Melatonin inhibits nitric oxide production by microvascular endothelial cells in vivo and in vitro.
OBJECTIVE
We have previously shown that melatonin inhibits bradykinin-induced NO production by endothelial cells in vitro. The purpose of this investigation was to extend this observation to an in vivo condition and to explore the mechanism of action of melatonin.


METHODS
RT-PCR assays were performed with rat cultured endothelial cells. The putative effect of melatonin upon arteriolar tone was investigated by intravital microscopy while NO production by endothelial cells in vitro was assayed by fluorimetry, and intracellular Ca(2+) measurements were assayed by confocal microscopy.


RESULTS
No expression of the mRNA for the melatonin synthesizing enzymes, arylalkylamine N-acetyltransferase and hydroxyindole-O-methyltransferase, or for the melatonin MT(2) receptor was detected in microvascular endothelial cells. Melatonin fully inhibited L-NAME-sensitive bradykinin-induced vasodilation and also inhibited NO production induced by histamine, carbachol and 2-methylthio ATP, but did not inhibit NO production induced by ATP or alpha, beta-methylene ATP. None of its inhibitory effects was prevented by the melatonin receptor antagonist, luzindole. In nominally Ca(2+)-free solution, melatonin reduced intracellular Ca(2+) mobilization induced by bradykinin (40%) and 2-methylthio ATP (62%) but not Ca(2+) mobilization induced by ATP.


CONCLUSIONS
We have confirmed that melatonin inhibited NO production both in vivo and in vitro. In addition, the melatonin effect was selective for some G protein-coupled receptors and most probably reflects an inhibition of Ca(2+) mobilization from intracellular stores.
Silva
C L M
CL
Departamento de Fisiologia, Instituto de Biociências, Universidade de São Paulo, São Paulo, Brazil.
Tamura
E K
EK
Macedo
S M D
SM
Cecon
E
E
Bueno-Alves
L
L
Farsky
S H P
SH
Ferreira
Z S
ZS
Markus
R P
RP
D000121:Acetylserotonin O-Methyltransferase / Q000235:genetics; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D047068:Arylalkylamine N-Acetyltransferase / Q000235:genetics; D001783:Blood Flow Velocity / Q000187:drug effects; D002118:Calcium / Q000378:metabolism; D002217:Carbachol / Q000494:pharmacology; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D005470:Fluorometry; D015870:Gene Expression / Q000187:drug effects; D006632:Histamine / Q000494:pharmacology; D008297:Male; D008550:Melatonin / Q000494:pharmacology*; D008638:Mesenteric Arteries / Q000187:drug effects* / Q000502:physiology; D018613:Microscopy, Confocal; D018715:Microscopy, Video; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D009569:Nitric Oxide / Q000096:biosynthesis*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D044123:Receptor, Melatonin, MT2 / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014664:Vasodilation / Q000187:drug effects
D012333:RNA, Messenger; D044123:Receptor, Melatonin, MT2; D009569:Nitric Oxide; D006632:Histamine; D000255:Adenosine Triphosphate; D002217:Carbachol; D000121:Acetylserotonin O-Methyltransferase; D047068:Arylalkylamine N-Acetyltransferase; D008550:Melatonin; D002118:Calcium; D019331:NG-Nitroarginine Methyl Ester