Genetic targeting of Purkinje fibres by Sema3a-CreERT2

Overview & Synopsis

The maintenance of the heart rhythm and the conduction of excitatory signals require changing excitatory signals via electrical activity and coordination by communication between working and conductive cardiomyocytes. Understanding how the ventricular conduction system is established provides novel insights into the pathophysiological progress of cardiac arrhythmias. However, the major hurdle in this field is the lack of a specific genetic tool that targets the Purkinje fibres of the ventricular conduction system and no other types of cardiomyocytes or coronary vessels. Here, we generated a Sema3a-CreERT2 knock-in mouse line to test its specificity for genetically labelled Purkinje fibres. We found that Sema3a was expressed in the subendocardial layer of the trabecular myocardium in the embryonic heart and was restricted to the Purkinje fibres in the adult heart. A fate mapping study based on the Sema3a-CreERT2 line revealed that the Sema3a + cardiomyocytes were restricted to the fate of Purkinje fibres in the perinatal but not the embryonic stage. Collectively, our study provides a new genetic tool, i.e., Sema3a-CreERT2, for studying the molecular mechanisms that regulate the function of Purkinje fibres. The cardiac conduction system controls cardiac contraction by regulating the propagation of the electrical activity through the heart. Defects in the cardiac conduction system may lead to cardiac arrhythmias. Cardiac contraction begins in the right atrium, where it is initiated by the sinoatrial node (SA node). Then, the electrical activity is delayed at the atrioventricular node (AV node) and subsequently rapidly propagated throughout the ventricular wall through the fast-conducting atrioventricular bundle (the bundle of His and the bundle branches), which is localized at the crest of interventricular septum, and the peripheral left and right ventricular Purkinje fibres that are localized to the subendocardial surface of the myocardial wall 1. The Purkinje fibres ensure the rapid propagation of electric pulses that permit the efficient and effective pumping of blood from the ventricles. Understanding the molecular mechanisms that regulate Purkinje fibre development, formation, maintenance and proper function will provide insights into the pathophysiological process of cardiac arrhythmias. A recent study demonstrated that Hcn4-CreERT2 labels the conduction system including the sinoatrial node, a subset of the atri-oventricular node, the bundle of His, the bundle branches and the Purkinje fibres 2,3. Additionally, Hcn4-CreERT2 also labels a subset of endothelial cells in the developing heart 2,3. Cntn2-Cre is another genetic tool for the cardiac conduction system that specifically targets the SA node, AV node, left and right bundle branches and the Purkinje fibres 4. Previous research has revealed that connexin 40 (Cx40) is largely restricted to the ventricular conduction system (VCS), and it is also expressed in atrial cardiomyocytes and coronary arterial endothelial cells 5,6. The genetic tool Cx40-CreERT2 targets both atrial and VCS cardiomyocytes, as well as coronary endothelial cells 1,7. Therefore, the generation of a new tool that specifically targets the Purkinje fibres without targeting other cells may be valuable for specifically studying the function of Purkinje fibres. Sema3a is a member of the semaphorin family that is conserved from invertebrates to vertebrates. Recent studies indicate that Sema3a has an important function in the development of the nervous system 8,9. Sema3a could be a chemorepellent that inhibits the outgrowth of axons, especially peripheral axons, to maintain a normal neuronal pattern 10. The overexpression of Sema3a can lead to nerve diseases, such as schizophrenia, but