Abstract
SummarySinoatrial node (SAN) cells are the pacemakers of the heart. This study describes a method for culturing and infection of adult mouse SAN cells with FRET-based biosensors that can be exploited to examine signaling events. SAN cells cultured in media with blebbistatin or (S)-nitro-blebbistatin retain their morphology, protein distribution, action potential (AP) waveform, and cAMP dynamics for at least 40 h. SAN cells expressing targeted cAMP sensors show distinct β-adrenergic-mediated cAMP pools. Cyclic GMP, protein kinase A, Ca2+/CaM kinase II, and protein kinase D in SAN cells also show unique dynamics to different stimuli. Heart failure SAN cells show a decrease in cAMP and cGMP levels. In summary, a reliable method for maintaining adult mouse SAN cells in culture is presented, which facilitates studies of signaling networks and regulatory mechanisms during physiological and pathological conditions.
Highlights
Sinoatrial node (SAN) cells are the intrinsic pacemakers responsible for normal heart rhythm
SAN cells cultured in media with blebbistatin or (S)-nitro-blebbistatin retain their morphology, protein distribution, action potential (AP) waveform, and cyclic adenosine monophosphate (cAMP) dynamics for at least 40 h
Heart failure SAN cells show a decrease in cAMP and cyclic guanosine monophosphate (cGMP) levels
Summary
Sinoatrial node (SAN) cells are the intrinsic pacemakers responsible for normal heart rhythm. The cross talk and synergy between these clocks are modulated by second messengers and signaling molecules acting on coupled-clock proteins (Behar and Yaniv, 2016; Han et al, 1995; Liao et al, 2010; Shimizu et al, 2002; van Borren et al, 2010; Vinogradova et al, 2000; Yaniv et al, 2013, 2015a, 2015b). Addressing this issue is important to gain insight into how complex signaling networks regulate SAN pacemaking activity in health and disease (MacDonald et al, 2020)
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