Phosphoinositide 3-kinase regulates excitation-contraction coupling in neonatal cardiomyocytes

• Published in: American journal of physiology. Heart and circulatory physiology 2004-02, Vol.286 (2), p.H796-H805
• Main Authors: McDowell, Susan A, McCall, Eileen, Matter, William F, Estridge, Thomas B, Vlahos, Chris J
• Format: Article
• Language: English
• Subjects: 3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester - pharmacology; Animals; Animals, Newborn; Calcium - metabolism; Calcium Channels, L-Type - drug effects; Calcium Channels, L-Type - physiology; Calcium-Transporting ATPases - metabolism; Cells, Cultured; Chromones - pharmacology; Endoplasmic Reticulum - drug effects; Endoplasmic Reticulum - physiology; Enzyme Inhibitors - pharmacology; Heart Ventricles - drug effects; Kinetics; Morpholines - pharmacology; Myocardial Contraction - physiology; Myocytes, Cardiac - enzymology; Myocytes, Cardiac - physiology; Phosphatidylinositol 3-Kinases - antagonists & inhibitors; Phosphatidylinositol 3-Kinases - physiology; Piperazines - pharmacology; Platelet-Derived Growth Factor - pharmacology; Rats; Rats, Sprague-Dawley; Ryanodine Receptor Calcium Release Channel - physiology; Sarcoplasmic Reticulum - drug effects; Sarcoplasmic Reticulum - physiology; Sarcoplasmic Reticulum Calcium-Transporting ATPases; Sodium-Calcium Exchanger - metabolism; Ventricular Function
• ISSN: 0363-6135; 1522-1539
• DOI: 10.1152/ajpheart.00546.2003

Cardiovascular Research, Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana 46285 Submitted 10 June 2003 ; accepted in final form 7 October 2003 The phosphoinositide 3-kinase (PI3K) inhibitor LY-294002 decreased steady-state contraction in neonatal rat ventricular myocytes (NRVM). To determine whether the effect on steady-state contraction could be due to decreased intracellular Ca 2+ content, Ca 2+ content was assessed with fluorescent plate reader analysis by using the caffeine-releasable Ca 2+ stores as an index of sarcoplasmic reticulum (SR) Ca 2+ content. Caffeine-releasable Ca 2+ content was diminished in a dose-dependent manner with LY-294002, suggesting that the decrease in steady-state contraction was due to diminished intracellular Ca 2+ content. Activation of the L-type Ca 2+ channel by BAY K 8644 was attenuated by LY-294002, suggesting the effect of LY-294002 is to reduce Ca 2+ influx at this channel. To investigate whether additional proteins involved in excitation-contraction (EC) coupling are likewise regulated by PI3K activity, the effects of compounds acting at sarco(endo)plasmic reticulum Ca 2+ -ATPase (SERCA2a), the ryanodine receptor, and the Na/Ca exchanger (NCX) were compared with LY-294002. Inhibition of SERCA2a by thapsigargin increased basal Ca 2+ levels in contrast to LY-294002, indicating that SERCA2a activity is sustained in the presence of LY-294002. Ryanodine decreased SR Ca 2+ content. The additive effect with coadministration of LY-294002 could be attributed to a decrease in Ca 2+ influx at the L-type Ca 2+ channel. The NCX inhibitor Ni 2+ was used to investigate whether the decrease in intracellular Ca 2+ content with LY-294002 could be due to inhibition of the NCX reverse-mode activity. The minimal effect of LY-294002 with Ni 2+ suggests that the primary effect of LY-294002 on EC coupling occurs through inhibition of PI3K-mediated L-type Ca 2+ channel activity. LY-294002; L-type calcium channel; fluorescent plate reader Address for reprint requests and other correspondence: C. J. Vlahos, Cardiovascular Research, Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, IN 46285-0520 (E-mail: Vlahos_Chris_J{at}Lilly.com ).