title: Mitochondrial and myoplasmic [Ca2+] in single fibres from mouse limb muscles during repeated tetanic contractions.
abstract: Previous studies on single fast-twitch fibres from mouse toe muscles have shown marked fatigue-induced changes in the free myoplasmic [Ca2+] ([Ca2+]i), while mitochondrial [Ca2+] remained unchanged. We have now investigated whether muscle fibres from the legs of mice respond in a similar way. Intact, single fibres were dissected from the soleus and extensor digitorum longus (EDL) muscles of adult mice. To measure [Ca2+]i, indo-1 was injected into the isolated fibres. Mitochondrial [Ca2+] was measured using Rhod-2 and confocal laser microscopy. Fatigue was induced by up to 1000 tetanic contractions (70 Hz) given at 2 s intervals. In soleus fibres, there was no significant decrease in tetanic [Ca2+]i at the end of the fatiguing stimulation, whereas tetanic force was significantly reduced by about 30 %. In 10 out of 14 soleus fibres loaded with Rhod-2 and subjected to fatigue, mitochondrial [Ca2+] increased to a maximum after about 50 tetani; this increase was fully reversed within 20 min after the end of stimulation. The force-frequency curve of the non-responding soleus fibres was shifted to higher frequencies compared to that of the responding fibres. In addition, eight out of nine Rhod-2-loaded EDL fibres showed similar changes in mitochondrial [Ca2+] during and after a period of fatiguing stimulation. The stimulation-induced increase in mitochondrial [Ca2+] was reduced when mitochondria were depolarised by application of carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone, whereas it was increased by application of an inhibitor of the mitochondrial Na+/Ca2+ exchange (CGP-37157). In conclusion, isolated slow-twitch muscle fibres show only modest changes in tetanic force and [Ca2+]i during repeated contractions. The increase in mitochondrial Ca2+ does not appear to be essential for activation of mitochondrial ATP production, nor does it cause muscle damage.
authors.affiliation: Department of Physiology and Pharmacology, Karolinska Institutet, 171 77 Stockholm, Sweden. joseph.bruton@fyfa.ki.se
authors.forename: Joseph
authors.initials: J
authors.lastname: Bruton
authors.forename: Pasi
authors.initials: P
authors.lastname: Tavi
authors.forename: Jan
authors.initials: J
authors.lastname: Aydin
authors.forename: Håkan
authors.initials: H
authors.lastname: Westerblad
authors.forename: Jan
authors.initials: J
authors.lastname: Lännergren
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone / Q000494:pharmacology; D002998:Clonazepam / Q000031:analogs & derivatives* / Q000494:pharmacology; D004558:Electric Stimulation; D005121:Extremities; D005456:Fluorescent Dyes; D006575:Heterocyclic Compounds, 3-Ring; D007425:Intracellular Membranes / Q000378:metabolism; D007476:Ionophores / Q000494:pharmacology; D008297:Male; D051379:Mice; D008815:Mice, Inbred Strains; D018613:Microscopy, Confocal; D008931:Mitochondria, Muscle / Q000378:metabolism*; D009119:Muscle Contraction / Q000502:physiology*; D018763:Muscle Fatigue / Q000502:physiology; D018657:Muscle Fibers, Slow-Twitch / Q000378:metabolism*; D018482:Muscle, Skeletal / Q000378:metabolism / Q000502:physiology*; D009994:Osmolar Concentration; D013841:Thiazepines / Q000494:pharmacology
chemical_list: D005456:Fluorescent Dyes; D006575:Heterocyclic Compounds, 3-Ring; D007476:Ionophores; D013841:Thiazepines; C068483:rhod-2; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; D002998:Clonazepam; C079032:CGP 37157; D002118:Calcium