Preferential role of intracellular Ca2+ stores in regulation of isometric force in NIH 3T3 fibroblast fibres.
Fibroblast contraction plays a major role in wound repair, but the regulatory mechanisms are not well known. We investigated the relations between isometric force and intracellular calcium concentration ([Ca2+]i) in fibroblast fibres. These fibres were made with mouse NIH 3T3 fibroblasts cultured with native collagen in a three-dimensional matrix. Calf serum (CS; 30%) elicited a monotonic increase in force that attained a maximum within 15 min and could be sustained indefinitely. In contrast, [Ca2+]i increased to a peak at 3 min after CS stimulation, then returned to baseline levels by 10 min. Pretreatment with Ca2+-free medium or the Ca2+-channel antagonist nicardipine (10 microM) blocked the CS-induced [Ca2+]i increase, but force was not affected. KCl (50 mM) stimulation on the other hand, elicited a prolonged increase in [Ca2+]i but did not increase force. Inhibition of the endoplasmic reticulum Ca2+ release with Ca2+-ATPase inhibitors cyclopiazonic acid (5 microM) or thapsigargin (5 microM) nearly abolished (<20% control) the increase in [Ca2+]i and force response to CS. Treatment with ryanodine (10 microM) and caffeine (20 mM) had a similar effect. The phospholipase C inhibitor U73122 (3 microM) reduced the CS-induced increases in [Ca2+]i and force by 70 and 40%, respectively. We conclude that fibroblast isometric force is not coupled to Ca2+ arising from transmembrane influx but is correlated with the transient [Ca2+]i increase due to release from intracellular stores. Store-released Ca2+ may initiate activation pathways for fibroblast force development, but is not required for force maintenance.
Nobe
K
K
Department of Molecular and Cellular Physiology, University of Cincinnati College of Medicine, Cincinnati, OH 45267-0576, USA.
Nobe
H
H
Obara
K
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Paul
R J
RJ
D016475:3T3 Cells; D000818:Animals; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D002121:Calcium Channel Blockers / Q000494:pharmacology; D015220:Calcium Channels / Q000187:drug effects / Q000378:metabolism; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors; D002417:Cattle / Q000097:blood; D004357:Drug Synergism; D004721:Endoplasmic Reticulum / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004963:Estrenes / Q000494:pharmacology; D005347:Fibroblasts / Q000187:drug effects / Q000502:physiology*; D007211:Indoles / Q000494:pharmacology; D007425:Intracellular Membranes / Q000378:metabolism; D051379:Mice; D009529:Nicardipine / Q000494:pharmacology; D011189:Potassium Chloride / Q000494:pharmacology; D011760:Pyrrolidinones / Q000494:pharmacology; D012433:Ryanodine / Q000494:pharmacology; D019284:Thapsigargin / Q000494:pharmacology; D010738:Type C Phospholipases / Q000037:antagonists & inhibitors
D002121:Calcium Channel Blockers; D015220:Calcium Channels; D004791:Enzyme Inhibitors; D004963:Estrenes; D007211:Indoles; D011760:Pyrrolidinones; C060229:1-(6-((3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione; D012433:Ryanodine; D002110:Caffeine; D011189:Potassium Chloride; D019284:Thapsigargin; D009529:Nicardipine; D010738:Type C Phospholipases; D000252:Calcium-Transporting ATPases; D002118:Calcium; C000543:cyclopiazonic acid