Determination of carbon monoxide (CO) in rodent tissue: Effect of heme administration and environmental CO exposure

Carbon monoxide (CO), produced endogenously during heme degradation, is considered a messenger molecule in vascular and neurologic tissues. To study this role, it is important to determine CO concentration in target tissues pre- and post-perturbations. Here, we describe a sensitive and reproducible method, which is linear and accurate, and provide some examples of its application for quantitation of CO concentrations in tissues pre- and post-perturbations. Tissues from adult rats and mice were sonicated (20% w/w), and volumes representing 0.04-8 mg fresh weight (FW) were incubated at 0 °C for ⩾30 min with sulfosalicylic acid. CO liberated into the headspace was quantitated by gas chromatography. Tissue CO concentrations (mean ± SD, pmol CO/mg FW) were as follows: blood (47 ± 10, 45 ± 5), muscle (4 ± 4, 10 ± 1), kidney (5 ± 2, 7 ± 2), heart (6 ± 3, 6 ± 1), spleen (11 ± 3, 6 ± 1), liver (4 ± 1, 5 ± 1), intestine (2 ± 1, 4 ± 2), lung (2 ± 1, 3 ± 1), testes (1 ± 1, 2 ± 1), and brain (2 ± 1, 2 ± 0) in untreated rat (n = 3) and mouse (n = 5), respectively. Between the rat and the mouse, only CO concentrations in the muscle and spleen were significantly different (p ⩽ 0.05). Endogenous CO generation, after administration of heme arginate to mice (n = 3), increased CO concentrations by 0-43 pmol/mg FW. Exposure of mice (n = 3) to 500 ppm CO for 30 min yielded significantly elevated CO concentrations by 4-2603 pmol/mg FW in all tissues over the native state. While blood had the highest CO concentration for all conditions, muscle, kidney, heart, spleen, and liver, all rich in hemoglobin and/or other CO-binding hemoproteins, also contained substantial CO concentrations. Intestine, lung, testes, and brain contained the lowest CO concentrations.