Persistent tubular conduction in vacuolated amphibian skeletal muscle following osmotic shock.
The transverse (T-)tubules primarily function in conducting the action potentials that initiate excitation contraction coupling in skeletal muscle but may additionally subserve longer-term roles in volume regulation, membrane fusion and other trafficking processes. Osmotic shock thus both electrically detaches the T-tubules from surface membrane ('detubulation') and produces tubular vacuolation. The present experiments separated these effects. An established, reference osmotic shock protocol that exposed muscles to Ca2+/Mg2+-Ringer and gradual cooling to 10 degrees C after 18 min in glycerol-Ringer accomplished significant detubulation (77.5+/-13.15%, mean +/- SEM; n = 4). In contrast, a test protocol conducted entirely at room temperature using Mg2+-rather than Ca2+/ Mg2+-Ringer yielded reduced (P < 0.05, post hoc Duncan's multiple range test) detubulation indices (1.67+/-1.67%, n = 6) statistically indistinguishable from findings in fibres spared osmotic shock. Yet both osmotic shocks caused a formation of closed vacuoles, demonstrated by Sulphorhodamine B trapping, that occupied statistically similar fractions of total fibre volume (reference procedure: 14.38+/-2.7%, n = 6; test procedure: 13.36+/-2.00%, n = 22) in turn higher than determinations in control fibres (P < 0.05). The findings reconcile reports associating detubulation with vacuolation in osmotically shocked muscle [S. Nik-Zainal et al. (1999) J Muscle Res Cell Motil 20: 45-53; K.N. Khan et al. (2000) J Muscle Res Cell Motil 21: 79-90] with the persistence of tubular electrical activity in extensively vacuolated amphibian fibres following fatigue [J. Lannergren and H. Westerblad (1987) Acta Physiol Scand 129: 311-318; J. Lannergren et al. (1999) J Muscle Res Cell Motil 20: 19-32]. Furthermore test protocols produced higher densities of open vacuoles (13.38+/-2.33%, n = 9) than did reference protocols (6.66+/-1.63%, n = 20) contrary to their possible involvement in the electrophysiological changes. Abolition of tubular electrophysiological activity thus either follows or is independent of tubular vacuolation whilst sharing some of its underlying osmotic mechanisms.
Devlin
C M
CM
Physiological Laboratory, Multi-Imaging Centre, University of Cambridge, UK.
Chawl
S
S
Skepper
J N
JN
Huan
C L
CL
D000200:Action Potentials; D000818:Animals; D046508:Culture Techniques; D004553:Electric Conductivity; D005990:Glycerol / Q000494:pharmacology; D007552:Isotonic Solutions / Q000494:pharmacology; D018613:Microscopy, Confocal; D018485:Muscle Fibers, Skeletal / Q000187:drug effects / Q000502:physiology / Q000648:ultrastructure; D018482:Muscle, Skeletal / Q000502:physiology* / Q000648:ultrastructure*; D009997:Osmotic Pressure; D011896:Rana temporaria; D000077331:Ringer's Solution; D012519:Sarcoplasmic Reticulum / Q000502:physiology* / Q000648:ultrastructure*; D013696:Temperature; D013997:Time Factors; D014617:Vacuoles / Q000648:ultrastructure*
D007552:Isotonic Solutions; D000077331:Ringer's Solution; D005990:Glycerol