title: Effect of sarcoplasmic reticulum Ca2+ content on action potential-induced Ca2+ release in rat skeletal muscle fibres.
abstract: This study examined the relationship between the level of Ca2+ loading in the sarcoplasmic reticulum (SR) and the amount of Ca2+ released by an action potential (AP) in fast-twitch skeletal muscle fibres of the rat. Single muscle fibres were mechanically skinned and electric field stimulation was used to induce an AP in the transverse-tubular system and a resulting twitch response. Responses were elicited in the presence of known amounts (0-0.38 mM) of BAPTA, a fast Ca2+ buffer, with the SR Ca2+ pump either functional or blocked by 50 microM 2,5-di-tert-butyl-1,4-hydroquinone (TBQ). When Ca2+ reuptake was blocked, an estimate of the amount of Ca2+ released by an AP could be derived from the size of the force response. In a fibre with the SR loaded with Ca2+ at the endogenous level (approximately 1.2 mM, expressed as total Ca2+ per litre fibre volume; approximately one-third of maximal loading), a single AP triggered the release of approximately 230 microM Ca2+. If a second AP was elicited 10 ms after the first, only a further approximately 60 microM Ca2+ was released, the reduction probably being due to Ca2+ inactivation of Ca2+ release. When Ca2+ reuptake was blocked, APs applied 15 s apart elicited similar amounts of Ca2+ release (approximately 230 microM) on the first two or three occasions and then progressively less Ca2+ was released until the SR was fully depleted after a total of approximately eight APs. When the SR was loaded to near-maximal capacity (approximately 3-4 mM), each AP (or pair of APs 10 ms apart) still only released approximately the same amount of Ca2+ as that released when the fibre was endogenously loaded. Consistent with this, successive APs (15 s apart) elicited similar amounts of Ca2+ release approximately 10-16 times before the amount released declined, and the SR was fully depleted of Ca2+ after a total release calculated to be approximately 3-4 mM. When the SR was loaded maximally, increasing the [BAPTA] above 280 microM resulted in an increase in the amount of Ca2+ released per AP, probably because the greater level of cytoplasmic Ca2+ buffering prevented Ca2+ inactivation from adequately limiting Ca2+ release. These results show that the amount of Ca2+ released by AP stimulation in rat fast-twitch fibres normally stays virtually constant over a wide range of SR Ca2+ content, in spite of the likely large change in the electrochemical gradient for Ca2+. This was also found to be the case in toad twitch fibres. This constancy in Ca2+ release should help ensure precise regulation of force production in fast-twitch muscle in a range of circumstances.
authors.affiliation: Department of Zoology, La Trobe University, Melbourne, Victoria 3086, Australia.
authors.forename: G S
authors.initials: GS
authors.lastname: Posterino
authors.forename: G D
authors.initials: GD
authors.lastname: Lamb
mesh_terms: D000200:Action Potentials / Q000502:physiology; D000818:Animals; D001001:Anura; D002021:Buffers; D002118:Calcium / Q000378:metabolism*; D002614:Chelating Agents / Q000494:pharmacology; D003593:Cytoplasm; D004533:Egtazic Acid / Q000031:analogs & derivatives* / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006873:Hydroquinones / Q000494:pharmacology; D066298:In Vitro Techniques; D008297:Male; D009119:Muscle Contraction / Q000502:physiology; D018485:Muscle Fibers, Skeletal / Q000378:metabolism*; D018482:Muscle, Skeletal / Q000378:metabolism*; D051381:Rats; D020318:Rats, Long-Evans; D012519:Sarcoplasmic Reticulum / Q000378:metabolism*
chemical_list: D002021:Buffers; D002614:Chelating Agents; D004791:Enzyme Inhibitors; D006873:Hydroquinones; C019359:2,5-di-tert-butylhydroquinone; D004533:Egtazic Acid; C025603:1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid; D002118:Calcium