INDUCTION BY ENDOTOXIN OF NITRIC-OXIDE SYNTHASE IN THE RAT MESENTERY - LACK OF EFFECT ON ACTION OF VASOCONSTRICTORS

1 Male Sprague-Dawley or Wistar rats were injected with bacterial lipo polysacharride (LPS; 5 mg kg-1 i.p) and killed after 1, 3, 6, 15, and 24 h. The brains, mesenteries, spleens, lungs, livers, kidneys, hearts , aortae and diaphragms were removed and frozen immediately. Control r ats were injected with sterile saline and killed after 6 h. 2 The orga ns were homogenized in a semi-frozen state and NO synthase (NOS) activ ity measured in tissues from both LPS-treated and saline-treated group s by the ability of homogenates to convert [H-3]-L-arginine to [H-3]-L -citrulline in a NADPH-dependent manner. 3 The NOS activity in all org ans taken from control animals was found to be calcium-dependent, with the highest activity being in the brain. After LPS-treatment an induc ed calcium-independent NOS was detected in all tissues tested, with th e exception of the brain. The spleen, lung, mesentery and liver had th e highest amounts of LPS-induced NOS activity. No induction of calcium -dependent NOS was detected. 4 Induction of NOS was maximum 6 h after administration of LPS and had returned to control levels in 24 h. 5 Th e constitutive NOS in brain and mesentery and the LPS-induced activiti es in the spleen, lung, liver and mesentery were inhibited by N(G)-mon omethyl-L-arginine (L-NMMA) or N(G)-nitro-L-arginine methyl ester (L-N AME) according to concentration. The IC50 for L-NAME was 2.5 mum again st the constitutive NOS from brain, and 20-25 mum against the inducibl e NOS. For L-NMMA the IC50 was 20-25 mum against either NOS isoform. 7 The vascular responses to endothelin-1 (ET-1), the thromboxane A2-mim etic 11alpha,9alpha-epoxymethano prostaglandin F2alpha (U46619), pheny lephrine (PE) or 5-hydroxytryptamine (5-HT) were measured in the simul taneously perfused arterial and venous mesenteric vascular beds from b oth control and LPS-treated (6 h) rats. Vasoconstrictor responses to a ll agonists tested were unaffected by LPS treatment. In the presence o f L-NAME (100 mum) vasoconstrictor responses were potentiated in both the arterial and venous portion of the mesenteric beds from both contr ol and LPS-treated rats. The potentiation of responses to U46619 was s ignificantly greater in beds from LPS-treated rats. 8 Injection of LPS i.p. is associated with induction of NOS in all organs tested, except for the brain. In the mesentery this is not accompanied by a hyporesp onsiveness to constrictor agents suggesting an increased sensitivity, particularly to U46619. This may explain the poor perfusion and tissue damage in the splanchnic circulation associated with sepsis.