Perivascular nitric oxide and superoxide in neonatal cerebral hypoxia-ischemia.
Decreased cerebral blood flow (CBF) has been observed following the resuscitation from neonatal hypoxic-ischemic injury, but its mechanism is not known. We address the hypothesis that reduced CBF is due to a change in nitric oxide (NO) and superoxide anion O(2)(-) balance secondary to endothelial NO synthase (eNOS) uncoupling with vascular injury. Wistar rats (7 day old) were subjected to cerebral hypoxia-ischemia by unilateral carotid occlusion under isoflurane anesthesia followed by hypoxia with hyperoxic or normoxic resuscitation. Expired CO(2) was determined during the period of hyperoxic or normoxic resuscitation. Laser-Doppler flowmetry was used with isoflurane anesthesia to monitor CBF, and cerebral perivascular NO and O(2)(-) were determined using fluorescent dyes with fluorescence microscopy. The effect of tetrahydrobiopterin supplementation on each of these measurements and the effect of apocynin and N(omega)-nitro-L-arginine methyl ester (L-NAME) administration on NO and O(2)(-) were determined. As a result, CBF in the ischemic cortex declined following the onset of resuscitation with 100% O(2) (hyperoxic resuscitation) but not room air (normoxic resuscitation). Expired CO(2) was decreased at the onset of resuscitation, but recovery was the same in normoxic and hyperoxic resuscitated groups. Perivascular NO-induced fluorescence intensity declined, and O(2)(-)-induced fluorescence increased in the ischemic cortex after hyperoxic resuscitation up to 24 h postischemia. L-NAME treatment reduced O(2)(-) relative to the nonischemic cortex. Apocynin treatment increased NO and reduced O(2)(-) relative to the nonischemic cortex. The administration of tetrahydrobiopterin following the injury increased perivascular NO, reduced perivascular O(2)(-), and increased CBF during hyperoxic resuscitation. These results demonstrate that reduced CBF follows hyperoxic resuscitation but not normoxic resuscitation after neonatal hypoxic-ischemic injury, accompanied by a reduction in perivascular production of NO and an increase in O(2)(-). The finding that tetrahydrobiopterin, apocynin, and L-NAME normalized radical production suggests that the uncoupling of perivascular NOS, probably eNOS, due to acquired relative tetrahydrobiopterin deficiency occurs after neonatal hypoxic-ischemic brain injury. It appears that both NOS uncoupling and the activation of NADPH oxidase participate in the changes of reactive oxygen concentrations seen in cerebral hypoxic-ischemic injury.
Fabian
Roderic H
RH
Department of Neurology, Baylor College of Medicine and the Michael E. DeBakey Veterans Affairs Medical Center Stroke Program, Houston 77030, USA. rfabian@bcm.tmc.edu
Perez-Polo
J Regino
JR
Kent
Thomas A
TA
D000098:Acetophenones / Q000494:pharmacology; D000818:Animals; D000831:Animals, Newborn; D001708:Biopterins / Q000031:analogs & derivatives / Q000494:pharmacology; D002245:Carbon Dioxide / Q000378:metabolism; D002540:Cerebral Cortex / Q000098:blood supply*; D002560:Cerebrovascular Circulation / Q000187:drug effects; D004195:Disease Models, Animal; D004730:Endothelium, Vascular / Q000187:drug effects / Q000378:metabolism* / Q000503:physiopathology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D045853:Exhalation; D020925:Hypoxia-Ischemia, Brain / Q000378:metabolism* / Q000473:pathology / Q000503:physiopathology / Q000628:therapy; D017078:Laser-Doppler Flowmetry; D008856:Microscopy, Fluorescence; D019255:NADPH Oxidases / Q000037:antagonists & inhibitors / Q000378:metabolism; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D009569:Nitric Oxide / Q000378:metabolism*; D052247:Nitric Oxide Synthase Type II / Q000037:antagonists & inhibitors / Q000378:metabolism; D052250:Nitric Oxide Synthase Type III / Q000037:antagonists & inhibitors / Q000378:metabolism; D010102:Oxygen Inhalation Therapy / Q000009:adverse effects; D051381:Rats; D017208:Rats, Wistar; D012151:Resuscitation / Q000009:adverse effects; D013481:Superoxides / Q000378:metabolism*; D013997:Time Factors
D000098:Acetophenones; D004791:Enzyme Inhibitors; D013481:Superoxides; D002245:Carbon Dioxide; D001708:Biopterins; D009569:Nitric Oxide; C056165:acetovanillone; D052247:Nitric Oxide Synthase Type II; D052250:Nitric Oxide Synthase Type III; C496316:Nos3 protein, rat; D019255:NADPH Oxidases; C003402:sapropterin; D019331:NG-Nitroarginine Methyl Ester