and histones but not with non-peptidic polycations) is the activation of RyR2 in the presence of nanomolar cytosolic Ca2+ and millimolar Mg2+ levels. Our results suggest that RyRs would be subject to dual modulation (activation and block) by polycationic domains of neighboring proteins via electrostatic interactions. Understanding these interactions could be important as such anomalies may be associated with the increased RyR2-mediated Ca2+ leak observed in cardiac diseases.

Ethanol Extract ofLycopus lucidusElicits Positive Inotropic Effect Via Activation of Ca2+Entry and Ca2+Release in Beating Rabbit Atria

“…19 RyR, as a SR Ca 2 + release channel, is an essential component of ECC in cardiac contractility and is modulated by several factors such as Ca 2 + , Mg 2 + , and protein kinase A (PKA). 20,21 Further, phospholamban is a phosphoprotein modulating Ca 2 + -ATPase 2a through which ELT (0.3 mg/mL), n = 6; propranolol (1 lM) + ELT (0.3 mg/mL), n = 4; propranolol control, n = 3. *P < .001 vs. control period; # P < .01 vs. the values before the addition of ELT; ++ P < .001 vs. control period; $ P < .001 vs. control group; & P < .001 vs. control group.…”

Modulation of Cardiac Ryanodine Receptor Channels by Alkaline Earth Cations

“…Moreover, the binding of M 2+ to luminal sites differentially affected RyR2 gating kinetics and voltage-dependence as well as RyR2 sensitivity to cytosolic Ca 2+ and cytosolic caffeine. Some of the results have been presented in a preliminary form [21], [22].…”

Coupled gating of skeletal muscle ryanodine receptors is modulated by Ca2+, Mg2+, and ATP

“…In this regard, we were able to mimic the followers behavior (including a decrease in the current amplitude of single RyRs) by exposing the channels to polycationic peptides, including protamine (12,49). In this regard, we were able to mimic the followers behavior (including a decrease in the current amplitude of single RyRs) by exposing the channels to polycationic peptides, including protamine (12,49).…”