Abstract
In vivo measurement of oxygen concentration in various tissues is crucial for understanding the adaptation of mitochondrial oxidative metabolism in hypoxia. In these oxygen measurements, a submicron spatial resolution is required because significant heterogeneities in oxygen concentration may be established within a cell. Now, we propose that GFP can be used as an in situ oxygen probe if it is efficiently expressed in cells. In cultured COS7 cell transiently expressing GFP (enhanced GFP), a brief exposure of the cell to 470-490 nm light elicited a red fluorescence (excitation and emission wavelengths, 520-550 nm and >580 nm, respectively) when the cell was exposed to anoxic gas. This red shift was quickly abolished upon reoxygenation. Then, using this technique, we examined whether hypoxia/anoxia can be detected in single cardiomyocytes in a submicron spatial resolution. We used transgenic mice in which GFP was stably expressed (green mouse). All the ventricular myocyte isolated from the green mice showed significant green fluorescence although its intensity was ~1/200 of the transiently GFP expressing COS7 cells. Photoactivation in anoxia certainly produced the red fluorescence in these cells but the magnitude of the increase was much smaller than expected. In summary, GFP can be used as an in situ probe for hypoxia/anoxia. In GFP expressing transgenic animals, in vivo imaging of anoxic loci with a submicron spatial resolution may be possible.
Supported by JSPS KAKENHI 15390061. [Jpn J Physiol 54 Suppl:S108 (2004)]
