Na_v1.5, the pore forming α-subunit of the voltage-dependent cardiac Na⁺ channel, is an integral membrane protein involved in the initiation and conduction of action potentials. Mutations in the gene-encoding Na_v1.5, SCN5A, have been associated with a variety of arrhythmic disorders, including long QT, Brugada, and sick sinus syndromes as well as progressive cardiac conduction defect and atrial standstill. Moreover, alterations in the Na_v1.5 expression level and/or sodium current density have been frequently noticed in acquired cardiac disorders, such as heart failure. The molecular mechanisms underlying these alterations are poorly understood, but are considered essential for conception of arrhythmogenesis and the development of therapeutic strategies for prevention or treatment of arrhythmias. The unravelling of such mechanisms requires critical molecular insight into the biology of Na_v1.5 expression and function. Therefore, the aim of this review is to provide an up-to-date account of molecular determinants of normal Na_v1.5 expression and function. The parts of the Na_v1.5 life cycle that are discussed include (i) regulatory aspects of the SCN5A gene and transcript structure, (ii) the nature, molecular determinants, and functional consequences of Na_v1.5 post-translational modifications, and (iii) the role of Na_v1.5 interacting proteins in cellular trafficking. The reviewed studies have provided valuable information on how the Na_v1.5 expression level, localization, and biophysical properties are regulated, but also revealed that our understanding of the underlying mechanisms is still limited.