A role for nociceptive, myelinated nerve fibers in itch sensation

Abstract

Despite its clinical importance, the underlying neural mechanisms of itch sensation are poorly understood. In many diseases, pruritus is not effectively treated with antihistamines, indicating the involvement of nonhistaminergic mechanisms. To investigate the role of small myelinated afferents in nonhistaminergic itch, we tested, in psychophysical studies in humans, the effect of a differential nerve block on itch produced by intradermal insertion of spicules from the pods of a cowhage plant (Mucuna pruriens). Electrophysiological experiments in anesthetized monkey were used to investigate the responsiveness of cutaneous, nociceptive, myelinated afferents to different chemical stimuli (cowhage spicules, histamine, capsaicin). Our results provide several lines of evidence for an important role of myelinated fibers in cowhage-induced itch: (1) a selective conduction block in myelinated fibers substantially reduces itch in a subgroup of subjects with A-fiber-dominated itch, (2) the time course of itch sensation differs between subjects with A-fiber- versus C-fiber-dominated itch, (3) cowhage activates a subpopulation of myelinated and unmyelinated afferents in monkey, (4) the time course of the response to cowhage is different in myelinated and unmyelinated fibers, (5) the time of peak itch sensation for subjects with A-fiber-dominated itch matches the time for peak response in myelinated fibers, and (6) the time for peak itch sensation for subjects with C-fiber-dominated itch matches the time for the peak response in unmyelinated fibers. These findings demonstrate that activity in nociceptive, myelinated afferents contributes to cowhage-induced sensations, and that nonhistaminergic itch is mediated through activity in both unmyelinated and myelinated afferents.

Figure 1.

Psychophysical studies of itch in normal human subjects. A, Average time course of itch, pricking/stinging, and burning sensations produced by topical application of cowhage spicules to the back of the hand (n = 14). B, Ratings of cowhage-evoked sensations decreased during a selective blockade of myelinated fiber function. C, Changes in peak itch and itch-AUC during selective myelinated fiber block. Subjects were considered to have an A-fiber-dominated itch if their itch decreased during the block such that their normalized AUC was <0.3 (n = 6, solid circles). Subjects with AUC > 0.4 were considered to have C-fiber-dominated itch (n = 8, open circles). Data during the block were normalized by dividing by the response before the block. D, In the absence of a nerve block, the average time course of itch sensation differs between subjects whose itch responded well to the selective myelinated fiber block (n = 6) and those that did not (n = 8).

Figure 2.

Cowhage activates mechano-sensitive, nociceptive A-fibers. Electrophysiological recordings were performed from nociceptive, myelinated afferents in monkeys. A, This myelinated afferent responded during the application of cowhage (41 spicules) to the RF (gray bar), presumably due to the mechanical stimulation associated with insertion of the spicules. Approximately 1 min after application, the fiber was activated. The response slowly decreased over the 10 min observation period. The discharge rate was irregular throughout the observation period. The total response of this fiber during the 10 min after active cowhage application was 559 APs. Each dot corresponds to an AP. Insert, Expanded time course of response 5.5 to 6 min after spicule insertion (vertical line corresponds to time of AP). B, Response of the same afferent to the prior application of inactive cowhage (40 spicules). Spicule application induced a short-lasting activation of the afferent and also some activity during the 10 min observation period. The total response of this fiber during the 10 min after inactive cowhage application was 54 APs. Insert, Action potential waveforms for the first 20 sequential APs after spicule insertion. C, This myelinated afferent responded to cowhage (21 spicules) with a bursting discharge. The instantaneous frequency varied between ∼0.1 and 100 Hz. The total response during the 10 min observation period was 170 APs. Insert, Expanded time course of response at indicated times (vertical line corresponds to time of AP). The bursting nature of the response is evident. D, Response of the same afferent as in C to the application of inactive cowhage (16 spicules). Only a single AP occurred during the 10 min after inactive cowhage application. Insert, Action potential waveforms for the first 20 sequential APs obtained during spicule insertion. Gray bar indicates the cowhage-application period. E, Average response of myelinated fibers to cowhage. Only those fibers that met the criteria for an evoked response are included (n = 23). The response to active cowhage (solid circles) was substantially greater than to inactive cowhage (open circles; mean ± SEM, bin size = 30 s). F, Responses to cowhage are variable. The responses to inactive and subsequent responses to active cowhage are shown for the 23 cowhage-responsive fibers. Each line corresponds to the response of one fiber. The magnitude of the responses varied within fibers (i.e., across trials) and also between fibers. There was not a significant change in the median response (red line) between trials and thus no obvious signs of tachyphylaxis or sensitization of the response to active cowhage.

Figure 3.

Comparison of responses to cowhage, histamine, and capsaicin. A, Venn diagram indicating incidence of response to topical cowhage and intradermal injection (10 μl) of histamine (10 μg) and capsaicin (10 μg) for 50 high-threshold, myelinated fibers tested with all three substances. Number in parentheses indicates number of fibers that were insensitive to mechanical stimuli. B, Net response during the 5 min observation period following administration of cowhage, histamine, and capsaicin to the receptive field of myelinated fibers (shaded bars) and unmyelinated fibers (open bars; median with quartiles and 5/95% percentiles). In A-fibers, the median cowhage response (184 APs/5 min) was approximately seven times larger than the median response to histamine (25APs/5 min) or capsaicin (27.5 APs/5 min) (Kruskal–Wallis ANOVA, p < 0.001, followed by multiple Mann–Whitney U tests, corrected for multiple comparisons, *p < 0.05, **p < 0.01). Within the C-fiber class, the response to cowhage was significantly larger than the response to histamine (Kruskal–Wallis ANOVA, p < 0.05, followed by Mann–Whitney U test, #p < 0.05). In A- and C-fibers, the response magnitudes to histamine and capsaicin were similar, but the cowhage response was significantly larger in A- than in C-fibers (***p < 0.001, Mann–Whitney U test). For this analysis, mechano-sensitive and -insensitive afferents were pooled, and only responders were included. C, Cowhage and histamine responses are not correlated. The magnitude of the cowhage and histamine net response in units that were tested with both pruritic stimuli did not correlate. Units responsive to either cowhage or histamine had variable response magnitudes, the cowhage responses being larger overall (note different scales on the x- and y-axes). These data suggest that different cellular signaling mechanisms are engaged by histamine and cowhage. Net response: response to active agent minus response to control; negative results were assigned zero.

Figure 4.

Comparison of the time course of responses to cowhage. A, The peak response of the unmyelinated fibers (open circles) occurred earlier than in myelinated fibers (filled circles). For each fiber, the response at a given time point was normalized by dividing by that fiber's maximum response. B, The peak itch sensation in subjects with A-fiber-dominated itch occurred significantly later than in subjects with C-fiber-dominated itch. Similarly, the peak response to cowhage occurred later in myelinated fibers than in unmyelinated fibers (Kruskal–Wallis ANOVA, p < 0.001 followed by multiple Mann–Whitney U tests corrected for multiple comparisons, *p < 0.05, **p < 0.01). Medians, quartiles and 5/95% percentiles are plotted.