Calmodulin reverses rundown of L-type Ca 2+ channels in guinea pig ventricular myocytes

• Published in: American Journal of Physiology: Cell Physiology 2004-12, Vol.287 (6), p.C1717-C1724
• Main Authors: Xu, Jian-Jun, Hao, Li-Ying, Kameyama, Asako, Kameyama, Masaki
• Format: Article
• Language: English
• ISSN: 0363-6143; 1522-1563
• DOI: 10.1152/ajpcell.00105.2004

Calmodulin (CaM) is implicated in regulation of Ca 2+ channels as a Ca 2+ sensor. The effect of CaM on rundown of L-type Ca 2+ channels in inside-out patch form was investigated in guinea pig ventricular myocytes. Ca 2+ channel activity disappeared within 1–3 min and did not reappear when the patch was excised and exposed to an artificial intracellular solution. However, application of CaM (0.03, 0.3, 3 μM) + 3 mM ATP to the intracellular solution within 1 min after patch excision resulted in dose-dependent activation of channel activity. Channel activity averaged 11.2%, 94.7%, and 292.9%, respectively, of that in cell-attached mode. Channel activity in inside-out patch mode was induced by CaM + ATP at nanomolar Ca 2+ concentrations ([Ca 2+ ]); however, increase to micromolar [Ca 2+ ] rapidly inactivated the channel activity induced, revealing that the effect of CaM on the channel was Ca 2+ dependent. At the 2nd, 4th, 6th, 8th, and 10th minutes after patch excision, CaM (0.75 μM) + ATP induced Ca 2+ channel activity to 150%, 100%, 96.9%, 29.3%, and 16.6%, respectively, revealing a time-dependent action of CaM on the channel. CaM added with adenosine 5′-(β,γ-imido)triphosphate (AMP-PNP) also induced channel activity, although with much lower potency and shorter duration. Protein kinase inhibitors KN-62, CaM-dependent protein kinase (CaMK)II 281-309, autocamtide-related CaMKII inhibitor peptide, and K252a (each 1–10 μM) did not block the effect of CaM, indicating that the effect of CaM on the Ca 2+ channel was phosphorylation independent. Neither CaM nor ATP alone induced Ca 2+ channel activity, showing a cooperative effect of CaM and ATP on the Ca 2+ channel. These results suggest that CaM is a crucial regulatory factor of Ca 2+ channel basal activity.