A modified method for isolating sinoatrial node myocytes from adult mice

Sinoatrial node (SAN) is the pacemaker of the heart in charge of initiating spontaneous electronical activity and controlling heart rate. Myocytes from SAN can generate spontaneous rhythmic action potentials, which propagate through the myocardium, thereby triggering cardiac myocyte contraction. Acutely, the method for isolating sinoatrial node myocytes (SAMs) is critical in studying the protein expression and function of myocytes in SAN. Currently, the SAMs were isolated by transferring SAN tissue directly into the digestion solution, but it is difficult to judge the degree of digestion, and the system was unstable. Here, we present a modified protocol for the isolation of SAMs in mice, based on the collagenase II and protease perfusion of the heart using a Langendorff apparatus and subsequent dissociation of SAMs. The appearance and droplet flow rate of the heart could be significantly changed during enzymatic digestion via perfusion, which allowed us to easily judge the degree of digestion and avoid incomplete or excessive digestion. The SAMs with stable yield and viability achieved from our optimized approach would facilitate the follow-up experiments.