Nerve Cross-Bridging to Enhance Nerve Regeneration in a Rat Model of Delayed Nerve Repair

There are currently no available options to promote nerve regeneration through chronically denervated distal nerve stumps. Here we used a rat model of delayed nerve repair asking of prior insertion of side-to-side cross-bridges between a donor tibial (TIB) nerve and a recipient denervated common peroneal (CP) nerve stump ameliorates poor nerve regeneration. First, numbers of retrogradely-labelled TIB neurons that grew axons into the nerve stump within three months, increased with the size of the perineurial windows opened in the TIB and CP nerves. Equal numbers of donor TIB axons regenerated into CP stumps either side of the cross-bridges, not being affected by target neurotrophic effects, or by removing the perineurium to insert 5-9 cross-bridges. Second, CP nerve stumps were coapted three months after inserting 0-9 cross-bridges and the number of 1) CP neurons that regenerated their axons within three months or 2) CP motor nerves that reinnervated the extensor digitorum longus (EDL) muscle within five months was determined by counting and motor unit number estimation (MUNE), respectively. We found that three but not more cross-bridges promoted the regeneration of axons and reinnervation of EDL muscle by all the CP motoneurons as compared to only 33% regenerating their axons when no cross-bridges were inserted. The same 3-fold increase in sensory nerve regeneration was found. In conclusion, side-to-side cross-bridges ameliorate poor regeneration after delayed nerve repair possibly by sustaining the growth-permissive state of denervated nerve stumps. Such autografts may be used in human repair surgery to improve outcomes after unavoidable delays.     

Neuroplasticity in the smooth muscle of the myenterically and extrinsically denervated rat jejunum

The objective of this study was to examine the effects of two different denervation procedures on the distribution of nerve fibers and neurotransmitter levels in the rat jejunum. Extrinsic nerves were eliminated by crushing the mesenteric pedicle to a segment of jejunum. The myenteric plexus and extrinsic nerves were eliminated by serosal application of the cationic surfactant benzyldimethyltetradecylammonium chloride (BAC). The effects of these two denervation procedures were evaluated at 15 and 45 days. The level of norepinephrine in whole segments of jejunum was initially reduced by more than 76% after both denervation procedures, but by 45 days the level of norepinephrine was the same as in control tissue. Tyrosine hydroxylase (nor-adrenergic nerve marker) immunostaining was absent at 15 days, but returned by 45 days. However, the pattern of noradrenergic innervating axons was altered in the segment deprived of myenteric neurons. Immunohistochemical studies showed protein gene product 9.5 (PGP 9.5)-immunoreactive fibers in whole-mount preparations of the circular smooth muscle in the absence of the myenteric plexus and extrinsic nerves. At 45 days, the number of nerve fibers in the circular smooth muscle increased. Vasoactive intestinal polypeptide (VIP)-immunoreactive fibers, a subset of the PGP 9.5 nerve fibers, were present in the circular smooth muscle at both time points examined. Choline acetyltransferase (CAT) activity and VIP and leucine enkephalin levels were measured in separated smooth muscle and submucosa-musosal layers of the denervated jejunum. VIP and leucine-enkephalin levels were no different from control in tissue that was extrinsically denervated alone. However, the levels of these peptides were elevated two-fold in the smooth muscle 15 and 45 days after myenteric and extrinsic denervation. In the submucosa-mucosa, VIP and leucine enkephalin levels also were elevated two-fold at 15 days, but comparable to control at 45 days. CAT activity was equal to control in the smooth muscle but elevated two-fold in the submucosa-mucosa at both times. These results provide evidence for innervation of the circular smooth muscle by the submucosal plexus. Moreover, these nerve fibers originating from the submucosal plexus proliferate in the absence of the myenteric plexus. Furthermore, the myenteric neurons appear to be essential for normal innervation of the smooth muscle by the sympathetic nerve fibers. It is speculated that the sprouting of the submucosal plexus induced by myenteric plexus ablation is mediated by increased production of trophic factors in the hyperplastic smooth muscle.     

Long-term changes in neurotrophic factor expression in distal nerve stump following denervation and reinnervation with motor or sensory nerve

Several factors have been proposed to account for poor motor recovery after prolonged denervation, including motor neuron cell death and incomplete or poor regeneration of motor fibers into the muscle. Both may result from failure of the muscle and the distal motor nerve stump to continue expression of neurotrophic factors following delayed muscle reinnervation. This study investigated whether regenerating motor or sensory axons modulate distal nerve neurotrophic factor expression. We found that transected distal tibial nerve up-regulated brain-derived neurotrophic factor (BDNF) and glial cell line-derived neurotrophic factor (GDNF) mRNA, down-regulated neurotrophin-3 and ciliary neurotrophic factor mRNA, and that although these levels returned to normal with regeneration, the chronically denervated distal nerve stump continued to express these neurotrophic factors for at least 6 months following injury. A sensory nerve (the cutaneous saphenous nerve) sutured to distal tibial nerve lowered injury-induced BDNF and GDNF mRNA levels in distal stump, but repair with a mixed nerve (peroneal, containing muscle and cutaneous axons) was more effective. Repair with sensory or mixed nerves did not affect nerve growth factor or neurotrophin-3 expression. Thus, distal nerve contributed to a neurotrophic environment for nerve regeneration for at least 6 months, and sensory nerve repair helped normalize distal nerve neurotrophic factor mRNA expression following denervation. Furthermore, as BDNF and GDNF levels in distal stump increased following denervation and returned to control levels following reinnervation, their levels serve as markers for the status of regeneration by either motor or sensory nerve.     

Episodic hypoxia induces long-term facilitation of neural drive to tongue protrudor and retractor muscles.

Hypoxic episodes can evoke a prolonged augmentation of inspiratory motor output called long-term facilitation (LTF). Hypoglossal (XII) LTF has been assumed to represent increased tongue protrudor muscle activation and pharyngeal airway dilation. However, recent studies indicate that tongue protrudor and retractor muscles are coactivated during inspiration, a behavior that promotes upper airway patency by reducing airway compliance. These experiments tested the hypothesis that XII LTF is manifest as increased inspiratory drive to both tongue protrudor and retractor muscles. Neurograms were recorded in the medial XII nerve branch (XIIMED; contains tongue protrudor motor axons), the lateral XII nerve branch (XIILAT; contains tongue retractor motor axons), and the phrenic nerve in anesthetized, vagotomized, paralyzed, ventilated male rats. Strict isocapnia was maintained for 60 min after five 3-min hypoxic episodes (arterial Po(2) = 35 +/- 2 Torr) or sham treatment. Peak inspiratory burst amplitude showed a persistent increase in XIIMED, XIILAT, and phrenic nerves during the hour after episodic hypoxia (P < 0.05 vs. sham). This effect was present regardless of the quantification method (e.g., % baseline vs. percent maximum); however, comparisons of the relative magnitude of LTF between neurograms (e.g., XIIMED vs. XIILAT) varied with the normalization procedure. There was no persistent effect of episodic hypoxia on inspiratory burst frequency (P > 0.05 vs. sham). These data demonstrate that episodic hypoxia induces LTF of inspiratory drive to both tongue protrudor and retractor muscles and underscore the potential contribution of tongue muscle coactivation to regulation of upper airway patency.     

Competitive reinnervation of salamander skin by regenerating and intact mechanosensory nerves

We have investigated in the salamander the possibility that regenerating mechanosensory nerves might prefer the epidermal Merkel cells (their specific targets) that are located within their segmental domain to those within a "foreign" domain. Since regerating nerves cross domain boundaries with no evidence of the marked delay exhibited by intact sprouting nerves, we examined situations in which the regenerating axons of one segmental nerve were effectively in equal competition for denervated skin with those of another segmental nerve. Additionally, we investigated whether there were differences between regenerating axons and intact sprouting axons of the same segmental nerve, in their ability to innervate available skin both inside and outside the parent domain. No preference was detected of any type of nerve, regenerating or intact, for particular skin regions, or for Merkel cells as indicated by the numbers of mechanosensory thresholds of the touch spots that developed in reinnervated skin. Neither was there any indicating of displacement of "foreign" nerves from a particular region by appropriate axons. When regenerating and intact (sprouting) axons invaded denervated skin more or less simultaneously, the former appeared to have a slight advantage since a significantly greater proportion of skin was innervated by regenerated fibres. With this one exception, all the results were explained most simply by assuming that the axon that first arrives at a denervated Merkel cell establishes a permanent association with that cell and at the same time causes it to lose its "target character" for other axons.     

Peptide-containing neurons intrinsic to the gut wall

Summary Nerve fibers containing substance P, VIP, enkephalin or somatostatin are numerous in the porcine gut wall. They are particularly numerous in the submucosal and myenteric plexuses where peptide-containing cell bodies are also observed. Peptide-containing nerve fibers occur also in the vagus nerves, suggesting that the gut receives an extrinsic supply of peptidergic nerves. The extrinsic contribution to the peptide-containing nerve supply of the gut wall has not yet been quantitatively assessed. In an attempt to clarify this question pigs were subjected to bilateral subdiaphragmatic vagotomy. Another group of animals was subjected to complete extrinsic denervation by autotransplantation of a jejunal segment. The pigs were killed at various time intervals after the operations; the longest time interval studied was four months. Following vagotomy the innervation pattern of the jejunum appeared completely unaffected. Following complete extrinsic denervation the adrenergic nerve fibers disappeared, while peptide-containing and acetylcholinesterase-positive nerve fibers remained apparently unaltered. This was confirmed chemically in the case of substance P.The motor activity of smooth muscle from the jejunum was studied in vitro. At low stimulation frequencies the smooth muscle from control jejunum responded by relaxation; upon cessation of stimulation a contraction occurred. With increasing stimulation frequencies the duration of the relaxation decreased; at high frequency stimulation only a contraction was recorded. In the autotransplant low frequency stimulation induced no or only a weak relaxation; high frequency stimulation induced contraction. After cholinergic and adrenergic blockade, the muscle responded with relaxation at all frequencies; the response was similar in innervated and denervated specimens. On the whole, the effects of extrinsic denervation on the motor activity of smooth muscle from porcine jejunum were minor, possibly reflecting the high degree of autonomy of the gut.     

Cutaneous Collateral Axonal Sprouting Re-Innervates the Skin Component and Restores Sensation of Denervated Swine Osteomyocutaneous Alloflaps

Reconstructive transplantation such as extremity and face transplantation is a viable treatment option for select patients with devastating tissue loss. Sensorimotor recovery is a critical determinant of overall success of such transplants. Although motor function recovery has been extensively studied, mechanisms of sensory re-innervation are not well established. Recent clinical reports of face transplants confirm progressive sensory improvement even in cases where optimal repair of sensory nerves was not achieved. Two forms of sensory nerve regeneration are known. In regenerative sprouting, axonal outgrowth occurs from the transected nerve stump while in collateral sprouting, reinnervation of denervated tissue occurs through growth of uninjured axons into the denervated tissue. The latter mechanism may be more important in settings where transected sensory nerves cannot be re-apposed. In this study, denervated osteomyocutaneous alloflaps (hind- limb transplants) from Major Histocompatibility Complex (MHC)-defined MGH miniature swine were performed to specifically evaluate collateral axonal sprouting for cutaneous sensory re-innervation. The skin component of the flap was externalized and serial skin sections extending from native skin to the grafted flap were biopsied. In order to visualize regenerating axonal structures in the dermis and epidermis, 50um frozen sections were immunostained against axonal and Schwann cell markers. In all alloflaps, collateral axonal sprouts from adjacent recipient skin extended into the denervated skin component along the dermal-epidermal junction from the periphery towards the center. On day 100 post-transplant, regenerating sprouts reached 0.5 cm into the flap centripetally. Eight months following transplant, epidermal fibers were visualized 1.5 cm from the margin (rate of regeneration 0.06 mm per day). All animals had pinprick sensation in the periphery of the transplanted skin within 3 months post-transplant. Restoration of sensory input through collateral axonal sprouting can revive interaction with the environment; restore defense mechanisms and aid in cortical re-integration of vascularized composite allografts.     

Contribution of the submentalis muscle to feeding mechanics in the leopard frog, Rana pipiens

This study investigated the functional contributions of the submentalis muscle to the coordination of feeding behavior in the leopard frog, Rana pipiens. Additionally, the anatomical origins of the motor neurons innervating this muscle are identified and described. The m. submentalis is a small muscle connecting the distal mandibular tips. Depending upon the anuran species studied, this muscle contributes to mandibular bending and the degree to which the tongue is protracted, or has little or no role in feeding biomechanics. High-speed videography was used to quantify feeding attempts before versus after bilateral denervation of the m. submentalis. Additionally, the terminal branch of the trigeminal nerve prior to innervating the m. submentalis was retrogradely labeled to identify the origins of motor neurons innervating the muscle. For the kinematic analyses, denervation of the submentalis resulted in significant increases in the time to maximum tongue protrusion, and the duration of tongue protrusion. Neither mandibular bending, nor tongue length variables differed significantly between normal conditions and deafferented conditions. However, when unsuccessful feeding attempts were quantified following the denervation, failed attempts were nearly always due to the tongue not reaching the prey. None of the unsuccessful feedings prior to denervation were due to inadequate tongue protrusion. Anatomical data show a much larger rostral-caudal distribution of the trigeminal motor neurons than previously described for anurans. These data suggest a larger role for the submentalis muscle in Rana than in previously studied anurans with long protrusible tongues, and suggests a feedback mechanism from the trigeminal nerve to the nerves coordinating tongue protraction and retraction.     

Mechanisms controlling axonal sprouting at the neuromuscular junction

This review considers the relative roles of sprouting stimuli, perisynaptic Schwann cells and neuromuscular activity in axonal sprouting at the neuromuscular junction in partially denervated muscles. A number of sprouting stimuli, including insulin-like growth factor II, which are generated from inactive muscle fibers in partially denervated and paralyzed skeletal muscles, has been considered. There is also evidence that perisynaptic Schwann cells induce and guide axonal sprouting in adult partially denervated muscles. Excessive neuromuscular activity significantly reduces bridging of perisynaptic Schwann cell processes between innervated and denervated endplates and thereby inhibits axonal sprouting in partially denervated adult muscles. Elimination of neuromuscular activity is also detrimental to sprouting in these muscles, suggesting that calcium influx into the nerve is crucial for axonal sprouting. The role of neuromuscular activity in axonal sprouting will be considered critically in the context of the roles of sprouting stimuli and perisynaptic Schwann cells in the process of axonal sprouting.     

The role of sensory input in motor neuron sprouting control

Primary sensory neurons project to motor neurons directly or through interneurons and affect their activity. In our previous paper we showed that intramuscular sprouting can be affected by changing the sensory synaptic input to motor neurons. In this work, motor axon sprouting within a peripheral nerve (extramuscular sprouting) was induced by nerve injury at such a distance from muscle so as not to allow nerve-muscle trophic interactions. Two different procedures were carried out: (1) sciatic nerve crush and (2) sciatic nerve crush with homosegmental ipsilateral L3-L5 dorsal rhizotomy. The number of regenerating motor axons innervating extensor digitorum longus muscle was determined by in vivo muscle tension recordings and an index of their individual conduction rate was obtained by in vitro intracellular recordings of excitatory postsynaptic end-plate potentials in muscle fibers. The main findings were: (1) there are more regenerated axons distally from the lesion than parent axons proximally to the lesion (sprouting at the lesion); (2) sprouting at the lesion was negatively affected by homosegmental ipsilateral dorsal rhizotomy; (3) the number of motor axons innervating extensor digitorum longus muscle extrafusal fibers counted proximally to the lesion increased following nerve injury and regeneration but this did not occur when sensory input was lost. A transient innervation of extrafusal fibers by &#110 motor neurons may explain the increase of motor axons counted proximally to the lesion.     

The role of sensory input in motor neuron sprouting control

Primary sensory neurons project to motor neurons directly or through interneurons and affect their activity. In our previous paper we showed that intramuscular sprouting can be affected by changing the sensory synaptic input to motor neurons. In this work, motor axon sprouting within a peripheral nerve (extramuscular sprouting) was induced by nerve injury at such a distance from muscle so as not to allow nerve-muscle trophic interactions. Two different procedures were carried out: (1) sciatic nerve crush and (2) sciatic nerve crush with homosegmental ipsilateral L3-L5 dorsal rhizotomy. The number of regenerating motor axons innervating extensor digitorum longus muscle was determined by in vivo muscle tension recordings and an index of their individual conduction rate was obtained by in vitro intracellular recordings of excitatory postsynaptic end-plate potentials in muscle fibers. The main findings were: (1) there are more regenerated axons distally from the lesion than parent axons proximally to the lesion (sprouting at the lesion); (2) sprouting at the lesion was negatively affected by homosegmental ipsilateral dorsal rhizotomy; (3) the number of motor axons innervating extensor digitorum longus muscle extrafusal fibers counted proximally to the lesion increased following nerve injury and regeneration but this did not occur when sensory input was lost. A transient innervation of extrafusal fibers by gamma motor neurons may explain the increase of motor axons counted proximally to the lesion.     

Sprouting and regeneration of frog motoneurons during synapse elimination

The ability of frog axons to sprout and reinnervate during the period of synapse elimination was examined in the cutaneous pectoris muscle from young postmetamorphic frogs using histological staining of nerve terminals and postsynaptic acetylcholinesterase. Partial denervation of the cutaneous pectoris muscle during the period of synapse elimination produced rapid sprouting of the intact axons. The majority of denervated endplates were being reinnervated by sprouts within 3 days. In addition, total denervation performed by either sectioning or crushing the nerve was followed by functional reinnervation. Approximately 98% of the endplates were being reinnervated within 7 days after a nerve crush and 10 days after sectioning the nerve.     

Impulse Activity Evokes Precocious Sprouting of Nociceptive Nerves into Denervated Skin

We have studied the sprouting of intact high-threshold mechanosensory nerves into adjacent denervated trunk skin in adult rats behaviorally, histologically, and electrophysiologically. In the anesthetized animal, stimulation of high-threshold endings in back skin by localized pinching elicits a bilateral reflex excitation of the underlying skeletal muscle, the cutaneous trunci muscle (CTM), visible as a twitch-like puckering of the skin. The reflex was also evoked by electrical excitation of Aδ and of C fibers in the dorsal cutaneous nerves (DCNs), with characteristic latencies of 7–20 msec and 40–60 msec, respectively; excitation of low-threshold (Aα) fibers was ineffective. After cutting selected DCNs, the deprived skin became insensible, but pinch responsiveness gradually recovered over the following 2 weeks. Regeneration of cut axons was not responsible for this recovery; when neighboring intact DCNs were cut, however, all responses were abolished in the recovered skin that had been initially denervated. By 3–5 days after denervation, axons in the dermis were all histologically absent or degenerating; when pinch sensitivity was restored to such skin, silver-stainable axons reappeared in the formerly empty Schwann tubes. During the work we noticed that the periodic examination by pinching, used to follow the time course of recovery of function in individual animals, led to an earlier development of this recovery than in animals that were examined only once at a specified time after denervation, This apparent acceleration in the redevelopment of pinch sensitivity was correlated with the appearance of axons in the recovered skin, and was shown to be due to the impulse activity evoked in the remaining nerves by the periodic pinching; it did not occur when the nerves were blocked by tetrodotoxin (TTX), and it was mimicked by a brief (10-min) period of electrical excitation of the Aδ fibers in a remaining nerve carried out at the time when the denervation of skin was done. The time course of the phenomenon suggested that the principal effect of the impulses was to shorten the latency to the onset of sprouting in the activated Aδ axons; that is, they induced precocious sprouting. The impulses needed to be conducted centrally for the effect to occur, and precocious sprouting failed to occur if the impulses were allowed to proceed only distally toward the skin.     

Transient inability of neonatal rat motoneurons to reinnervate muscle

We studied the reinnervation of internal intercostal muscles of newborn rats. The distal halves were denervated by nerve section at various ages between birth and 6 weeks. Regardless of the age at denervation, neither evoked nor spontaneous nerve-muscle transmission reappeared until the animals were at least 3 weeks old. Older rats recovered a substantial degree of function within 7 days of nerve section. Normally the motor units in this muscle are narrowly distributed, so most axotomized motoneurons lost their entire synaptic periphery. Reinnervation was by axons which had been sectioned, and regenerated motor units were of normal size and number. There was no collateral sprouting from end plates left intact. Motoneurons axotomized at birth did regenerate axons the full length of the muscle within 7 days of operation. Their failure to reinnervate the muscle was due to delay in forming functional end plates. Nerve section in animals aged 1 month or older resulted in an abnormal pattern of reinnervation; reinnervated motor units were diffusely spread through large portions of the muscle, although they still did not overlap with the region left intact. This indicates that thoracic motoneurons respond to axotomy differently in neonatal rats than they do in adults.     

Efficiency of reinnervation of neonatal rat muscle of original and foreign nerves

Formation of neuromuscular connections in mammals may involve a hierarchy of efficiency of synapse formation at a stage when motor nerves have already contacted muscle fibers and during the transitional period of multiple innervation. In an attempt to test for such a hierarchy, we examined, in neonatal rats, the relative efficiency of reinnervation by foreign or original nerves implanted simultaneously in a large muscle so that competition for muscle fibers was minimized. The tibial nerve, containing gastrocnemius nerve fibers, and the “foreign” peroneal nerve were implanted into the denervated lateral gastrocnemius muscle. One to five months later, indirect tetanic tensions obtained upon stimulating the implanted nerves were measured by isometric techniques and were compared to contralateral control muscles. When both nerves were implanted side by side at the end-plate region, approximately equal tetanic tensions were obtained at the time of testing. The same result was also obtained when the tibial and common peroneal nerves were implanted into non-end-plate and end-plate regions, respectively. However, in the reverse experiment, the tibial nerve implanted at the end-plate region produced significantly higher tetanic tension than the peroneal nerve at the non-end-plate site in the same muscle. Thus, the original nerve, compared to a foreign nerve, appeared to reinnervate neonatal muscle more effectively, but this was only revealed under conditions where access to former end-plate regions was unequal.     

Neuromuscular activity impairs axonal sprouting in partially denervated muscles by inhibiting bridge formation of perisynaptic Schwann cells

Following partial denervation of rat hindlimb muscle, terminal Schwann cells extend processes from denervated endplates to induce and guide sprouting from the remaining intact axons. Increased neuromuscular activity significantly reduces motor unit enlargement and sprouting during the acute phase of sprouting. These findings led to the hypothesis that increased neuromuscular activity perturbs formation of Schwann cell bridges and thereby reduces sprouting. Adult rat tibialis anterior (TA) muscles were extensively denervated by avulsion of L4 spinal root and were immediately subjected to normal caged activity or running exercise (8 h daily) for 3, 7, 14, 21, and 28 days. Combined silver/cholinesterase histochemical staining revealed that the progressive reinnervation of denervated endplates by sprouts over a 1 month period in the extensively partially denervated TA muscles was completely abolished by increased neuromuscular activity. Immunohistochemical staining and triple immunofluorescence revealed that the increased neuromuscular activity did not perturb the production of Schwann cell processes, but prevented bridging between Schwann cell processes at innervated and denervated endplates. Our findings suggest that failure of Schwann cell processes to bridge between endplates accounts, at least in part, for the inhibitory effect of increased neuromuscular activity on sprouting.     

The effect of two episodes of denervation and reinnervation on skeletal muscle contractile function.

Sensory or motor "baby-sitting" has been proposed as a clinical strategy to preserve muscle integrity if motion-specific axons must regenerate over a long distance to reach denervated target muscles. Denervated muscles are innervated temporarily by using axons from nearby sensory or motor nerves. After motion specific motor axons have reached the target, the baby-sitter nerve is severed and motion-specific axons are directed to the target. Although this strategy minimizes denervation time, the requisite second episode of denervation and reinnervation might be deleterious to muscle contractile function. This study was designed to test the hypothesis that two sequential episodes of skeletal muscle denervation and reinnervation result in greater force and power deficits than a single peripheral nerve injury and repair. Adult Lewis rats underwent either transection and epineurial repair or sham exposure of the left peroneal nerve. After a 4-month recovery period, the contractile properties of the extensor digitorum longus muscle of the sham exposure group (control, n = 9) and one of the nerve division and repair groups (repair group 1, n = 9) were evaluated with measurements of the maximum tetanic isometric force, peak power, and maximal sustained power. A third group of rats underwent a second cycle of nerve division and repair (repair group 2, n = 9) at this same time point. Four months postoperatively, contractile properties of the extensor digitorum longus muscles were evaluated. Maximum tetanic isometric force and peak power were significantly reduced in repair group 2 rats as compared with repair group 1 and control rats. Maximal sustained power was not significantly different between the groups. These data support our working hypothesis that skeletal muscle contractile function is adversely affected by two cycles of denervation and reinnervation as compared with a single episode of nerve division and repair.     

Mitochondrial heat shock protein 70, a molecular chaperone for proteins encoded by mitochondrial DNA

The protracted absence of muscle activation initiates complex cellular and molecular reactions aimed at restoring functional neuromuscular transmission and preventing degenerative processes. A central aspect of these reactions is the sprouting of intramuscular nerves in the vicinity of inactivated muscle fibers. Sprouts emerging from terminal nerve branches and nodes of Ranvier can reestablish functional contacts with inactive muscle fibers, and this is an essential restorative process in pathological conditions of the neuromuscular system. Due to their rapid upregulation in inactive skeletal muscle fibers and their ability to induce nerve sprouting in adult muscle, insulin-like growth factors (IGFs) are candidate signaling molecules to promote restorative reactions in the neuromuscular system. In this study we have exploited the high affinity and specificity of IGF-binding protein 4 (IGF-BP4) and IGF-BP5 for IGF1 and IGF2 to determine whether these growth factors are involved in the nerve sprouting reaction in paralyzed skeletal muscle. In tissue culture experiments with sensory- and motoneurons we demonstrate that the neurite promoting activity of IGF1 is blocked by IGF-BP4, and that a similar IGF-BP-sensitive activity is detected in muscle extracts from paralyzed, but not from control muscle. In in vivo experiments, we show that local delivery of IGF-BP4 to Botulinum toxin A-paralyzed skeletal muscle effectively prevents nerve sprouting in that muscle. Our findings indicate that muscle IGFs play an essential role in intramuscular nerve sprouting. In addition, these findings suggest that IGFs are major signaling factors from inactivated muscle to promote local restorative reactions, including interstitial cell proliferation and nerve sprouting.     

sidestep encodes a target-derived attractant essential for motor axon guidance in Drosophila

At specific choice points in the periphery, subsets of motor axons defasciculate from other axons in the motor nerves and steer into their muscle target regions. Using a large-scale genetic screen in Drosophila, we identified the sidestep (side) gene as essential for motor axons to leave the motor nerves and enter their muscle targets. side encodes a target-derived transmembrane protein (Side) that is a novel member of the immunoglobulin superfamily (IgSF). Side is expressed on embryonic muscles during the period when motor axons leave their nerves and extend onto these muscles. In side mutant embryos, motor axons fail to extend onto muscles and instead continue to extend along their motor nerves. Ectopic expression of Side results in extensive and prolonged motor axon contact with inappropriate tissues expressing Side.     

Lower cranial nerve motor function in unilateral vascular lesions of the cerebral hemisphere.

Motor function subserved by cranial nerves V, VII, X, XI, and XII was assessed in 100 patients with hemiparesis due to a unilateral vascular lesion of the cerebral hemisphere. Several of the findings were not described clearly in many of the standard textbooks of neurology. Weakness of sternomastoid when present was always contralateral to the hemiparesis. This emphasises the principle that the cerebral hemisphere controls movement of the body parts in or towards the contralateral half of the body rather than simply the contralateral muscle groups. An apparent exception to this was seen, however, in the small group of patients who had unilateral weakness of the tongue. In those patients deviation of the tongue was towards the hemiparetic side--that is, the cerebral hemisphere controlled the contralateral half of the tongue and hence protrusion towards the ipsilateral side. Mild dysarthria was common with both right and left sided hemiparesis.