A bioluminescence method for the mapping of local ATP concentrations within the arterial wall, with potential to assess the in vivo situation

Abstract

According to the anoxemia theory of atherosclerosis, an imbalance between the demand for and supply of oxygen and nutrients in the arterial wall is a key factor in atherogenesis. However, the energy metabolic state of the arterial tissue in vivo is largely unknown. We applied a bioluminescence method, metabolic imaging, to study local ATP concentrations in cryosections of normal pig and atherosclerotic and normal rabbit aorta. Some vessels were subjected to energy metabolic restrictions by incubation at different oxygen and glucose concentrations and others were rapidly frozen in liquid nitrogen to reflect the in vivo situation. Local ATP concentrations and the ATP distribution at a microscale was dependent on oxygen as well as glucose concentrations during incubation. ATP depletion was seen in the mid media of pig aorta in all incubations, but only at low oxygen concentration without glucose in the media of the thinner rabbit aorta. ATP-depleted zones were seen deep in pig media (>750 μm from the lumen) and in rabbit plaques (>300 μm from the lumen) even at high oxygen (pig 75% O2 and rabbit 21% O2) and glucose concentrations (5.6 mmol/L glucose). This observation probably illustrates an insufficient diffusion of glucose, which highlights the importance of studying the conditions for diffusion not only of oxygen but also of other metabolites in the arterial wall. In rapidly frozen vessels the medial ATP concentration was shown to be 0.6 to 0.8 μmol/g wet weight (both pig and rabbit aorta) and in pig aorta a gradient could be seen indicating higher ATP concentrations at the lumenal side. We propose that metabolic imaging, as applied to snap-frozen tissue, may be used to assess the energy metabolic situation in the arterial wall in vivo. The spatial resolution allows the detection of local variations within the arterial tree. However, steep concentration gradients (eg, near the border of the tissue) will be underestimated. The method may be extended to include determinations of glucose and lactate concentrations and will be used in parallel with an established method to assess hypoxia in the arterial wall in vivo.