Angiotensin-(1-7) increases neuronal potassium current via a nitric oxide-dependent mechanism

Yang R, Yin J, Li Y, Zimmerman MC, Schultz HD. Angiotensin(1-7) increases neuronal potassium current via a nitric oxide-dependent mechanism. Am J Physiol Cell Physiol 300: C58-C64, 2011. First published October 27, 2010; doi: 10.1152/ajpcell.00369.2010.-Actions of angiotensin-(1-7) [Ang-(1-7)], a heptapeptide of the renin-angiotensin system, in the periphery are mediated, at least in part, by activation of nitric oxide (NO) synthase (NOS) and generation NO center dot. Studies of the central nervous system have shown that NO center dot acts as a sympathoinhibitory molecule and thus may play a protective role in neurocardiovascular diseases associated with sympathoexcitation, such as hypertension and heart failure. However, the contribution of NO in the intraneuronal signaling pathway of Ang-(1-7) and the subsequent modulation of neuronal activity remains unclear. Here, we tested the hypothesis that neuronal NOS (nNOS)-derived NO center dot mediates changes in neuronal activity following Ang-(1-7) stimulation. For these studies, we used differentiated catecholaminergic (CATH.a) neurons, which we show express the Ang-(1-7) receptor (Mas R) and nNOS. Stimulation of CATH. a neurons with Ang-(1-7) (100 nM) increased intracellular NO levels, as measured by 4-amino-5-methylamino- 2',7' -difluorofluoresceindiacetate (DAF-FM) fluorescence and confocal microscopy. This response was significantly attenuated in neurons pretreated with the Mas R antagonist (A-779), a nonspecific NOS inhibitor (nitro-L-arginine methyl ester), or an nNOS inhibitor (S-methyl-L-thiocitrulline, SMTC), but not by endothelial NOS (eNOS) or inhibitory NOS (iNOS) inhibition {L-N-5-(1-iminoethyl) ornithine (L-NIO) and 1400W, respectively}. To examine the effect of Ang-(1-7)-NO center dot signaling on neuronal activity, we recorded voltage-gated outward K+ current (I-Kv) in CATH. a neurons using the whole cell configuration of the patch-clamp technique. Ang-(1-7) significantly increased I-Kv, and this response was inhibited by A-779 or S-methyl-L-thiocitrulline, but not L-NIO or 1400W. These findings indicate that Ang-(1-7) is capable of increasing nNOS-derived NO center dot levels, which in turn, activates hyperpolarizing I-Kv in catecholaminergic neurons.