title: Alpha-adrenergic agonist and endothelin-1 induced intracellular Ca2+ response in the presence of a Ca2+ entry blocker in cultured rat ventricular myocytes.
abstract: Previously we demonstrated that stimulation of cultured neonatal rat ventricular myocytes by either alpha 1-adrenergic agonist or endothelin-1 resulted in a rapid formation of total inositolphosphates, although the levels of inositol 1,4,5-trisphosphate and inositol 1,3,4,5-tetrakisphosphate did not rise significantly. The aim of this study was to examine whether stimulation by alpha 1-adrenergic agonist and endothelin-1 could still elicit phosphatidylinositol cycle mediated intracellular Ca2+ mobilization in these cells. The intracellular free Ca2+ concentration ([Ca2+]i) was measured by single cell imaging dual wavelength fluorescence microscopy in Fura-2-loaded cardiomyocytes. The interference of agonist induced [Ca2+]i responses by the beat to beat variation of [Ca2+]i was prevented by arresting the cells with the Ca2+ entry blocker diltiazem (10 microM). The [Ca2+]i response (expressed as % of baseline ratio of fluorescence intensities of Fura-2 at 340 nm and 380 nm excitation wavelength), induced by phenylephrine (10(-4) M) and endothelin-1 (10(-8) M) was small, up to 20% of baseline after 9-20 min. In contrast, Ca(2+)-influx induced by incubation in Na(+)-free buffer caused a steep increase of [Ca2+]i up to 150% of baseline after 30 s. Analysis of single cells following stimulation with phenylephrine or endothelin-1 showed heterogeneity with respect to a rise in [Ca2+]i. However, if rapid Ca(2+)-influx was induced by incubation in Na(+)-free buffer, [Ca2+]i responses in individual myocytes occurred homogeneously. It is concluded that the alpha 1-adrenergic agonist and endothelin-1 induced [Ca2+]i responses are delayed in time, small and quite heterogeneous among cells. The findings are in agreement with earlier observations which revealed no detectable overall increase of the Ca2+ releasing inositolphosphates under these conditions and suggest that other second messengers, such as 1,2-diacylglycerol, are involved in the agonist mediated Ca2+ signals.
authors: Department of Biochemistry, Faculty of Medicine and Health Sciences, Erasmus University Rotterdam, The Netherlands.
authors: H W
authors: HW
authors: De Jonge
authors: D E
authors: DE
authors: Atsma
authors: E J
authors: EJ
authors: van der Valk-Kokshoorn
authors: H A
authors: HA
authors: van Heugten
authors: A
authors: A
authors: van der Laarse
authors: J M
authors: JM
authors: Lamers
mesh_terms: D000316:Adrenergic alpha-Agonists / Q000494:pharmacology*; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology*; D002478:Cells, Cultured / Q000187:drug effects / Q000378:metabolism; D004110:Diltiazem / Q000494:pharmacology*; D016232:Endothelins / Q000494:pharmacology*; D005456:Fluorescent Dyes; D016257:Fura-2; D006352:Heart Ventricles / Q000166:cytology; D015544:Inositol 1,4,5-Trisphosphate / Q000096:biosynthesis; D007295:Inositol Phosphates / Q000096:biosynthesis; D008856:Microscopy, Fluorescence; D009206:Myocardium / Q000166:cytology*; D010656:Phenylephrine / Q000494:pharmacology*; D051381:Rats; D017208:Rats, Wistar; D015290:Second Messenger Systems / Q000502:physiology; D012964:Sodium / Q000494:pharmacology; D014316:Tritium / Q000378:metabolism
chemical_list: D000316:Adrenergic alpha-Agonists; D002121:Calcium Channel Blockers; D016232:Endothelins; D005456:Fluorescent Dyes; D007295:Inositol Phosphates; D014316:Tritium; C081655:inositol-1,3,4,5-tetrakisphosphate; D010656:Phenylephrine; D015544:Inositol 1,4,5-Trisphosphate; D012964:Sodium; D004110:Diltiazem; D002118:Calcium; D016257:Fura-2