Regional distribution of T-tubule density in left and right atria in dogs

Abstract

Background: The peculiarities of transverse tubule (T-tubule) morphology and distribution in the atrium-and how they contribute to excitation-contraction coupling-are just beginning to be understood.

Objectives: The objectives of this study were to determine T-tubule density in the intact, live right and left atria in a large animal and to determine intraregional differences in T-tubule organization within each atrium.

Methods: Using confocal microscopy, T-tubules were imaged in both atria in intact, Langendorf-perfused normal dog hearts loaded with di-4-ANEPPS. T-tubules were imaged in large populations of myocytes from the endocardial surface of each atrium. Computerized data analysis was performed using a new MatLab (Mathworks, Natick, MA) routine, AutoTT.

Results: There was a large percentage of myocytes that had no T-tubules in both atria with a higher percentage in the right atrium (25.1%) than in the left atrium (12.5%) (P < .02). The density of transverse and longitudinal T-tubule elements was low in cells that did contain T-tubules, but there were no significant differences in density between the left atrial appendage, the pulmonary vein-posterior left atrium, the right atrial appendage, and the right atrial free wall. In contrast, there were significant differences in sarcomere spacing and cell width between different regions of the atria.

Conclusion: There is a sparse T-tubule network in atrial myocytes throughout both dog atria, with significant numbers of myocytes in both atria-the right atrium more so than the left atrium-having no T-tubules at all. These regional differences in T-tubule distribution, along with differences in cell width and sarcomere spacing, may have implications for the emergence of substrate for atrial fibrillation.

Keywords: Atrium; Left atrial appendage; Posterior left atrium; T-tubules.

Figure 1

Examples of T-tubule staining in intact rat left ventricle (A), dog right atrium (B) and in dog right atrium at high magnification (C).

Figure 2

Representative examples of dog right atrial myocytes showing one cell with extensive T-tubules (left panels) and another with no discernible T-tubules (right panels). The top panel shows the cell outline with the cell membrane as presented by AutoTT. The middle panels show the region actually analyzed (magenta) and the cell membrane excluded (green). The bottom panels show the thresholded, skeletonized T-tubules after image processing. The table shows some of the measurements made by AutoTT for these myocytes. T-ED – percent cell volume for transverse tubules; L-ED – same for longitudinal tubules; Spacing measures mean distance between T-tubules; TTi refers to total percentage of cell volume for all tubules.

Figure 3

Staining of T-tubules in an intact rat LV for purposes of comparison to dog atrium in Figure 2. Top panel shows original image and middle panel shows outlined and threshholded image from AutoTT. Table summarizes T-tubule characteristics in this myocyte.

Figure 4

Summary of T-tubule density as percent of total cell volume (left column), sarcomere spacing (middle column) and cell width (right column) in myocytes from LAA, PV-PLA and total in whole left atrium. Graphs show events histograms summarizing results from many cells in all 5 dog atria.

Figure 5

Summary of T-tubule density as percent of total cell volume (left column), sarcomere spacing (middle column) and cell width (right column) in myocytes from RAA, RA free wall and total in whole right atrium. Graphs show events histograms summarizing results from many cells in all dog 5 atria.

Figure 6

Summary of regional differences in T-tubule density (panels A–B) and in right vs. left atrium in T-tubule containing cells only (panel C) and in all myocytes from RA and LA (panel D).

Figure 7

Summary of regional differences in cell width (panels A) and in RA and LA (panel B) and regional differences in sarcomere spacing (panel C) and in all myocytes from RA and LA (panel D).