Inhibitory actions of the phosphatidylinositol 3-kinase inhibitor LY294002 on the human Kv1.5 channel.
OBJECTIVE
Kv1.5 channels conduct the ultra-rapid delayed rectifier potassium current (I(Kur)), and in humans, Kv1.5 channels are highly expressed in cardiac atria but are scarce in ventricles. Pharmacological blockade of human Kv1.5 (hKv1.5) has been regarded as effective for prevention and treatment of re-entry-based atrial tachyarrhythmias. Here we examined blockade of hKv1.5 channels by LY294002, a well-known inhibitor of phosphatidylinositol 3-kinase (PI3K).


METHODS
hKv1.5 channels were heterologously expressed in Chinese hamster ovary cells. Effects of LY294002 on wild-type and mutant (T462C, H463C, T480A, R487V, A501V, I502A, I508A, L510A and V516A) hKv1.5 channels were examined by using the whole-cell patch-clamp method.


RESULTS
LY294002 rapidly and reversibly inhibited hKv1.5 current in a concentration-dependent manner (IC(50) of 7.9 micromol.L(-1)). In contrast, wortmannin, a structurally distinct inhibitor of PI3K, had little inhibitory effect on hKv1.5 current. LY294002 block of hKv1.5 current developed with time during depolarizing voltage-clamp steps, and this blockade was also voltage-dependent with a steep increase over the voltage range for channel openings. The apparent binding (k(+1)) and unbinding (k(-1)) rate constants were calculated to be 1.6 micromol.L(-1) (-1).s(-1) and 5.7 s(-1) respectively. Inhibition by LY294002 was significantly reduced in several hKv1.5 mutant channels: T480A, R487V, I502A, I508A, L510A and V516A.


CONCLUSIONS
LY294002 acts directly on hKv1.5 currents as an open channel blocker, independently of its effects on PI3K activity. Amino acid residues located in the pore region (Thr480, Arg487) and the S6 segment (Ile502, Ile508, Leu510, Val516) appear to constitute potential binding sites for LY294002.
Wu
J
J
Pharmacology Department, Medical School of Xi'an Jiaotong University, Xi'an, Shaanxi, China.
Ding
W-G
WG
Matsuura
H
H
Tsuji
K
K
Zang
W-J
WJ
Horie
M
M
D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D001665:Binding Sites; D016466:CHO Cells; D002867:Chromones / Q000494:pharmacology*; D006224:Cricetinae; D003412:Cricetulus; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005260:Female; D006801:Humans; D051666:Kv1.5 Potassium Channel / Q000037:antagonists & inhibitors* / Q000235:genetics; D008564:Membrane Potentials / Q000187:drug effects; D009025:Morpholines / Q000494:pharmacology*; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D018408:Patch-Clamp Techniques; D000081082:Phosphoinositide-3 Kinase Inhibitors; D013997:Time Factors; D000077191:Wortmannin
D000730:Androstadienes; D002867:Chromones; D004791:Enzyme Inhibitors; D051666:Kv1.5 Potassium Channel; D009025:Morpholines; D000081082:Phosphoinositide-3 Kinase Inhibitors; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D000077191:Wortmannin