Nitric oxide modulates both pre-sinusoidal and sinusoidal responses to phenylephrine in normal rat liver

Abstract

Methods• Animals were studied 4 weeks after sham surgery or bile duct ligation (BDL). NOS activity was assessed by measuring L-NAME inhibited L-citrulline generation from liver lysates in the presence of a standard concentration ot calmodutin {0.1 /~M) or alternatively lO-fold excess in calmodulin (1.0 /~M). eNOS/iNOS protein levels were assessed by quantitative immunoprecipitation/Western blot analysis. Results, NOS activity was significantly reduced in liver lysates from BDL rat livers in the presence of 0.1 /~M calmodulin, the concentration routinely used in this assay [see Figure (white bars); *P<O.05; sham vs 6DL at 0.1 /~M calmodulin; n = 6]. Western blot analysis demonstrated no reduction in eNOS protein levels or iNOS protein expression in BDL liver lysates. Importantly, supplementation of lO-fold excess calmodulin (1.0/~M) did not affect NOS activity from sham animals, but significantly increased NOS catalytic activity in BDL lysates, thereby correcting deficient hepatic NOS activity (see Figure, **P<O.O5; 0.1 /LM VS. 1.0 ~M calmodulin in BDL; n=6). Conclusion. Provision of excess calmodulin corrects deficient NOS catalytic activity detected from BDL liver lysates in an experimental cell-free assay.