Endothelial nitric oxide synthase decreases beta-adrenergic responsiveness via inhibition of the L-type Ca2+ current.
Signaling via endothelial nitric oxide synthase (NOS3) limits the heart's response to beta-adrenergic (beta-AR) stimulation, which may be protective against arrhythmias. However, mechanistic data are limited. Therefore, we performed simultaneous measurements of action potential (AP, using patch clamp), Ca2+ transients (fluo 4), and myocyte shortening (edge detection). L-type Ca2+ current (ICa) was directly measured by the whole cell ruptured patch-clamp technique. Myocytes were isolated from wild-type (WT) and NOS3 knockout (NOS3-/-) mice. NOS3-/- myocytes exhibited a larger incidence of beta-AR (isoproterenol, 1 microM)-induced early afterdepolarizations (EADs) and spontaneous activity (defined as aftercontractions). We also examined ICa, a major trigger for EADs. NOS3-/- myocytes had a significantly larger beta-AR-stimulated increase in ICa compared with WT myocytes. In addition, NOS3-/- myocytes had a larger response to beta-AR stimulation compared with WT myocytes in Ca2+ transient amplitude, shortening amplitude, and AP duration (APD). We observed similar effects with specific NOS3 inhibition [L-N5-(1-iminoethyl)-ornithine (l-NIO), 10 microM] in WT myocytes as with NOS3 knockout. Specifically, l-NIO further increased isoproterenol-stimulated EADs and aftercontractions. l-NIO also further increased the isoproterenol-stimulated ICa, Ca2+ transient amplitude, shortening amplitude, and APD (all P < 0.05 vs isoproterenol alone). l-NIO had no effect in NOS3-/- myocytes. These results indicate that NOS3 signaling inhibits the beta-AR response by reducing ICa and protects against arrhythmias. This mechanism may play an important role in heart failure, where arrhythmias are increased and NOS3 expression is decreased.
Wang
Honglan
H
Department of Physiology and Cell Biology, The Ohio State University, Columbus, OH 43210, USA.
Kohr
Mark J
MJ
Wheeler
Debra G
DG
Ziolo
Mark T
MT
D000200:Action Potentials / Q000187:drug effects; D000318:Adrenergic beta-Agonists / Q000494:pharmacology; D000818:Animals; D001145:Arrhythmias, Cardiac / Q000503:physiopathology; D020746:Calcium Channels, L-Type / Q000187:drug effects / Q000502:physiology*; D020013:Calcium Signaling / Q000187:drug effects; D007545:Isoproterenol / Q000494:pharmacology; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D009200:Myocardial Contraction / Q000187:drug effects; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000502:physiology; D052250:Nitric Oxide Synthase Type III / Q000235:genetics / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D011943:Receptors, Adrenergic, beta / Q000502:physiology*
D000318:Adrenergic beta-Agonists; D020746:Calcium Channels, L-Type; D011943:Receptors, Adrenergic, beta; D052250:Nitric Oxide Synthase Type III; D007545:Isoproterenol