Sinoatrial node myocytes (
SAMs) act as the natural pacemakers of the heart, initiating each heart beat by generating spontaneous action potentials (APs). These pacemaker APs reflect the coordinated activity of numerous membrane currents and intracellular
calcium cycling. However the precise mechanisms that drive spontaneous pacemaker activity in
SAMs remain elusive. Acutely isolated
SAMs are an essential preparation for experiments to dissect the molecular basis of cardiac pacemaking. However, the indistinct anatomy, complex microdissection, and finicky enzymatic digestion conditions have prevented widespread use of acutely isolated
SAMs. In addition, methods were not available until recently to permit longer-term culture of
SAMs for
protein expression studies. Here we provide a step-by-step protocol and video demonstration for the isolation of
SAMs from adult mice. A method is also demonstrated for maintaining adult mouse
SAMs in vitro and for expression of exogenous
proteins via adenoviral
infection. Acutely isolated and cultured
SAMs prepared via these methods are suitable for a variety of electrophysiological and imaging studies.