, Volume 63, Issue 6, pp 959-966
Date: 04 Feb 2009

Nitric oxide production: an easily measurable condition index for vertebrates

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Nitric oxide (NO) is a multifunctional signalling molecule, acting as a vasodilator, neurotransmitter, and modulator of inflammatory processes. It also participates in killing parasites, virus-infected cells, and tumor cells by formation of peroxynitrite, one of the most important initiators of the free radical damage. Uncontrolled production of NO can lead to nitrosative stress, causing damages to proteins and DNA and cell injury and death. Determination of NO production in animals is potentially informative for description of individual variation in physiological condition, health state, and work load; however, it’s potential in ecophysiological research has remained almost totally unexplored. Here, we describe application of a simple, precise, and inexpensive spectrophotometric assay for determination of NO production from 5 to 10 μL plasma samples of passerine birds. The method is based on estimation of concentrations of nitrate and nitrite—the stable end products of nitric oxide oxidation. The principle of the assay is reduction of nitrate to nitrite by copper-coated cadmium granules, followed by color development with Griess reagent. NO production in captive greenfinches (Carduelis chloris L.) was significantly repeatable over 6-day period (r = 0.35). Injection of an inflammatory agent phytohemagglutinin into wing web resulted in 21% higher levels of NO production at the third day after treatment as compared to saline-injected birds. These findings, consistently with further evidence from veterinary and biomedical literature, suggest that measuring NO production appears an efficient and robust tool for monitoring individual condition and assessment of the magnitude of innate immune response, pathogenicity of infections, and physical effort. We propose that this assay, which is easily applicable in field studies, has an excellent potential in ecophysiological research, particularly in the rapidly developing fields of immunoecology and conservation physiology.

Communicated by J. Graves