In vivo imaging of cerebral energy metabolism with two-photon fluorescence lifetime microscopy of NADH
Spotlight summary: Imaging of endogenous tissue chromophores has the potential to link the biochemical and morphological properties of tissue aiding in understanding of normal and diseased tissue processes. There are a number of endogenous tissue chromophores including amino acids, structural proteins, porphyrins, lipids, and enzymes. Changes in these fluorescent tissue chromophores as well as changes in hemoglobin and water can be used to interpret tissue metabolism, proliferation, differentiation, and disease states. Of particular interest over the past 50 years has been the relationship between the endogenous fluorescence from the reduced form of the enzyme nicotinamide adenine dinucleotide (NADH) and energy metabolism. In metabolism NAD+/NADH is involved in oxidation-reduction (redox) reactions where NADH is the ubiquitous electron carrier that plays a crucial roll in both glycolysis and oxidative metabolism. However detection in vivo can prove technically challenging as its peak absorption and emission lie in the ultraviolet region of the electromagnetic spectrum, where photon penetration is low, tissue light scattering is high, and emitted NADH signal is convolved with other tissue chromophores.
The ability to monitor NADH in vivo remained limited until the development of two-photon microscopy techniques, which utilize the two-photon cross section of chromophores to enable excitation at double the wavelength increasing tissue penetration depth. Additionally, methods to couple two-photon microscopy with time resolved fluorescence lifetime imaging microscopy (FLIM) have been developed and used to probe the microenvironment of tissue chromophores. In the current work by Yaseen et al the first use of two-photon FLIM to detect NADH fluorescence in cerebral tissues in vivo is demonstrated. The authors demonstrated detection of four species of NADH using two-photon FLIM, which define the different microenvironments within the tissue where NADH resides. A multi-component fitting algorithm is developed and utilized to model the measured NADH in vivo fluorescence lifetime. The microenvironmental difference between the components was tested by generating brief periods of anoxia, where the four NADH fluorescence components were found to respond differently likely indicating different enzymatic formulations. Thus, the ability to measure individual components of the NADH fluorescence lifetime may enable detection of the specific molecular pathways involved in oxidative metabolism in future studies.
ToC Category: Functional Imaging
|OCIS Codes:||(170.0170) Medical optics and biotechnology : Medical optics and biotechnology|
|(170.3650) Medical optics and biotechnology : Lifetime-based sensing|
|(180.4315) Microscopy : Nonlinear microscopy|
03/14/2013 11:55 AM posted by zhao H.
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