| 18057120 |
White CN, Hamilton EJ, Garcia A, Wang D, Chia KK, Figtree GA, Rasmussen HH: Opposing effects of coupled and uncoupled NOS activity on the Na+-K+ pump in cardiac myocytes. Am J Physiol Cell Physiol. 2008 Feb;294(2):C572-8. Epub 2007 Dec 5.
Pharmacological delivery of nitric oxide (NO) stimulates the cardiac Na (+)-K (+) pump. However, effects of NO synthesized by NO synthase (NOS) often differ from the effects of NO delivered pharmacologically. In addition, NOS can become "uncoupled" and preferentially synthesize O (2)(.-), which often has opposing effects to NO. We tested the hypothesis that NOS-synthesized NO stimulates Na (+)-K (+) pump activity, and uncoupling of NOS inhibits it. To image NO, we loaded isolated rabbit cardiac myocytes with 4,5-diaminofluorescein-2 diacetate (DAF-2 DA) and measured fluorescence with confocal microscopy. L-arginine (L-arg; 500 micromol/l) increased DAF-2 DA fluorescence by 51% compared with control (n = 8; P < 0.05). We used the whole cell patch-clamp technique to measure electrogenic Na (+)-K (+) pump current (I (p)). Mean I (p) of 0.35 +/- 0.03 pA/pF (n = 44) was increased to 0.48 +/- 0.03 pA/pF (n = 7, P < 0.05) by 10 micromol/l L-Arg in pipette solutions. This increase was abolished by NOS inhibition with radicicol or by NO-activated guanylyl cyclase inhibition with 1H-[1,2,4] oxadiazole [4,3-a] quinoxalin-1-one. We next examined the effect of uncoupling NOS using paraquat. Paraquat (1 mmol/l) induced a 51% increase in the fluorescence intensity of O (2)(.-)-sensitive dye dihydroethidium compared with control (n = 9; P < 0.05). To examine the functional effects of uncoupling, we measured I (p) with 100 micromol/l paraquat included in patch pipette solutions. This decreased I (p) to 0.28 +/- 0.03 pA/pF (n = 12; P < 0.001). The paraquat-induced pump inhibition was abolished by superoxide dismutase (in pipette solutions). We conclude that NOS-mediated NO synthesis stimulates the Na (+)-K (+) pump, whereas uncoupling of NOS causes O (2)(.-)-mediated pump inhibition. |
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