Abstract:
LIB-stimulation has proved to be effective in maintaining and
restoring contraction force of fast rabbit muscle following denervation The
present study in white New Zealand rabbits was done to investigate the
influence of LIB-stimulation on the changes of contractile proteins and energy
metabolism following denervation.
Without stimulation, TA showed
a clear reduction of fibre diameter (19.9 µ) as compared to 41.1 µ after
stimulation; {Normal: 52.7 µ}. While, after denervation alone, no major changes
were found in fibre type distribution, stimulation induced a shift towards type
IIB (2.2 % I, 3.2 % IIA, 94.6 % IIB). - In SOL, fibre diameter was 23.5 µ
without and 34.3 µ with stimulation; {62.0 µ}. Without stimulation, 53.9 % type
I fibres were found, and 45.8 % after stimulation; {97.1 %}. In TA, denervation
alone induced no significant changes in MHC distribution (0 {3.2} % I, 14.8
{17.6} % IIA, 80.3 {79.2} % IIB, 4.9 % undefined isoforms). Following
stimulation, we found an increase of MHC IIB (96.8 %). - In SOL, type I
decreased after denervation (61.0 {94.8} %) as well as after stimulation (51.4
%). Following denervation, we found an overall decrease of enzymes, except for
hexokinase and G6PDH. In TA, stimulation induced a severalfold increase of
mitochondrial enzymes (citrate synthase 7x, cytochrome-c-oxidase 2-4x and ketoacid-CoA-
transferase 2x), while the glycolytic enzymes (phosphorylase,
phosphofructokinase) showed only small changes. In SOL, these enzymes increased
(6x, 4x), while the mitochondrial enzymes tended to decrease.
The present results show that electrical stimulation with long bidirectional rectangular balanced impulses (LIB-stimulation) has a clear beneficial effect on histochemical and biochemical features of denervated fast contracting skeletal muscle of rabbit, and only a small effect on the slow muscle. For the different reactions of small and fast muscles, stimulation parameters are thought to be responsive.
1.
Introduction
Background
In previous studies of our group (Ref. 1-3), electrical stimulation with
balanced bidirectional rectangular impulses of high intensity and long impulse
duration (LIB-stimulation) has proved to be effective in maintaining and
restoring muscle contraction force in fast muscles of rabbit. Additionally, the
morphological sequelae of denervation atrophy was stopped and muscle bulk was
restored.
LIB-Stimulation has also proved its efficacy in a reasonable number of
patients with complete and irreversible states of denervation following
destruction of brachial and/or lumbosacral plexus. Tetanic contraction force
was increased up to 400% and more, at maximum up to 30% of normal). Additionally,
it was shown that the histological signs of denervation atrophy could be
avoided and that muscle bulk even could be restored after a long time course of
denervation.
The response
of a fast skeletal rabbit muscle to different stimulus patterns, varying
frequency, impulse duration and total amount of stimulation, has shown some unexpected
findings before (same author, ifess 2002): A shorter impulse duration resulted
in a slower muscle than a longer impulse width (20 ms vs. 10 ms) three months
after denervation, following daily electrical stimulation for several minutes. Stimulation
with a higher frequency (50 Hz vs. 25 Hz) resulted in a slower contracting
muscle.
Additionally,
the stimulated muscle proved to be fast contracting but at the same time
fatigue resistent.
The question
was whether these unexpected findings of contractile properties correspond to morphological
and biochemical findings. The present study in white New Zealand rabbits was
done to investigate the influence of LIB-stimulation with variation of stimulus
patterns on the changes of contractile proteins and energy metabolism following
denervation.
1.1.
Previous
Work
Design/Methods
Animals/Denervation
Thirty adult
white New Zealand rabbits were denervated reaching a complete sensory-motor
loss of the right hindlimb by transsection of the sciatic nerve, femoral nerve,
obturator nerve and the lateral cutaneous nerve of the thigh. By means of
electrophysiology, care was taken that no reinnervation occurred during the
time of the overall experiment and the lack of reinnervation then was verified
in a final histological evaluation.
Electrical
stimulation
A painless
electrical stimulation was performed twice daily with a total stimulation time
of 9 minutes each, using surface electrodes over a period of three months. The
usual training regime included a tetanic contraction time of 30 seconds,
followed by a break of 2.5 minutes. The impulse was of a bidirectional
rectangular shape which was balanced in charge, with a duration of 20 ms,
followed by a break of the same length, from which resulted a frequency of 25
Hz.
Three months
after denervation, histochemical investigation of tibialis anterior muscle (TA)
and soleus muscle (SOL) included NADH-TR and ATPase stainings (following
preincubation at pH 4.3, 4.6 and 9.4). Biochemical investigation included
SDS-PAGE of myosin heavy chains and activities of mitochondrial (citrate synthase
{CS}, cytochrome-c-oxidase {CCO} and ß-oxidation of fatty acids:
ketoacid-CoA-transferase {KCT}) and glycolytic or glycogenolytic (phosphorylase
{PHO} and phosphofructokinase {PFK}) enzymes.
Results
Histochemical
findings
Without
stimulation, TA showed a clear reduction of fibre diameter (19.9 µ) as compared
to 41.1 µ after stimulation; {Normal: 52.7 µ}. While, after denervation alone,
no major changes were found in fibre type distribution, stimulation induced a
shift towards type IIB (2.2 % I, 3.2 % IIA, 94.6 % IIB). - In SOL, fibre
diameter was 23.5 µ without and 34.3 µ with stimulation; {62.0 µ}. Without
stimulation, 53.9 % type I fibres were found, and 45.8 % after stimulation;
{97.1 %}.
Biochemical findings
In TA, denervation alone induced no significant
changes in MHC distribution (0 {3.2} % I, 14.8 {17.6} % IIA, 80.3 {79.2} % IIB,
4.9 % undefined isoforms). Following stimulation, we found an increase of MHC
IIB (96.8 %). - In SOL, type I decreased after denervation (61.0 {94.8} %) as
well as after stimulation (51.4 %). See Fig. 1
Fig. 1:
Changes of myosin heavy chain in fast and slow rabbit muscle following
denervation and 3 months of stimulation with variation of stimulus parameters
Following denervation, we found an overall decrease of enzymes, except for hexokinase and G6PDH. In TA, stimulation induced a severalfold increase of mitochondrial enzymes (citrate synthase 7x, cytochrome-c-oxidase 2-4x and ketoacid-CoA- transferase 2x), while the glycolytic enzymes (phosphorylase, phosphofructokinase) showed only small changes. In SOL, these enzymes increased (6x, 4x), while the mitochondrial enzymes tended to decrease (Fig. 2)
Fig. 2: Changes of mitochondrial and glycolytic enzyme patterns in fast and slow rabbit muscle following denervation and different types of electrical stimulation
2. Summary and Conclusions
This type of
stimulation (LIB) has a clear beneficial effect on the fast contracting muscle,
and only a small effect on the slow muscle concerning contraction force. There
are, however, major changes in the enzyme patterns, different in fast and slow
muscles, that parralel the changes of contractile properties.. For the
different reactions of small and fast muscles, these variations of stimulation
parameters are thought to be responsible. The clinical meaning of these
findings might be to be able to chose whatever type of stimulation pattern to
influence the contractile patterns of the stimulated muscle. During therapeutic
stimulation, sometimes it might be of some clinical importance to be able to
chose a particular stimulation pattern to receive a more or less fast
contracting muscle (as there are, for example, fast contracting finger
muscles), or to have a more fatigue resistant muscle group. Further studies
have to show to what extent this choice can be realized.
Acknowledgment
The author wishes to acknoledge the financial support of the Deutsche Gesellschaft für Elektrostimulation und Elektrotherapie e.V. (GESET)
References
[1] K.F. Eichhorn, W. Schubert, E. David, Maintenance,
Training and Functional Use of Denervated Muscle, J Biomed Eng, Vol. 6, pp.
205-11, 1984
[2] T. Mokrusch, B. Neundörfer, Electrotherapy
of Permanently Denervated Muscle Long-Term Experience with a New Method, PMR,
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[3] T. Mokrusch, A. Engelhardt, K.F. Eichhorn, G. Prischenk, H. Prischenk, G. Sack, B. Neundörfer, Effects of Long Impulse Electrical Stimulation on Atrophy and Fibre Type Composition of Chronically Denervated Fast Rabbit Muscle, J Neurol, Vol. 237, pp. 29-34, 1990
[4] D. Pette, The Dynamic State of Muscle Fibers, De Gruyter, 1990