Crossbridge properties during force enhancement by slow stretching in single intact frog muscle fibres

Abstract

The mechanism of force enhancement during lengthening was investigated on single frog muscle fibres by using fast stretches to measure the rupture tension of the crossbridge ensemble. Fast stretches were applied to one end of the activated fibre and force responses were measured at the other. Sarcomere length was measured by a striation follower device. Fast stretching induced a linear increase of tension that reached a peak and fell before the end of the stretch indicating that a sudden increase of fibre compliance occurred due to forced crossbridge detachment induced by the fast loading. The peak tension (critical tension, Pc) and the sarcomere length needed to reach Pc (critical length, Lc) were measured at various tensions during the isometric tetanus rise and during force enhancement by slow lengthening. The data showed that Pc was proportional to the tension generated by the fibre under both isometric and slow lengthening conditions. However, for a given tension increase, Pc was 6.5 times greater during isometric than during lengthening conditions. Isometric critical length was 13.04 +/- 0.17 nm per half-sarcomere (nm hs(-1)) independently of tension. During slow lengthening critical length fell as the force enhancement increased. For 90% enhancement, Lc reduced to 8.19 +/- 0.039 nm hs(-1). Assuming that the rupture force of the individual crossbridge is constant, these data indicate that the increase of crossbridge number during lengthening accounts for only 15.4% of the total force enhancement. The remaining 84.6% is accounted for by the increased mean strain of the crossbridges.

Figure 1. Force and sarcomere length responses induced by a fast stretch applied at tension of about 0.5 P₀ on the tetanus rise (A) and during slow stretching (B)

The stretch (17.5 nm hs⁻¹ amplitude, 950 μs duration, corresponding to 17.54 l₀ s⁻¹) in B was applied at the same relative tension as in A but during slow lengthening (at 0.75 l₀ s⁻¹), which increased the tension by 20% with respect to the isometric tension. Upper traces: sarcomere length; lower traces: force. Note that the sarcomere elongation needed to reach the crossbridge rupture, is smaller in B than in A. Interruptions on the traces indicate the change from slow to fast time base. The arrow in B shows the start of slow ramp stretching. Vertical dashed line shows the critical tension and the corresponding critical length.

Figure 2. Force and sarcomere length responses induced by fast stretches applied during the tetanus rise with and without slow lengthening

Upper traces: sarcomere length; lower traces: force. A, force responses at two different isometric force levels (0.2 and 0.5 P₀; thin and thick line, respectively); B, force responses at 0.2 P₀, during isometric (thin line) and at 0.5 P₀ during lengthening (thick line). Lengthening velocity, 1.65 l₀ s⁻¹. Force enhancement, 150%. In A the timing of stimulation is adjusted so that the isometric tension reaches 0.2 P₀ or 0.5 P₀ at the time of the fast stretch. In B the tension at 0.5 P₀ is obtained by slow lengthening of the fibre before the fast stretch. The arrow on the sarcomere length trace in B shows the start of slow lengthening. Note that for the same tension, P_c, is much greater during the isometric rise than during slow lengthening.

Figure 3. Relative critical tension against relative tension under isometric (filled symbols) or during lengthening (open and half-filled symbols) in two fibres

Fast stretches were applied at various times during the tetanus rise (isometric data) and at time when the isometric tension was 0.29 and 0.40 P₀ (in one fibre, squares), and 0.57 and 0.83 P₀ (in the other fibre, circles), preceded by slow lengthening at speeds between 0.08 and 2.4 l₀ s⁻¹, which induced various amount of force enhancement (lengthening data). The arrow shows the force (P_in) at the intercept between slow lengthening and isometric values for one set of data.

Figure 4. Normalized relations between critical tension and tension developed under isometric (filled symbols) and slow stretching conditions (open symbols)

Pooled data from 6 fibres. Each point represents the mean value ± s.e.m. for 0.25 P/P_in class averaging. The continuous lines fitted to the filled and open symbols represent the best fittings. The star represents the mean value ± s.e.m. from 5 experiments made at the tetanus plateau under condition of steady force enhancement (see later).

Figure 5. Force and sarcomere length responses to slow and fast stretches applied at tetanus plateau

A, force response to slow stretching (0.29 l₀ s⁻¹) which enhances the force towards an almost steady value corresponding to 1.6 P₀. B, force response to a fast stretch (29.75 l₀ s⁻¹) applied during the steady force enhancement. It can be seen that the shape of the force response is the same as during the tetanus rise. P_c during force enhancement is 1.95 times the tension developed at the time of the stretch. The average of the results obtained in all fibres at plateau is plotted in Fig. 4 as a single data point. The arrows show the start of slow stretching. The interruptions on the traces indicate the switch from slow to fast time base. Vertical dashed line shows the critical tension and the corresponding critical length.

Figure 6. Critical length as a function of tension under isometric and slow lengthening conditions

Pooled data from the same fibres as Fig. 4. Each point represents the mean ± s.e.m. for 0.25 P/P_in class averaging. Tensions are expressed relatively to P_in. Data on the isometric tetanus rise (filled symbols) show that L_c is independent of tension. On the contrary, during lengthening (open symbols), L_c decreases as the force enhancement increases. The averaged data, (obtained at plateau during steady force enhancement, represented with the star, show no significant difference from other data.