Nkx2.5 cell-autonomous gene function is required for the postnatal formation of the peripheral ventricular conduction system.

The ventricular conduction system is responsible for rapid propagation of electrical activity to coordinate ventricular contraction. To investigate the role of the transcription factor Nkx2.5 in the morphogenesis of the ventricular conduction system, we crossed Nkx2.5(+/-) mice with Cx40(eGFP/+) mice in which eGFP expression permits visualization of the His-Purkinje conduction system. Major anatomical and functional disturbances were detected in the His-Purkinje system of adult Nkx2.5(+/-)/Cx40(eGFP/+) mice, including hypoplasia of eGFP-positive Purkinje fibers and the disorganization of the Purkinje fiber network in the ventricular apex. Although the action potential properties of the individual eGFP-positive cells were normal, the deficiency of Purkinje fibers in Nkx2.5 haploinsufficient mice was associated with abnormalities of ventricular electrical activation, including slowed and decremented conduction along the left bundle branch. During embryonic development, eGFP expression in the ventricular trabeculae of Nkx2.5(+/-) hearts was qualitatively normal, with a measurable deficiency in eGFP-positive cells being observed only after birth. Chimeric analyses showed that maximal Nkx2.5 levels are required cell-autonomously. Reduced Nkx2.5 levels are associated with a delay in cell cycle withdrawal in surrounding GFP-negative myocytes. Our results suggest that the formation of the peripheral conduction system is time- and dose-dependent on the transcription factor Nkx2.5 that is cell-autonomously required for the postnatal differentiation of Purkinje fibers.

Architectural and functional asymmetry of the His-Purkinje system of the murine heart.

OBJECTIVE: The aim of this work was to target a vital reporter gene in the mouse cardiac conduction system (CS) to distinguish this tissue from the surrounding myocardium in the adult heart. METHODS: A transgenic mouse line has been created in which EGFP is expressed under the control of the Cx40 gene. Correlative investigations associating EGFP imaging and electrophysiological techniques were carried out on the adult heart and isolated cardiomyocytes. RESULTS: In the heart of the Cx40(EGFP/+) mice, EGFP signal was seen in the coronary arteries, the atria, the atrioventricular (AV) node and the His-Purkinje system. The latter was found to be structurally and functionally asymmetrical. The anatomical asymmetry was apparent in both the number of strands or fasciculi making up the His bundle branches (BBs) (1 strand on the right, 20 or so on the left), and the density (low on the right, high on the left) of the network of Purkinje fibers (PFs) that extends over the ventricular wall surfaces. The profiles of the electrical activation patterns recorded on the right and left flanks of the septum were also asymmetrical, mirroring the architecture of the branches. EGFP made it easy to identify the Purkinje cells in populations of dissociated cardiomyocytes and they were investigated using the patch-clamp technique. The hyperpolarization-activated current (If) was recorded in all spontaneously active Purkinje cells. CONCLUSIONS: This investigation provides positive evidence of the asymmetry of the His-Purkinje system of the adult mouse, and the first patch-clamp recording data on murine cardiac Purkinje cells. This mouse model opens up new perspectives for investigating the contribution of specific genes to the morphology and function of the His-Purkinje system.

Genetically modified mice: tools to decode the functions of connexins in the heart-new models for cardiovascular research.

It has long been known that gap junctions are required for the propagation of electrical impulse in the heart. A good deal later, the connexins (Cxs), which are probably exclusive components of the junctional channels that constitute the gap junctions, were identified. More recently, the in vivo functions of cardiac Cxs have been investigated by the analysis of genetically modified mice. These studies have confirmed that Cxs are involved in cardiac impulse conduction, and, unexpectedly, in heart morphogenesis. In addition, cardiac abnormalities described in mice genetically modified for Cx genes, and those observed in certain human cardiac diseases, have been proven to be similar.