Capsaicin responses in heat-sensitive and heat-insensitive A-fiber nociceptors

Abstract

The recently cloned vanilloid receptor (VR1) is postulated to account for heat and capsaicin sensitivity in unmyelinated afferents. We sought to determine whether heat and capsaicin sensitivity also coexist in myelinated nociceptive afferents. Action potential (AP) activity was recorded from single A-fiber nociceptors that innervated the hairy skin in monkey. Before intradermal injection of capsaicin (10 microg/10 microl) into the receptive field, nociceptors were classified as heat-sensitive (threshold, </=53 degrees C, 1 sec) or heat-insensitive afferents and as mechanically sensitive (von Frey threshold, <6 bar) or mechanically insensitive afferents. All heat-sensitive afferents (n = 16) were insensitive to mechanical stimuli but responded to the intradermal injection of capsaicin (69 +/- 7 APs in 10 min). Responsiveness to mechanical stimuli, thermal stimuli, and capsaicin varied in their receptive fields; the majority of receptive field sites (24 of 36) were responsive to only one or two stimulus modalities, whereas only eight sites responded to all three modalities. For most heat-insensitive afferents, the activity induced by the capsaicin injection did not exceed the activity induced by needle insertion alone. However, the largest response to capsaicin (314 +/- 98 APs in 10 min) was observed for five afferents that were insensitive to heat as well as mechanical stimuli and therefore may be classified as cutaneous chemoreceptors. These results suggest that A-fiber nociceptors play a role in the pain and hyperalgesia associated with capsaicin injection. Our finding that a subgroup of capsaicin-sensitive A-fiber nociceptors are insensitive to heat predicts the existence of heat-insensitive capsaicin receptors.

Fig. 1.

Specimen recordings of a response to capsaicin. A, Response to intradermal injection of capsaicin (10 μg/10 μl) for a type II heat-sensitive afferent (heat threshold, 49°C, 1 sec; mechanical threshold, 8.6 bar). The capsaicin was injected into an area responsive to von Frey hair stimulation. The needle insertion induced some activity in the fiber that was followed by a strong discharge after injection of capsaicin. Activity could be observed for 9 min after the injection. B, Response for a type I heat-insensitive afferent (heat threshold, 53°C, 4 sec; mechanical threshold, 4.6 bar). The capsaicin was injected into the mechanical receptive field. The needle insertion itself induced a strong activation. The excitation caused by the injection of capsaicin was short-lived (<5 sec), and no further action potentials were observed for 10 min after the injection. Top panels,Time course of action potential activity. Every vertical line marks the time of occurrence of an action potential.Middle panels, Response to needle insertion and capsaicin injection illustrated on an expanded time scale.Bottom panels, Instantaneous frequency. Everycircle represents the instantaneous frequency and time of occurrence of an action potential. Open trianglesindicate the time point of tone from computer to prompt needle insertion. Filled triangles indicate the time of tone for capsaicin injection.

Fig. 2.

Average time course of capsaicin-evoked responses.A, Fibers with a type II heat response (n = 16). Type II fibers showed a strong response to capsaicin injection and only a weak response to needle insertion.B, Subset of heat-insensitive afferents with a vigorous response to capsaicin. These afferents (n = 5) were not type II heat-sensitive afferents but responded with >100 action potentials per 10 min to capsaicin injection. Open barsindicate the response to needle insertion, and filled bars represent the response to capsaicin (bin size = 1 min). Inset, Expanded display at time of injection (bin size = 15 sec).

Fig. 3.

Comparison of the responses to needle insertion and the responses during the first 15 sec after the capsaicin injection. A, Type II heat-sensitive afferents. The response after capsaicin injection was significantly larger than the response to needle insertion alone (p < 0.001; paired t test; n = 16).B, Heat-insensitive afferents. In these afferents, capsaicin injection did not lead to a significantly larger response than needle insertion alone (n = 24). The five heat-insensitive afferents that did not respond to capsaicin are excluded.

Fig. 4.

Distribution of the total response to capsaicin in 10 min. A, Type II heat-sensitive afferents. All type II fibers gave a response to capsaicin that was >20 action potentials in 10 min (dashed line). In fact, the majority of fibers exhibited a strong response to capsaicin (>50 action potentials in 10 min). B, Heat-insensitive afferents. The majority of fibers categorized as HTM, type I, or not type II had very weak responses to capsaicin (≤20 action potentials per 10 min). However, five fibers not showing a type II response to heat showed the most vigorous responses to capsaicin (>100 action potentials per 10 min). Note: stacked histogram.

Fig. 5.

Vigorous response to capsaicin in a heat-insensitive afferent. A, Response to vehicle.B, Response to capsaicin. This afferent (mechanical threshold = 2.46 bar) did not respond to short-duration heat stimuli (1 sec) up to 49°C. However, the afferent responded to a long-lasting stimulus (49°C, 30 sec) and was therefore classified as a type I, heat-insensitive afferent. Both injections were performed inside the mechanosensitive receptive field, and the injection of vehicle preceded the injection of capsaicin. Insertion of the needle induced activity in the afferent for both injections. However, only the injection of capsaicin induced lasting activation of the afferent. Same format as Figure 1.

Fig. 6.

Time course of the capsaicin response in mechanically sensitive (gray columns) and mechanically insensitive (black columns) afferents. In mechanically sensitive afferents (n = 18), needle insertion itself produced the largest response. In contrast, the largest discharge in mechanically insensitive afferents (n = 22) was observed during the first minute after injection of capsaicin. The response to capsaicin was significantly larger in mechanically insensitive than mechanically sensitive afferents (p < 0.001; two-way ANOVA with repeated measures followed by multiple t tests with Bonferroni correction). The six afferents (three MIAs, three MSAs) that did not respond at all to the capsaicin injection are excluded.

Fig. 7.

Comparison of capsaicin responses in mechanically sensitive and mechanically insensitive afferents. A,Histogram for the total number of action potentials observed in 10 min after the injection of capsaicin. Only the largest response to capsaicin obtained in each fiber was included (n = 46). Most of the vigorous responses (>50 action potentials per 10 min) were observed in fibers classified as mechanically insensitive.B, Average response of the population during the 10 min. Mechanically insensitive afferents exhibited a significantly greater response to capsaicin than mechanically sensitive afferents (p < 0.05; t test).Numbers in parentheses give the number of fibers in each group.

Fig. 8.

Responsiveness to different stimulus modalities varies across the receptive field of a type II fiber. The mechanosensitive receptive field (gray area) and the electroreceptive field (dotted line) are indicated. Multiple test sites indicated by the circles were tested with different stimulus modalities (heat, mechanical, capsaicin). The size of the circle corresponds to size of the laser beam used for the heat testing. Capsaicin induced a response at all injection sites, whereas only two test sites were heat-sensitive. Responses to needle insertion were observed in two locations inside the continuous mechanoreceptive field. This afferent was characterized as type II heat-sensitive with a heat threshold of 49°C, 1 sec and a mechanical threshold of 11.2 bar.

Fig. 9.

Polymodality of different sites within the receptive field of type II afferents. Venn diagram for the different stimulus modalities to which a test site inside the electroreceptive field of a type II fiber was sensitive. Only sites tested with all three modalities are included. As for Figure 8, a test site was classified as mechanosensitive when it responded to von Frey hair stimulation. Most test sites were responsive to capsaicin (29 of 32), and many of these were sensitive to capsaicin only (11 of 29). Only eight capsaicin-sensitive sites were also sensitive to heat and mechanical stimuli. Four sites were insensitive to each of the stimulus modalities. Data come from 15 type II afferents.