AM-36 modulates the neutrophil inflammatory response and reduces breakdown of the blood brain barrier after endothelin-1 induced focal brain ischaemia.
OBJECTIVE
Following transient focal stroke, rapid accumulation and activation of neutrophils in the ischaemic region is deleterious due to release of reactive oxygen species and myeloperoxidase (MPO). The purpose of this study was to examine whether AM-36, both a Na+ channel blocker and an antioxidant, afforded neuroprotection by modulating neutrophil accumulation into brain, following endothelin-1 (ET-1) induced middle cerebral artery occlusion (MCAo) in conscious rats.


METHODS
AM-36 was administered at 3 and 24 h after ET-1-induced MCAo. Functional recovery was determined using grid-walking and cylinder tests. Image analysis of brain sections was used to determine infarct volume. The effect of AM-36 on neutrophil infiltration and their interaction with macrophages was examined in rats at 48 h following MCAo by both an MPO assay and double-label immunofluorescence. Blood brain barrier (BBB) breakdown was measured by the area stained by intravenous Evans Blue.


RESULTS
AM-36 reduced functional deficits in both tests such that no difference existed from pre-ischaemic values at 48 h. Neutrophil infiltration, assessed by MPO activity, and infarct volume were significantly reduced following AM-36 administration by 54 and 60% respectively. Similarly, immunofluorescence revealed that AM-36 reduced neutrophil infiltration by approximately 50% in selected brain regions, when compared to controls, and also modulated macrophage phagocytosis of neutrophils. Breakdown of the BBB was significantly reduced by 60% following AM-36 treatment.


CONCLUSIONS
These findings suggest that AM-36 can directly modulate the neutrophil inflammatory response and reduce BBB breakdown following MCAo.
Weston
R M
RM
Department of Pharmacology, Monash University, Clayton, Australia.
Jarrott
B
B
Ishizuka
Y
Y
Callaway
J K
JK
D000818:Animals; D001812:Blood-Brain Barrier / Q000187:drug effects*; D002545:Brain Ischemia / Q000139:chemically induced / Q000473:pathology / Q000503:physiopathology*; D019332:Endothelin-1 / Q000494:pharmacology*; D005455:Fluorescent Antibody Technique; D007249:Inflammation / Q000517:prevention & control*; D008297:Male; D018613:Microscopy, Confocal; D009504:Neutrophils / Q000187:drug effects*; D009195:Peroxidase / Q000378:metabolism; D010879:Piperazines / Q000494:pharmacology*; D051381:Rats; D020318:Rats, Long-Evans
D019332:Endothelin-1; D010879:Piperazines; C401121:AM 36; D009195:Peroxidase