Neuronal Nitric Oxide Synthase Activation Is Involved in Insulin-Mediated Cardiovascular Effects in the Nucleus Tractus Solitarii of Rats

H. T. Chiang, W. H. Cheng, P. J. Lu, H. N. Huang, W. C. Lo, Y. C. Tseng, J. L. Wang, M. Hsiao, C. J. Tseng

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Abstract

Neuronal nitric oxide synthases (nNOS) is distributed throughout the central nervous system (CNS) and has been proposed to modulate neuronal activity in the nucleus tractus solitarii (NTS). Here, we investigated whether the activation of nNOS is involved in insulin-induced cardiovascular responses in the NTS. Insulin (100 IU/ml) was unilaterally microinjected into the NTS, and the cardiovascular effects were evaluated before and after microinjection of the nNOS inhibitors 7-nitroindazole (7-NI) (5 pmol) and N(5)-(1-imino-3-butenyl)-l-ornithine (vinyl-L-NIO) (600 pmol). Western blot and immunohistochemical analyses were performed to determine nNOS phosphorylation levels after insulin or phosphoinositide 3-kinase (PI3K) inhibitor LY294002 microinjection into the NTS. Unilateral microinjection of insulin into the NTS produced prominent depressor and bradycardic effects in WKY rats. Pretreatment with the nNOS inhibitors 7-NI and Vinyl-L-NIO attenuated the cardiovascular response evoked by insulin in Wistar-Kyoto (WKY) rats. Moreover, Western blot analysis showed a significant increase in nNOS (16.5±0.4-fold; P<0.05; n=4) phosphorylation after insulin injection, whereas the PI3K inhibitor LY294002 abolished the insulin-induced effects. In situ nNOS phosphorylation was found to be increased in the NTS after insulin injection. Furthermore, co-immunoprecipitation assay showed Akt and nNOS can bind to each other as detected by phospho-AktS473 and phospho-nNOSS1416 antibodies. In vitro kinase assay showed insulin activated Akt can directly phosphorylate nNOSS1416. These results demonstrated that nNOS may couple with the activation of the insulin receptor, via the liberation of NO, in order to participate in central cardiovascular regulation of WKY rats.

Original languageEnglish
Pages (from-to)727-734
Number of pages8
JournalNeuroscience
Volume159
Issue number2
DOIs
Publication statusPublished - 2009 Mar 17

Fingerprint

Nitric Oxide Synthase Type I
Solitary Nucleus
Insulin
Inbred WKY Rats
Microinjections
2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one
1-Phosphatidylinositol 4-Kinase
Phosphorylation
Western Blotting
Injections
Insulin Receptor
Immunoprecipitation
Phosphotransferases
Central Nervous System
Antibodies

All Science Journal Classification (ASJC) codes

  • Neuroscience(all)

Cite this

Chiang, H. T. ; Cheng, W. H. ; Lu, P. J. ; Huang, H. N. ; Lo, W. C. ; Tseng, Y. C. ; Wang, J. L. ; Hsiao, M. ; Tseng, C. J. / Neuronal Nitric Oxide Synthase Activation Is Involved in Insulin-Mediated Cardiovascular Effects in the Nucleus Tractus Solitarii of Rats. In: Neuroscience. 2009 ; Vol. 159, No. 2. pp. 727-734.
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abstract = "Neuronal nitric oxide synthases (nNOS) is distributed throughout the central nervous system (CNS) and has been proposed to modulate neuronal activity in the nucleus tractus solitarii (NTS). Here, we investigated whether the activation of nNOS is involved in insulin-induced cardiovascular responses in the NTS. Insulin (100 IU/ml) was unilaterally microinjected into the NTS, and the cardiovascular effects were evaluated before and after microinjection of the nNOS inhibitors 7-nitroindazole (7-NI) (5 pmol) and N(5)-(1-imino-3-butenyl)-l-ornithine (vinyl-L-NIO) (600 pmol). Western blot and immunohistochemical analyses were performed to determine nNOS phosphorylation levels after insulin or phosphoinositide 3-kinase (PI3K) inhibitor LY294002 microinjection into the NTS. Unilateral microinjection of insulin into the NTS produced prominent depressor and bradycardic effects in WKY rats. Pretreatment with the nNOS inhibitors 7-NI and Vinyl-L-NIO attenuated the cardiovascular response evoked by insulin in Wistar-Kyoto (WKY) rats. Moreover, Western blot analysis showed a significant increase in nNOS (16.5±0.4-fold; P<0.05; n=4) phosphorylation after insulin injection, whereas the PI3K inhibitor LY294002 abolished the insulin-induced effects. In situ nNOS phosphorylation was found to be increased in the NTS after insulin injection. Furthermore, co-immunoprecipitation assay showed Akt and nNOS can bind to each other as detected by phospho-AktS473 and phospho-nNOSS1416 antibodies. In vitro kinase assay showed insulin activated Akt can directly phosphorylate nNOSS1416. These results demonstrated that nNOS may couple with the activation of the insulin receptor, via the liberation of NO, in order to participate in central cardiovascular regulation of WKY rats.",
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Neuronal Nitric Oxide Synthase Activation Is Involved in Insulin-Mediated Cardiovascular Effects in the Nucleus Tractus Solitarii of Rats. / Chiang, H. T.; Cheng, W. H.; Lu, P. J.; Huang, H. N.; Lo, W. C.; Tseng, Y. C.; Wang, J. L.; Hsiao, M.; Tseng, C. J.

In: Neuroscience, Vol. 159, No. 2, 17.03.2009, p. 727-734.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Neuronal Nitric Oxide Synthase Activation Is Involved in Insulin-Mediated Cardiovascular Effects in the Nucleus Tractus Solitarii of Rats

AU - Chiang, H. T.

AU - Cheng, W. H.

AU - Lu, P. J.

AU - Huang, H. N.

AU - Lo, W. C.

AU - Tseng, Y. C.

AU - Wang, J. L.

AU - Hsiao, M.

AU - Tseng, C. J.

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