Prevention of a hypoxic Ca(2+)(i) response by SERCA inhibitors in cerebral arterioles.
1. The aim of the study was to investigate the mechanism of a novel effect of hypoxia on intracellular Ca(2+) signalling in rabbit cerebral arteriolar smooth muscle cells, an effect that was resistant to the L-type Ca(2+) channel antagonist methoxyverapamil (D600). 2.[Ca(2+)](i) of smooth muscle cells in intact arteriolar fragments was measured using the Ca(2+)-indicator dye fura-PE3. Hypoxia (PO(2) 10 - 20 mmHg) lowered basal [Ca(2+)](i) but did not inhibit Ca(2+) entry pathways measured by Mn(2+)-quenching of fura-PE3. 3. The effect of hypoxia was completely prevented by thapsigargin or cyclopiazonic acid, selective inhibitors of sarcoplasmic reticulum Ca(2+) ATPase (SERCA). Since these inhibitors do not block Ca(2+) extrusion or uptake via the plasma membrane, the data indicate that the effect of hypoxia depends on a functional sarcoplasmic reticulum. 4. Because actions of nitric oxide (NO) on vascular smooth muscle are also prevented by SERCA inhibitors it was explored whether the effect of hypoxia occurred via modulation of endogenous NO release. Residual NOS-I and NOS-III were detected by immunostaining, and there were NO-dependent effects of NOS inhibitors on Ca(2+)(i)-signalling. Nevertheless, inhibition of endogenous NO production did not prevent the effect of hypoxia on [Ca(2+)](i). 5. The experiments reveal a novel nitric oxide-independent effect of hypoxia that is prevented by SERCA inhibitors.
Guibert
C
C
School of Biomedical Sciences, Worsley Building, University of Leeds, UK.
Flemming
R
R
Beech
D J
DJ
D000818:Animals; D001160:Arterioles; D001921:Brain / Q000098:blood supply*; D020746:Calcium Channels, L-Type / Q000187:drug effects / Q000502:physiology; D020013:Calcium Signaling / Q000187:drug effects*; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors*; D015687:Cell Hypoxia; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005455:Fluorescent Antibody Technique; D066298:In Vitro Techniques; D007211:Indoles / Q000494:pharmacology; D007424:Intracellular Fluid / Q000378:metabolism; D015640:Ion Channel Gating; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D009569:Nitric Oxide / Q000096:biosynthesis; D019001:Nitric Oxide Synthase / Q000378:metabolism; D052248:Nitric Oxide Synthase Type I; D052250:Nitric Oxide Synthase Type III; D011817:Rabbits; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D019284:Thapsigargin / Q000494:pharmacology
D020746:Calcium Channels, L-Type; D004791:Enzyme Inhibitors; D007211:Indoles; D009569:Nitric Oxide; D019284:Thapsigargin; D019001:Nitric Oxide Synthase; D052248:Nitric Oxide Synthase Type I; D052250:Nitric Oxide Synthase Type III; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D000252:Calcium-Transporting ATPases; C000543:cyclopiazonic acid