R. Mejia-alvarez et al., Unitary Ca2+ current through cardiac ryanodine receptor channels under quasi-physiological ionic conditions, J GEN PHYSL, 113(2), 1999, pp. 177-186
Single canine cardiac ryanodine receptor channels were incorporated into pl
anar lipid bilayers. Single-channel currents were sampled at 1-5 kHz and fi
ltered at 0.2-1.0 kHz. Channel incorporations were obtained in symmetrical
solutions (20 mM HEPES-Tris, pH 7.4, and pCa 5). Unitary Ca2+ currents were
monitored when 2-30 mM Ca2+ was added to the lumenal side of the channel.
The relationship between the amplitude of unitary Ca2+ current (at 0 mV hol
ding potential) and lumenal [Ca2+] was hyperbolic and saturated at similar
to 4 pA. This relationship was then defined in the presence of different sy
mmetrical CsCH3SO3 concentrations (5, 50, and 150 mM). Under these conditio
ns, unitary current amplitude was 1.2 +/- 0.1, 0.65 +/- 0.1, and 0.35 +/- 0
.1 pA in 2 mM lumenal Ca2+; and 3.3 +/- 0.4, 2.4 +/- 0.2, and 1.63 +/- 0.2
pA in 10 mM lumenal Ca2+ (n > 6). Unitary Ca2+ current was also defined in
the presence of symmetrical [Mg2+] (1 mM) and low [Cs+] (5 mM). Under these
conditions, unitary Ca2+ current in 2 and 10 mM lumenal Ca2+ was 0.66 +/-
0.1 and 1.52 +/- 0.06 pA, respectively. In the presence of higher symmetric
al [Cs+] (50 mM), Mg2+ (1 mM), and lumenal [Ca2+] (10 mM), unitary Ca2+ cur
rent exhibited an amplitude of 0.9 +/- 0.2 pA (n = 3). This result indicate
s that the actions of Cs+ and Mg2+ on unitary Ca2+ current were additive. T
hese data demonstrate that physiological levels of monovalent cation and Mg
2+ effectively compete with Ca2+ as charge carrier in cardiac ryanodine rec
eptor channels. If lumenal free Ca2+ is 2 mM, then our results indicate tha
t unitary Ca2+ current under physiological conditions should be <0.6 pA.