Endothelin A and B receptors of preglomerular vascular smooth muscle cells.
BACKGROUND
The endothelin (ET) receptors are subclassified into ET(A,) which are purely vasoconstrictive, and ET(B). The ET(B) receptors may cause either vasodilation by stimulating the release of nitric oxide from endothelial cells, or vasoconstriction of vascular smooth muscle cells (VSMC). The relative contribution of ET(A) and ET(B) receptors to calcium signaling and vasoconstriction in the renal microcirculation is not clear. Our goal was to study the cytosolic calcium concentration ([Ca(2+)](i)) responses of fresh rat preglomerular VSMC and afferent arterioles to agonists and antagonists of ET(A) and ET(B) receptors in rats.


METHODS
Fresh VSMC and afferent arterioles were isolated using the magnetized microsphere/sieving technique, followed by gentle collagenase digestion. [Ca(2+)](i) was measured with fura-2 ratiometric fluorescence.


RESULTS
Afferent arterioles and VSMC responded to ET-1 stimulation with a rapid peak increase in [Ca(2+)](i) (Delta= 287 +/- 81 and 342 +/- 55 nmol/L, respectively). The ET(B) receptor agonist IRL 1620 stimulated a rise in [Ca(2+)](i) in afferent arterioles (106 +/- 35 nmol/L); subsequent addition of ET-1 at the IRL 1620 nadir to stimulate ET(A) receptors caused a second peak that was twice as large (213 +/- 44 nmol/L). In VSMC, the ET(B) agonist peak increase was 99 +/- 12 nmol/L; addition of ET-1 then increased [Ca(2+)](i) by 294 +/- 23 nmol/L. The ET(B) inhibitor BQ-788 prevented stimulation of [Ca(2+)](i) by IRL 1620 in afferent arterioles and VSMC; subsequent stimulation of ET(A) receptors with ET-1 caused an increase in [Ca(2+)](i) (239 +/- 17 and 248 +/- 22 nmol/L). Pretreatment with the selective ET(A) inhibitor PD 156707 attenuated but did not abolish the responses to ET-1, suggesting that the residual [Ca(2+)](i) response was caused by ET(B) stimulation.


CONCLUSIONS
These results indicate that fresh preglomerular VSMC as well as afferent arterioles have both ET(A) and ET(B) receptors, and that the rapid peak [Ca(2+)](i) responses to the ET(B) agonist IRL 1620 are less than half that of subsequent stimulation of ET(A) receptors with ET-1. The similarity of findings in isolated VSMC and afferent arterioles suggests that responses in VSMC in our arteriolar preparation overshadow any potential contribution of endothelial cells when reagents are administered abluminally.
Fellner
Susan K
SK
Department of Cell and Molecular Physiology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina 27599-7545, USA. sfellner@med.unc.edu
Arendshorst
William J
WJ
D000818:Animals; D001160:Arterioles / Q000187:drug effects / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling / Q000187:drug effects; D003600:Cytosol / Q000378:metabolism; D004149:Dioxoles / Q000494:pharmacology; D065130:Endothelin A Receptor Antagonists; D065131:Endothelin B Receptor Antagonists; D019332:Endothelin-1 / Q000494:pharmacology; D016232:Endothelins / Q000494:pharmacology; D066298:In Vitro Techniques; D007668:Kidney / Q000098:blood supply* / Q000378:metabolism*; D007678:Kidney Glomerulus / Q000098:blood supply / Q000378:metabolism; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D009842:Oligopeptides / Q000494:pharmacology; D010446:Peptide Fragments / Q000494:pharmacology; D010880:Piperidines / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D044022:Receptor, Endothelin A / Q000378:metabolism*; D044023:Receptor, Endothelin B / Q000819:agonists / Q000378:metabolism*
D004149:Dioxoles; D065130:Endothelin A Receptor Antagonists; D065131:Endothelin B Receptor Antagonists; D019332:Endothelin-1; D016232:Endothelins; D009842:Oligopeptides; C093321:PD 156707; D010446:Peptide Fragments; D010880:Piperidines; D044022:Receptor, Endothelin A; D044023:Receptor, Endothelin B; C074308:sovateltide; C086539:BQ 788; D002118:Calcium