An ultrafast system for signaling mechanical pain in human skin

Abstract

The canonical view is that touch is signaled by fast-conducting, thickly myelinated afferents, whereas pain is signaled by slow-conducting, thinly myelinated ("fast" pain) or unmyelinated ("slow" pain) afferents. While other mammals have thickly myelinated afferents signaling pain (ultrafast nociceptors), these have not been demonstrated in humans. Here, we performed single-unit axonal recordings (microneurography) from cutaneous mechanoreceptive afferents in healthy participants. We identified A-fiber high-threshold mechanoreceptors (A-HTMRs) that were insensitive to gentle touch, encoded noxious skin indentations, and displayed conduction velocities similar to A-fiber low-threshold mechanoreceptors. Intraneural electrical stimulation of single ultrafast A-HTMRs evoked painful percepts. Testing in patients with selective deafferentation revealed impaired pain judgments to graded mechanical stimuli only when thickly myelinated fibers were absent. This function was preserved in patients with a loss-of-function mutation in mechanotransduction channel PIEZO2. These findings demonstrate that human mechanical pain does not require PIEZO2 and can be signaled by fast-conducting, thickly myelinated afferents.

Fig. 1. Humans are equipped with high-threshold and very fast conducting primary afferents.

(A) Location of myelinated HTMR receptive fields from recordings in the peroneal and radial nerves. Each red dot represents the location of an individual A-HTMR (n = 18). The pattern of receptive field spots, mapped with a Semmes-Weinstein monofilament, is shown for two A-HTMRs (marked by arrows; top, radial; bottom, peroneal). Receptive field spots were redrawn on photographic images so they can easily be seen. The horizontal lines represent the medial-lateral dimension, and the vertical lines represent the proximal-distal dimension. The average size of an A-HTMR receptive field, mapped using a filament force six times higher than that for an A-LTMR, was 26.8 mm² (±6.9; n = 9). This was significantly smaller than the receptive field of field afferents (100.5 ± 7.2 mm²; n = 51; P < 0.0001, Dunnett’s test) but was not different from that of SA1 afferents (41.6 ± 7.5 mm²; n = 17; P > 0.05, Dunnett’s test). (B) Brush responses of an A-HTMR and a field afferent. Using a soft or a coarse brush, the skin area centered on the receptive field of the recorded afferent was gently stroked at 3 cm/s. The field afferent responded vigorously to soft brush stroking. The A-HTMR did not respond to soft brush stroking, but it did respond to coarse brush stroking. The mechanical threshold of this A-HTMR was 4 mN, and the conduction velocity was 52 m/s. It responded to pinching, and its receptive field moved with skin translocation. Freq, frequency. (C) Mechanical threshold distribution of HTMRs and LTMRs in the recorded sample. For RA1 afferents, the preferred stimulus is hair movement, so monofilament thresholds were not measured. For A-HTMR, the median mechanical threshold was 10.0 mN (Q, 5.5–20.0; n = 18). This was significantly higher than the mechanical thresholds of all tested A-LTMR types (at least P < 0.001 for all individual comparisons, Dunn’s test) but was not different from C-HTMRs (10.0 mN; Q, 10.0–27.0; n = 5). (D) Spike activity of an A-HTMR to electrical and mechanical stimulations of the receptive field. Individual electrically and mechanically evoked spikes were superimposed on an expanded time scale to show that the electrically stimulated spike (used for latency measurement) was from the same unit as the one that was mechanically probed at the receptive field. (E) Conduction velocities of HTMRs and LTMRs to surface electrical stimulation (and monofilament tapping in case of one HTMR, conducting at 30 m/s). The data show individual and average (±SEM) conduction velocities of single afferents from peroneal (circles) and radial (diamonds) nerves. Conduction velocities of peroneal A-HTMRs (33.5 ± 2.1; n = 13) were statistically indistinguishable from peroneal A-LTMRs [SA1: 39.8 ± 2.3, n = 10; SA2: 38.6 ± 4.0, n = 4; RA1: 36.8 ± 2.8, n = 6; field: 34.3 ± 1.3, n = 18; F(4,46) = 1.70; P = 0.17, one-way analysis of variance (ANOVA)]. All three peroneal C-HTMRs were conducting at 0.9 m/s. In comparison to the peroneal nerve, conduction velocities of A-fiber types were faster in the radial nerve as expected (A-HTMR: 54.5 ± 2.4, n = 4; SA1: 56.8 m/s, n = 1; SA2: 53.0 ± 3.3, n = 3; RA1: 48.7 ± 1.6, n = 3; field: 47.3 ± 0.2, n = 2) (46). Both radial C-HTMRs were conducting at 1.1 m/s. Conduction velocity of RA2 afferents was not measured. (F) Slowly adapting properties of an A-HTMR at higher indentation forces. Spike activity of a field afferent and an A-HTMR during monofilament stimulation at three different forces, applied using electronic filaments with force feedback. Compared to the field afferent, the A-HTMR showed a sustained response at a lower indentation force (see also fig. S1).

Fig. 2. Neural discharge of A-HTMRs and perception of mechanical pain in response to punctate forces.

(A) Peak discharge rates of human A-HTMRs for monofilament stimulation. The data show individual and average (±SEM) responses of nine A-HTMRs (≥30 m/s) to 5-s monofilament stimulation at eight different indentation forces. A significant linear fit was displayed (R² = 0.6981, P = 0.0098), and the response at the highest indentation force (3000 mN) was significantly higher than responses to all weaker indentation forces (at least P < 0.05, Dunnett’s test). Individual units are color-coded. (B) Psychophysical data for pain intensity to graded monofilament stimulation. Psychophysical pain ratings were collected from 16 healthy participants (dorsal foot: 8 participants, 12 trial sets; dorsal toe: 8 participants, 9 trial sets) from the microneurography sample. The monofilament force had a significant effect on psychophysical pain ratings [F(8,171) = 29.96; P < 0.0001, two-way ANOVA], with a significant pain of ~14 of 100 emerging at 260 mN, and higher pain ratings with increasing forces (Tukey’s multiple comparisons test). No difference in pain ratings was found between the foot and toe stimulation sites [F(1,171) = 0.1733; P = 0.6777, two-way ANOVA]; hence, the data were pooled for a subsequent analysis. (C) Psychophysical pain ratings as a function of neural discharge in A-HTMRs. A comparison of the average peak discharge rates of nine A-HTMRs and average psychophysical pain ratings for skin indentations (eight forces, 4 to 3000 mN) revealed a significant positive correlation (Pearson r = 0.8944, R² = 0.8, P = 0.0027). (D) Number of spikes produced by monofilament stimulation in human A-HTMRs. Individual and average (±SEM) responses of nine A-HTMRs to 5-s monofilament stimulation at eight different indentation forces. A significant linear fit was displayed (R² = 0.6361, P = 0.0177). Individual units are color-coded. (E) Spike activity of an A-HTMR during the first 0.5 s of monofilament stimulation (with force feedback). Top: Neural discharge rates. Middle: Spike markers and indentation force markers. Bottom: Neural recording. The first 0.5 s was selected as the onset period (dynamic phase) of monofilament stimulation (total duration, 5 s). (F) Mean discharge rates of human A-HTMRs for monofilament onset. Individual and average (±SEM) responses of nine A-HTMRs to the onset period (0.5 s) of monofilament stimulation at eight different indentation forces. A significant linear fit was displayed (R² = 0.6784, P = 0.0120). Individual units are color-coded.

Fig. 3. Mechanical pain sensitivity is dependent on Aβ afferents but not PIEZO2 stretch-gated ion channels.

(A) Psychophysical pain ratings to graded monofilament stimulation of the peroneal foot for two patients with Aβ deafferentation, two patients with HSAN-V and six healthy participants (pooled means ± SEM and individual data comprising mean of triplicates). A comparison of the pain ratings between patients with HSAN-V and healthy participants revealed no effect of condition—healthy versus HSAN-V [F(1,197) = 1.372; P = 0.2428, two-way ANOVA]. In contrast, a comparison of patients with HSAN-V and healthy participants, on one hand, and patients with Aβ deafferentation, on the other hand, revealed a significant effect of condition—healthy versus Aβ deafferentation [F(1,198) = 15.72, P = 0.0001] and HSAN-V versus Aβ deafferentation [F(1,89) = 13.01, P = 0.0005]. ABD, Aβ deafferentation. (B) Psychophysical pain ratings to graded monofilament stimulation of the radial forearm for two patients with PIEZO2^LOF, in addition to average pain ratings from four healthy participants (pooled means ± SEM and individual data comprising mean of triplicates). A comparison of the pain ratings between patients with PIEZO2^LOF and four healthy participants revealed slightly higher ratings in the patients with PIEZO2^LOF compared to healthy participants [F(1,144) = 4.079; P = 0.0453, two-way ANOVA]. The two patients with Aβ deafferentation reported no pain to graded monofilament stimulation of the forearm.