Alterations of Ca2+ signaling and Ca2+ release sites in cultured ventricular myocytes with intact internal Ca2+ storage.
Biochem Biophys Res Commun
; 527(2): 379-386, 2020 06 25.
Article
em En
| MEDLINE
| ID: mdl-32321644
Although cultured adult cardiac myocytes in combination with cell-level genetic modifications have been adopted for the study of protein function, the cellular alterations caused by the culture conditions themselves need to be clarified before we can interpret the effects of genetically altered proteins. We systematically compared the cellular morphology, global Ca2+ signaling, elementary Ca2+ release (sparks), and arrangement of ryanodine receptor (RyR) clusters in short-term (2 days)-cultured adult rat ventricular myocytes with those of freshly isolated myocytes. The transverse (t)-tubules were remarkably decreased (to â¼25%) by culture, and whole-cell capacitance was reduced by â¼35%. The magnitude of depolarization-induced Ca2+ transients decreased to â¼50%, and Ca2+ transient decay was slowed by culture. The culture did not affect sarcoplasmic reticulum (SR) Ca2+ loading. Therefore, fractional Ca2+ release was attenuated by culture. In the cultured cells, the L-type Ca2+ current (ICa) was smaller (â¼50% of controls) and its inactivation was slower. In cultured myocytes, there were significantly fewer (â¼50% of control) Ca2+ sparks, the local Ca2+ releases through RyR clusters, compared with in freshly isolated cells. Amplitude and kinetics (duration and time-to-peak) of individual sparks were similar, but they showed greater width in cultured cells. Immunolocalization analysis revealed that the cross-striation of RyRs distribution became weaker and less organized, and that the density of RyR clusters decreased in cultured myocytes. Our data suggest that the loss of t-tubules and generation of compromised Ca2+ transients and ICa in short-term adult ventricular cell culture are independent of SR Ca2+ loading status. In addition, the deteriorated arrangement of the RyR-clusters and their decreased density after short-term culture may be partly responsible for fewer Ca2+ sparks and a decrease in global Ca2+ release.
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Texto completo:
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Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Sinalização do Cálcio
/
Miócitos Cardíacos
/
Ventrículos do Coração
Limite:
Animals
Idioma:
En
Revista:
Biochem Biophys Res Commun
Ano de publicação:
2020
Tipo de documento:
Article
País de afiliação:
Coréia do Sul
País de publicação:
Estados Unidos