title: Neuroprotective FK506 does not alter in vivo nitric oxide production during ischemia and early reperfusion in rats.
abstract: OBJECTIVE
  Previous studies have demonstrated that the immunosuppressant FK506 provides neuroprotection in experimental brain injury and suggest that this action may be mediated by suppression of neuronal nitric oxide synthase activation that occurs after ischemic depolarization. We sought to determine whether FK506 reduces histological injury after middle cerebral artery occlusion (MCAO) in the rat and whether the neuroprotective effect is mediated via suppression of in vivo nitric oxide (NO) production during ischemia or early reperfusion.
  
  
  METHODS
  Under controlled conditions of normoxia, normocarbia, and normothermia, halothane-anesthetized male Wistar rats were subjected to 2 hours of MCAO by the intraluminal occlusion technique in a blinded, randomized experimental trial. Ipsilateral parietal cortical laser-Doppler flowmetry was monitored throughout ischemia. Animals were randomly assigned to 4 pretreatment groups: intravenous FK506 0.3 mg/kg or 1. 0 mg/kg, vehicle (cremaphor), or an equivalent volume of saline administered 30 minutes before MCAO. Infarction volume was assessed by a triphenyltetrazolium chloride staining at 22 hours of reperfusion. In separate experiments, microdialysis probes were placed bilaterally into the striatum. Rats were perfused with artificial cerebrospinal fluid containing 3 micromol/L [14C]- L-arginine for 3 hours and then subjected to 2 hours of right MCAO. Intravenous 0.3 mg/kg FK506 or cremaphor was given 30 minutes before right MCAO. Right-left differences between [14C]-L-citrulline in the effluent were assumed to reflect differences in NO production.
  
  
  RESULTS
  All values are mean+/-SE. FK506 at 0.3 mg/kg reduced infarction volume in cortex: 40+/-12 mm3 compared with saline (109+/-15 mm3) and cremaphor vehicle (148+/-23) (P<0.05). Striatal infarction was also reduced by low-dose FK506: 16+/-4 mm3 versus 36+/-4 mm3 and 34+/-4 mm3 in saline and vehicle groups, respectively (P<0.05). High-dose treatment reduced infarction volume in cortex (61+/-14 mm3, P<0.05 from saline and vehicle groups) and in striatum (22+/-5 mm3, P<0.05 from saline and vehicle groups). [14C]-L-citrulline recovery via microdialysis was markedly enhanced in ischemic compared with nonischemic striatum. However, ischemia-evoked [14C]-L-citrulline recovery was not different in FK506-treated rats compared with vehicle-treated animals.
  
  
  CONCLUSIONS
  These data demonstrate that FK506 provides robust neuroprotection against transient focal cerebral ischemia in the rat. The mechanism of protection in vivo is not through attenuation of ischemia-evoked NO production during MCAO and early reperfusion.
authors.affiliation: Departments of Anesthesiology/Critical Care Medicine, Neurology, and Neuroscience, Johns Hopkins University School of Medicine, Baltimore, MD, USA.
authors.forename: T J
authors.initials: TJ
authors.lastname: Toung
authors.forename: A
authors.initials: A
authors.lastname: Bhardwaj
authors.forename: V L
authors.initials: VL
authors.lastname: Dawson
authors.forename: T M
authors.initials: TM
authors.lastname: Dawson
authors.forename: R J
authors.initials: RJ
authors.lastname: Traystman
authors.forename: P D
authors.initials: PD
authors.lastname: Hurn
mesh_terms: D000818:Animals; D002544:Cerebral Infarction / Q000473:pathology; D003342:Corpus Striatum / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D002546:Ischemic Attack, Transient / Q000378:metabolism*; D008297:Male; D018696:Neuroprotective Agents / Q000494:pharmacology*; D009569:Nitric Oxide / Q000096:biosynthesis*; D019001:Nitric Oxide Synthase / Q000378:metabolism; D052248:Nitric Oxide Synthase Type I; D051381:Rats; D017208:Rats, Wistar; D015427:Reperfusion Injury / Q000378:metabolism*; D016559:Tacrolimus / Q000494:pharmacology*; D013997:Time Factors
chemical_list: D018696:Neuroprotective Agents; D009569:Nitric Oxide; D019001:Nitric Oxide Synthase; D052248:Nitric Oxide Synthase Type I; C496322:Nos1 protein, rat; D016559:Tacrolimus