Determination of carbon monoxide (CO) in rodent tissue: Effect of heme administration and environmental CO exposure
ABSTRACT 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 degrees 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 (p0.05). 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.
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ABSTRACT: Background: Metabolic syndrome is a collection of ailments resulting in a higher risk of cardiovascular disease and type II diabetes mellitus. It also results in prolonged endothelial dysfunction which promotes hypertension. Objective: The current study examines the acute effect of carbon monoxide (CO) inhibition and nitric oxide (NO) stimulation in septal coronary arteries. Methods: These studies were conducted in inactin anesthetized obese and lean Zucker rats (13-14 weeks of age). Coronary arteries were isolated from obese and lean Zucker rats and in vitro experiments were conducted. Isolated coronary arteries were pre-treated with chromium mesoporphyrin (CrMP) which is a heme oxygenase inhibitor and L-arginine, a NO precursor. Results: Blood pressure, non-fasting blood glucose, HBCO, CO levels and Arginase I expression were higher in obese Zucker rats (ZR) as compared to the lean (L) group. Obese ZR had higher body, kidney and heart weights as compared to the LZR. Acetylcholine induced vasodilation was greatly attenuated in Obese ZR compared to the lean group. No differences in the diameters of the septal coronary artery were observed in both groups when treated with CrMP. However, pretreatment with L-arginine, abolished the differences between the groups. Conclusion: This study demonstrates the potential of NO induction to improve coronary blood flow during metabolic syndrome induced endothelial dysfunction, where alterations in CO levels appeared to have no significant coronary effects.03/2014; 2(1):8. DOI:10.14419/ijm.v2i1.1941
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ABSTRACT: It has been recognized that gaseous molecules and their signaling cascades play a vital role in alterations of metabolic systems in physiologic and pathologic conditions. Contrary to this awareness, detailed mechanisms whereby gases exert their actions, in particular in vivo, have been unclear because of several reasons. Gaseous signaling involves diverse reactions with metal centers of metalloproteins and thiol modification of cysteine residues of proteins. Both the multiplicity of gas targets and the technical limitations in accessing local gas concentrations make dissection of exact actions of any gas mediator a challenge. However, a series of advanced technologies now offer ways to explore gas-responsive regulatory processes in vivo. Imaging mass spectrometry combined with quantitative metabolomics by capillary-electrophoresis/mass spectrometry reveals spatio-temporal profiles of many metabolites. Comparing the metabolic footprinting of murine samples with a targeted deletion of a specific gas-producing enzyme makes it possible to determine sites of actions of the gas. In this review, we intend to elaborate on the ideas how small gaseous molecules interact with metabolic systems to control organ functions such as cerebral vascular tone and energy metabolism in vivo.Respiratory Physiology & Neurobiology 03/2012; 184(2). DOI:10.1016/j.resp.2012.03.016 · 1.97 Impact Factor
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ABSTRACT: Purpose. Studies were performed to examine hemodynamic and renal function before and after acute induction of the endogenous CO system with delta-aminolevulinic acid (DALA), which drives HO activity. Methods. In vivo studies were conducted on Inactin-anesthetized male Sprague Dawley rats (250-300 g) either with or without chronic pretreatment with L-NAME (50 mg/Kg, q12 hours x4d). Results. DALA (80 μmol/Kg, IV bolus) administration acutely increased endogenous CO production and HO-1 protein. In untreated and L-NAME-pretreated rats, DALA did not alter BP, GFR, or RBF but increased UF, U(Na)V, and U(K)V (untreated: Δ108.8 ± 0.28%, 172.1 ± 18.4%, and 165.2 ± 45.9%; pretreated: Δ109.4 ± 0.29%, 187.3 ± 26.9%, and 197.2 ± 45.7%). Acute administration of biliverdin (20 mg/kg, IV) and bilirubin (30 mg/kg, IV) to similarly treated animals did not alter UF, U(Na)V, and U(K)V. Conclusion. These results demonstrate that heme oxygenase induction increases urine and electrolyte excretion and suggest a direct tubular action of endogenous carbon monoxide.01/2012; 2012(2090-0384):690973. DOI:10.1155/2012/690973