InsP 3 Triggered Calcium Release Events in Mouse Atrial Myocytes

Abstract

In the heart, the dominant mechanism of intracellular Ca2+ release is Ca2+-induced Ca2+ release (CICR) via sarcoplasmic reticulum (SR) Ca2+ release channels (RyR's). Recently, a second mechanism, Ca2+ release through channels sensitive to the intracellular second messenger Inositol-1,4,5-trisphosphate (InsP3) has been described. The contribution and significance of InsP3-induced Ca2+ release in cardiac excitation-contraction coupling (EC-coupling) is still a matter of debate. However, recent reports have emphasized the importance of InsP3 signaling for EC-coupling in atrial myocytes, for excitation transcription coupling and embryogenesis. Investigating InsP3 signaling is challenging because adequately selective pharmacological tools or fluorescent indicators are not available and the specific activation of highly localized intracellular InsP3 receptors (InsP3R) is hampered due to experimental inaccessibility. We are using UV-flash uncaging approaches of caged InsP3 to study the interplay of InsP3R Ca2+ release and CICR based on the activation of RyR under whole-cell conditions. UV-flash photolysis of caged InsP3 was accompanied by an increase in the number of local Ca2+ release events that show larger FDHM and/or smaller amplitude. In the presence of InsP3R blocker xestospongin C the frequency of InsP3 evoked Ca2+ events was reduced and suggest coexistence of spontaneous SR-Ca2+ release events (Ca2+ sparks) and InsP3 evoked SR-Ca2+ release events (Ca2+ puffs). In addition, photorelease of InsP3 in PM loaded cells induced global Ca2+ release events suggesting that InsP3 facilitation of Ca2+ release may be linked to RyR mediated Ca2+ release in atrial cells. Two-photon excitation photolysis (TPP) of caged InsP3 was used to study InsP3 signaling in a highly targeted way. TPP triggered InsP3-induced Ca2+ release exhibiting spatio-temporal characteristics corresponding to elementary Ca2+ signals, such as Ca2+ waves and Ca2+ “puffs” and seems to be promising tools for studying InsP3 signaling on the sub-cellular scale. Supported by SNF.