The Electrophysiological Properties of Spontaneously Beating Pacemaker Cells Isolated from Mouse Sinoatrial Node

Abstract

In order to examine the phenotypic consequences of genetic manipulation in the function of the sinoatrial (SA) node, it is a prerequisite to record the electrical activities of a single pacemaker cell of the SA node of mouse heart. In the present study, we isolated spontaneously beating pacemaker cells from the SA node by enzymatic digestion. The rate of spontaneous action potential firing was 294 ± 59 min−1 at 33–34 °C. The maximal diastolic potential (MDP) was −56.7 ± 7.4 mV and the overshoot was 22.7 ± 6.2 mV. With hyperpolarizing voltage clamp pulses, the hyperpolarization‐activated, cyclic nucleotide‐sensitive cation current (Ih or If) was recorded. Ih started to activate at −70 to ≈−80 mV, more negative than MDP. The half‐maximal activation of Ih was obtained at −107.9 ± 10.4 mV. The inward‐rectifier K+ current (IK1) was also recorded in spontaneously beating myocytes. However, the amplitude of outward IK1 was negligibly small (9.1 ± 3.4 pA at −60 mV). With depolarization, voltage‐gated Na+ current, L‐type Ca2+ current and T‐type Ca2+ current were consistently observed. The sustained inward current (Ist) was also recorded in spontaneously beating pacemaker cells. E4031‐sensitive, rapidly activating delayed rectifier K+ current (IKr) was activated by depolarization, although the amplitude was no more than 38.3 ± 22.2 pA at 0 mV. The chromanol 293B‐sensitive, slowly activating delayed rectifier K+ current (IKs) was not present. The major repolarizing current was the slowly inactivating, 4‐aminopyridine‐insensitive outward current. We concluded that mouse pacemaker cells possess similar membrane currents, including Ist, to those of other species.

Pacemaker Mechanism of Porcine Sino-atrial Node Cells.

“…Typical pacemaker cells of the porcine SAN are morphologically similar to those of other species (Irisawa et al, 1993;Ono et al, 2000;Ono and Ito, 1995;Cho et al, 2003). They were visually identified as long, spindle shaped cells in control Tyrode's solution.…”

“…It has been reported that IKr exists in SAN cells of a small species such as the rabbit Ono and Ito, 1995;Verheijck et al, 1995), guinea-pig (Anumonwo et al, 1992;Guo et al, 1997) and mouse (Cho et al, 2003), and that it plays an important role in repolarization and the resulting slow diastolic depolarization. Although IKr was not recorded in porcine SAN cells that showed little evidence of striations, it did exist in large, rod-shaped quiescent cells in the same preparations (Ono and Iijima, unpublished data).…”

Creation of a Biopacemaker: Lessons from the Sinoatrial Node

“…Not only do these patients exhibit prolonged QT intervals, causing arrhythmias during exercise due to prolonged repolarization in ventricular tissue, they also have a diminished increase in heart rate upon exercise (Haapalahti et al, 2006). No I Ks has been found in mouse SA node (Cho et al, 2003), and, as expected, KCNQ1 knock out mice do not show prolongation of cycle length (Knollmann et al, 2004). Since I Ks is the only I K present in pig, spontaneous activity ceased on blocking I Ks with chromanol 293B (Ono et al, 2000).…”

“…Consequently, I K1 will stabilize the membrane potential around E K . This current, which at first glance appears to antagonize spontaneous activity, is not present in rabbit SA node (Irisawa et al, 1993), and is negligibly small in murine and rat SA node cells (Cho et al, 2003;Shinagawa et al, 2000). Figure 4A, right panel, shows typical current traces in a rabbit SA node cell measured with a similar voltage clamp protocol and solutions as in the ventricular myocyte (left panel).…”

“…-Adrenergic stimulation shifts the threshold for activation and the potential of maximum amplitude to more negative potentials (Cho et al, 2003;Huang et al, 2008;Toyoda et al, 2005). Acetylcholine (ACh) does not have an inhibitory effect in an unstimulated situation, but, after stimulation with isoproterenol, ACh causes a reduction in amplitude (Toyoda et al, 2005).…”