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Influence of vasoactive agents on cytoplasmic free calcium in vascular endothelial cells
U S Ryan1, P V Avdonin, E Y Posin
1Department of Medicine, University of Miami School of Medicine, Florida 33101.
Insights
Endothelial cells (EC) showed increased cytoplasmic free calcium concentration ([Ca2+]i) in response to histamine and thrombin. Histamine primarily mobilizes intracellular calcium stores via H1-receptors, with protein kinase C inhibiting this response.
Area of Science:
- Cellular Biology
- Physiology
- Pharmacology
Background:
- Cytoplasmic free calcium concentration ([Ca2+]i) is crucial for endothelial cell (EC) function.
- Understanding the regulation of [Ca2+]i is vital for various physiological processes.
Purpose of the Study:
- To investigate the regulation of [Ca2+]i in EC from different human and bovine blood vessels.
- To elucidate the receptor-mediated mechanisms underlying [Ca2+]i increases.
Main Methods:
- Utilized the fluorescent calcium indicator indo-1 to measure [Ca2+]i.
- Stimulated EC with various agents including histamine, thrombin, ATP, bradykinin, and platelet-activating factor (PAF).
- Investigated the role of extracellular calcium, intracellular signaling pathways (cAMP, cGMP), and protein kinase C (PKC).
Main Results:
- Histamine and thrombin induced rapid [Ca2+]i elevation in human EC.
- ATP and bradykinin affected aortic EC, while PAF and thrombin affected bovine pulmonary artery EC.
- Histamine's effect was H1-receptor mediated and primarily involved mobilization of intracellular calcium stores, independent of extracellular calcium.
- Elevating cAMP or cGMP levels did not affect histamine-induced [Ca2+]i increase.
- Phorbol myristate acetate (PMA), a PKC activator, strongly inhibited the histamine-induced [Ca2+]i elevation.
Conclusions:
- Histamine triggers a rapid increase in EC [Ca2+]i mainly through intracellular calcium release via H1-receptors.
- Protein kinase C activation appears to mediate a negative feedback mechanism inhibiting receptor-mediated [Ca2+]i elevation in EC.
- These findings contribute to understanding calcium signaling in endothelial cells and its modulation.
Abstract:
The regulation of cytoplasmic free calcium concentration [( Ca2+]i) in endothelial cells (EC) derived from human umbilical vein, aorta, and pulmonary artery, or from bovine pulmonary artery, was studied by means of the fluorescent Ca2+ indicator indo-1. Histamine and thrombin caused a rapid transient elevation in [Ca2+]i in the EC of all the human blood vessels tested. In aortic EC, [Ca2+]i also rose in response to ATP and bradykinin. It was shown that in bovine pulmonary artery EC [Ca2+]i rises in response to platelet-activating factor (PAF) and thrombin. For a more detailed investigation of the receptor-mediated mechanism of [Ca2+]i increase in EC we used histamine as a stimulating agent. Histamine effects were seen at concentrations ranging from 5 X 10(-7) to 10(-4) M [50% effective dose (ED50) approximately 2-4 microM)] and were mediated by H1-receptors. The histamine-induced increase in [Ca2+]i was not markedly diminished when the extracellular calcium was bound by excess ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA). The data obtained indicate that the histamine effect is best explained by Ca2+ mobilization from intracellular stores. The histamine-induced increase in [Ca2+]i was not influenced by elevating the intracellular levels of adenosine 3',5'-cyclic monophosphate (cAMP) or cyclic guanylic acid (cGMP) by use of isobutylmethylxanthine and forskolin or by nitroprusside preincubation, respectively. However, the protein kinase C stimulator, phorbol myristate acetate (PMA), strongly inhibits [Ca2+]i elevation. It is assumed that a negative feedback mechanism that blocks receptor-mediated [Ca2+]i increase is triggered as a result of the activation of protein kinase C.