Subcellular biochemical investigation of purkinje neurons using synchrotron radiation fourier transform infrared spectroscopic imaging with a focal plane array detector
Coupling Fourier transform infrared spectroscopy with focal plane array detectors at synchrotron radiation sources (SR-FTIR-FPA) has provided a rapid method to simultaneously image numerous biochemical markers in situ at diffraction limited resolution. Since cells and nuclei are well resolved at thi...
| Main Authors: | , , , , , , , , |
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| Format: | Journal Article |
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American Chemical Society
2013
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| Online Access: | http://hdl.handle.net/20.500.11937/30173 |
| _version_ | 1848753012830371840 |
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| author | Hackett, Mark Borondics, F. Brown, D. Hirschmugl, C. Smith, S. Paterson, P. Nichol, H. Pickering, I. George, G. |
| author_facet | Hackett, Mark Borondics, F. Brown, D. Hirschmugl, C. Smith, S. Paterson, P. Nichol, H. Pickering, I. George, G. |
| author_sort | Hackett, Mark |
| building | Curtin Institutional Repository |
| collection | Online Access |
| description | Coupling Fourier transform infrared spectroscopy with focal plane array detectors at synchrotron radiation sources (SR-FTIR-FPA) has provided a rapid method to simultaneously image numerous biochemical markers in situ at diffraction limited resolution. Since cells and nuclei are well resolved at this spatial resolution, a direct comparison can be made between FTIR functional group images and the histology of the same section. To allow histological analysis of the same section analyzed with infrared imaging, unfixed air-dried tissue sections are typically fixed (after infrared spectroscopic analysis is completed) via immersion fixation. This post fixation process is essential to allow histological staining of the tissue section. Although immersion fixation is a common practice in this filed, the initial rehydration of the dehydrated unfixed tissue can result in distortion of subcellular morphology and confound correlation between infrared images and histology. In this study, vapor fixation, a common choice in other research fields where postfixation of unfixed tissue sections is required, was employed in place of immersion fixation post spectroscopic analysis. This method provided more accurate histology with reduced distortions as the dehydrated tissue section is fixed in vapor rather than during rehydration in an aqueous fixation medium. With this approach, accurate correlation between infrared images and histology of the same section revealed that Purkinje neurons in the cerebellum are rich in cytosolic proteins and not depleted as once thought. In addition, we provide the first direct evidence of intracellular lactate within Purkinje neurons. This highlights the significant potential for future applications of SR-FTIR-FPA imaging to investigate cellular lactate under conditions of altered metabolic demand such as increased brain activity and hypoxia or ischemia. © 2013 American Chemical Society. |
| first_indexed | 2025-11-14T08:17:45Z |
| format | Journal Article |
| id | curtin-20.500.11937-30173 |
| institution | Curtin University Malaysia |
| institution_category | Local University |
| last_indexed | 2025-11-14T08:17:45Z |
| publishDate | 2013 |
| publisher | American Chemical Society |
| recordtype | eprints |
| repository_type | Digital Repository |
| spelling | curtin-20.500.11937-301732018-03-29T09:08:49Z Subcellular biochemical investigation of purkinje neurons using synchrotron radiation fourier transform infrared spectroscopic imaging with a focal plane array detector Hackett, Mark Borondics, F. Brown, D. Hirschmugl, C. Smith, S. Paterson, P. Nichol, H. Pickering, I. George, G. Coupling Fourier transform infrared spectroscopy with focal plane array detectors at synchrotron radiation sources (SR-FTIR-FPA) has provided a rapid method to simultaneously image numerous biochemical markers in situ at diffraction limited resolution. Since cells and nuclei are well resolved at this spatial resolution, a direct comparison can be made between FTIR functional group images and the histology of the same section. To allow histological analysis of the same section analyzed with infrared imaging, unfixed air-dried tissue sections are typically fixed (after infrared spectroscopic analysis is completed) via immersion fixation. This post fixation process is essential to allow histological staining of the tissue section. Although immersion fixation is a common practice in this filed, the initial rehydration of the dehydrated unfixed tissue can result in distortion of subcellular morphology and confound correlation between infrared images and histology. In this study, vapor fixation, a common choice in other research fields where postfixation of unfixed tissue sections is required, was employed in place of immersion fixation post spectroscopic analysis. This method provided more accurate histology with reduced distortions as the dehydrated tissue section is fixed in vapor rather than during rehydration in an aqueous fixation medium. With this approach, accurate correlation between infrared images and histology of the same section revealed that Purkinje neurons in the cerebellum are rich in cytosolic proteins and not depleted as once thought. In addition, we provide the first direct evidence of intracellular lactate within Purkinje neurons. This highlights the significant potential for future applications of SR-FTIR-FPA imaging to investigate cellular lactate under conditions of altered metabolic demand such as increased brain activity and hypoxia or ischemia. © 2013 American Chemical Society. 2013 Journal Article http://hdl.handle.net/20.500.11937/30173 10.1021/cn4000346 American Chemical Society restricted |
| spellingShingle | Hackett, Mark Borondics, F. Brown, D. Hirschmugl, C. Smith, S. Paterson, P. Nichol, H. Pickering, I. George, G. Subcellular biochemical investigation of purkinje neurons using synchrotron radiation fourier transform infrared spectroscopic imaging with a focal plane array detector |
| title | Subcellular biochemical investigation of purkinje neurons using synchrotron radiation fourier transform infrared spectroscopic imaging with a focal plane array detector |
| title_full | Subcellular biochemical investigation of purkinje neurons using synchrotron radiation fourier transform infrared spectroscopic imaging with a focal plane array detector |
| title_fullStr | Subcellular biochemical investigation of purkinje neurons using synchrotron radiation fourier transform infrared spectroscopic imaging with a focal plane array detector |
| title_full_unstemmed | Subcellular biochemical investigation of purkinje neurons using synchrotron radiation fourier transform infrared spectroscopic imaging with a focal plane array detector |
| title_short | Subcellular biochemical investigation of purkinje neurons using synchrotron radiation fourier transform infrared spectroscopic imaging with a focal plane array detector |
| title_sort | subcellular biochemical investigation of purkinje neurons using synchrotron radiation fourier transform infrared spectroscopic imaging with a focal plane array detector |
| url | http://hdl.handle.net/20.500.11937/30173 |