https://hdl.handle.net/20.500.12412/4336 2026-05-04T13:27:48Z 2026-05-04T13:27:48Z Ruíz-Hurtado, G Gomez-Hurtado, N Fernandez-Velasco, M Calderón-Sánchez, E Smani, T Ordoñez, A Cachofeiro, V Boscá, L Díez, J Gómez, AM Delgado, C https://hdl.handle.net/20.500.12412/7226 2026-04-27T21:00:23Z 2012-01-01T00:00:00Z

Cardiotrophin-1 induces sarcoplasmic reticulum Ca2+ leak and arrhythmogenesis in adult rat ventricular myocytes Ruíz-Hurtado, G; Gomez-Hurtado, N; Fernandez-Velasco, M; Calderón-Sánchez, E; Smani, T; Ordoñez, A; Cachofeiro, V; Boscá, L; Díez, J; Gómez, AM; Delgado, C Aims Plasma levels of cardiotrophin-1 (CT-1) are elevated in several cardiovascular diseases and are correlated with the severity of the pathology. However, the mechanisms by which this inflammatory cytokine participates in the pathology of the heart are not completely understood. It is well established that alterations in intracellular calcium ([Ca2+]i) handling are involved in cardiac dysfunction during heart failure, but it is unknown whether CT-1 modulates [Ca2+]i handling in adult cardiomyocytes. Here we have analyzed for the first time the effects of CT-1 on [Ca2+]i homeostasis in adult rat cardiomyocytes. Methods and results L-type calcium current (ICaL) was recorded using patch-clamp techniques, and [Ca2+]i transients and Ca2+ sparks were viewed by confocal microscopy. Treatment of cardiomyocytes with 1 nM CT-1 for 20–60 min induced a significant increase in ICaL density, [Ca2+]i transients, and cell shortening compared with control cells. Our study reveals that CT-1 increases ICaL by a protein kinase A-dependent mechanism, and Ca2+ sparks by a Ca2+/calmodulin kinase II-dependent and protein kinase A-independent mechanism. Cardiomyocytes treated with CT-1 exhibited a higher occurrence of arrhythmogenic behaviour, manifested as spontaneous Ca2+ waves and aftercontractions. Conclusion Our findings provide evidence that cardiomyocytes treated with CT-1 present high spontaneous Ca2+ release during diastole, a mechanism linked to arrhythmogenicity in the pathologic heart.

2012-01-01T00:00:00Z Domínguez-Rodríguez, A Díaz, I Rodríguez-Moyano, M Calderón-Sánchez, EM Rosado, JA Ordóñez, A Smani, T https://hdl.handle.net/20.500.12412/7225 2026-04-27T21:00:14Z 2012-01-01T00:00:00Z

Urotensin-II signaling mechanism in rat coronary artery: Role of STIM1 and Orai1-dependent store operated calcium influx in vasoconstriction Domínguez-Rodríguez, A; Díaz, I; Rodríguez-Moyano, M; Calderón-Sánchez, EM; Rosado, JA; Ordóñez, A; Smani, T Objective-: Human urotensin-II (UII) is considered the most potentendogenous vasoconstrictor discovered to date, although the precise mechanism activated downstream of its receptor UTS2R in blood vessels remains elusive. The aim of this study was to determine the role of the store operated Ca 2+ entry (SOCE) signaling pathway in UII-induced coronary artery vasoconstriction. Methods And Results-: We used a combination of isometric tension measurement, Ca 2+ imaging, pharmacology, and molecular approaches to study UII-mediated rat coronary artery vasoconstriction and intracellular Ca 2+ mobilization in coronary smooth muscle cells. We found that UII promoted dose-dependent vasoconstriction and elicited Ca 2+ and Mn influx, which were sensitive to classical SOCE inhibitors. In addition, knockdown of either STIM1 or Orai1 essentially inhibited UII-mediated SOCE and prevented UII but not high-KCL evoked contraction in transfected coronary artery. Moreover, we found that Ca 2+-independent phospholipase A2β was involved in UII effects and that is colocalized with STIM1 in different submembrane compartments. Importantly, STIM1 but not Orai1 downregulation inhibits significantly independent phospholipase A 2 activation. Furthermore, lysophosphatidylcholine, an independent phospholipase A 2 product, activated Orai1 but not STIM1-dependent contraction and SOCE. Conclusion-: Here, we demonstrated that different critical players of SOCE signaling pathway are required for UII-induced vasoconstriction of rat coronary artery.

2012-01-01T00:00:00Z Calderón-Sánchez, E Ruíz-Hurtado, Gema Smani, T Delgado, C Benitah, JP Gómez, AM Ordóñez, A https://hdl.handle.net/20.500.12412/7224 2026-04-27T21:00:28Z 2011-01-01T00:00:00Z

Cardioprotective action of urocortin in postconditioning involves recovery of intracellular calcium handling. Calderón-Sánchez, E; Ruíz-Hurtado, Gema; Smani, T; Delgado, C; Benitah, JP; Gómez, AM; Ordóñez, A Ischemia/reperfusion (I/R) damage in the heart occurs mainly during the first minutes of reperfusion. Urocortin (Ucn) is a member of the corticotrophin-releasing factor that has been identified as a potent endogenous cardioprotector peptide when used in pre- and postconditioning protocols. However, the underlying mechanisms are not completely elucidated. Here, we focused on intracellular calcium ([Ca(2+)](i)) handling by Ucn when applied in early reperfusion. We used Langendorff-perfused rat hearts to determine hemodynamic parameters, and confocal microscopy to study global [Ca(2+)](i) transients evoked by electrical stimulation in isolated cardiomyocytes loaded with fluorescence Ca(2+) dye fluo-3AM. We found that the acute application of Ucn at the onset of reperfusion, in isolated hearts submitted to ischemia, fully recovered the hearts contractility and relaxation. In isolated cardiac myocytes, following ischemia we observed that the diastolic [Ca(2+)](i) was increased, the systolic [Ca(2+)](i) transients amplitude were depressed and sarcoplasmic reticulum (SR) Ca(2+) load was reduced. These effects were correlated to a decrease in the Na(+)/Ca(2+) exchanger (NCX) activity. Importantly, Ucn applied at reperfusion produced a complete recovery in diastolic [Ca(2+)](i) and global [Ca(2+)](i) transient amplitude, which were due to NCX activity improvement. In conclusion, we demonstrated that [Ca(2+)](i) handling play an essential role in postconditioning action of Ucn.

2011-01-01T00:00:00Z Fernández Tenorio, Miguel Porras Gonzalez, Cristina Castellano, Antonio del Valle Rodríguez, Alberto López Barneo, José Ureña, Juan https://hdl.handle.net/20.500.12412/7221 2026-04-24T21:00:16Z 2011-04-14T00:00:00Z

Metabotropic Regulation of RhoA/Rho-Associated Kinase by L-type Ca2+ Channels New Mechanism for Depolarization-Evoked Mammalian Arterial Contraction Fernández Tenorio, Miguel; Porras Gonzalez, Cristina; Castellano, Antonio; del Valle Rodríguez, Alberto; López Barneo, José; Ureña, Juan Background: Sustained vascular smooth muscle contraction is mediated by extracellular Ca2 influx through L-type voltage-gated Ca2 channels (VGCC) and RhoA/Rho-associated kinase (ROCK)-dependent Ca2 sensitization of the contractile machinery. VGCC activation can also trigger an ion-independent metabotropic pathway that involves G-protein/phospholipase C activation, inositol 1,4,5-trisphosphate synthesis, and Ca2 release from the sarcoplasmic reticulum (calcium channel-induced Ca2 release). We have studied the functional role of calcium channel-induced Ca2 release and the inter-relations between Ca2 channel and RhoA/ROCK activation. Methods and Results: We have used normal and genetically modified animals to study single myocyte electrophys iology and fluorimetry as well as cytosolic Ca2 and diameter in intact arteries. These analyses were complemented with measurement of tension and RhoA activity in normal and reversibly permeabilized arterial rings. We have found that, unexpectedly, L-type Ca2 channel activation and subsequent metabotropic Ca2 release from sarcoplasmic reticulum participate in depolarization-evoked RhoA/ROCK activity and sustained arterial contraction. We show that these phenomena do not depend on the change in the membrane potential itself, or the mere release of Ca2 from the sarcoplasmic reticulum, but they require the simultaneous activation of VGCC and the downstream metabotropic pathway with concomitant Ca2 release. During protracted depolarizations, refilling of the stores by a residual extracellular Ca2 influx through VGCC helps maintaining RhoA activity and sustained arterial contraction. Conclusions: These findings reveal that calcium channel-induced Ca2 release has a major role in tonic vascular smooth muscle contractility because it links membrane depolarization and Ca2 channel activation with metabotropic Ca2 release and sensitization (RhoA/ROCK stimulation)

2011-04-14T00:00:00Z