K201 (JTV519), a benzothiazepine derivative, has been shown to possess 
anti-arrhythmic and cardioprotective properties, but the mechanism of its action 
is both complex and controversial. It is believed to stabilize the closed state 
of the RyR2 (cardiac ryanodine receptor) by increasing its affinity for the 
FKBP12.6 (12.6 kDa FK506 binding protein) [Wehrens, Lehnart, Reiken, Deng, Vest, 
Cervantes, Coromilas, Landry and Marks (2004) Science 304, 292-296]. In the 
present study, we investigated the effect of K201 on spontaneous Ca2+ release 
induced by Ca2+ overload in rat ventricular myocytes and in HEK-293 cells (human 
embryonic kidney cells) expressing RyR2 and the role of FKBP12.6 in the action 
of K201. We found that K201 abolished spontaneous Ca2+ release in cardiac 
myocytes in a concentration-dependent manner. Treating ventricular myocytes with 
FK506 to dissociate FKBP12.6 from RyR2 did not affect the suppression of 
spontaneous Ca2+ release by K201. Similarly, K201 was able to suppress 
spontaneous Ca2+ release in FK506-treated HEK-293 cells co-expressing RyR2 and 
FKBP12.6. Furthermore, K201 suppressed spontaneous Ca2+ release in HEK-293 cells 
expressing RyR2 alone and in cells co-expressing RyR2 and FKBP12.6 with the same 
potency. In addition, K201 inhibited [3H]ryanodine binding to RyR2-wt 
(wild-type) and an RyR2 mutant linked to ventricular tachycardia and sudden 
death, N4104K, in the absence of FKBP12.6. These observations demonstrate that 
FKBP12.6 is not involved in the inhibitory action of K201 on spontaneous Ca2+ 
release. Our results also suggest that suppression of spontaneous Ca2+ release 
and the activity of RyR2 contributes, at least in part, to the anti-arrhythmic 
properties of K201.