Mitochondrial Ca2+ homeostasis during Ca2+ influx and Ca2+ release in gastric myocytes from Bufo marinus.
1. The Ca(2+)-sensitive fluorescent indicator rhod-2 was used to monitor mitochondrial Ca2+ concentration ([Ca2+]m) in gastric smooth muscle cells from Bufo marinus. In some studies, fura-2 was used in combination with rhod-2, allowing simultaneous measurement of cytoplasmic Ca2+ concentration ([Ca2+]i) and [Ca2+]m, respectively. 2. During a short train of depolarizations, which causes Ca2+ influx from the extracellular medium, there was an increase in both [Ca2+]i and [Ca2+]m. The half-time (t1/2) to peak for the increase in [Ca2+]m was considerably longer than the t1/2 to peak for the increase in [Ca2+]i. [Ca2+]m remained elevated for tens of seconds after [Ca2+]i had returned to its resting value. 3. Stimulation with caffeine, which causes release of Ca2+ from the sarcoplasmic reticulum (SR), also produced increases in both [Ca2+]i and [Ca2+]m. The values of t1/2 to peak for the increase in [Ca2+] in both cytoplasm and mitochondria were similar; however, [Ca2+]i returned to baseline values much faster than [Ca2+]m. 4. Using a wide-field digital imaging microscope, changes in [Ca2+]m were monitored within individual mitochondria in situ, during stimulation of Ca2+ influx or Ca2+ release from the SR. 5. Mitochondrial Ca2+ uptake during depolarizing stimulation caused depolarization of the mitochondrial membrane potential. The mitochondrial membrane potential recovered considerably faster than the recovery of [Ca2+]m. 6. This study shows that Ca2+ influx from the extracellular medium and Ca2+ release from the SR are capable of increasing [Ca2+]m in smooth muscle cells. The efflux of Ca2+ from the mitochondria is a slow process and appears to be dependent upon the amount of Ca2+ in the SR.
Drummond
R M
RM
Department of Physiology, University of Massachusetts Medical Center, Worcester 01605, USA. robert.drummond@umassmed.edu
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Walsh
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D000818:Animals; D002024:Bufo marinus; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D003593:Cytoplasm / Q000378:metabolism; D004558:Electric Stimulation; D005456:Fluorescent Dyes / Q000493:pharmacokinetics; D006575:Heterocyclic Compounds, 3-Ring; D006706:Homeostasis / Q000502:physiology*; D007425:Intracellular Membranes / Q000187:drug effects / Q000378:metabolism; D008564:Membrane Potentials / Q000187:drug effects; D018715:Microscopy, Video; D008928:Mitochondria / Q000378:metabolism*; D009130:Muscle, Smooth / Q000166:cytology / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D012519:Sarcoplasmic Reticulum / Q000187:drug effects / Q000378:metabolism; D013270:Stomach
D005456:Fluorescent Dyes; D006575:Heterocyclic Compounds, 3-Ring; C068483:rhod-2; D002110:Caffeine; D002118:Calcium