Elsevier

Neuroscience

Volume 152, Issue 4, 9 April 2008, Pages 877-887
Neuroscience

Cellular neuroscience
Facilitated sprouting in a peripheral nerve injury

https://doi.org/10.1016/j.neuroscience.2008.01.060Get rights and content

Abstract

During regeneration of injured peripheral nerves, local conditions may influence how regenerative axon sprouts emerge from parent axons. More extensive lesions might be expected to disrupt such growth. In this work, we discovered instead that long segmental crush injuries facilitate the growth and maturation of substantially more axon sprouts than do classical short crush injuries (20 mm length vs. 2 mm). At identical distances from the proximal site of axon interruption there was a 45% rise in the numbers of neurofilament labeled axons extending through a long segmental crush zone by 1 week. By 2 weeks, there was a 35% greater density of regenerating myelinated axons in long compared with short crush injuries just beyond (5 mm) the proximal injury site. Moreover, despite the larger numbers of axons, their maturity was identical and they were regular, parallel, associated with Schwann cells (SCs) and essentially indistinguishable between the injuries. Backlabeling with Fluorogold indicated that despite these differences, the axons arose from similar numbers of parent motor and sensory neurons. Neither injury was associated with ischemia. Both injuries were associated with rises in GFAP (glial acidic fibrillary protein) and p75 mRNAs, markers of SC plasticity but p75, GFAP and brain-derived neurotrophic factor mRNAs did not differ between the injuries. There was a higher local mRNA level of GAP43/B50 at 7 days following injury and a higher sonic hedgehog protein (Shh) mRNA at 24 h in long crush zones. GAP43/B50 protein and SHH protein both had prominent localization within regenerating axons.

Long segmental nerve trunk crush injuries do not impair regeneration but instead generate greater axon plasticity that results in larger numbers of mature myelinated axons. The changes occur without apparent change in SC activation, overall nerve architecture or nerve blood flow. While the mechanism is uncertain, the findings indicate that manipulation of the nerve microenvironment can induce substantial changes in regenerative sprouting.

Section snippets

Animals

The experiments used male Sprague–Dawley rats of initial weight 200–330 g maintained in cages with shavings covered plastic flooring and exposed to normal day-light cycles. Procedures were carried out under pentobarbital anesthesia (65 mg/kg) and postoperative meperidine or butorphanol were provided for analgesia. The number of rats used was minimized and their suffering minimized using appropriate anesthesia and analgesia. All procedures were reviewed an approved by the University of Calgary

Heightened axon outgrowth

We studied longitudinal sections of nerves, taken through the crush zone and labeled with an antibody directed against a Nf subunit (Nf200). These sections were taken at an early time point, 7 days after injury, to address whether enhanced sprouting occurred soon after the onset of regrowth. In our experience, a 7 day time point after crush is optimal for identifying new axons that express Nf and that can be distinguished from breakdown Nf products during Wallerian-like degeneration. At

Overall findings

In this work, we describe an interesting, unexpected and novel response of regenerating peripheral nerve axons to their microenvironment. Injuries with a simple alteration in the extent of direct injury had dramatically different impacts on axon behavior. We believe that longer segmental crush injuries may actually be more common in complex human traumatic injuries than narrow circumscribed “surgical” focal crushes traditionally applied by experimentalists. Both injuries are classified as type

Conclusion

In summary, we have identified a form of facilitated axon sprouting and maintenance associated with a simple alteration in the microenvironment of an injured peripheral nerve. Understanding the signals for this phenomenon may allow it to be exploited when this kind of plasticity is desirable during regeneration.

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