Dataset Information

Deciphering cellular signals in adult mouse sinoatrial node cells.

ABSTRACT: Sinoatrial 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.

SUBMITTER: Reddy GR

PROVIDER: S-EPMC8749457 | biostudies-literature | 2022 Jan

REPOSITORIES: biostudies-literature

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Deciphering cellular signals in adult mouse sinoatrial node cells.

Reddy Gopireddy R GR Ren Lu L Thai Phung N PN Caldwell Jessica L JL Zaccolo Manuela M Bossuyt Julie J Ripplinger Crystal M CM Xiang Yang K YK Nieves-Cintrón Madeline M Chiamvimonvat Nipavan N Navedo Manuel F MF

iScience 20211225 1

Sinoatrial 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. ...[more]

PMID: 35036877

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Project description:Phosphodiesterases (PDEs) are critical regulators of cyclic nucleotides in the heart. In ventricular myocytes, the L-type Ca(2+) current (I(Ca,L)) is a major target of regulation by PDEs, particularly members of the PDE2, PDE3 and PDE4 families. Conversely, much less is known about the roles of PDE2, PDE3 and PDE4 in the regulation of action potential (AP) properties and I(Ca,L) in the sinoatrial node (SAN) and the atrial myocardium, especially in mice. Thus, the purpose of our study was to measure the effects of global PDE inhibition with Isobutyl-1-methylxanthine (IBMX) and selective inhibitors of PDE2, PDE3 and PDE4 on AP properties in isolated mouse SAN and right atrial myocytes. We also measured the effects of these inhibitors on I(Ca,L) in SAN and atrial myocytes in comparison to ventricular myocytes. Our data demonstrate that IBMX markedly increases spontaneous AP frequency in SAN myocytes and AP duration in atrial myocytes. Spontaneous AP firing in SAN myocytes was also increased by the PDE2 inhibitor erythro-9-[2-hydroxy-3-nonyl] adenine (EHNA), the PDE3 inhibitor milrinone (Mil) and the PDE4 inhibitor rolipram (Rol). In contrast, atrial AP duration was increased by EHNA and Rol, but not by Mil. IBMX also potently, and similarly, increased I(Ca,L) in SAN, atrial and ventricular myocytes; however, important differences emerged in terms of which inhibitors could modulate I(Ca,L) in each myocyte type. Consistent with our AP measurements, EHNA, Mil and Rol each increased I(Ca,L) in SAN myocytes. Also, EHNA and Rol, but not Mil, increased atrial I(Ca,L). In complete contrast, no selective PDE inhibitors increased I(Ca,L) in ventricular myocytes when given alone. Thus, our data show that the effects of selective PDE2, PDE3 and PDE4 inhibitors are distinct in the different regions of the myocardium indicating important differences in how each PDE family constitutively regulates ion channel function in the SAN, atrial and ventricular myocardium.

Dataset Information

Deciphering cellular signals in adult mouse sinoatrial node cells.

Publications

Deciphering cellular signals in adult mouse sinoatrial node cells.

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