Elevation of intracellular calcium levels contributes to the inhibition of nitric oxide production by atrial natriuretic peptide.
Atrial natriuretic peptide (ANP) attenuates LPS-induced inducible nitric oxide synthase (iNOS) expression in murine macrophages by destabilizing iNOS mRNA. Because elevated intracellular free Ca2+ levels [Ca2+]i reduce iNOS mRNA stability, the aim of the present study was to determine whether inhibition of iNOS by ANP is due to alterations in intracellular calcium. As determined by fluorescence photometry, ANP (10(-7) and 10(-6) mol/L) was shown to elevate intracellular calcium levels in bone marrow-derived macrophages. This effect seemed to be mediated via the guanylate cyclase-coupled A receptor, because dibutyryl-cGMP mimicked and the A-receptor antagonist HS-142-1 partially abrogated the effect of ANP. Because the Ca2+ increase was also observed in Ca2+-free buffer, it is suggested that the liberation of intracellular calcium pools contributes to the elevation of [Ca2+]i by ANP. The B-receptor ligand C-type natriuretic peptide (CNP), which does not alter iNOS expression, had no effect on [Ca2+]i. The Ca2+-ionophore 4-Br-A23187 and thapsigargin, a compound known to liberate Ca2+ from intracellular stores, were further demonstrated to reduce LPS-induced NO production in macrophages (Griess assay), confirming a functional link for elevated [Ca2+]i and iNOS inhibition. These effects were abrogated by coincubation with extra- as well as intracellular Ca2+ chelators (EGTA, 1,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA)). The inhibitory effect of ANP on NO production was also abrogated by Ca2+ chelation. These findings support a causal relationship between reduced iNOS induction and elevation of [Ca2+]i. Taken together, the data indicate that intracellular Ca2+ elevation by ANP is involved in the inhibition of LPS-induced nitric oxide production in macrophages.
Kiemer
A K
AK
Institute of Pharmacy, Center of Drug Research, Pharmaceutical Biology, University of Munich, Munich, Germany. Alexandra.Kiemer@cup.uni-muenchen.de
Vollmar
A M
AM
D000814:Aniline Compounds / Q000378:metabolism; D000818:Animals; D009320:Atrial Natriuretic Factor / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D000001:Calcimycin / Q000031:analogs & derivatives* / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D003995:Dibutyryl Cyclic GMP / Q000494:pharmacology; D005456:Fluorescent Dyes / Q000378:metabolism; D006162:Guanylate Cyclase / Q000037:antagonists & inhibitors; D008070:Lipopolysaccharides / Q000494:pharmacology; D008264:Macrophages / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D051379:Mice; D009569:Nitric Oxide / Q000096:biosynthesis* / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors* / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II; D011134:Polysaccharides / Q000494:pharmacology; D017461:Receptors, Atrial Natriuretic Factor / Q000037:antagonists & inhibitors; D019284:Thapsigargin / Q000494:pharmacology; D014966:Xanthenes / Q000378:metabolism
D000814:Aniline Compounds; D005456:Fluorescent Dyes; C072551:HS 142-1; D008070:Lipopolysaccharides; D011134:Polysaccharides; D014966:Xanthenes; C059715:Fluo-3; D009569:Nitric Oxide; D003995:Dibutyryl Cyclic GMP; D000001:Calcimycin; D019284:Thapsigargin; C033170:4-bromo-A-23187; D009320:Atrial Natriuretic Factor; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496317:Nos2 protein, mouse; D006162:Guanylate Cyclase; D017461:Receptors, Atrial Natriuretic Factor; C091715:atrial natriuretic factor receptor A; D002118:Calcium