Biomedical subjects

G Vrbová

Publications and source records attributed to G Vrbová.

At least 19 recordsLinked to original sources

PubMed2001

Treatment of the neuromuscular junction with 4-aminopyridine results in improved reinnervation following nerve injury in neonatal rats.

During early postnatal development, nerve injury results in the death of a large proportion of motoneurones and poor recovery of muscle function. Our previous results have shown that premature enhancement of transmitter release from nerve terminals prevents the death of motoneurones following neonatal nerve injury. Whether this increase in motoneurone survival is reflected in an improvement in the reinnervation of muscle was studied here. The muscles in one hindlimb of newborn rats were treated with 4-aminopyridine. Three days later, the sciatic nerve was crushed in the treated leg. When the animals were seven, 14 and 21days of age, the soleus and extensor digitorum longus muscles were removed and processed for GAP-43 (a 43-kDa growth-associated protein) and synaptophysin immunocytochemistry. Both GAP-43 and synaptophysin were expressed in normal soleus and extensor digitorum longus muscles at seven days. Synaptophysin was still expressed at 14 days, but GAP-43 expression had declined. Following nerve injury at three days of age, there was no GAP-43 or synaptophysin immunoreactivity in nerve terminals at seven days. By 21 days, there were 17.3+/-2.1 GAP-43-positive terminals per section in the soleus and 17.7+/-1.4 in the extensor digitorum longus, with mean terminal areas of 47.5+/-3.3 and 49.8+/-2.6 microm(2), respectively. In animals in which nerve crush was preceded by 4-aminopyridine treatment, at 21 days there were 32.9+/-2.6 GAP-43-immunoreactive terminals in the soleus and 44.9+/-2.3 in the extensor digitorum longus, with a mean area of 122.7+/-6.6 microm(2) in the soleus and 136.2+/-9.7 microm(2) in the extensor digitorum longus. These results indicate that in muscles pretreated with 4-aminopyridine, prior to nerve crush at three days, there are significantly more terminals, which occupy a larger area than in untreated muscles. Thus, increasing transmitter release prior to nerve injury significantly improved the ability of axons to reinnervate muscle.

4-Aminopyridine↗

PubMed2001

The effect of riluzole treatment in rats on the survival of injured adult and grafted embryonic motoneurons.

The effect of riluzole on the survival of injured motoneurons was studied. The L4 ventral root was avulsed and reimplanted into the spinal cord. Immediately after the operation, 4 animals were treated with riluzole for 3 weeks while another 4 animals received no treatment after the operation. Three months later the fluorescent dyes, Fast Blue and Diamidino Yellow, were applied to the cut ventral ramus of the L4 spinal nerve, for retrograde labelling of neurons. Three days later, the spinal cords were processed to reveal the retrograde-labelled cells. In untreated animals, there were 20 +/- 2.1 labelled neurons (+/- SEM), while in animals treated with riluzole there were 723 +/- 26. Thus, treatment with riluzole dramatically enhanced the survival of injured motoneurons. In another series of experiments, after avulsion of the L4 ventral root and its reinsertion, embryonic spinal cord pieces were grafted into the host cord. Five animals received riluzole treatment and 4 were left untreated. In the untreated animals, 125 +/- 5.1 retrograde-labelled cells of both graft and host origin were detected. In rats treated with riluzole, 645 +/- 35.7 retrograde-labelled cells were seen and almost all of these were of host origin. Thus, treatment with riluzole enhanced the survival of injured host motoneurons, and by doing so, (i) reduced the ability of grafted neurons to extend their axons into the reimplanted L4 ventral root, and (ii) reduced the survival of the grafted cells.

Animals↗

PubMed2000

Reducing transmitter release from nerve terminals influences motoneuron survival in developing rats.

Motoneurons in neonatal rats die following injury to the peripheral nerve. However, this vulnerability to nerve injury declines rapidly so that nerve injury at five days of age results in little if any motoneuron death. We have proposed that the role of the target during this critical period of development is to up-regulate the release of transmitter from developing motor nerve terminals. Here we show that reducing the release of acetylcholine from nerve terminals in neonatal rats can affect motoneuron maturation and survival. The soleus muscle in neonatal rats was treated with either magnesium or hemicholinium, and the number of motoneurons that survived was established 10 weeks later by retrograde labelling. Following treatment with magnesium, only 58.1% (+/-10.4 S.E.M., n=5) of the motoneurons in the soleus motor pool survived, although hemicholinium had no effect on motoneuron survival. However, those motoneurons that survived following treatment with either magnesium or hemicholinium did not develop normally since they remained susceptible to axotomy-induced cell death for longer than normal. In adult animals in which the sciatic nerve was crushed at five days of age following prior treatment with either magnesium or hemicholinium, only 27.6% (+/-6.2 S.E.M., n=5) and 44% (+/-6.1 S.E.M., n=4) of motoneurons in the sciatic motor pool survived, respectively, although no motoneurons died following injury alone or when injury was preceded by treatment with control implants containing NaCl. These results indicate that the release of acetylcholine from motor nerve terminals plays an important role in the development and survival of motoneurons.

Acetylcholine↗

PubMed2000

Repeated stimuli for axonal growth causes motoneuron death in adult rats: the effect of botulinum toxin followed by partial denervation.

Axons of motoneurons to tibialis anterior and extensor digitorum longus muscles of adult rats were induced to sprout by injecting botulinum toxin into them, by partial denervation or by a combination of the two procedures. Ten weeks later, the number of motoneurons innervating the control and operated tibialis anterior and extensor digitorum longus muscles was established by retrograde labelling with horseradish peroxidase. In the same preparations, the motoneurons were also stained with a Nissl stain (gallocyanin) to reveal motoneurons in the sciatic pool. Examination of the spinal cords from animals treated with botulinum toxin showed that the number of retrogradely labelled cells and those stained with gallocyanin in the ventral horn on the treated compared to the control side was unchanged. In rats that had their L4 spinal nerve sectioned on one side, the number of retrogradely labelled cells on the operated side was 48+/-3% (n = 5) of that present in the control unoperated ventral horn. Thus, just over half the innervation was removed by cutting the L4 spinal nerve. Counts made from gallocyanin-stained sections showed that 94+/-4% (n = 5) of motoneurons were present in the ventral horn on the operated side. Thus, section of the L4 spinal nerve did not lead to any death of motoneurons. In rats that had their muscles injected with botulinum toxin three weeks prior to partial denervation, the number of retrogradely labelled cells was reduced from 48+/-3% (n = 5) to 35+/-4% (n = 5). Moreover, only 67+/-5% (n = 5) of motoneurons stained with gallocyanin, suggesting that a proportion of motoneurons died after this combined procedure. This result was supported by experiments in which motor unit numbers in extensor digitorum longus muscles were determined by measurements of stepwise increments of force in response to stimulation of the motor nerve with increasing stimulus intensity. In partially denervated extensor digitorum longus muscles, 16.6+/-0.7 (n = 5) motor units could be identified, and in animals treated with botulinum toxin prior to partial denervation only 13.3+/-0.9 (n = 3) motor units were present. Taken together, these results show that treatment with botulinum toxin followed by partial denervation causes motoneuron death in adult rats.

Animals↗

PubMed1999

What does chronic electrical stimulation teach us about muscle plasticity?

The model of chronic low-frequency stimulation for the study of muscle plasticity was developed over 30 years ago. This protocol leads to a transformation of fast, fatigable muscles toward slower, fatigue-resistant ones. It involves qualitative and quantitative changes of all elements of the muscle fiber studied so far. The multitude of stimulation-induced changes makes it possible to establish the full adaptive potential of skeletal muscle. Both functional and structural alterations are caused by orchestrated exchanges of fast protein isoforms with their slow counterparts, as well as by altered levels of expression. This remodeling of the muscle fiber encompasses the major, myofibrillar proteins, membrane-bound and soluble proteins involved in Ca2+ dynamics, and mitochondrial and cytosolic enzymes of energy metabolism. Most transitions occur in a coordinated, time-dependent manner and result from altered gene expression, including transcriptional and posttranscriptional processes. This review summarizes the advantages of chronic low-frequency stimulation for studying activity-induced changes in phenotype, and its potential for investigating regulatory mechanisms of gene expression. The potential clinical relevance or utility of the technique is also considered.

Animals↗

PubMed1999

Neurotrophic factors.

Explore the source record for details and available documents.

Brain-Derived Neurotrophic Factor↗

PubMed1999

Stabilizing neuromuscular contacts reduces motoneuron death caused by paralysis of muscles in neonatal rats.

Transient paralysis of the soleus muscle in neonatal rats leads to permanent muscle weakness, loss of muscle fibres and motoneuron death. Application of leupeptin, an inhibitor of a calcium-activated neutral protease, to the neuromuscular junction is known to enhance the maintenance of neuromuscular contacts during development and axonal sprouting. Here, we show that treatment of soleus muscles with leupeptin as they recover from a period of paralysis rescues motoneurons that would otherwise die. The number of motoneurons to the soleus muscle was established by retrograde labelling with horseradish peroxidase eight to 10 weeks after recovery from paralysis. There were only 38.4 (+/-2.8 S.E.M., n=5) motoneurons innervating the soleus muscle that had been paralysed with alpha-bungarotoxin, compared to 58.2 (+/-3.1 S.E.M., n=5) to the control untreated soleus. Thus, the number of motoneurons to the soleus muscle on the alpha-bungarotoxin-treated side was 66.9% (+/-6.2% S.E.M., n=5) of the control side. In those animals where paralysis of the soleus muscle was followed three days later by treatment with leupeptin, the number of labelled motoneurons on the treated side of the spinal cord was 61.5 (+/-4.6 S.E.M., n=4) and that on the contralateral untreated control side was 59 (+/-3.8 S.E.M., n=4). This improvement in motoneuron survival in the leupeptin-treated animals is also confirmed by counts of the number of motor units in the soleus muscle obtained by recording muscle tension. In animals that had their soleus muscles paralysed at birth, only 21 (+/-0.7 S.E.M., n=5) motor units were present, compared to 30 motor units in control muscles. When the paralysed soleus muscle was subsequently treated with leupeptin, the number of remaining motor units in the muscle was 29.8 (+/- 1.0 S.E.M., n=5). In addition, the force output of the soleus muscles that had undergone a period of neonatal paralysis was calculated for both the NaCl- and leupeptin-treated animals. The results showed that paralysis at birth results in a reduction in weight and force output of the soleus muscle, which is not improved following treatment with leupeptin. This study shows that application of leupeptin to the soleus muscle after alpha-bungarotoxin-induced paralysis rescues motoneurons to the soleus that would otherwise die. This effect is most likely due to stabilization of their neuromuscularjunctions.

Animals↗

PubMed1999

Overexpression of GAP-43 induces prolonged sprouting and causes death of adult motoneurons.

In neurodegenerative diseases, neurons undergo prolonged periods of sprouting. Whether this sprouting compromises these neurons is unknown. Here, we examined the effect of axotomy on adult motoneurons undergoing prolonged sprouting in transgenic mice that overexpress GAP-43 (growth-associated protein). Sciatic nerve injury in these adult mice results in motoneuron death, but has no effect in non-transgenic mice. Thus, continued growth of motor axons renders adult motoneurons susceptible to nerve injury and compromises their long-term survival. The progressive nature of neurodegenerative diseases may therefore be caused by prolonged sprouting.

Animals↗

PubMed1999

Integrins at the neuromuscular junction are important for motoneuron survival.

During development motoneurons depend on target contact for their survival. Following injury to the sciatic nerve in neonatal rats, a large proportion of motoneurons die. However, the same injury inflicted at 5 days of age results in no loss of motoneurons. This critical period of postnatal development coincides with the time during which there is a significant increase in the release of transmitter from the nerve terminals at the neuromuscular junction. We have proposed that the role of the target muscle cell during this period is to induce this up-regulation of transmitter release from motor nerve terminals. It has been shown that stretch-induced increase in transmitter release from frog motor nerve terminals is accomplished via an integrin-dependent mechanism. In this study we examined the role of integrins at the rat neuromuscular junction in motoneuron survival. We found that blocking integrin binding at the developing neuromuscular junction delayed the increase in choline acetyltransferase activity that normally takes place during the early postnatal period, and resulted in motoneuron death. Furthermore, the maturation of those motoneurons that survived was delayed so they remained susceptible to subsequent nerve injury. These results support the possibility that integrins, by their involvement in modulating transmitter release, can influence motoneuron survival.

Animals↗

PubMed1999

The age dependent effect of partial denervation of rat fast muscles on their activity.

In 3 or 18 day old Wistar rats the hindlimb muscles were partially denervated by cutting the L4 spinal nerve. Three months later, the effects of partial denervation of the fast extensor digitorum longus (EDL) muscle on the activity of its remaining motor units were studied using electromyographic (EMG) recordings in freely moving animals. In spite of a reduced number of motor units the amount of aggregate EMG activity was greater in the partially denervated EDL muscle in all experimental conditions. This increase was more obvious at rest than during exploratory behaviour, and was significantly greater in muscles that were partially denervated at 3 days than at 18 days of age. On the other hand, the effect of partial denervation on the EMG activity pattern during locomotion was similar in animals partially denervated at 3 or 18 days of age. Unlike in intact EDL, in the partially denervated EDL muscle the duration of the bursts was influenced by the step cycle duration. Thus, we conclude that although partial denervation of EDL muscle influences the amount and pattern of activity of the remaining undamaged motor units in all animals, some of the alterations of EMG activity were more pronounced in animals denervated at younger age.

Aging↗

PubMed1998

Mechanical activity is necessary for the elimination of polyneuronal innervation of developing rat soleus muscles.

During early development, rat soleus muscle fibres are innervated by several axons. Neuromuscular activity is involved in the elimination of all but one terminal, but it is not clear whether electrical or mechanical activity is important. Here, we reduced mechanical activity only, by interfering with excitation-contraction coupling. Muscles treated with dantrolene sodium at 9 days produced significantly less force at 13 days of age than normal muscles, and their sensitivity to ACh was greater than that of controls. The elimination of polyneuronal innervation occurs between days 9-12, but in muscles treated with dantrolene, the loss of synapses was slower. Thus, reducing mechanical activity by interfering with excitation-contraction coupling, (a) delays muscle development and (b) reduces the rate of elimination of polyneuronal innervation.

Acetylcholine↗

PubMed1998

Increased activity improves recovery of partially denervated fast rat muscles.

Partial denervation of the neonatal rat extensor digitorum longus muscles by removing the L4 spinal nerve and thus 80% of its innervation [A.L. Connold, T.J. Fisher, S. Maudarbocus, G. Vrbová, Response of developing fast muscles to partial denervation, Neuroscience 46 (1992) 981-988; F. Tyc, G. Vrbová, The effect of partial denervation of developing rat fast muscles on their motor unit properties, J. Physiol. 482 (1995) 651-660] results in its permanent weakness. The possibility that the weakness that follows partial denervation is due to the effects of reducing activity of the muscle during a critical stage of development was studied here. Partial denervation was carried out in 3-day-old pups by removing the L4 spinal nerve. To enhance motor activity two days later some animals had injections of l-Dopa twice a day for 8 days. This treatment induced locomotor activity for at least 2 h/day. The muscles from treated and untreated animals were examined 2 months later. There was a significantly smaller reduction of weight and force production in the muscles from l-Dopa treated animals. Both twitch and tetanic force developed by the EDL muscle from the treated group was twice that of the control untreated group. This effect was due mainly to the larger size of the motor units (MUs) in the l-Dopa treated muscles compared to the controls. The mean motor unit force in the untreated group was 69% of that in the control muscle, whereas this value was 127% in the l-Dopa treated animals. Thus it appears that the activity induced by treatment with l-Dopa could to some extent prevent the loss of weight and force output seen after partial denervation of young fast muscles.

Animals↗

PubMed1998

Organization of common peroneal nerve motoneurones in the rat and monkey: a comparative study.

The motoneurones with axons in the common peroneal nerve (CPN) of the rat and monkey were examined using retrograde labelling with horseradish peroxidase (HRP). In both species, the CPN motoneurone pool was localized in the dorsolateral part of the ventral horn of the spinal cord. In the rat, the labelled motoneurones were located between the L3 and L6 spinal segments whereas in the monkey, they extended from the caudal end of L4 to the L6 spinal segments. In both species the majority of the labelled neurones were located within the L5 segment. The mean number of the CPN motoneurones in the rat and monkey was 458 and 1148, respectively. A bimodal size distribution of motoneurones was found in both species.

Animals↗

PubMed1998

Recovery of rat skeletal muscles after partial denervation is enhanced by treatment with nifedipine.

Following partial denervation of adult rat skeletal muscle intact axons sprout to reinnervate denervated muscle fibres and increase their territory. The extent of this increase is limited and may depend on the ability of axon terminals to form and maintain synaptic contacts with the denervated muscle fibres. Here we tested the possibility whether reducing Ca2+ entry into presynaptic nerve terminals through dihydropyridine sensitive channels may allow more nerve-muscle contacts to be formed and maintained. Hindlimb muscles of adult Wistar rats were partially denervated by removing a small segment of the L4 or L5 spinal nerve on one side. A nifedipine-containing silastic rubber strip was subsequently implanted close to the partially denervated soleus or extensor digitorum longus (EDL) muscles in some animals. In control experiments silastic strips which did not contain nifedipine were used. Several weeks later isometric contractions were recorded, to determine the effect of (a) partial denervation and (b) nifedipine treatment on force output and motor unit numbers. The tension produced by nifedipine treated partially denervated muscles was 82% and 79% of the unoperated contralateral value for soleus and EDL, respectively. This was significantly greater than in untreated muscles, which only produced 61% and 48%, respectively. Mean motor unit force was also significantly larger with nifedipine treatment. Histological analysis revealed that a significantly larger proportion of the total number of muscle fibres remained in nifedipine-treated partially denervated muscles (soleus, 90% and EDL, 101%) compared with untreated muscles (soleus, 51% and EDL, 66%). Thus the number of neuromuscular contacts was increased with nifedipine treatment.

Animals↗

PubMed1998

Time course of changes in EMG activity of fast muscles after partial denervation.

After partial denervation, the remaining motor units (MUs) of adult fast extensor digitorum longus muscle (EDL) expand their peripheral field. The time course of this event was studied using tension measurement and recordings of electromyographic (EMG) activity. The results show that after section of the L4 spinal nerve, when only 5.3 +/- 0.63 of the 40 MUs normally supplying EDL muscle remain, the force of individual motor units starts to increase between the 1st and 2nd week after the operation and continues to do so for a further week. The drastic reduction of the number of motoneurones supplying the fast EDL leads to an increase in activity of the remaining MUs. In the 1st week after partial denervation, there was a sharp increase in the EMG activity of remaining motor units. During the next 12 days, this increase became less marked, but EMG activity remained nevertheless significantly higher than that of the unoperated EDL muscle. Many MUs became tonically active during posture. The EMG activity pattern during locomotion was also altered, so that the burst duration was positively correlated with the step cycle duration. Moreover, shortly after partial denervation, the interlimb coordination was disturbed but returned to its original symmetrical use 1-2 weeks later.

Animals↗

PubMed1998

Motoneurons innervating partially denervated rat hindlimb muscles remain susceptible to axotomy-induced cell death.

Tibialis anterior and extensor digitorum longus muscles were partially denervated by cutting the L4 spinal nerve in three-day-old rats. The ultrastructure of the intact axons to these muscles in the L5 spinal nerve was examined in nine-day-old rats. In the control L5 spinal nerve, myelinated and unmyelinated axons were intermingled throughout the cross-section of the nerve, while on the operated side the nerve contained areas with predominantly small unmyelinated immature axons. The number of motoneurons innervating the partially denervated muscles was established by retrograde labelling with Diamidino Yellow. In nine- and 21-day-old rats, the number of labelled motoneurons on the partially denervated side, expressed as a percentage of the control side, was 26.1 +/- 5.5% and 20.7 +/- 3.0%, respectively. The response of these uninjured motoneurons to axotomy was tested. The axons of the motoneurons to the partially denervated muscles were crushed at nine days and the numbers of labelled motoneurons in the spinal cord of these rats counted at 21 days of age. Only 4.9 +/- 2.0% labelled motoneurons were seen on the operated side, as opposed to 20.7 +/- 3.0% present in animals without sciatic nerve injury. In normal animals, nerve injury at nine days does not cause motoneuron death. Thus, motoneurons to partially denervated muscles (i) have axons with several immature features and (ii) remain susceptible to axotomy-induced death for much longer than normal.

Animals↗

PubMed1998

Motoneurones that innervate the rat soleus muscle mature later than those to the tibialis anterior and extensor digitorum longus muscles.

The response of motoneurones that innervate either the soleus or tibialis anterior (TA) and extensor digitorum longus (EDL) muscles to increased locomotor activity or to nerve injury at different stages after birth was examined. Increased locomotor activity of rat pups was induced by daily treatment with L-dopa during the first 12 days after birth, and the number of surviving motoneurones to the soleus or TA/EDL muscles was established by retrograde labelling. Treatment with L-dopa resulted in the loss of a significant number of motoneurones within the soleus motor pool but had no effect on the survival of those motoneurones innervating the TA/EDL. Furthermore, following nerve injury during the first few days postnatally, more motoneurones within the soleus motor pool die than in the TA/EDL pool. These results indicate that motoneurones to the soleus muscle mature later than those to the TA/EDL muscles.

Animals↗

PubMed1997

Neonatal paralysis of the rat soleus muscle selectively affects motoneurones from more caudal segments of the spinal cord.

Transient paralysis of the rat soleus muscle shortly after birth leads to permanent muscle weakness, loss of muscle fibres and the death of motoneurones. The soleus muscle receives its innervation from motoneurones whose axons exit the spinal cord either via the L4 ventral ramus or in the more caudal part of the cord via the L5 ventral ramus. Whether both populations of motoneurones are equally affected by neonatal paralysis was studied here. In soleus muscles paralysed with alpha-bungarotoxin shortly after birth and examined 8-10 weeks later, there is no loss of force or muscle fibres in the part of the muscle supplied by axons in the L4 ventral ramus. Loss of force and muscle fibre numbers occurs only in the part of the muscle supplied by axons in the L5 ventral ramus. In a normal adult soleus 30.3 +/- 2.4% of muscle force is produced by stimulating the L4 ventral ramus and 69.0 +/- 5.5% by stimulating the L5 ventral ramus. In soleus muscles treated with alpha-bungarotoxin 28 +/- 1.4% of the force produced by the contralateral control soleus was generated by axons in the L4 ventral ramus and only 20.3 +/- 5.6% by stimulating the L5 spinal nerve. The number of muscle fibres supplied by either ventral ramus in control and experimental muscles confirmed that the decrease of force after treatment with alpha-bungarotoxin can be accounted for by loss of muscle fibres supplied by axons in the L5 ventral ramus. The reduced force production and muscle fibre numbers was due to a selective loss of motoneurones that had their axons in the L5 ventral ramus. The number of axons to soleus in the L4 ventral ramus was 9.3 +/- 0.7 in controls and 10.3 +/- 0.9 in the experimental animals, whereas the L5 ventral ramus contained 17.2 +/- 0.7 in controls and only 4.7 +/- 1.7 in the experimental animals. Thus paralysis of the soleus muscle at birth selectively affects motoneurones in the more caudal part of the spinal cord, suggesting that the more cranial motoneurones are more mature and less likely to be influenced by lack of neuromuscular interaction at the time of birth.

Acute Disease↗