Fibre-related nitric oxide synthase (NOS) in Duchenne muscular dystrophy.

Nitric oxide (NO) mediates fundamental physiological actions on skeletal muscle. The loss of NO synthase (NOS) from the sarcolemma was assumed to be associated with development of Duchenne muscular dystrophy (DMD). We have, however, recently reported that, in contrast to the commonly accepted view, NOS expression in DMD myofibres is up-regulated. This poses the question of the fibre type-specific NOS expression in DMD muscles and how the NOS expression is related to the regeneration or degeneration status. To address this issue, we examined localization of NOS isoforms I, II and III in skeletal muscles of DMD patients employing immunohistochemical labelling with tyramide signal amplification complemented with enzyme histochemistry. We found that NOS immunolabelling as well as metabolic enzyme activity in DMD muscles were heterogeneously distributed along the fibre length of DMD muscle fibres revealing regenerating and degenerate (hypercontracted) fibres as well as normal segments. Like in normal muscles, positive NOS immunoreactivity was found to be associated with fast-oxidative glycolytic (FOG) phenotype. The regeneration status of NOS-positive segments was deduced from the presence of neonatal and developmental myosin heavy chains. High NOS expression in regenerating DMD muscle fibres can be well reconciled with reports about the protective role of endogenous NO in inflammatory diseases and in muscle repair.

Effects of caffeine at different temperatures on contractile properties of slow-twitch and fast-twitch rat muscles.

The slow-twitch soleus muscle (SOL) exhibits decreased twitch tension (cold depression) in response to a decreased temperature, whereas the fast-twitch extensor digitorum longus (EDL) muscle shows enhanced twitch tension (cold potentiation). On the other hand, the slow-twitch SOL muscle is more sensitive to twitch potentiation and contractures evoked by caffeine than the fast-twitch EDL muscle. In order to reveal the effects of these counteracting conditions (temperature and caffeine), we have studied the combined effects of temperature changes on the potentiation effects of caffeine in modulating muscle contractions and contractures in both muscles. Isolated muscles, bathed in a Tyrode solution containing 0.1-60 mM caffeine, were stimulated directly and isometric single twitches, fused tetanic contractions and contractures were recorded at 35 degrees C and 20 degrees C. Our results showed that twitches and tetani of both SOL and EDL were potentiated and prolonged in the presence of 0.3-10 mM caffeine. Despite the cold depression, the extent of potentiation of the twitch tension by caffeine in the SOL muscle at 20 degrees C was by 10-15 % higher than that at 35 degrees C, while no significant difference was noted in the EDL muscle between both temperatures. Since the increase of twitch tension was significantly higher than potentiation of tetani in both muscles, the twitch-tetanus ratio was enhanced. Higher concentrations of caffeine induced contractures in both muscles; the contracture threshold was, however, lower in the SOL than in the EDL muscle at both temperatures. Furthermore, the maximal tension was achieved at lower caffeine concentrations in the SOL muscle at both 35 degrees C and 20 degrees C compared to the EDL muscle. These effects of caffeine were rapidly and completely reversed in both muscles when the test solution was replaced by the Tyrode solution. The results have indicated that the potentiation effect of caffeine is both time- and temperature-dependent process that is more pronounced in the slow-twitch SOL than in the fast-twitch EDL muscles.

The effect of a unilateral muscle transplantation on the muscle fiber type and the MyHC isoform content in unoperated hind limb slow and fast muscles of the inbred Lewis rats.

To reveal the effect of foreign innervation and altered thyroid status on fiber type composition and the myosin heavy chain (MyHC) isoform expression in the rat slow soleus (SOL) and fast extensor digitorum longus (EDL) muscles, a method of heterochronous isotransplantation was developed. In this experimental procedure, the SOL or EDL muscles of young inbred Lewis rats are grafted either into the host EDL or SOL muscles of adult rats of the same strain with normal or experimentally altered thyroid status. To estimate the extent of fiber type transitions in the transplanted muscles, the SOL and EDL muscle from the unoperated leg and unoperated muscles from the operated leg could be legitimately used as controls, but only when the experimental procedure itself does not affect these muscles. To verify this assumption, we have compared the fiber type composition and the MyHC isoform content of unoperated contralateral SOL and EDL muscles and ipsilateral unoperated SOL muscle of experimental rats after unilateral isotransplantation into the host EDL muscle with corresponding muscles of the naive rats of the same age and strain. We provide compelling evidence that the unilateral heterochronous isotransplantation has no significant effect on the fiber type composition and the MyHC isoform content of unoperated muscles of experimental animals. Hence, these muscles can be used as controls in our grafting experiments.

Electrophoretic analysis of myosin heavy chain isoform patterns in extraocular muscles of the rat.

Six oculorotatory muscles and the levator palpebrae muscle of the rat were analysed by SDS-PAGE for their myosin heavy chain (MHC) isoform patterns. Oculorotatory muscles display a marked predominance of fast MHC isoforms. They contain, in addition to the slow (MHCI) and fast (MHCIIb, MHCIId, MHCIIa) skeletal MHCs, the neonatal MHCneo and the extraocular MHCeom. The levator palpebrae, generally assumed to be a member of the extraocular muscle group because of its innervation by the oculomotor nerve, does not contain MHCneo and MHCeom. It resembles a fast-twitch skeletal muscle with a predominance of MHCIId.

Site-dependent effects of experimental hypo- and hyperthyroidism on resident macrophages in extraocular muscles of rats: a quantitative immunohistochemical study.

It has been suggested that the effects of dysthyroidism on resident immunocompetent cells of the extraocular muscles may play a role in the pathogenesis of Graves' ophthalmopathy. The distribution of such cells was therefore studied in extraocular muscles of rats that were made hyper- or hypothyroid by the oral administration of thyroxine or propylthiouracil respectively. Skeletal muscles were studied for comparison. The cell distributions were analysed in cryostat cross-sections subjected to a two-step immunoperoxidase method using well-characterized monoclonal antibodies against T cells, B cells, macrophages and MHC class II antigens. The extraocular muscles of control (euthyroid) rats contained numerous macrophages, fewer MHC-II positive cells and T cells and no B cells. Differences in the distribution of immunocompetent cells were found in control rats, between skeletal and extraocular muscles as well as within the various recti eye muscles. This particular tissue distribution resembles that previously reported for human extraocular and skeletal muscles. Quantitative analysis showed that experimental dysthyroidism only affected cell populations in the extraocular muscles. Significant effects on the number of macrophages were observed in the inferior rectus muscle of both hypo- and hyperthyroid rats, this was most pronounced in the orbital layer of the muscles. Both hyper- and hypothyroidism appear to affect local cell distributions in a tissue-specific manner. The presently observed site-dependent effects of dysthyroidism on local immunocompetent cell populations may have relevance for the differential involvement of muscular tissues in Graves' ophthalmopathy.

Anabolic steroids (metenolone) improve muscle performance and hemodynamic characteristics in cardiomyoplasty.

The loss of force and mass in the conditioned latissimus dorsi muscle are principal reasons for the poor improvement in hemodynamic functioning attained by cardiomyoplasty. Using 24 sheep, we investigated the effect of anabolic steroids on the hemodynamic, histologic, and myophysiologic characteristics in the setting of cardiomyoplasty. In 12 of the animals (group A), the latissimus dorsi muscles were electrically conditioned with an Itrel pulse generator; in the remaining 12 animals (group B), the electrical conditioning was combined with the administration of an anabolic hormone (metenolone; 100 mg/week). The hemodynamic measurements were performed during isolated perfusion of the subclavian artery (maintenance of pressure in the muscles), while all other circulation variables were held at the exact and reproducible value of zero by inducing ventricular fibrillation. Maximum force and muscle mass showed a significant increase in group B (maximum force: group A, 4.23 +/- 0.55 kp, and group B, 6.0 +/- 3.14 kp; muscle mass: group A, +11.07% +/- 1.06%, and group B, +79.9% +/- 40.8%). The ratio of type I to type II fibers after 12 weeks was 65.2% to 34.8% in group A and 96.7% to 3.3% in group B, as opposed to 19.9% to 80.1% in the control group. No side effects of the anabolic steroids were observed during the experiment. In the hemodynamic studies, we were able to demonstrate a further significant increase in the left ventricular pressure, fractional fiber shortening value, ejection fraction, stroke volume, cardiac output, and stroke work when using conditioned latissimus dorsi muscles that were additionally treated with metenolone.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of an anabolic steroid (Metenolon) on contractile performance of the chronically stimulated latissimus dorsi in sheep.

In 12 sheep the left latissimus dorsi muscles (LD) were conditioned by chronic electrostimulation with a pulse generator (Itrel, Medtronic). Six animals (group B) received a weekly intramuscular injection of an anabolic steroid (Metenolon). After 14 weeks the contraction parameters of the left LDs (group A and B) and right LDs (control group) were investigated. The increase in weight of the conditioned LDs was 11.07% (+/- 1.06%) in group A and 79.97% (+/- 40.8; P < 0.05) in group B. The force capacity under stimulation patterns which were just tetanic was 1.15 kp in group A and 4.13 kp in group B (P < 0.05); under supramaximal stimulation patterns it was 4.23 kp (A) and 6.0 kp (B) (P = ns). The force time relation (dF/dt) was 6.7 kp/s for the left LDs in group A versus 16.4 kp/s for the right LDs (P < 0.01); in group B it was 5.13 kp/s for the left LDs versus 15.8 kp/s for the control muscles (P < 0.05). The maximal force (Fmax) per 100 g muscle weight did not differ significantly (A: 2.42 kp/100 g; B: 2.52 kp/100 g). In conclusion, the LD muscles which were subjected to both anabolic therapy and electrical stimulation showed a significant increase in their force capacity due to an enormous increase in mass. Fibre type transformation was complete only in group B. No fibre deterioration was observable in either group. No anabolic side effects were detected in the animals. With the use of anabolic steroids, therefore, a clearer direct increase in contractility on the left ventricle should be expected ("squeezing" theory), as well as a contribution to reduction in wall tension and myocardial oxygen consumption, respectively, according to Laplace's Law (via the considerable increase in thickness).

Actions of lyotropic anions on the mechanical properties of fast and slow twitch rat muscles at different temperatures.

The effects of lyotropic (swelling) anions (Cl(-), Br(-), NO(3)(-) and I(-)) on contractile properties of fast-twitch extensor digitorum longus (EDL) and slow-twitch soleus (SOL) muscles were investigated in vitro at 20 degrees C and 35 degrees C. Isolated muscles bathed in anionic Tyrode solution were stimulated directly and isometric single twitches and fused tetanic contractions were recorded. In a Cl(-)Tyrode solution a decrease of the bathing temperature led to a cold potentiation of the twitch tension (P(t)) in EDL muscles, however, to a cold depression in SOL muscles, in both muscles combined with a prolongation of contraction (CT) and half relaxation (HRT) times. The extent and order of the potentiating effect of lyotropic anions on the P(t), CT and HRT in EDL and SOL were quite similar and increased in the order: Cl(-)< Br(-)< NO(3)(-)< I(-). Since the lyotropic anions did not influence tetanic tensions, the twitch-tetanus ratio (TTR) was increased in NO(3)(-) and I(-)solutions. All effects of the anions were rapidly and completely reversed in both muscles when the test solution was replaced by the normal one. The temperature decrease caused no significant alteration in the potentiation capacity of the anions or in the kinetics of their action and reversibility.

Effects on skeletal muscle fibres of diabetes and Ginkgo biloba extract treatment.

Combined cytophotometric and morphometric analysis of muscle fibre properties and myosin heavy chain electrophoresis were performed on extensor digitorum longus and soleus muscles from healthy rats and rats with streptozotocin-induced diabetes. Moreover, the protective effect of Ginkgo biloba extract, a potent oxygen radical scavenger, on diabetic muscles was investigated. Changes in fibre type-related enzyme activities, fibre type distribution, fibre cross areas and myosin isoforms were found. In muscles of diabetic rats, a metabolic shift was measured mainly in fibres with oxidative metabolism. Fast-oxidative glycolytic fibres showed a shift to more glycolytic metabolism and about a third transformed into fast-glycolytic fibres. Slow-oxidative fibres became more oxidative. Fibre atrophy was measured in diabetic muscles dependent on fibre type and muscle. Different fibre types atrophied to a different degree. Therefore, a decreased area percentage of slow fibres and an increased area percentage of fast fibres of the whole muscle cross section in both muscles were found. This is supported by reduced slow and increased fast myosin heavy chain isoforms. These alterations of diabetic muscle fibres could be due to less motion of diabetic rats and diabetic neuropathy. After treatment with Ginkgo biloba extract, enzyme activities were increased mainly in oxidative fibres of diabetic muscles, which was interpreted as protective effect. Generally, the soleus muscle with predominant oxidative metabolism was more vulnerable to diabetic alterations and Ginkgo biloba extract treatment than the extensor digitorum longus muscle with predominant glycolytic metabolism.

Effects of artificial obstruction on the function of the upper urinary tract of Guinea pigs, rats and pigs.

PURPOSE: Four different experiments in animals were performed to evaluate the influence of pyelo-ureteral surgery on the function of the upper urinary tract. METHODS: Experiment I: In 17 female guinea pigs pyelo-ureteral anastomosis was performed microsurgically. Three months later, the ureteral peristalsis was investigated by measuring the intraureteral pressure and the in vitro activity of the renal pelvic and ureteric wall was analysed. Experiment II: 10 rats were used for microsurgical uretero-ureteral anastomosis. One month after surgery the pyelo-ureteral peristalsis was examined by videomicroscopy while simultaneously measuring the renal pelvic and intravesical pressure. Subsequently the kidneys were removed for histological examination. Experiment III: In 2 pigs unilateral pyeloplasty was performed. Using an implanted transmitter the intravesical and the renal pelvic pressures were recorded continuously over a time interval of 3 months. Five months after surgery the pyelo-ureteral peristalsis was investigated by pyelography. The kidneys were then removed for histological and biomechanical examinations. Experiment IV: A partial artificial obstruction was performed in 16 guinea pigs by implanting the ureter into the psoas muscle. Two to six months following surgery their upper urinary tracts were removed for analysis of in vitro activity as well as histological and immunohistochemical investigations of the ureter and renal pelvis. RESULTS: Experiment I: Ultrasound investigation showed in all cases a significant dilation of the renal pelvis. The ureteral contraction frequency distally was decreased in vivo as well as in vitro (p <0.05) compared with the controls. Experiment II: Videomicroscopic imaging showed in eight out of nine cases an interruption of the peristaltic wave below the anastomosis; the ureteral peristalsis was restored distally by ureteral contractions with a decreased frequency. Retroperistalsis was seen in the lower part of the ureter. The frequency of renal pelvic and ureteral contractions were decreased (p <0.05). Renal pelvic baseline pressure as well as contraction amplitude were irregularly changed. Histological examinations showed increased connective tissue within the renal pelvic wall in all cases. Experiment III: In both pigs an intermittent change in contraction frequency of the renal pelvis was found, associated with a changing contraction amplitude. Five months after surgery an interruption of the peristaltic wave was detected in both pigs. Histological examinations showed increased connective tissue within the renal pelvic wall. The stiffness of caliceal and pelvic tissue was lower following the pyeloplasty compared to the controls. Experiment IV: Following artificial partial ureteral obstruction in all guinea pigs the in vitro investigations showed an increased spontaneous activity of the upper urinary tract except in the proximal part of the ureter. Ureteral obstruction produced a change in contraction pattern of the proximal ureter and a decrease in contraction frequency of the distal ureter. Immunohistochemical investigations revealed rarefication and disorientation of nerve fibres within the proximal ureteric wall. CONCLUSIONS: Surgical interruption of the ureteral continuity and re-anastomosis cause a temporary disruption of the peristaltic wave at the anastomosis site. Ureteral peristalsis is restored by ureteral contractions associated with retroperistalsis as well as a decreased contraction frequency. Uretero-ureteral anastomosis in rats, pyelo-ureteral anastomosis in guinea pigs and pyeloplasty in pigs seem to influence the upper urinary tract similarly to a chronical functional obstruction, causing changes in pyelo-ureteral motility and spontaneous muscular activity of the renal pelvic and ureteral wall as well as biomechanical and histological characteristics.

The properties of the extraocular muscles of the frog. I. Mechanical properties of the isolated superior oblique and superior rectus muscles.

The mechanical properties of two extraocular muscles (superior oblique and superior rectus muscles) of the frog were studied and compared with those of a frog's skeletal muscle (iliofibularis muscle) which contains the same types of muscle fibres as the oculorotatory muscles. The extraocular muscles are very fast twitching muscles. They exhibit a smaller contraction time, a smaller half-relaxation time, a higher fusion frequency, and a lower twitch-tetanus ratio than the skeletal muscles. The maximum isometric tetanic tension produced per unit cross-sectional area is lower in the extraocular muscles than in skeletal muscles. However, the extraocular muscles show a higher fatigue resistance than the skeletal muscles. With respect to the dynamic properties there are some differences between the various oculorotatory muscles of the frog. The superior rectus muscle exhibits a faster time-course of the contraction, a higher fusion frequency, and a higher fatigability than the superior oblique muscle. An increase of the extracellular K+-concentration evokes sustained contractures not only in the extraocular muscles but also in the iliofibularis muscle; between these muscles there are no striking differences in the mechanical threshold of the whole muscle preparation. The mechanical threshold depends on the Ca++-concentration of the bathing solution and it is found in a range between 12.5 and 17.5 mM K+ in a normal Ringer solution containing 1.8 mM Ca++. The static-mechanical properties of the extraocular muscles of the frog and the dependence of the active developed tension on the muscle extension are very similar to those which are known to exist in the extraocular muscles of other vertebrates. In tetanic activated frog's oculorotatory muscles a linear relationship exists between length and tension. A variation of the stimulation frequency does not change the slope of this curve but causes parallel shifts of the curve. The peculiar properties of the extraocular muscles of the frog are discussed with respect to the muscle fibre types in these muscles and to the diameter of the muscle fibres.

The properties of the extraocular muscles of the frog. II. Pharmacological properties of the isolated superior oblique and superior rectus muscles.

The pharmacological properties of the superior oblique and the superior rectus muscles of the frog's eye were investigated in comparison with those of a skeletal muscle (iliofibularis muscle) of the same animal. Acetylcholine causes sustained contractures of the extraocular muscles; this effect is increased by physostigmine and decreased or abolished by d-tubocurarine. Also the applications of succinylcholine, choline or caffeine are able to evoke contractures. There are no striking differences in pharmacological properties between extraocular and skeletal muscles of the frog. The time-course of the contractures and the sensitivity of the muscle preparations to the drugs which evoke contractures are identical in extraocular and iliofibularis muscles. In comparison with skeletal muscles there is no higher sensitivity of the extraocular muscles against curare-like drugs. The existence of adrenergic receptors could not be found neither in extraocular nor in skeletal muscles of the frog. It is concluded that in frogs no pharmacological differences exist between the muscle fibre types which compose the extraocular and the skeletal muscles.

The properties of the extraocular muscles of the frog. III. Morphological, mechanical and pharmacological properties of the isolated retractor bulbi muscle.

Some morphological, physiological, and pharmacological properties of the retractor bulbi muscle of the frog were tested. The enzyme-histochemical investigation shows that the retractor bulbi muscle contains twitch muscle fibres only. Two types of twitch muscle fibres, which are especially different in their diameter and in the content of mitochondria, build the muscle in an irregular arrangement; tonic muscle fibres were not observed. On the average, the isolated retractor bulbi muscle has at room temperature a contraction time of 26 ms, a half-relaxation time of 28 ms, a fusion frequency of 75 stimuli/s, and a twitch-tetanus ratio of 0.28. The fatigability of this muscle is higher than in oculorotatory eye muscles but lower than in skeletal muscles of the frog. An increase of the extracellular K+-concentration elicits in retractor bulbi muscles a quickly transient contracture; the mechanical threshold of the muscle fibres is found in a range between 20 and 25 mM K+ in Ringer solution. Similar short-lasting contractures, which are probably caused by twitch fibres, rich in mitochondria, are also evoked by application of depolarizing drugs like acetylcholine. The properties of the retractor bulbi muscle are compared with those of the sartorius muscle of the frog, which likewise contains twitch muscle fibres only.

Arrest of developmental conversion of type II to type I fibres after suspension hypokinesia.

The effects of hypokinesia and of the lack of gravity on muscle fibres, fibre type composition and myosin light chain pattern, as well as on muscle mechanoreceptors were investigated in the slow-twitch soleus (SOL) and fast-twitch extensor digitorum longus (EDL) muscles of young growing and adult rats after suspension hypokinesia (SH) of their hind limbs. The animals were suspended by their tail so that their hind limbs were relieved of their normal weight-bearing function for 3-6 weeks. In normal 3- to 4-week-old rats the SOL contained about 50% type I fibres and their percentage increased up to about 80% until the 10th week, with simultaneous reduction of type IIA fibres. After 3 to 6 weeks of suspension treatment maintained from 3- to 4-week-old rats up to 6 to 10 weeks of age, the SOL still only contained about 50% of type I fibres. The content of fast LC1 and LC2 in the SOL of 6-week-old rats after 3 weeks of suspension was higher than that of control litter-mates reflecting the higher occurrence of IIA fibres in the suspended solei. No changes in fibre type composition were observed after SH performed in adult rats. SH thus leads, in young animals, to the arrest of conversion of type IIA to type I fibres resulting in the persistence of the fibre type composition and of the myosin light chain pattern corresponding to those present in the SOL at the time of the onset of suspension. In both young and adult rats, SH markedly decreased the mass and the mean cross-sectional area of the SOL, mainly due to the severe atrophy of type I fibres. We observed no signs indicating conversion of type I back to type IIA muscle fibres due to the SH either in young or adult animals. In contrast to profound changes in the SOL, no significant differences were found in the EDL in any of the parameters studied. No changes in the investigated parameters of muscle spindles and tendon organs were observed after SH, performed either in young or in adult rats. We thus conclude that SH leads to muscle atrophy and that it influences mainly or exclusively type I fibres in muscles with a postural function such as the SOL.(ABSTRACT TRUNCATED AT 400 WORDS)

[Contractile properties of the striated muscle fibres of esophageal muscle in comparison with skeletal muscle in rats]. / Kontraktile Eigenschaften der quergestreiften Muskelfasern des Osophagus im Vergleich mit ausgewählten Skelettmuskeln der Ratte.

The contractile properties of the striated oesophageal muscle of the rat were studied in vitro at 35 degrees C and 25 degrees C. They were compared with those of the slow-twitch soleus muscle and the fast-twitch extensor digitorum longus (EDL) muscle of the same animals. The contraction time of the oesophageal muscle is 30 msec (+/- 2.5 msec S.E.) at 35 degrees C. It is little but significantly longer than those of the soleus muscle. It is remarkable that the twitch-tetanus ratio of the oesophageal muscle fibres is approximately twice that of the skeletal muscles. Immediately after a tetanic stimulation the amplitude of a single twitch of the fast EDL is increased (posttetanic potentiation); in the slow soleus muscle it is unchanged or decreased, however. Also the muscle fibres of the oesophagus exhibit a post-tetanic-potentiation, but this is smaller than in EDL. A decrease of the temperature of the bathing solution causes in preparations of the fast EDL a higher tension developed by a single twitch (cold potentiation). This is not detectable in preparations of the slow soleus muscle. The oesophageal muscle shows also a remarkable cold potentiation, but it is smaller than in EDL. An increase of the extracellular K+-concentration evokes a short-lasting contracture in the investigated rat muscles. The mechanical thresholds of the oesophageal muscle fibres are clearly higher than those of the soleus muscle but lower than those of the EDL. The striated oesophageal muscle fibres of the rat in spite of their low speed of contraction show some properties of muscle fibres of the fast twitch type.

Mechanical properties of the isolated inferior oblique muscle of the rabbit.

The passive-mechanical and dynamic properties of the rabbit inferior oblique muscle IO were studied in vitro at 35 degrees C. The influence of length on the resting tension and isometric contractions were determined. Maximum twitch tension and fusion tension were developed at optimum length (Lo) an extension of the muscle to about 1.15 times LR, the resting length of the IO in situ. A linear relation was found between length and tension in the activated muscle. An increase in stimulation frequency induced a parallel shift in the curves to higher tension but the slope of the curves remained unchanged. On an average the IO had in response to direct massive stimulation a twitch contraction time of 6.4 ms and a half-relaxation time of 7.0 ms. At stimulation with 300 Hz or above the tetanus fused. Stimulus frequencies above fusion frequency increased the rate of tension rise but not the maximum tetanic tension. The maximum tetanic tension was about 6.4 N/cm2, and the twitch:tetanus ratio was 0.1. To prolonged tetanic stimulations the IO exhibited a high fatigue resistance. Cooling the muscle to 25 degrees C was followed by an increase in the time parameters of single twitches and tetanic contractions, a decrease of the tension developed in a fused tetanus and a small potentiation of the twitch. Following a repetitive stimulation a small post-tetanic potentiation of the twitch was observed.

Contractures in normal and denervated inferior oblique muscle of the rabbit.

Isometric contracture responses of normal and denervated inferior oblique muscles (IO) of the rabbit have been investigated in vitro at 35 degrees C. The threshold concentration for eliciting potassium contractures was about 20 mM K+. In normal IU low potassium concentrations up to about 50 mM K+ evoked only sustained contractures, higher concentrations were responded by contractures with an initial transient component. The transient tension development was maximal at about 100 mM K+ the sustained component at 80 mM K+. After denervation the characteristic time course of the contractures was not changed, but the tension output of the preparation was diminished and long-term denervated IO have a somewhat lowered threshold. In normal IO acetylcholine (ACh), succinylcholine (SCh) and choline (Ch) caused also sustained contractures, the threshold doses were about 5 microM for ACh and SCh and 500 microM for Ch. The ACh sensitivity of the preparations was increased by physostigmine and decreased or abolished by d-tubocurarine. Denervation increased the drug sensitivity but the shape of the contractures was hardly influenced. The properties of slow tonic muscle fibres in mammalian extraocular muscles (EOM) probably responsible for sustained contractures and their changes after denervation are discussed.

Changes in mechanical properties of the inferior oblique muscle of the rabbit after denervation.

The inferior oblique muscle (IO) of the rabbit was denervated. The mechanical properties of the muscles were determined at 35 degrees C in vitro 2--76 days after the operation. After denervation the muscles exhibited a considerable hypertrophy. The cross-sectional area of muscles denervated 30 days or longer grew to about twice as innervated controls. The length-tension relationship of passive or activated long-term denervated IO differed from normal in a higher stiffness. After denervation the time parameters of single twitches and tetanic contractions evoked by massive stimulation were prolonged, the fusion frequency was decreased, and the fatigue resistance was increased. During the first week after cutting the motor nerve both twitch and tetanic tensions decreased drastically. A minimum was reached at the end of the second weak. In the subsequent weeks the tension development was increased again, but per unit cross-sectional area it was always smaller than in innervated muscles. After denervation the twitch:tetanus ratio was increased. Cooling to 25 degrees C was followed by an increase in time parameters of single twitches and tetanic contractions and by a depression of twitch and tetanic tensions. Following a repetitive stimulation denervated IO showed a posttetanic depression of the single twitch.

Motoneuronal location and acetylcholine sensitivity of developing rat slow and fast twitch muscle fibres.

The postnatal changes of the extrajunctional acetylcholine (ACh) sensitivity of fast twitch (extensor digitorum longus muscle, EDL) and slow twitch (soleus muscle, SOL) of the rat were investigated in vitro. As a measure of the ACh-sensitivity serves the threshold dose of ACh necessary to evoke a contracture. Immediately after birth rat muscles show a high extrajunctional ACh-sensitivity, but the SOL exhibits a somewhat higher sensitivity (threshold dose 0.5-5 microM ACh/l) than the EDL (threshold dose 5-20 microM ACh/l). These sensitivities are unchanged up to the 8th (EDL) or 10th (SOL) day of life, afterwards the ACh-sensitivity decreases. Values of adult EDL (unsensitivity against ACh in the bathing solution) and SOL (threshold dose 1-5 mM ACh/l) are reached around the 16th (EDL) and 20th (SOL) day after birth. Fast twitch muscles innervated by motoneurons of the cervical spinal cord (sternomastoideus muscle or biceps brachii muscle) show an accelerated decrease of ACh-sensitivity in comparison to fast twitch EDL innervated by motoneurons located in the lumbar spinal cord according to the cranio-caudal relation of the development of the spinal cord.