Effect of a 17 day spaceflight on contractile properties of human soleus muscle fibres

Abstract

1. Soleus biopsies were obtained from four male astronauts 45 days before and within 2 h after a 17 day spaceflight. 2. For all astronauts, single chemically skinned post-flight fibres expressing only type I myosin heavy chain (MHC) developed less average peak Ca2+ activated force (Po) during fixed-end contractions (0.78 +/- 0. 02 vs. 0.99 +/- 0.03 mN) and shortened at a greater mean velocity during unloaded contractions (Vo) (0.83 +/- 0.02 vs. 0.64 +/- 0.02 fibre lengths s-1) than pre-flight type I fibres. 3. The flight-induced decline in absolute Po was attributed to reductions in fibre diameter and/or Po per fibre cross-sectional area. Fibres from the astronaut who experienced the greatest relative loss of peak force also displayed a reduction in Ca2+ sensitivity. 4. The elevated Vo of the post-flight slow type I fibres could not be explained by alterations in myosin heavy or light chain composition. One alternative possibility is that the elevated Vo resulted from an increased myofilament lattice spacing. This hypothesis was supported by electron micrographic analysis demonstrating a reduction in thin filament density post-flight. 5. Post-flight fibres shortened at 30 % higher velocities than pre-flight fibres at external loads associated with peak power output. This increase in shortening velocity either reduced (2 astronauts) or prevented (2 astronauts) a post-flight loss in fibre absolute peak power (microN (fibre length) s-1). 6. The changes in soleus fibre diameter and function following spaceflight were similar to those observed after 17 days of bed rest. Although in-flight exercise countermeasures probably reduced the effects of microgravity, the results support the idea that ground-based bed rest can serve as a model of human spaceflight. 7. In conclusion, 17 days of spaceflight decreased force and increased shortening velocity of single Ca2+-activated muscle cells expressing type I MHC. The increase in shortening velocity greatly reduced the impact that impaired force production had on absolute peak power.

Figure 1. Superimposed force records of a maximally Ca²⁺-activated pre-flight (A) and post-flight (B) soleus fibre obtained immediately following slack steps of 200, 250, 300, 350 and 400 μm

The slack step caused force to drop to baseline after which the fibre shortened under no load. Unloaded shortening ended with the re-development of force. The force records have been superimposed to illustrate the longer duration of unloaded shortening following the longer slack steps. In C, the time required for the re-development of force has been plotted vs. the imposed slack step distance. The slope of the least-squares regression line, after normalization to the length of the fibre, defines V_o. In this example, the V_o of the pre-flight (•) and post-flight (○) fibre was 0.62 and 0.85 FL s⁻¹, respectively. Compliance, determined from the y axis intercepts, was < 3 % of fibre length for each fibre. Gel electrophoresis indicated that each fibre expressed only type I MHC.

Figure 2. Force-velocity relationship of a human pre-flight soleus fibre

Fibre length and force records obtained during a series of three isotonic contractions are illustrated in the upper right portion of the figure. Each isotonic step was 100 ms in duration. Total fibre shortening across all three steps was < 20 % of the initial fibre length. Force and shortening velocity were determined over the second half of each record with shortening velocity normalized to fibre length. The three isotonic steps illustrated in this figure represented loads of 7, 28 and 78 % of peak isometric force (P_o) and are denoted in the force-velocity curve by the filled symbols. In all, 15 force-velocity data points were obtained and fitted by the Hill equation (Hill, 1938) yielding a V_max (determined by extrapolation of the curve to a load of zero) of 0.44 FL s⁻¹ and a/P_o of 0.050 for this particular fibre. Subsequent gel electrophoresis indicated that this fibre expressed solely type I MHC.

Figure 3. Force-pCa relationship (A) and Hill plot analysis (B) of a single pre-flight soleus fibre

The fibre was activated in solutions with free Ca²⁺ concentrations ranging from pCa 6.8 to 4.7. The initial and final contraction and every 4th or 5th intervening contraction, was performed at a pCa of 4.5. In A, forces have been plotted relative to peak force at pCa 4.5, i.e. P_r= (force at sub-maximal activation)/(force at pCa 4.5). Hill plots were fitted to the data (B). Half-maximal activation, or pCa₅₀, was defined as the mean abscissal intercept of the Hill plots fitted to data above and below P_r= 0.5. The Ca²⁺ activation threshold, calculated from the plot of P_r < 0.5, was defined as the pCa where log(P_r/(1 - P_r)) equalled -2.5. For this particular fibre, activation threshold and half-maximal activation occurred at pCa values of 7.09 and 6.02, respectively. The slope of the force-calcium relationship was determined for forces greater (n₁) and less (n₂) than 50 % of the maximal Ca²⁺-activated force. Subsequent gel electrophoresis indicated that this fibre expressed type I MHC.

Figure 4. Representative SDS-polyacrylamide gels illustrating MHC and MLC expression in single human soleus fibres

Left: 5 % gel demonstrating separation of type I and IIa MHC in two single soleus fibres. Right: 12 % gel illustrating MLC composition of the same two single fibres. Fibre V_o was 1.09 FL s⁻¹ for the type I fibre in lane A and 4.09 FL s⁻¹ for the type IIa fibre in lane B.

Figure 5. Electron micrographs of longitudinal sections of soleus muscle fibres obtained from subject D before (upper photo) and after the flight (lower photo)

The paucity of sarcoplasmic reticulum membranes indicates that both fibres are slow fibres. Thin myofibrils and short Z lines (brackets) indicate atrophy of the post-flight fibre. The mitochondria and glycogen-like particles are similar in both fibres but the higher content of lipid droplets (arrows) is characteristic of post-flight muscle. Scale bars, 0.5 μm.

Figure 6. High magnification cross-sectional views of the thick and thin filament overlap region in the A-bands of a soleus muscle fibre biopsied before (upper photo) and after (lower figure) spaceflight (subject C)

Myofilament packing density is reduced post-flight because of a disproportionate loss of thin filaments. Scale bars, 0.1 μm.

Figure 7. Relationships between fibre diameter and peak Ca²⁺-activated isometric force (P_o)

Each symbol represents the results of a single soleus fibre. Type I fibres, shaded circles. Type IIa fibres, filled triangles. Type I/IIa fibres, open triangles.

Figure 8. Force-pCa relationships for type I fibres obtained before (• and continuous lines) and after (○ and dashed lines) spaceflight

Forces observed during sub-maximal activation (P_r) were expressed relative to force obtained at pCa 4.5. Symbols and error bars represent means ± 1 s.e.m. Total number of fibres studied were (pre, post): subject A: 15, 23; subject B: 10, 21; subject C: 14, 20; subject D: 13, 16. All fibres expressed type I MHC on 5 % gels.

Figure 9. V_o histograms of pre- and post-flight fibres from each subject

Pre-flight distributions are indicated by the shaded bars. Post-flight distributions are indicated by the hatched bars.

Figure 10. Force-velocity and force-power relationships of pre- and post-flight soleus fibres

Continuous lines represent composite pre-flight force-velocity and force-power relationships. Dashed lines represent composite post-flight force-velocity and force-power relationships.

Figure 11. Effects of 17 days of spaceflight or ground-based bed rest on the diameter and functional properties of single type I soleus fibres

The symbols represent the mean relative change (i.e. (post-flight mean value/pre-flight mean value) × 100 %) experienced by each individual astronaut (circles) and bed rest subject (triangles). Bars represent the mean (±s.e.m.) of the individual relative changes. Bed rest data obtained from Widrick et al. 1997a, . Responses of one bed rest subject have been omitted from this analysis (see Widrick et al. 1997a and for justification).