Evaluation of ADMA-DDAH-NOS axis in specific brain areas following nitroglycerin administration: study in an animal model of migraine.
BACKGROUND
Nitric oxide (NO) is known to play a key role in migraine pathogenesis, but modulation of NO synthesis has failed so far to show efficacy in migraine treatment. Asymmetric dimethylarginine (ADMA) is a NO synthase (NOS) inhibitor, whose levels are regulated by dimethylarginine dimethylaminohydrolase (DDAH). Systemic administration of nitroglycerin (or glyceryl trinitrate, GTN) is a NO donor that consistently induces spontaneous-like headache attacks in migraneurs. GTN administration induces an increase in neuronal NOS (nNOS) that is simultaneous with a hyperalgesic condition. GTN administration has been used for years as an experimental animal model of migraine. In order to gain further insights in the precise mechanisms involved in the relationships between NO synthesis and migraine, we analyzed changes induced by GTN administration in ADMA levels, DDHA-1 mRNA expression and the expression of neuronal and endothelial NOS (nNOS and eNOS) in the brain. We also evaluated ADMA levels in the serum.


METHODS
Male Sprague-Dawley rats were injected with GTN (10 mg/kg, i.p.) or vehicle and sacrificed 4 h later. Brain areas known to be activated by GTN administration were dissected out and utilized for the evaluation of nNOS and eNOS expression by means of western blotting. Cerebral and serum ADMA levels were measured by means of ELISA immunoassay. Cerebral DDAH-1 mRNA expression was measured by means of RT-PCR. Comparisons between experimental groups were performed using the Mann Whitney test.


RESULTS
ADMA levels and nNOS expression increased in the hypothalamus and medulla following GTN administration. Conversely, a significant decrease in DDAH-1 mRNA expression was observed in the same areas. By contrast, no significant change was reported in eNOS expression. GTN administration did not induce any significant change in serum levels of ADMA.


CONCLUSIONS
The present data suggest that ADMA accumulates in the brain after GTN administration via the inhibition of DDAH-1. This latter may represent a compensatory response to the excessive local availability of NO, released directly by GTN or synthetized by nNOS. These findings prompt an additional mediator (ADMA) in the modulation of NO axis following GTN administration and offer new insights in the pathophysiology of migraine.
Laboratory of Neurophysiology of Integrative Autonomic Systems, Headache Science Centre, "C. Mondino" National Neurological Institute, Pavia, Italy, rosaria.greco@mondino.it.
Rosaria
R
Greco
Andrea
A
Ferrigno
Chiara
C
Demartini
Annamaria
A
Zanaboni
Antonina Stefania
AS
Mangione
Fabio
F
Blandini
Giuseppe
G
Nappi
Mariapia
M
Vairetti
Cristina
C
Tassorelli
D000581:Amidohydrolases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000818:Animals; D001120:Arginine / Q000031:analogs & derivatives* / Q000378:metabolism; D001921:Brain / Q000187:drug effects / Q000378:metabolism*; D004195:Disease Models, Animal; D004353:Drug Evaluation, Preclinical / Q000379:methods; D008297:Male; D008881:Migraine Disorders / Q000378:metabolism*; D052248:Nitric Oxide Synthase Type I / Q000037:antagonists & inhibitors / Q000378:metabolism*; D005996:Nitroglycerin / Q000494:pharmacology*; D051381:Rats; D017207:Rats, Sprague-Dawley
C018524:N,N-dimethylarginine; D001120:Arginine; D052248:Nitric Oxide Synthase Type I; D000581:Amidohydrolases; C059999:dimethylargininase; D005996:Nitroglycerin