Ascorbate Inhibits Reduced Arteriolar Conducted Vasoconstriction in Septic Mouse Cremaster Muscle

Abstract

Objective: The mechanism of neuronal nitric oxide synthase (nNOS)-dependent reduction in arteriolar conducted vasoconstriction in sepsis, and the possible protection by antioxidants, are unknown. The authors hypothesized that ascorbate inhibits the conduction deficit by reducing nNOS-derived NO production.

Methods: Using intravital microscopy and the cecal ligation and perforation (CLP) model of sepsis (24 h), arterioles in the cremaster muscle of male C57BL/6 wild-type mice were locally stimulated with KCl to initiate conducted vasoconstriction. The authors used the ratio of conducted constriction (500 μ m upstream) to local constriction as an index of conduction (CR₅₀₀). Cremaster muscle NOS enzymatic activity and protein expression, and plasma nitrite/nitrate levels were determined in control and septic mice. Intravenous ascorbate bolus (200 mg/kg in 0.1 ml of saline) was given early (0 h) or delayed at 23 h post CLP.

Results: Sepsis reduced CR₅₀₀ from 0.73 ± 0.03 to 0.21 ± 0.03, increased nNOS activity from 87 ± 9 to 220 ± 29 pmol/mg/h and nitrite/nitrate from 16 ± 1 to 39 ± 3 μ M, without affecting nNOS protein expression. Ascorbate at 0 and 23 h prevented/reversed the conduction deficit and the increases in nNOS activity and nitrite/nitrate level. NO donor SNAP (S-nitroso-N-acetylpenicillamine) reestablished the conduction deficit in ascorbate-treated septic mice. Superoxide scavenger MnTBAP (Mn(III)tetrakis(4-benzoic acid)porphyrin chloride) did not affect this deficit. Conclusion: These data indicate that early and delayed intravenous boluses of ascorbate prevent/reverse sepsis-induced deficit in arteriolar conducted vasoconstriction in the cremaster muscle by inhibiting nNOS-derived NO production.

Key words:

• arteriolar conducted response
• ascorbate
• nitric oxide
• sepsis

The authors thank Drs. S. J. Dixon, S. Metha, and Y. Ouellette for discussion of this research project; Ms. F. Li and Mr. M. Bolon for technical assistance; Heart and Stroke Foundation of Ontario (research grant HSFO NA 5941 to K.T.); Canadian Institutes of Health Research (salary award to D.L. and research grant CIHR 14794 to K.T.); Natural Sciences and Engineering Research Council of Canada; and Ontario Graduate Scholarship in Science and Technology (salary awards to R.L.M.).