Intradermal injection of capsaicin in humans produces degeneration and subsequent reinnervation of epidermal nerve fibers: correlation with sensory function

Abstract

The ability of capsaicin to excite and subsequently to desensitize a select group of small sensory neurons has made it a useful tool to study their function. For this reason, application of capsaicin to the skin has been used for a variety of painful syndromes. We examined whether intradermal injection of capsaicin produced morphological changes in cutaneous nerve fibers that would account for its analgesic properties by comparing cutaneous innervation in capsaicin-treated skin with psychophysical measures of sensation. At various times after capsaicin injection, nerve fibers were visualized immunohistochemically in skin biopsies and were quantified. In normal skin the epidermis is heavily innervated by nerve fibers immunoreactive for protein gene product (PGP) 9.5, whereas fibers immunoreactive for substance P (SP) and calcitonin gene-related peptide (CGRP) are typically associated with blood vessels. There was nearly complete degeneration of epidermal nerve fibers and the subepidermal neural plexus in capsaicin-treated skin, as indicated by the loss of immunoreactivity for PGP 9.5 and CGRP. The effect of capsaicin on dermal nerve fibers immunoreactive for SP was less obvious. Capsaicin decreased sensitivity to pain produced by sharp mechanical stimuli and nearly eliminated heat-evoked pain within the injected area. Limited reinnervation of the epidermis and partial return of sensation occurred 3 weeks after treatment; reinnervation of the epidermis was approximately 25% of normal, and sensation improved to 50-75% of normal. These data show that sensory dysfunction after capsaicin application to the skin results from rapid degeneration of intracutaneous nerve fibers.

Fig. 1.

Confocal images showing innervation of epidermis and superficial dermis for one subject at 72 hr after a single injection of vehicle (Veh) or capsaicin doses of 0.2, 2.0, or 20 μg. Nerve fibers (N) immunoreactive for PGP 9.5 appear yellow-green; the basement membrane (B) and vessels (V) appear red. Scale bars, 100 μm.

Fig. 2.

Somatic sensation and the number of ENFs for all five subjects at 72 hr after injection of capsaicin. Data are expressed as the mean (± SEM) percent change from the data for vehicle-treated skin. For heat pain and cold sensation, data represent the change in the magnitude of sensation. Mechanical pain sensation is represented as the change in the proportion of stimuli perceived as painful.

Fig. 3.

Confocal images showing denervation and reinnervation of epidermis and superficial dermis by PGP 9.5-immunoreactive nerve fibers for one subject. Biopsies were taken from capsaicin-treated (20 μg) skin at 1, 2, and 4 weeks after injection and from normal untreated skin. The appearance of nerve fibers immunoreactive for PGP 9.5 is the same as that described in Figure 1. Scale bars, 100 μm.

Fig. 4.

Confocal images of skin biopsies from one subject showing nerve fibers immunoreactive for CGRP and SP in normal skin and skin at 1 and 4 weeks after capsaicin injection.Right, CGRP-immunoreactive fibers were completely absent 1 week after capsaicin injection and reappeared 4 weeks after injection (arrows). Left, There was not a complete loss of SP-immunoreactive fibers at 1 week after capsaicin injection, and one fiber can be seen orientedhorizontally below the basement membrane (arrow). At 4 weeks after capsaicin injection, fibers were occasionally found deep in the dermis (arrow) and oriented verticallytoward the basement membrane. Scale bars, 100 μm.

Fig. 5.

The mean (± SEM) change in sensation and in the number of ENFs for all subjects through 1–6 weeks after injection of 20 μg of capsaicin. Data are presented as the percent change from normal untreated skin.

Fig. 6.

The localization of degeneration of ENFs after an intradermal injection of 20 μg of capsaicin as shown by confocal images of skin biopsy from one subject. Top, Montage of confocal images that span across the skin biopsy. Abundant ENFs are seen only at the far right portion of the biopsy, which was located outside the capsaicin injection site and presumably not exposed to the neurotoxin. Bottom, The two opposite ends of the skin biopsy, outlined by thesquares in the top image, shown at greater magnification to illustrate differences in epidermal innervation.