Incomplete rematching of nerve and muscle properties in motor units after extensive nerve injuries in cat hindlimb muscle

Abstract

1. Motor units were characterized in partially denervated or completely denervated and reinnervated cat medial gastrocnemius (MG) muscles where the number of innervating motor axons was severely reduced to determine (1) to what extent the nerve and muscle properties are rematched in enlarged motor units, (2) whether the normal size relationships between axon size, unit tetanic force and contractile speed are re-established, and (3) whether the type of nerve injury and/or repair affects the re-establishment of nerve and muscle properties. 2. Single MG units were sampled in (1) partially denervated muscles and in reinnervated muscles after either (2) crushing or (3) transecting the nerve and suturing its proximal end to either the distal nerve stump (N-N), or (4) directly to the muscle fascia (N-M). 3. The majority (75-88 %) of motor units in all muscles were classified as S (slow), FR (fast fatigue resistant), FI (fast fatigue intermediate) and FF (fast fatigable). However, there was an increased number of FI and unclassifiable motor units compared to normal. These results suggest that motor unit properties are not entirely regulated by the reinnervating motoneurone. 4. Despite more overlap in the range of unit force between different motor unit types the tetanic force of each type increased in all muscles when reinnervated by few (< 50 %) motor axons. This increase in unit force was due to an expansion in motor unit innervation ratio. 5. The normal relationships between axon size, unit tetanic force, and contractile speed were re-established in all muscles except when reinnervated by < 50 % of their normal complement of motor units after N-M suture. This lack of correlation was due to the reduced fast glycolytic (FG) fibre size and the proportionately greater increase in force of the S units. 6. After reinnervation the ranges in fibre cross-sectional area within single FF units were very similar to those found within the entire FG fibre population. 7. These results show that when few axons make functional connections in partially denervated or reinnervated muscles the normal relationships between axon size and motor unit contractile properties are re-established provided the nerves regenerate within the distal nerve sheath. This rematching of motoneurone size and motor unit contractile properties occurs primarily because the size of the motor axon governs the number of muscle fibres it supplies.

Figure 1. Single motor unit contractile properties in normal, partially denervated, or reinnervated MG muscle

MG motor units in 1 normal (A), 3 partially denervated (PD; B), and 2 reinnervated muscles after MG nerve crush (C) plotted as a function of twitch contraction time, fatigue index and tetanic force on 3-dimensional graphs. •, motor units that did not ‘sag’ during unfused tetanic contractions (type S units); ○, motor units that ‘sagged’ during unfused tetanic contractions (type F units). Partially denervated (B) and reinnervated (C) muscles contain ≈20 % of their original complement of motor units. Despite a continuous distribution of unit twitch contraction times and tetanic force, S units in normal muscles show little overlap with F units with respect to these contractile properties. The distribution of motor units is similar to normal in partially denervated and reinnervated muscles after nerve crush apart from a few large motor units that show atypical ‘no sag’ characteristics with fast contraction times and/or low fatigue indices.

Figure 2. Distribution of motor unit tetanic forces in normal and reinnervated muscles

Frequency histograms of tetanic force developed by FF, FI, FR and S units sampled in 2 normal and 2 partially denervated (PD) muscles innervated by ≈20 % (mean ± s.e.m., 19.8 ± 3.7 %) of its normal complement of motor units. The force of all 4 motor unit types increased significantly in partially denervated muscles, but the range remained the same. Mean values are indicated by the vertical line to show that S < FR < FI = FF in normal and partially denervated muscles.

Figure 3. Distribution of unit twitch contraction times in normal and reinnervated muscles

Frequency histograms of unit twitch contraction times (ms) in normal (A), partially denervated (B) and reinnervated muscles after nerve crush (C), nerve transection with N-N (D) or N-M sutures (E). Mean ( ± s.e.m.) number (%) of motor units in each experimental condition are: 73 ± 4.7 % (B),76 ± 6.9 % (C), 77 ± 5.2 % (D) and 80 ± 5.1 % (E). Motor units were sampled from 6, 10, 7, 9 and 7 cats in A, B, C, D and E, respectively. The mean and range in unit twitch contraction times are similar to normal in B-E. Means are shown by vertical lines and are ( ± s.e.m.; in ms): 42.2 ± 1.2 (A), 42 ± 2 (B), 41.6 ± 1.7 (C),42.4 ± 1.6 (D) and 46.3 ± 1.8 (E).

Figure 4. Distribution of motor unit half-fall times in normal and reinnervated muscles

Frequency histograms of motor unit half-fall times in normal (A), partially denervated (B) and reinnervated muscles after nerve crush (C), N-N (D) or N-M sutures (E). Motor units were sampled from the same muscles as in Fig. 3. Means are shown by vertical lines and are ( ± s.e.m.; in ms): 30.2 ± 2.1 (A), 38.2 ± 2.1 (B), 39.6 ± 3.3 (C), 40.7 ± 2.3 (D) and 42.7 ± 2.1 (E).

Figure 5. Distribution of motor unit fatigue index in normal and reinnervated muscles

Frequency histograms of motor unit fatigue index sampled from normal (A), partially denervated (B) and reinnervated muscles after nerve crush (C), N-N (D) or N-M sutures (E). The distribution of fatigue index values is bimodally distributed in normal, partially denervated and reinnervated muscles after nerve crush, but is less discretely divided into 2 modes in muscles reinnervated by completely transected nerves (i.e. N-N and N-M sutures). Mean fatigue index is indicated by a vertical line.

Figure 6. Single motor unit twitch and unfused tetanic contractions

An example of a single fast motor unit (contraction time, 31 ms), sampled from a normal muscle (A) showing a typical ‘sag’ response during an unfused tetanic contraction (B). An example of a single fast contracting motor unit (contraction time, 28 ms), sampled from a reinnervated muscle after N-M suture (C) showing an atypical ‘no sag’ response (D). These atypical fast contracting motor units cannot be classified as fast or slow on the basis of ‘sag’ of unfused tetanic contraction and speed of contraction. They are therefore considered ‘unclassifiable’.

Figure 7. Motor unit contractile properties in reinnervated muscles

Motor units sampled from reinnervated muscles after complete MG nerve transection and repair with either N-N (A) or N-M sutures (B) plotted as in Fig. 1. In each case, motor units were sampled from 2 muscles reinnervated by 36 ± 2.8 % (A) and 41 ± 6.7 % (B) of their normal complement of motor units. Unit force increased in reinnervated muscles after N-N, but not after N-M suture (note z-axis is 2 times greater in A than B). Generally, the distribution of motor units in reinnervated muscles is similar to normal with the exception of a marked increase in FI units (fatigue index < 0.25 < 0.75) and a number of atypical motor units that did not ‘sag’ but had relatively fast contraction times (< 40 ms) or low fatigue indices (< 0.75).

Figure 8. Distribution of unit tetanic force from reinnervated muscles reinnervated by few motoneurones

Frequency histogram of tetanic force developed by each of FF, FI, FR and S units in muscles reinnervated by ≈25 % of their normal complement of motor units after complete nerve transection and repair with N-N or N-M sutures. The force of the S units increased by a greater extent than the F units. Mean values are indicated by the vertical line to show that S < FR < FI = FF in normal and in reinnervated muscles after N-N and N-M suture.

Figure 9. Relationship between axon potential amplitude and twitch contraction time with unit tetanic force

Axon potential amplitude and twitch contraction time plotted as a function of tetanic force in motor units from normal (A and B), partially denervated muscle innervated by 25 % of its motor units (C and D), and muscles reinnervated after nerve crush by 25 % of its motor units (E and F). ○, S units; ▪, FR units; ▵, FI and FF units;, unclassifiable units. The slopes ( ± s.e.) of the regression lines for axon potential amplitude and tetanic force (0.12 ± 0.02, 0.15 ± 0.03 and 0.19 ± 0.06; r = 0.60, 0.83 and 0.53, respectively) are all significantly different from zero (P < 0.01-0.05) and not different from each other. The negative slopes of the regression lines for contraction time and tetanic force (0.14 ± 0.02, 0.16 ± 0.05 and 0.28 ± 0.07; r = 0.66, 0.68 and 0.70, respectively) are also significantly different from zero. Slopes of B and D are not different from each other; however, the slope in F is significantly steeper.

Figure 10. Ranges in cross-sectional area (CSA) of each muscle fibre type in normal and reinnervated muscles

Ranges in CSA of each muscle fibre type in normal, partially denervated (PD) and reinnervated muscles after nerve crush or nerve transection with N-N or N-M sutures. All experimental muscles were reinnervated by ≈25 % of their normal complement of motor units. Mean ( ± s.e.m.) CSAs (vertical bars) for FG (fast glycolytic), FOG (fast oxidative glycolytic) and SO (slow oxidative) fibres, respectively, are (10³μm²): 6.2 ± 0.1, 2.8 ± 0.07 and 2.8 ± 0.06 (Normal); 4.6 ± 0.06, 3.2 ± 0.05 and 3.3 ± 0.08 (Crush); 3.5 ± 0.14, 2.7 ± 1.1 and 2.5 ± 0.07 (N-N); 3.6 ± 0.04, 2.8 ± 0.05 and 3.0 ± 0.04 (N-M).

Figure 11. Size relationship between axon potential amplitude and unit tetanic force when reinnervated by few or many motoneurones

Axon potential amplitude plotted as a function of tetanic force in motor units from muscles reinnervated by > 50 % and < 50 % of their normal complement of motor units after N-N (A and B, respectively) or N-M sutures (C and D, respectively). Unit types denoted by same symbols as in Fig. 9. Muscles in A, B, C and D were reinnervated by 88, 32, 88 and 31 % of their motor units, respectively. The slopes ( ± s.e.) of the regression lines in A-C(0.13 ± 0.03, 0.13 ± 0.03 and 0.12 ± 0.02; r = 0.63, 0.58 and 0.67, respectively) are all significantly different from zero (P < 0.01-0.05) and not different from each other or from normal (Fig. 9A). The regression line fitted to the values in D is not significantly different from zero (P < 0.05) and therefore has not been drawn.

Figure 12. Ranges in cross-sectional area (CSA) from single motor units in normal and reinnervated muscles

Ranges in fibre CSA from glycogen-depleted muscle unit fibres (□) and non-depleted fibres of the same histochemical types (τ) for 5 normal (3 FF units; 2 S units) and 10 reinnervated motor units (6 N-N; 4 N-M). FG, fast-glycolytic; SO, slow oxidative.