An antibody to neural cell adhesion molecule impairs motor nerve terminal sprouting in a mouse muscle locally paralysed with botulinum toxin

doi:10.1016/0306-4522(90)90123-L

Neuroscience

Volume 35, Issue 1, 1990, Pages 85-91

https://doi.org/10.1016/0306-4522(90)90123-L Get rights and content

Abstract

We have investigated the possible role of neural cell adhesion molecule, in promoting motor nerve terminal sprouting in the paralysed mouse gluteus maximus muscle, using a rabbit polyclonal antiserum to mouse brain neural cell adhesion molecule (donated by Dr C. Goridis). This antiserum recognizes neural cell adhesion molecule in the basal lamina and extracellular matrix of denervated mouse muscles (1988, Booth C. M. and Brown M. C.,Neuroscience27, 699–709). As a single dose of antibody injectedin vivo over the surface of denervated gluteal muscles bound for 12–48 h to over 80% of fibres expressing neural cell-adhesion molecule, once daily injections were used to see if there was any action on terminal sprouting. Injections of antibody were made for four days, starting three days after muscles were paralysed with botulinum toxin. To avoid non-specific complement-mediated attack on neural cell adhesion molecule bearing motor nerve terminals, mice of the DBA/2 strain were used as these lack complement component C5. They were further complement depleted by daily intraperitoneal injections of colloidal inulin. The percentage of end plates bearing terminal sprouts (visualized in whole mounts of the gluteus maximus with zinc iodide-osmium staining) was reduced from a mean of60.5% ± 3.2 (n = 11) for mice treated with botulinum toxin alone and61.5% ± 1.2 (n = 8) for those treated with non-immune serum, to37.3% + 3.6 (n = 8) in the anti-neural cell adhesion molecule injected series. In addition the anti-neural cell adhesion molecule treated material showed a higher proportion of shorter sprouts than the two control groups. A polyclonal antibody to Thy-1 (donated by Dr A. N. Barclay) did not reduce terminal sprouting significantly when compared with non-immune serum-treated controls.

We conclude that the expression of neural cell adhesion molecule in inactivated muscles may be at least partially responsible for the effectiveness of the terminal sprouting response.

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Cited by (31)

Ciliary neurotrophic factor is not required for terminal sprouting and compensatory reinnervation of neuromuscular synapses: Re-evaluation of CNTF null mice
2007, Experimental Neurology
Citation Excerpt :
Hence, it seems possible that surface-bound molecules with neuritogenic activity primarily induce terminal sprouting, which provides a unifying mechanism for both paralysis- and denervation-elicited sprouting. Candidates include cell adhesion molecules such as NCAM and tenascin-C (Booth et al., 1990; Cifuentes-Diaz et al., 1998; Lee et al., 1999, Walsh et al., 2000; but see Moscoso et al., 1998). Our conclusion that endogenous CNTF is dispensable for motor nerve sprouting, however, does not challenge the neuroprotective and myoprotective effects of CNTF or its therapeutic potential for degenerative neuromuscular diseases (cf., Sendtner, 1996; Vergara and Ramirez, 2004; Pun et al., 2006).
Loss of synaptic activity or innervation induces sprouting of intact motor nerve terminals that adds or restores nerve–muscle connectivity. Ciliary neurotrophic factor (CNTF) and terminal Schwann cells (tSCs) have been implicated as molecular and cellular mediators of the compensatory process. We wondered if the previously reported lack of terminal sprouting in CNTF null mice was due to abnormal reactivity of tSCs. To this end, we examined nerve terminal and tSC responses in CNTF null mice using experimental systems that elicited extensive sprouting in wildtype mice. Contrary to the previous report, we found that motor nerve terminals in the null mice sprout extensively in response to major sprouting-stimuli such as exogenously applied CNTF per se, botulinum toxin-elicited paralysis, and partial denervation by L4 spinal root transection. In addition, the number, length and growth patterns of terminal sprouts, and the extent of reinnervation by terminal or nodal sprouts, were similar in wildtype and null mice. tSCs in the null mice were also reactive to the sprouting-stimuli, elaborating cellular processes that accompanied terminal sprouts or guided reinnervation of denervated muscle fibers. Lastly, CNTF was absent in quiescent tSCs in intact, wildtype muscles and little if any was detected in reactive tSCs in denervated muscles. Thus, CNTF is not required for induction of nerve terminal sprouting, for reactivation of tSCs, and for compensatory reinnervation after nerve injury. We interpret these results to support the notion that compensatory sprouting in adult muscles is induced primarily by contact-mediated mechanisms, rather than by diffusible factors.
Ectopic expression of NCAM in skeletal muscle of transgenic mice results in terminal sprouting at the neuromuscular junction and altered structure but not function
2000, Molecular and Cellular Neurosciences
The neuromuscular system provides an excellent model for the analysis of molecular interactions involved in the development and plasticity of synaptic contacts. The neural cell adhesion molecule (NCAM) is believed to be involved in the development and plasticity of the neuromuscular junction, in particular the axonal sprouting response observed in paralyzed and denervated muscle. In order to explore the role of myofiber NCAM in modulating the differentiation of motor neurons, we generated transgenic mice expressing a GPI-anchored NCAM isoform that is normally found in developing and denervated muscle, under the control of a skeletal muscle-specific promoter. This results in the constitutive expression of NCAM at postnatal ages, a time when the endogenous mouse NCAM is absent from the myofiber. We found that a significant number of neuromuscular junctions in adult transgenic animals displayed terminal sprouting (>20%) reminiscent of that elicited in response to cessation of neuromuscular activity. Additionally, a significant increase in the size and complexity of neuromuscular synapses as a result of extensive intraterminal sprouting was detected. Electrophysiological studies, however, revealed no significant alterations of neuromuscular transmission at this highly efficient synapse. Sprouting in response to paralysis or following nerve crush was also significantly enhanced in transgenic animals. These results suggest that in this ectopic expression model NCAM can directly modulate synaptic structure and motor neuron–muscle interactions. The results contrast with knockout experiments of the NCAM gene, where very limited changes in the neuromuscular system were observed.
Association of adhesive macromolecules with terminal sprouts at the neuromuscular junction after botulinum treatment
1999, Otolaryngology - Head and Neck Surgery
Small quantities of botulinum toxin (BTX) are useful in the treatment of certain movement disorders, such as laryngeal spasmodic dysphonia, blepharospasm, and cervical dystonia. However, the corrective paralytic effects of BTX are only temporary, in part because of the formation of remodeled neuromuscular junctions. Here, we questioned whether various factors within and near the neuromuscular junction could contribute to the remodeling seen after BTX treatment. BTX was injected subcutaneously in the region of the levator auris longus muscle. At 1-week intervals, levator auris longus muscles were removed and examined histochemically. As previously described, BTX treatment results in a progressive elongation of end plates. The neural cell adhesion molecule was not associated with the elongated end plates but was associated with the BTX-induced nerve sprouts after long intervals (3 to 4 weeks). Similarly, after BTX, laminin-1 (composed of α1, β1, and γ1 chains) reactivity was associated with the nerve sprouts, but not with the end plates. Laminin β2 reactivity at the end plate dispersed somewhat within 1 week but remained diffusely associated with the elongating end plates for up to 5 weeks. Together these results suggest that neural cell adhesion molecule and laminins may participate in the sprouting observed after BTX treatment and that alterations in laminin β2 expression may participate in initial loss of contacts. (Otolaryngol Head Neck Surg 1999;120:255-61.)
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The response of different types of skeletal muscle fibers to a snake venom PLA₂ myotoxin was tested in vivo by injecting ACL myotoxin (ACLMT) into mice. Both the soleus (slow-twitch) and gastrocnemius (fast-twitch) were examined at different time periods (3 h, 3 and 21 d) after the injection. All animals received 5 mg/kg myotoxin into the subcutaneous lateral region of the right hind limb, near the Achilles tendon; contralateral muscles were used as controls. Cross-sections (10 μm) of frozen muscle tissue were cut from the medial region of the muscle. Alternate serial sections were stained either with toluidine blue or for acid phosphatase, myofibrillar ATPase activity after alkali (pH 10.3) or acid preincubation (pH 4.3), succinate dehydrogenase or acetylcholinesterase. Several stages of necrosis were observed 3 h after ACLMT injection, in both superficial and deep regions of both muscles. In these same regions 3 d after injection, clusters of regenerated muscle fibers were present, and some of them presented AChE activity. Twenty-one days after ACLMT injection the muscle fibers of soleus and gastrocnemius presented only chronic signs of damage such as split fibers and centralized nuclei. Using m-ATPase reactions it was possible to determine that both muscle fiber types I and II were injured in both muscles. The number of type IIC fibers was significantly increased, and the number of type II fibers significantly decreased in the gastrocnemius 21 d after ACLMT injection, suggesting a change in muscle fiber type from type II to type I, through type IIC. The increased number of type IIC fibers and the presence of AChE activity in clusters of regenerating fibers and split fibers indicate that injury by ACLMT produces axonal remodeling and muscle fiber type change.
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The neuromuscular system provides an ideal model for the elucidation of the molecular mechanisms involved in neuron–target cell interactions, and the development and maintenance of synaptic connections. Many processes important in neuronal development occur during the formation of neuromuscular connections and synaptogenesis. These include the process of pathfinding by motor neurons, selection of target tissues and synaptogenesis. In the present paper we discuss the molecular mechanisms that probably play a role in the development of skeletal muscle innervation, with particular emphasis on cell adhesion molecules (CAMs).
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An antibody to neural cell adhesion molecule impairs motor nerve terminal sprouting in a mouse muscle locally paralysed with botulinum toxin

Abstract

Brain Res.

Neuroscience

Brain Res.

Molec. Immun.

Cell

Immunol. Lett.

Growth of nerve implants in voluntary muscle

J. Anat.

The density of Thy-1 on axonal membrane of different rat nerves

J. Neurochem.

Neurite outgrowth on muscle cell surfaces involves matrix receptors as well as Ca2+-dependent and -independent cell adhesion molecules

Motor nerve sprouting

A. Rev. Neurosci.