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Superior SA node myocytes exhibit elevated diastolic ATP and metabolic flux compared with the inferior region. (A) 3D segmented maximum-intensity projection of a whole-mount SA node immunolabeled for <t>CD31</t> (vasculature, red) and cyto-iATP (myocytes, green). The dashed line denotes the boundary between superior and inferior regions. (B) Image-processing workflow illustrating merged maximum-intensity projections, binary segmentation masks, and extraction of grayscale cyto-iATP signals used for quantitative analysis. (C) Mean cyto-iATP fluorescence intensity per myocyte, grouped by region ( N = 5 mice per region), reporting expression levels of the EGFP-tagged cyto-iATP sensor. (D) Live confocal imaging of cyto-iATP signals showing representative line-scan images and corresponding normalized fluorescence traces (F/F 0 ) from superior and inferior regions. (E and F) Summary quantification of cyto-iATP signal mass rate (E) and estimated diastolic [ATP] i (F). P values are shown above comparisons. Large circles denote per-animal means; small circles indicate individual biological replicates. N represents the number of independent mice.
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Superior SA node myocytes exhibit elevated diastolic ATP and metabolic flux compared with the inferior region. (A) 3D segmented maximum-intensity projection of a whole-mount SA node immunolabeled for <t>CD31</t> (vasculature, red) and cyto-iATP (myocytes, green). The dashed line denotes the boundary between superior and inferior regions. (B) Image-processing workflow illustrating merged maximum-intensity projections, binary segmentation masks, and extraction of grayscale cyto-iATP signals used for quantitative analysis. (C) Mean cyto-iATP fluorescence intensity per myocyte, grouped by region ( N = 5 mice per region), reporting expression levels of the EGFP-tagged cyto-iATP sensor. (D) Live confocal imaging of cyto-iATP signals showing representative line-scan images and corresponding normalized fluorescence traces (F/F 0 ) from superior and inferior regions. (E and F) Summary quantification of cyto-iATP signal mass rate (E) and estimated diastolic [ATP] i (F). P values are shown above comparisons. Large circles denote per-animal means; small circles indicate individual biological replicates. N represents the number of independent mice.
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Superior SA node myocytes exhibit elevated diastolic ATP and metabolic flux compared with the inferior region. (A) 3D segmented maximum-intensity projection of a whole-mount SA node immunolabeled for <t>CD31</t> (vasculature, red) and cyto-iATP (myocytes, green). The dashed line denotes the boundary between superior and inferior regions. (B) Image-processing workflow illustrating merged maximum-intensity projections, binary segmentation masks, and extraction of grayscale cyto-iATP signals used for quantitative analysis. (C) Mean cyto-iATP fluorescence intensity per myocyte, grouped by region ( N = 5 mice per region), reporting expression levels of the EGFP-tagged cyto-iATP sensor. (D) Live confocal imaging of cyto-iATP signals showing representative line-scan images and corresponding normalized fluorescence traces (F/F 0 ) from superior and inferior regions. (E and F) Summary quantification of cyto-iATP signal mass rate (E) and estimated diastolic [ATP] i (F). P values are shown above comparisons. Large circles denote per-animal means; small circles indicate individual biological replicates. N represents the number of independent mice.
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Superior SA node myocytes exhibit elevated diastolic ATP and metabolic flux compared with the inferior region. (A) 3D segmented maximum-intensity projection of a whole-mount SA node immunolabeled for <t>CD31</t> (vasculature, red) and cyto-iATP (myocytes, green). The dashed line denotes the boundary between superior and inferior regions. (B) Image-processing workflow illustrating merged maximum-intensity projections, binary segmentation masks, and extraction of grayscale cyto-iATP signals used for quantitative analysis. (C) Mean cyto-iATP fluorescence intensity per myocyte, grouped by region ( N = 5 mice per region), reporting expression levels of the EGFP-tagged cyto-iATP sensor. (D) Live confocal imaging of cyto-iATP signals showing representative line-scan images and corresponding normalized fluorescence traces (F/F 0 ) from superior and inferior regions. (E and F) Summary quantification of cyto-iATP signal mass rate (E) and estimated diastolic [ATP] i (F). P values are shown above comparisons. Large circles denote per-animal means; small circles indicate individual biological replicates. N represents the number of independent mice.
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Superior SA node myocytes exhibit elevated diastolic ATP and metabolic flux compared with the inferior region. (A) 3D segmented maximum-intensity projection of a whole-mount SA node immunolabeled for <t>CD31</t> (vasculature, red) and cyto-iATP (myocytes, green). The dashed line denotes the boundary between superior and inferior regions. (B) Image-processing workflow illustrating merged maximum-intensity projections, binary segmentation masks, and extraction of grayscale cyto-iATP signals used for quantitative analysis. (C) Mean cyto-iATP fluorescence intensity per myocyte, grouped by region ( N = 5 mice per region), reporting expression levels of the EGFP-tagged cyto-iATP sensor. (D) Live confocal imaging of cyto-iATP signals showing representative line-scan images and corresponding normalized fluorescence traces (F/F 0 ) from superior and inferior regions. (E and F) Summary quantification of cyto-iATP signal mass rate (E) and estimated diastolic [ATP] i (F). P values are shown above comparisons. Large circles denote per-animal means; small circles indicate individual biological replicates. N represents the number of independent mice.
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Superior SA node myocytes exhibit elevated diastolic ATP and metabolic flux compared with the inferior region. (A) 3D segmented maximum-intensity projection of a whole-mount SA node immunolabeled for <t>CD31</t> (vasculature, red) and cyto-iATP (myocytes, green). The dashed line denotes the boundary between superior and inferior regions. (B) Image-processing workflow illustrating merged maximum-intensity projections, binary segmentation masks, and extraction of grayscale cyto-iATP signals used for quantitative analysis. (C) Mean cyto-iATP fluorescence intensity per myocyte, grouped by region ( N = 5 mice per region), reporting expression levels of the EGFP-tagged cyto-iATP sensor. (D) Live confocal imaging of cyto-iATP signals showing representative line-scan images and corresponding normalized fluorescence traces (F/F 0 ) from superior and inferior regions. (E and F) Summary quantification of cyto-iATP signal mass rate (E) and estimated diastolic [ATP] i (F). P values are shown above comparisons. Large circles denote per-animal means; small circles indicate individual biological replicates. N represents the number of independent mice.
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Superior SA node myocytes exhibit elevated diastolic ATP and metabolic flux compared with the inferior region. (A) 3D segmented maximum-intensity projection of a whole-mount SA node immunolabeled for CD31 (vasculature, red) and cyto-iATP (myocytes, green). The dashed line denotes the boundary between superior and inferior regions. (B) Image-processing workflow illustrating merged maximum-intensity projections, binary segmentation masks, and extraction of grayscale cyto-iATP signals used for quantitative analysis. (C) Mean cyto-iATP fluorescence intensity per myocyte, grouped by region ( N = 5 mice per region), reporting expression levels of the EGFP-tagged cyto-iATP sensor. (D) Live confocal imaging of cyto-iATP signals showing representative line-scan images and corresponding normalized fluorescence traces (F/F 0 ) from superior and inferior regions. (E and F) Summary quantification of cyto-iATP signal mass rate (E) and estimated diastolic [ATP] i (F). P values are shown above comparisons. Large circles denote per-animal means; small circles indicate individual biological replicates. N represents the number of independent mice.

Journal: The Journal of General Physiology

Article Title: Beat-locked ATP microdomains in the sinoatrial node map a Ca 2+ -timed energetic hierarchy and regional pacemaker roles

doi: 10.1085/jgp.202513874

Figure Lengend Snippet: Superior SA node myocytes exhibit elevated diastolic ATP and metabolic flux compared with the inferior region. (A) 3D segmented maximum-intensity projection of a whole-mount SA node immunolabeled for CD31 (vasculature, red) and cyto-iATP (myocytes, green). The dashed line denotes the boundary between superior and inferior regions. (B) Image-processing workflow illustrating merged maximum-intensity projections, binary segmentation masks, and extraction of grayscale cyto-iATP signals used for quantitative analysis. (C) Mean cyto-iATP fluorescence intensity per myocyte, grouped by region ( N = 5 mice per region), reporting expression levels of the EGFP-tagged cyto-iATP sensor. (D) Live confocal imaging of cyto-iATP signals showing representative line-scan images and corresponding normalized fluorescence traces (F/F 0 ) from superior and inferior regions. (E and F) Summary quantification of cyto-iATP signal mass rate (E) and estimated diastolic [ATP] i (F). P values are shown above comparisons. Large circles denote per-animal means; small circles indicate individual biological replicates. N represents the number of independent mice.

Article Snippet: For immunolabeling, SA nodes were incubated for 48 h at 4°C with a goat anti-mouse CD31 primary antibody (1:50, AF3628; R&D Systems).

Techniques: Immunolabeling, Extraction, Fluorescence, Expressing, Imaging