The phosphoinositide 3-kinase inhibitor LY294002 enhances cardiac myocyte contractility via a direct inhibition of Ik,slow currents.
OBJECTIVE
Phosphoinositide 3-kinase (PI3K) is a key component in regulating myocardial growth, survival and contractility. LY294002 and wortmannin are two PI3K inhibitors used widely to establish the role of PI3K. The goal of this study was to examine the effects of acute application of LY294002 and wortmannin on cardiac myocyte contractility and underlying mechanisms.


METHODS
Patch-clamp, indo-1 epifluorescence and video-edge detection techniques were used to measure outward K(+) currents, action potentials (AP), Ca(2+) transients and shortening of myocytes isolated from mouse left ventricular free wall.


RESULTS
In field-stimulated myocytes, LY294002 (10 micromol/l) increased Ca(2+) transient amplitude by 23%, and cell shortening amplitude by 60% in the absence or presence of wortmannin, while wortmannin alone had no effect. LY294002 (but not wortmannin) prolonged AP duration by specifically inhibiting slowly inactivating K(+) currents (i.e., the 4-aminopyrydine-sensitive I(k,slow1) and the tetraethylammonium-sensitive I(k,slow2)), leading to an increase in sarcoplasmic reticular Ca(2+) levels. It appeared that the AP prolongation was responsible for elevated contractility since AP-clamp of myocytes with prolonged APs (recorded in LY294002-treated myocytes) induced a 29% increase in cell shortening compared with control APs, while LY294002 application did not increase contractility in voltage-clamp studies using either step or AP depolarizations.


CONCLUSIONS
The putative PI3K inhibitor LY294002 increases Ca(2+) release and myocyte contractility via direct inhibition of cardiac I(k,slow) and AP prolongation, thus limiting the usefulness of this agent in the analyses of the role of PI3K in heart function.
Sun
Hui
H
Department of Physiology and Medicine, Heart & Stroke/Richard Lewar Centre and Division of Cardiology, University Health Network, University of Toronto Fitzgerald Building, 150 College St., Toronto, Ontario, Canada M5S 3E2.
Oudit
Gavin Y
GY
Ramirez
Rafael J
RJ
Costantini
Danny
D
Backx
Peter H
PH
D000200:Action Potentials / Q000187:drug effects; D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000378:metabolism; D048429:Cell Size / Q000187:drug effects; D002867:Chromones / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D007211:Indoles; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008856:Microscopy, Fluorescence; D009025:Morpholines / Q000494:pharmacology*; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D024642:Potassium Channels, Voltage-Gated / Q000187:drug effects* / Q000378:metabolism; D012519:Sarcoplasmic Reticulum / Q000378:metabolism; D000077191:Wortmannin
D000730:Androstadienes; D002118:Calcium; D002867:Chromones; D007211:Indoles; D009025:Morpholines; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D024642:Potassium Channels, Voltage-Gated; D000077191:Wortmannin; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C048960:indo-1