抄録
L-type Cav1.2 Ca2+ channel is regulated by Ca2+-dependent facilitation (CDF) and inactivation (CDI). Although calmodulin (CaM) has been suggested to mediate both CDI and CDF, CaM-dependent protein kinase II (CaMKII) is also suggested to play an important role in CDF. In this study, we examined the roles of Ca2+, CaM and CaMKII in CDF and CDI using patch-clamp method in guinea-pig cardiomyocytes, in which run-down of the channel was controlled. Application of CaM (0.1-14 μM) + ATP ( [Ca2+]i <10 nM) to the intracellular side of the channels, within 1 min after patch excision, dose-dependently produced channel activity up to 250% of control. The relationship between [CaM] and channel activity was bell-shaped with a peak at ∼[CaM] 3 μM. Increasing [Ca2+]i (–500 nM) shifted the curve toward lower [CaM]. Thus, at a fixed [CaM] (e.g. 0.5μM), Ca2+ showed biphasic effects: facilitation at [Ca2+]i <500 nM, and inactivation at [Ca2+]i >500 nM. This Ca2+-dependent effect of CaM was considered to represent CDF and CDI and was not affected by protein kinase inhibitors KN-62 and K252a. The effects of CaM were attenuated, however, when the run-down time of the channels was >5 min. Application of CaMKII together with CaM + ATP, during the run-down period, restored the susceptibility of the channel to the modulation by CaM. These results suggested that CaM plays a key role in CDF and CDI, and that CaMKII and [Ca2+]i modulate the effect of CaM. [J Physiol Sci. 2007;57 Suppl:S14]
