Nuclear Ca2+ signaling to endothelin-1 in rat aortic smooth-muscle cells.
Vascular smooth-muscle cells (VSMCs) isolated from genetically hypertensive animals show increased intracellular free calcium levels ([Ca2+]i) in response to endothelin-1 (ET-1). The differences in time course and distribution of Ca2+ increase after addition of ET-1 within the VSMCs are unknown. Therefore, ET-1-evoked changes in fluo-3 fluorescence were determined using a confocal laser scanning microscope in primary cultures of aortic smooth-muscle cells (ASMCs) from 12-week-old male Sprague-Dawley (SD) rats, Wistar-Kyoto (WKY) rats, and spontaneously hypertensive rats (SHR). Syto-11 staining enabled the assessment of intracellular free [Ca2+]i changes in the cytosolic ([Ca2+]c), perinuclear ([Ca2+]p), and nuclear ([Ca2+]n) regions. In the basal state, [Ca2+] was evenly distributed throughout the rat ASMCs. There were no significant differences in basal fluorescence values among the three strains. ET-1 evoked a concentration-dependent increase in fluo-3 intensity. The peak [Ca2+]i rise to ET-1 was much more rapid in ASMCs from SHR and WKY strains. The changes in [Ca2+]n were greater than in [Ca2+]c. Pretreatment of rat ASMCs with BQ-123 (an ETA antagonist) and BQ-788 (an ETB antagonist) abolished the rapid peak rise and the slow sustained elevation in [Ca2+]i, respectively. The nonselective antagonist bosentan attenuated both phases of the ET-1 response in all three strains. The ETB-selective agonist IRL 1620 evoked a significant elevation in [Ca2+]n values at 2 min in the ASMCs of SHR. These data suggest that ETA activation is linked to initial rapid increases in [Ca2+]c and [Ca2+]n, whereas ETB activation promotes slow [Ca2+]n signaling, particularly in ASMCs of SHR.
Naik
R D
RD
Cardiovascular Risk Factor Reduction Unit, College of Medicine, University of Saskatchewan, Saskatoon, Canada.
McNeill
J R
JR
Wilson
T W
TW
Gopalakrishnan
V
V
D000814:Aniline Compounds; D000818:Animals; D001013:Aorta, Thoracic / Q000187:drug effects / Q000502:physiology / Q000648:ultrastructure; D002118:Calcium / Q000502:physiology*; D002467:Cell Nucleus / Q000187:drug effects / Q000502:physiology*; D065128:Endothelin Receptor Antagonists; D019332:Endothelin-1 / Q000494:pharmacology*; D005456:Fluorescent Dyes; D066298:In Vitro Techniques; D008297:Male; D018613:Microscopy, Confocal; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000502:physiology* / Q000648:ultrastructure; D009842:Oligopeptides / Q000494:pharmacology; D010456:Peptides, Cyclic / Q000494:pharmacology; D010880:Piperidines / Q000494:pharmacology; D051381:Rats; D011918:Rats, Inbred SHR; D011921:Rats, Inbred WKY; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction / Q000187:drug effects*; D014966:Xanthenes
D000814:Aniline Compounds; D065128:Endothelin Receptor Antagonists; D019332:Endothelin-1; D005456:Fluorescent Dyes; D009842:Oligopeptides; D010456:Peptides, Cyclic; D010880:Piperidines; D014966:Xanthenes; C059715:Fluo-3; C086539:BQ 788; C072247:cyclo(Trp-Asp-Pro-Val-Leu); D002118:Calcium