Mechanisms of tolerance to sodium nitroprusside in cultured rat aortic smooth muscle cells (SMC)

Andreas Papapetropoulos, C. Y. Go, F. Murad, J. D. Calravas

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Abstract

We investigated mechanisms responsible for the sodium nitroprusside (SNP)-induced tolerance to guanylate cyclase (sGC) activation in intact SMC. sGC activity was estimated from the accumulation of cGMP in response to lOpM SNP over a 15min period in the presence of a phosphodiesterase (PDE) inhibitor. Pretrealment of rat aortic SMC with 10500pM SNP led to a dose-dependent downregulation of cGMP accumulation upon subsequent SNP stimulation, which was blocked by coincubation with 30uM oxyhemoglobin. Pretreatment of SMC with the PDE inhibitor zaprinast resulted in downregulation of the SNP-induced cGMP accumulation. This downregulation was completely inhibited by the protein kinase A (PKA) inhibitor H89; tolerance to SNP was only partially reversed by H89. P, sGC steady state mRNA levels of S-nitroso Nacetylpenicillamine (SNAP)- or 8Br-cGMP-pretreated cells were unchanged, as indicated by Northern blot analysis. However, Western blot analysis revealed that a, protein levels were decreased in zaprinast, but not in SNP, SNAP or 8Br-cGMP pretreated cells. While thiol depletion did not prevent the development of tolerance, pretreatment of cells with SNP in the presence of reducing agents partially or completely restored the ability of cells to respond to SNP. Thus, tolerance to SNP results from two distinct mechanisms: an early onset, NO-mediated event that is reversed by reducing agents and a more delayed, PKA-sensitive process that is mediated through increased cGMP and decreased sGC protein levels. Supported by HL52958.

Original languageEnglish (US)
JournalFASEB Journal
Volume10
Issue number3
StatePublished - Dec 1 1996
Externally publishedYes

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ASJC Scopus subject areas

  • Agricultural and Biological Sciences (miscellaneous)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Biochemistry
  • Cell Biology

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