Endothelial Ca2+ waves preferentially originate at specific loci in caveolin-rich cell edges.
Stimulation of endothelial cells (ECs) with ATP evoked an increase in intracellular Ca2+ concentration ([Ca2+]i). In a single bovine aortic EC, the [Ca2+]i rise started at a specific peripheral locus and propagated throughout the entire cell as a Ca2+ wave. The initiation locus was constant upon repeated stimulation with ATP or other agonists (bradykinin and thrombin). The Ca2+ wave was unaffected by the removal of extracellular Ca2+, demonstrating its dependence on intracellular Ca2+ release. Microinjection of heparin into the cell inhibited the ATP-induced Ca2+ responses, indicating that the Ca2+ wave is at least partly mediated by the inositol 1,4, 5-trisphosphate receptor. Immunofluorescence staining revealed that caveolin, a marker protein for caveolae, is distributed heterogeneously in the cell and that Ca2+ waves preferentially originate at caveolin-rich cell edges. In contrast to caveolin, internalized transferrin and subunits of the clathrin-associated adaptor complexes such as adaptor protein-1 and -2 were diffusely distributed. Disruption of microtubules by Colcemid led to redistribution of caveolin away from the edges into the perinuclear center of the cell, and the ATP-induced [Ca2+]i increase was initiated on the rim of the centralized caveolin. Thus, caveolae may be involved in the initiation of ATP-induced Ca2+ waves in ECs.
Isshiki
M
M
Department of Biomedical Engineering, Graduate School of Medicine, University of Tokyo 113-0033, Japan. isshikim@tau.bekkoame.or.jp
Ando
J
J
Korenaga
R
R
Kogo
H
H
Fujimoto
T
T
Fujita
T
T
Kamiya
A
A
D033965:Adaptor Protein Complex alpha Subunits; D033942:Adaptor Proteins, Vesicular Transport; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000502:physiology*; D002417:Cattle; D051242:Caveolin 1; D022461:Caveolins; D002451:Cell Compartmentation / Q000187:drug effects; D002478:Cells, Cultured; D002966:Clathrin / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D003703:Demecolcine / Q000494:pharmacology; D011992:Endosomes / Q000378:metabolism; D004730:Endothelium, Vascular / Q000378:metabolism*; D019084:Fluorescent Antibody Technique, Indirect; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism; D007834:Lasers; D008565:Membrane Proteins / Q000378:metabolism / Q000502:physiology*; D018613:Microscopy, Confocal; D014168:Transferrin / Q000378:metabolism
D033965:Adaptor Protein Complex alpha Subunits; D033942:Adaptor Proteins, Vesicular Transport; D051242:Caveolin 1; D022461:Caveolins; D002966:Clathrin; D008565:Membrane Proteins; D014168:Transferrin; D015544:Inositol 1,4,5-Trisphosphate; D000255:Adenosine Triphosphate; D002118:Calcium; D003703:Demecolcine