Complex modulation of L-type Ca2+ current inactivation by sorcin in isolated rabbit cardiomyocytes

Modulation of the L-type Ca2+ channel (LTCC) by sorcin was investigated by measuring the L-type Ca2+ current (I (Ca,L)) in isolated rabbit ventricular myocytes using ruptured patch, single electrode voltage clamp in the absence of extracellular Na+. Fifty millimolars EGTA (170 nM Ca2+) in the pipette solution buffered bulk cytoplasmic [Ca2+], but retained rapid Ca2+-dependant inactivation of I (Ca,L,). Recombinant sorcin (3 mu M) in the pipette significantly slowed time-dependant inactivation (tau (fast): 8.8 +/- 0.9 vs. 15.1 +/- 1.7 ms). Sorcin had no significant effect on I (Ca,L,) after inhibition of the sarcoplasmic reticulum (SR). Using 10 mM 1,2-bis(o-N,N,N',N'-tetraacetic acid (170 nM Ca2+), I (Ca,L) inactivation was then determined by a Ca2+ -independent, voltage-dependant process. Under these conditions, 3 mu M sorcin speeded up inactivation. A similar effect was observed by substitution of Ca2+ with Ba2+. Down-regulation of endogenous sorcin to 27 +/- 7% using an RNAi adenoviral vector slowed inactivation of I (Ca,L) by similar to 42%. The effects of sorcin on voltage-dependant inactivation were mimicked by a truncated form of the protein containing only the Ca2+-binding domain. This data is consistent with two independent actions of sorcin on the LTCC: (1) slowing Ca2+-dependant inactivation and (2) stimulating voltage-dependant inactivation. The net effect of sorcin on the time-dependent inactivation of I (Ca,L) was a balance between these two effects. Under normal conditions, sorcin slows I (Ca,L) inactivation because the effects of Ca2+-dependant inactivation out-weigh the effects on voltage-dependant inactivation.