Peter P. Robinson

Sympathetic nerve sprouting fails to occur in the trigeminal ganglion after peripheral nerve injury in the rat

Peripheral nerve injury induces sprouting of sympathetic nerve fibers in dorsal root ganglia after spinal nerve injury. In the present study, we sought to determine the extent of intraganglionic noradrenergic sprouting in the trigeminal system. The inferior alveolar nerve, a major branch of the mandibular division, or the infraorbital nerve of the maxillary division was either ligated or chronically constricted in Sprague-Dawley rats and recovery permitted for either 2-3 or 6-9 weeks. In some animals both nerves were injured. Using immunohistochemistry with tyrosine hydroxylase antibodies, we found no signs of sympathetic nerve fiber sprouting in the trigeminal ganglion after injury. In contrast, sciatic nerve injury in rat littermates induced a widespread autonomic nerve outgrowth in affected DRGs. Thus, sensory ganglion sympathetic nerve sprouting does not seem to be a general outcome of PNS injury, but is restricted to certain specific locations. Sympathetic nerve fiber networks that surround primary sensory neurons have been suggested to form a structural basis for interactions between the sympathetic and sensory nervous systems after PNS injury. Such interactions, sometimes resulting in paraesthesia or dysaesthesia in patients, appear to be less common in territories innervated by the trigeminal nerve than in spinal nerve regions. The lack of injury-induced intraganglionic sympathetic sprouting in the trigeminal ganglion may help to explain this observation.

Nerve guides manufactured from photocurable polymers to aid peripheral nerve repair.

The peripheral nervous system has a limited innate capacity for self-repair following injury, and surgical intervention is often required. For injuries greater than a few millimeters autografting is standard practice although it is associated with donor site morbidity and is limited in its availability. Because of this, nerve guidance conduits (NGCs) can be viewed as an advantageous alternative, but currently have limited efficacy for short and large injury gaps in comparison to autograft. Current commercially available NGC designs rely on existing regulatory approved materials and traditional production methods, limiting improvement of their design. The aim of this study was to establish a novel method for NGC manufacture using a custom built laser-based microstereolithography (μSL) setup that incorporated a 405 nm laser source to produce 3D constructs with ∼ 50 μm resolution from a photocurable poly(ethylene glycol) resin. These were evaluated by SEM, in vitro neuronal, Schwann and dorsal root ganglion culture and in vivo using a thy-1-YFP-H mouse common fibular nerve injury model. NGCs with dimensions of 1 mm internal diameter × 5 mm length with a wall thickness of 250 μm were fabricated and capable of supporting re-innervation across a 3 mm injury gap after 21 days, with results close to that of an autograft control. The study provides a technology platform for the rapid microfabrication of biocompatible materials, a novel method for in vivo evaluation, and a benchmark for future development in more advanced NGC designs, biodegradable and larger device sizes, and longer-term implantation studies.

Spontaneous and mechanically evoked afferent activity originating from myelinated fibres in ferret inferior alveolar nerve neuromas

&NA; The inferior alveolar nerve is a predominantly sensory branch of the trigeminal nerve which runs within a bony canal, and is frequently damaged in patients. A small proportion of these patients develop neuropathic pain, and this may result from neural activity generated at the injury site. To investigate this abnormality we have used electrophysiological techniques in an animal model to determine the level of spontaneous activity and mechanical sensitivity of myelinated fibres ending in a neuroma of the inferior alveolar nerve. In 20 anaesthetised adult male ferrets the left inferior alveolar nerve was ligated in the region of the third premolar tooth, cut distally, and recovery permitted for periods of 3–113 days prior to making single unit recordings from the nerve central to the injury. The proportion of units which were spontaneously active ranged from 0% to 26%, with discharge rates of 0.3–12.9 Hz. Discharge in response to mechanical stimulation of the neuroma was found in 0–36% of the units. Both spontaneous activity and mechanical sensitivity were significantly higher after shorter recovery periods and the majority of the spontaneously active units was also mechanically sensitive. These data reveal that the inferior alveolar nerve responds to injury in a similar way to some other peripheral nerves, and the neural activity generated at the injury site may play a role in the development of dysaesthesia.

Interleukin-10 reduces scarring and enhances regeneration at a site of sciatic nerve repair.

Abstract  Axonal regeneration at a site of peripheral nerve repair can be impeded by the formation of scar tissue, which creates a mechanical barrier and initiates the development of multiple branched axonal sprouts that form a neuroma. We have investigated the hypothesis that the application of a scar‐reducing agent to the nerve repair site would permit better axonal regeneration. In anaesthetised C57 Black‐6 mice, the left sciatic nerve was sectioned and immediately re‐approximated using four epineurial sutures. In five groups of eight mice, we injected transforming growth factor‐β3 (50 or 500 ng), interleukin‐10 (IL‐10) (125 or 500 ng), or saline into and around the repair site, both before and after the nerve section. Another group of eight animals acted as sham‐operated controls. After 6 weeks, the outcome was assessed by recording compound action potentials (CAPs), measuring collagen levels using picrosirius red staining, and counting the number of myelinated axons proximal and distal to the repair. CAPs evoked by electrical stimulation distal to the repair were significantly smaller in all repair groups except for the low‐dose IL‐10 group, where they were not significantly different from that in controls. The area of staining for collagen had significantly increased in all repair groups except for the low‐dose IL‐10 group, which was not significantly different from that in controls. The myelinated fibre counts were always higher distal to the repair site, but there were no significant differences between groups. We conclude that administration of a low‐dose of IL‐10 to a site of sciatic nerve repair reduces scar formation and permits better regeneration of the damaged axons.

Changes in vanilloid receptor 1 (TRPV1) expression following lingual nerve injury.

We have investigated a possible role for vanilloid receptor 1 (TRPV1), a transducer of noxious stimuli, in the development of neuropathic pain following injury to a peripheral branch of the trigeminal nerve. In nine adult ferrets the left lingual nerve was sectioned and recovery permitted for 3 days, 3 weeks or 3 months (3 ferrets per group). A retrograde tracer, fluorogold, was injected into the damaged nerve to identify associated cell bodies in the trigeminal ganglion. Three further ferrets, receiving only tracer injection, served as uninjured controls. Indirect immunofluorescence for TRPV1 and image analysis was used to quantify the percentage area of staining (PAS) of TRPV1 in the left and right lingual nerves. Additionally, the proportion of fluorogold positive and fluorogold negative cells expressing TRPV1 in the ganglion was determined. TRPV1 expression increased significantly at the injury site of damaged nerves 3 days after injury and this was matched by a reduction in the proportion of fluorogold positive cells expressing TRPV1 in the ganglion. At 3 weeks TRPV1 expression at the injury site was still high, while in the ganglion was significantly greater than in the controls. In the 3‐month recovery group TRPV1 expression in both nerve fibres and ganglion cells, was not significantly different from controls and there were no changes in expression in the fluorogold negative cells in the ganglion at any time point studied. These data suggest that after injury there is an increase in the axonal transport of TRPV1 from the cell bodies to the damaged axons and this is followed by an increase in synthesis in the ganglion. These changes in expression may be involved in development of sensory disturbances or dysaesthesia after injury.

Surgical anatomy of the buccal nerve

The buccal nerve may be damaged during surgical procedures which require an incision along the external oblique ridge of the mandible and this study was undertaken to clarify its surgical anatomy. The course and relationships of the nerve were determined in 20 formalin-fixed cadaver specimens. The number of major branches of the nerve ranged from 4 to 8 and a mean of 3 branches was present as the nerve crossed the external oblique ridge. In 14 dissections the main trunk of the nerve crossed the external oblique ridge within 3 mm of the deepest concavity, but in the other 6 it was 7-12 mm below this point. We conclude that incisions even 12 mm below the deepest concavity in the external oblique ridge could result in buccal nerve damage.

Afferent activity from myelinated inferior alveolar nerve fibres in ferrets after constriction or section and regeneration

&NA; To investigate possible peripheral mechanisms for post‐injury sensory disorders in the trigeminal system, we have made electrophysiological recordings from myelinated fibres in the inferior alveolar nerve (IAN) which have previously sustained an injury. In earlier experiments we have shown that axons in ligature‐induced neuromas of the IAN develop spontaneous activity and mechanical sensitivity. The present study has investigated these responses after two different types of injury. In 24 anaesthetised adult male ferrets the left IAN was either chronically constricted by four loose chromic gut ligatures (12 animals) or sectioned and regeneration permitted (12 animals). After recovery periods of 3 days, 1, 3, 6, 12 or 24 weeks, single unit recordings were made from the nerve proximal to the injury site. The proportion of units which were spontaneously active ranged from 0% to 19% after constriction injury and from 0% to 10% after nerve section and regeneration. Both groups revealed a marked variability between individual animals at similar time periods. Mechanical sensitivity was found in 0–42% of units after constriction and 0–25% of units after nerve section; both groups showed a significant negative correlation between mechanical sensitivity and recovery period. None of the fibres which had regained peripheral receptive fields was either spontaneously active or mechanically sensitive. There was no significant difference between the levels of spontaneous activity or mechanical sensitivity in the two groups or that previously found in ligature‐induced neuromas. Thus we conclude that widely differing types of peripheral nerve injury are capable of initiating similar raised levels of afferent activity in myelinated inferior alveolar nerve fibres.

Ectopic neural activity from myelinated afferent fibres in the lingual nerve of the ferret following three types of injury

Sensory disturbances following nerve injury may result from abnormal neural activity initiated at the injury site. We have studied the activity generated in the lingual nerve after three types of injury which may have variable potentials for the initiation of sensory disturbances. We have also compared the results with those found after damage to the inferior alveolar nerve, another branch of the trigeminal nerve, to determine whether differences in nerve fibre type or location affect the level of abnormal activity. In anaesthetised adult male ferrets the left lingual nerve was either ligated and cut distally, chronically constricted, or sectioned and allowed to regenerate. Following recovery periods of 3 days-6 months, single unit electrophysiological recordings were made from central to the injury site. After all three types of injury, some of the damaged axons at the injury site developed spontaneous activity (up to 36% of units) and mechanical sensitivity (up to 35% of units). There were significantly fewer spontaneously active units after ligation than after the other two types of injury but the level of mechanical sensitivity was not significantly different between the three types of injury. There was a significant increase in the spontaneous activity between 3 weeks and later recovery periods following both ligation and section injuries, and this late increase was not seen in our previous studies on the inferior alveolar nerve. Differences in the time-course of ectopic activity in adjacent branches of the trigeminal nerve suggest that the fibre types or anatomical relationships affect the outcome of injury.

The role of nerve growth factor in collateral reinnervation by cutaneous C-fibres in the rat

We have investigated whether chronic nerve growth factor (NGF) depletion affects the development of transmedian collateral reinnervation by C-fibres. Using a dye-labelled plasma extravasation technique in rats, the extent of transmedian innervation of the skin by C-fibres in the inferior alveolar nerve (IAN) was determined 8-10 weeks after sectioning and preventing regeneration of the contralateral IAN. Another group of animals were immunised against NGF prior to the nerve section and a third group acted as unoperated controls. A small but significant transmedian collateral reinnervation by C-fibres developed after contralateral denervation alone, but was not found in the animals also immunised against NGF. These results suggest that NGF is essential for the development of collateral reinnervation from cutaneous C-fibres.

The effect of injury on the properties of afferent fibres in the lingual nerve

The receptor properties of mechanosensitive afferent fibres in the lingual nerve have been studied using electrophysiological techniques in cats. In normal animals some fibres responded only to mechanical stimulation of filiform or fungiform papillae but others also responded when a cold stimulus was applied to the receptive field. Twelve weeks after crushing the lingual nerve, the regenerated fibres had slower conduction velocities but the receptor properties were not significantly different from normal. Twelve weeks after sectioning the lingual nerve, there was a greater reduction in conduction velocities and in addition the mechanoreceptive fields were smaller, force thresholds were higher, adaptation times longer and a smaller proportion of fibres responded to a cold stimulus. These results suggest that section injuries are more likely to result in persistent sensory abnormalities than crush injuries.