Pressure and stretch mechanosensitivity of peripheral nerve fibres following local inflammation of the nerve trunk

Abstract

Patients with non-specific limb pain often show signs of nerve mechanosensitivity, i.e. local tenderness over nerve trunks and pain in response to limb movements that cause nerve stretch. In such patients a nerve lesion is not apparent, and it has been suggested that local neural inflammation may be a key factor. The present study examines the extent to which nerve fibres in regions of local inflammation respond to small stretches, and whether functional changes occur throughout the primary afferent neurone. A local neuritis was induced in adult rats by wrapping oxidised cellulose saturated in complete Freund's adjuvant (CFA) around the peroneal or sciatic nerves. A small cut was made in the perineurium of some of the peroneal lesioned animals. A- and C-fibre recordings were made 2-10 days post-surgery from filaments dissected proximal to the lesion. Local mechanosensitivity was assessed using a glass probe and by small stretches. Responses to stretch and local pressure were recorded in 7% of C- and 8% of A-fibres from the peroneal nerve following CFA treatment with the sheath opened. A smaller proportion of stretch sensitive fibres were seen in sciatic and peroneal nerves after CFA treatment alone (2% of C- and 3% of A-fibres), but such fibres were not seen in control preparations. The most responsive fibres fired to 3% stretch, which is within the range of nerve stretch seen during normal limb movements. Less than 1% of stretch sensitive fibres had peripheral fields, indicating that most had probably degenerated distally.

Fig. 1

Diagram of electrophysiology setup in animals with (a) a peroneal nerve and (b) a sciatic nerve lesion. In (a) peroneal nerve fibres are stimulated proximal (S1) and distal (S2) to the lesion site (grey ellipse). In (b) sural nerve fibres are stimulated distal (S1) to the lesion site (grey ellipse). In both preparations, recordings are made from fine filaments teased from the sciatic nerve proximal to the lesion site. S1: stimulus 1, S2: stimulus 2; Rec: recording electrodes.

Fig. 2

Summary of local pressure responses following neuritis. (a) Proportion of A- and C-fibres responding to local pressure at the lesion in the CFA-treated peroneal lesion with the sheath opened (CFA peroneal (cut)) and intact (CFA peroneal), CFA-treated sciatic lesion, untreated animals, saline-treated sciatic lesion and saline-treated peroneal lesion with the sheath opened (saline peroneal (cut)). (b and c) Typical responses to local pressure at the treatment site for an A-fibre (b) and a C-fibre (c). The total number of units sampled and the number responding to pressure are shown for each group.

Fig. 3

Summary of stretch responses following neuritis. (a) Proportion of A- and C-fibres responding to stretch across the lesion in the CFA-treated peroneal lesion with the sheath opened (CFA peroneal (cut)) and intact (CFA peroneal), CFA-treated sciatic lesion, untreated animals and saline-treated sciatic lesion and saline-treated peroneal lesion with the sheath opened (saline peroneal (cut)). A typical (b) A-fibre and (c) C-fibre response to stretch at the lesion site. The erratic firing in (b) is caused by small fluctuations in the amount of stretch applied to the nerve. The total number of units sampled and the number responding to stretch are shown for each group.

Fig. 4

Proportion of A- and C-fibres failing to conduct through the lesion site in the CFA-treated peroneal lesioned animals with the sheath opened (CFA peroneal (cut)), intact (CFA peroneal), in untreated animals and in the saline-treated peroneal lesion with the sheath opened (saline peroneal (cut)). The total number of units sampled and the number not conducting through the lesion site are shown for each group.

Fig. 5

Proportion of conducting A- and C-fibres without peripheral fields in the CFA-treated peroneal lesioned animals with the sheath opened (CFA peroneal (cut)) and intact (CFA peroneal), in untreated animals and in the saline-treated peroneal lesion with the sheath opened (saline peroneal (cut)). The total number of units sampled and the number without peripheral fields are shown for each group.

Fig. 6

Proportion of spontaneously active A- and C-fibres following different nerve treatments or in untreated (naïve) nerves.

Fig. 7

Light microscopic appearance of the peroneal nerve at the treatment site (b–d) or the equivalent part of an untreated nerve (a). Treatments were CFA with the sheath intact (b and c) and CFA with the sheath opened (d). In the untreated nerve (a) myelin profiles appear normal. In (b) the myelinated profiles appear relatively normal with only a few absent profiles, whereas in (c) myelinated profiles appear sparse. Thus (b) and (c) give an indication of the variability of changes seen from animal to animal. In most CFA-treated animals with the sheath opened, there were very few surviving myelinated profiles at the lesion site, as can be seen in (d). Bar=50 μm.

Fig. 8

Relationship between lesion severity and A- and C-fibre conduction failure across the lesion site (Lowest rank=least damaged). The number of days post surgery is shown for each group. Trend lines have been fitted to all data points (rank correlation coefficient (r) for A-fibres=0.91, correlation coefficient (r) for C-fibres=0.87). Note that in a single CFA-treated animal with the sheath cut there was no A- or C-fibre conduction block and correspondingly this animal had one of the lowest rank values.

Fig. 9

Immunofluorescence staining of DRG following CFA treatment of the sciatic nerve (a). The nuclei of DAPI positive neuronal cells appear as large circular pale blue structures compared to the smaller elongated brighter blue nuclei of the satellite cells; nuclei of ATF3 positive cells appear bright green (arrows). ATF3 and DAPI images were overlaid using Photoshop (Adobe, USA). ATF3 expression following CFA treatment was 15.3%. Scale bars=100 μm. (b) Summary of ATF3 expression following nerve transection (n=5), CFA treatment (n=6) and saline treatment (n=7). Error bars=SEM.