nitroarginine and Subarachnoid-Hemorrhage

We designed the present experiments to investigate the involvement of endogenous nitric oxide synthase (NOS) inhibitors, dimethylarginine dimethylaminohydrolase (DDAH) as a hydrolyzing enzyme of the NOS inhibitors, NOS, arginase which shares l-arginine as a common substrate with NOS, and endothelin-1 (ET-1) in the pulmonary dysfunction after induction of experimental subarachnoid hemorrhage (SAH) in the rabbit. SAH was induced by injecting autologous blood into the cisterna magna, and controls were injected with saline. On day 2, pulmonary arteries were isolated for determinations. A significant impairment of the endothelium-dependent relaxation (EDR) caused by acetylcholine was found in 20 cases (43.5%) out of 46 SAH animals, and the same animals exhibited accompanying the significantly impaired cyclic GMP production, accumulated endogenous NOS inhibitors, attenuated DDAH activity, enhanced arginase activity and accumulated ET-1 within the vessel wall. Meanwhile, there were no differences in endothelial NOS activity per se and sodium nitroprusside-induced relaxation between the animals with an impaired EDR and those without such a change. ET-1 content within aortic wall was increased with concomitant decrease in cyclic GMP production after the intraperitoneal application of authentic monomethylarginine as a NOS inhibitor in the rat. The current results suggest that accumulated endogenous NOS inhibitors and enhanced arginase activity possibly bring about the impaired NO production, thereby attenuating the EDR and contributing to the accumulation of ET-1 within the vessel wall. The accumulated endogenous NOS inhibitors at least partly result from the decreased DDAH activity. These alterations may be relevant to the pulmonary dysfunction after induction of SAH.

Topics: Acetylcholine; Amidohydrolases; Animals; Arginase; Cyclic GMP; Cyclooxygenase Inhibitors; Dose-Response Relationship, Drug; Endothelin-1; Indomethacin; Lung Diseases; Male; Models, Biological; Nitric Oxide Donors; Nitric Oxide Synthase; Nitroarginine; Nitroprusside; omega-N-Methylarginine; Oxadiazoles; Pulmonary Artery; Quinoxalines; Rabbits; Rats; Rats, Sprague-Dawley; Subarachnoid Hemorrhage; Vasoconstriction

The mechanisms of hemolysate-induced cerebral injury following subarachnoid hemorrhage are just beginning to be clarified. This study examined the injurious effects of hemolysate on endothelial cells derived from bovine middle cerebral arteries, and evaluated the roles of lipid peroxidation and nitric oxide production in this type of damage. Cultured endothelial cells were grown to confluency on gelatin-coated plates. The cells were characterized as endothelial cells on the basis of morphology. Factor VIII-related antigen staining, and low density lipoprotein (LDL) uptake. Additional cells were grown to confluency on collagen-coated well inserts, and were treated with hemolysate for 24 hours. Prior to hemolysate exposure, cells were treated with: a) an inhibitor of iron-dependent lipid peroxidation (tirilazad mesylate 100 microM), or b) an inhibitor of nitric oxide synthase (either N-nitro-L-arginine: NLA 300 microM, or aminoguanidine: AG at 1.5, 7.5, 15 or 150 microM). Permeability of the tracer, U-14C-sucrose, across the layer of endothelial cells was examined over a 24 hour period. Hemolysate induced a significant increase in the permeability across the endothelial cell layer. Pretreatment with tirilazad mesylate, NLA, or AG attenuated significantly hemolysate-induced changes in the endothelial cell barrier. These findings indicate that free radical generation and lipid peroxidation are critical participants in hemolysate-induced injury to the barrier function of the cerebrovascular endothelium. In addition, the results indicate that endothelial cells provide an adequate source of nitric oxide to damage their own cellular function. Finally, these findings strongly implicate free radical mechanisms in endothelial damage associated with subarachnoid hemorrhage.

Topics: Animals; Antioxidants; Brain; Cattle; Cell Membrane Permeability; Cells, Cultured; Endothelium, Vascular; Enzyme Inhibitors; Free Radicals; Guanidines; Lipid Peroxidation; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Pregnatrienes; Rabbits; Reactive Oxygen Species; Subarachnoid Hemorrhage

We hypothesize that the interaction between protein kinase C (PKC) and nitric oxide (NO) plays a role in the modulation of cerebral vascular tone, and the disturbance of this interaction following subarachnoid hemorrhage (SAH) results in vasospasm. To prove this hypothesis with direct evidence, PKC activities of smooth muscle cells of canine basilar arteries in the control and in the SAH groups were measured by an enzyme immunoassay method. N omega-nitro-L arginine (L-NA), an inhibitor of NO production, enhanced PKC activity. This enhancement was inhibited neither by 8-bromo-guanosine 3',5'-cyclic monophosphate (8-bromo-cGMP) nor SIN-1, a NO releasing agent. PKC activity in the SAH was significantly higher than in the control; however, no further enhancement was produced with L-NA. In the SAH, PKC activity was not inhibited either by 8-bromo-cGMP or SIN-1. We conclude that NO maintains an appropriate vascular tone through inactivation of PKC, and that this effect is disturbed following SAH, resulting in PKC-dependent vascular contraction, such as vasospasm. On the other hand, once PKC has been activated, NO precursors do not inhibit PKC. These facts indicate NO inactivates PKC through the inhibition of phosphatidylinositol breakdown.

Topics: Animals; Basilar Artery; Cyclic GMP; Dogs; Enzyme Activation; Enzyme Inhibitors; Female; Immunoenzyme Techniques; Ischemic Attack, Transient; Male; Molsidomine; Muscle, Smooth, Vascular; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Protein Kinase C; Reference Values; Subarachnoid Hemorrhage