Visualization of left ventricular Purkinje fiber distribution using widefield optical coherence microscopy

Abstract

Background: The distribution and connection of ventricular Purkinje fibers are known to be associated with idiopathic left ventricular arrhythmias. Unusual anatomy is one of the important factors associated with catheter ablation success rate. With the widefield high-speed, swept-source optical coherence microscopy (OCM) and light microscope, we visualized the left ventricular Purkinje fiber distribution.

Methods: Left ventricular walls of five adult ovine hearts were incised from the mitral annulus to the apex. Using the widefield OCM technique and light microscopy, we observed the distribution, direction, depth, and dividing patterns of the Purkinje network with multiple tangential angles and without tissue destruction.

Results: Widefield OCM was used to characterize the ovine heart Purkinje network system in a 4 × 4 mm² field. Left ventricular Purkinje fibers traveled in the sub-endocardial area near the left-sided peri-membranous septal area and ran like a wide hair bundle. The distal branching fibers penetrated to the endocardium and connected to the contractile muscle. In this distal area, Purkinje fibers were connected to each other, forming multiple layers. Some Purkinje fibers were directly connected within the false tendon between the papillary muscles or between the trabeculations. Some free-running Purkinje fibers were directly connected to the papillary muscle from the left bundle.

Conclusion: Using widefield OCM, we were able to observe the left bundle and its branching patterns in ovine left ventricle without tissue destruction. This might be applied to future cardiac ablation procedures.

Keywords: Purkinje fibers; cardiac arrhythmia; heart conduction system; optical microscopy.

Figure 1

OCM system. This new optical coherence microscopy had lateral resolution of 7.8 μm and axial resolution of 11.4 μm in tissue. Field of view (FOV) from galvanometer scanning was 4 mm × 4 mm and wide FOV images up to 20 mm × 20 mm were generated by stitching and reconstructing the multiple galvanometer scanning FOV images acquired by moving the two-axis translation stage. FC, fiber coupler. C, circulator. PC, polarization controller. CL, collimation lens. GM, Galvanometer. RM, reference mirror. FL, focusing lens. OL, objective lens.

Figure 2

OCM and conventional light microscopic observations of HPS in the same spot. A. OCM image. Purkinje fibers can be seen as low density and reticular sheath is observed as high density. It is suitable for observing the running direction of fibers in a wide area. B. Light microscope image. Purkinje fibers stained by hematoxylin and eosin have fewer myofibrils compared to other nearby myocardial tissues, have glycogen around the nucleus, and are larger and longer than cardiac myocytes.

Figure 3

Serial optical coherence microscopic images. With optical coherence microscopy, the region of interest can be observed at various desired depths without tissue destruction.

Figure 4

OCM images of left bundle branch and Purkinje bundles. A. Left bundle branch. The left bundle runs in the apical direction like a waterfall emerging from the myocardial peri-membranous septal wall. B. Bundle of Purkinje fibers. The direction of this bundle is different from that of the working myocardium in the endocardial area. Each fiber splits into a thinner fascicle according to its destination.

Figure 5

Free-running Purkinje fibers. A. Gross findings. B. OCM findings. C. Histology. Some Purkinje bundles are free running without attaching with ventricular surface or directly connect between trabeculae and the papillary muscle. The Purkinje fibers are seen at most of the false tendon at the apical part of the septum. Only Purkinje cells without cardiac myocytes are in these false tendons.

Figure 6

Schematic diagram of left ventricular Purkinje system. A. The left bundle branch is divided into two or four major fascicles. B. Main fascicles are divided into branches, but some branches meet again. C. Some fibers in the main fascicle are separated from the subendocardial area and connect directly to the papillary muscle. D. There are some free-running fibers connecting branched fascicles. E. Free-running fibers also branch or meet each other.