Laser-evoked potentials mediated by mechano-insensitive nociceptors in human skin.

OBJECTIVES: Laser-evoked potentials (LEP) were assessed after peripheral nerve block of the lateral femoral cutaneous nerve (LFCN) in healthy volunteers from partially anesthetized skin areas to differentially stimulate mechano-insensitive nociceptors. METHODS: An ultrasound-guided nerve block of the LFCN was performed in 12 healthy male subjects with Ropivacain 1%. After 30 min, the nerve block induced significantly larger anesthetic areas to mechanical stimuli than to electrical stimuli revealing an area of differential sensitivity. LEPs, reaction times and pain ratings were recorded in response to the laser stimuli of (1) completely anesthetic skin, (2) mechano-insensitive, but electrically excitable skin ('differential sensitivity'), (3) normal skin. RESULTS: LEP latencies in the area of differential sensitivity were increased compared to unaffected skin (228 ± 8.5 ms, vs. 181 ± 3.6 ms, p < 0.01) and LEP amplitudes were reduced (14.8 ± 1.2 µV vs. 24.6 ± 1.7 µV, p < 0.01). Correspondingly, psychophysically assessed response latencies in the differentially anesthetic skin were increased (649 ms vs. 427 ms, p < 0.01) and pain ratings reduced (1.5/10 vs. 5/10 NRS, p < 0.01). CONCLUSION: The increase in LEP latency suggests that mechano-insensitive heat-sensitive Aδ nociceptors (MIA, type II) have a slower conduction velocity or higher utilization time than mechano-sensitive type II Aδ nociceptors. Moreover, widely branched, slowly conducting and mechano-insensitive branches of Aδ nociceptors can explain our finding. LEPs in the differentially anesthetized skin provide specific information about a mechanically insensitive but heat-sensitive subpopulation of Aδ nociceptors. These findings support the concept that A-fibre nociceptors exhibit a similar degree of modality specificity as C-fibre nociceptors.

Mechano-sensitive nociceptors are required to detect heat pain thresholds and cowhage itch in human skin.

BACKGROUND: Mechano-sensitive and mechano-insensitive C-nociceptors in human skin differ in receptive field sizes and electrical excitation thresholds, but their distinct functional roles are yet unclear. METHODS: After blocking the lateral femoral cutaneous nerve (NCFL) in eight healthy male subjects (3-mL Naropin(®) 1%), we mapped the skin innervation territory being anaesthetic to mechanical pin prick but sensitive to painful transcutaneous electrical stimuli. Such 'differentially anaesthetic zones' indicated that the functional innervation with mechano-sensitive nociceptors was absent but the innervation with mechano-insensitive nociceptors remained intact. In these areas, we explored heat pain thresholds, low pH-induced pain, cowhage- and histamine-induced itch, and axon reflex flare. RESULTS: In differentially anaesthetic skin, heat pain thresholds were above the cut-off of 50°C (non-anaesthetized skin 47 ± 0.4°C). Pain ratings to 30 µL pH 4 injections were reduced compared to non-anaesthetized skin (48 ± 9 vs. 79 ± 6 VAS; p < 0.01). The axon reflex flare area did not differ between these zones (7.8 ± 1.4 cm(2) vs. 8.3 ± 0.5 cm(2) ). Histamine iontophoresis still caused pruritus in differentially anaesthetized skin in five of eight subjects (VAS 26 ± 14), whereas itch upon cowhage spicules was absent (VAS 0 vs. 29 ± 11 in non-anaesthetized skin). CONCLUSIONS: We conclude that activation of mechano-insensitive nociceptors is sufficient to provoke itch by histamine- and acid-induced pain. The mechano-sensitive nociceptors are crucial for cowhage-induced itch and for the assessment of heat pain thresholds.

Selective expression of Narp in primary nociceptive neurons: role in microglia/macrophage activation following nerve injury.

Neuronal activity regulated pentraxin (Narp) is a secreted protein implicated in regulating synaptic plasticity via its association with the extracellular surface of AMPA receptors. We found robust Narp immunostaining in dorsal root ganglia (DRG) that is largely restricted to small diameter neurons, and in the superficial layers of the dorsal horn of the spinal cord. In double staining studies of DRG, we found that Narp is expressed in both IB4- and CGRP-positive neurons, markers of distinct populations of nociceptive neurons. Although a panel of standard pain behavioral assays were unaffected by Narp deletion, we found that Narp knockout mice displayed an exaggerated microglia/macrophage response in the dorsal horn of the spinal cord to sciatic nerve transection 3days after surgery compared with wild type mice. As other members of the pentraxin family have been implicated in regulating innate immunity, these findings suggest that Narp, and perhaps other neuronal pentraxins, also regulate inflammation in the nervous system.

Analgesic properties of loperamide differ following systemic and local administration to rats after spinal nerve injury.

BACKGROUND: The analgesic properties and mechanisms of loperamide hydrochloride, a peripherally acting opioid receptor agonist, in neuropathic pain warrant further investigation. METHODS: We examined the effects of systemic or local administration of loperamide on heat and mechanical hyperalgesia in rats after an L5 spinal nerve ligation (SNL). RESULTS: (1) Systemic loperamide (0.3-10 mg/kg, subcutaneous in the back) dose dependently reversed heat hyperalgesia in SNL rats, but did not produce thermal analgesia. Systemic loperamide (3 mg/kg) did not induce thermal antinociception in naïve rats; (2) systemic loperamide-induced anti-heat hyperalgesia was blocked by pretreatment with intraperitoneal naloxone methiodide (5 mg/kg), but not by intraperitoneal naltrindole (5 mg/kg) or intrathecal naltrexone (20 µg/10 µL); (3) local administration of loperamide (150 µg), but not vehicle, into plantar or dorsal hind paw tissue induced thermal analgesia in SNL rats and thermal antinociception in naïve rats; (4) the analgesic effect of intraplantar loperamide (150 µg/15 µL) in SNL rats at 45 min, but not 10 min, post-injection was blocked by pretreatment with an intraplantar injection of naltrexone (75 µg/10 µL); (5) systemic (3.0 mg/kg) and local (150 µg) loperamide reduced the exaggerated duration of hind paw elevation to noxious pinprick stimuli in SNL rats. Intraplantar injection of loperamide also decreased the frequency of pinprick-evoked response in naïve rats. CONCLUSIONS: These findings suggest that both systemic and local administration of loperamide induce an opioid receptor-dependent inhibition of heat and mechanical hyperalgesia in nerve-injured rats, but that local paw administration of loperamide also induces thermal and mechanical antinociception.

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

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,

Early onset of spontaneous activity in uninjured C-fiber nociceptors after injury to neighboring nerve fibers.

Ligation and transection of the L5 spinal nerve in the rat lead to behavioral signs of pain and hyperalgesia. Discharge of injured nociceptors has been presumed to play a role in generating the pain. However, A fibers, but not C fibers, in the injured L5 spinal nerve have been shown to develop spontaneous activity. Moreover, an L5 dorsal root rhizotomy does not reverse this pain behavior, suggesting that signals from other uninjured spinal nerves are involved. We asked if abnormal activity develops in an adjacent, uninjured root. Single nerve fiber recordings were made from the L4 spinal nerve after ligation and transection of the L5 spinal nerve. Within 1 d of the lesion, spontaneous activity developed in approximately half of the C fiber afferents. This spontaneous activity was at a low level (median rate, seven action potentials/5 min), originated distal to the dorsal root ganglion, and was present in nociceptive fibers with cutaneous receptive fields. The incidence and level of spontaneous activity were similar 1 week after injury. The early onset of spontaneous activity in uninjured nociceptive afferents could be the signal that produces the central sensitization responsible for the development of mechanical hyperalgesia. Because L4 afferents comingle with degenerating L5 axons in the peripheral nerve, we hypothesize that products associated with Wallerian degeneration lead to an alteration in the properties of the adjacent, uninjured afferents.

Electrophysiological assessment of the cutaneous arborization of Adelta-fiber nociceptors.

Little is known about the relationship between the branching structure and function of physiologically identified cutaneous nociceptor terminals. The axonal arborization itself, however, has an impact on the afferent signal that is conveyed along the parent axon to the CNS. We therefore developed electrophysiological techniques to investigate the branching structure of cutaneous nociceptors. Single-fiber recordings were obtained from physiologically identified nociceptors that innervated the hairy skin of the monkey. Electrodes for transcutaneous stimulation were fixed at two separate locations inside the receptive field. For 32 Adelta-fiber nociceptors, distinct steps in latency of the recorded action potential were observed as the intensity of the transcutaneous electrical stimulus increased, indicating discrete sites for action potential initiation. The number of discrete latencies at each stimulation location ranged from 1 to 9 (3.7 +/- 0. 2; mean +/- SE) and the mean size of the latency step was 9.9 +/- 1. 0 ms (range: 0.4-89.1 ms). For seven Adelta fibers, collision techniques were used to locate the position of the branch point where the daughter fibers that innervated the two locations within the receptive field join the parent axon. To correct for changes in electrical excitability at the peripheral terminals, collision experiments between the two skin locations and between each skin location and a nerve trunk electrode were necessary. Nine branch points were studied in the seven Adelta fibers; the mean propagation time from the action potential initiation site to the branch point was 31 +/- 5 ms corresponding to a distance of 54 +/- 10 mm. Almost half of the daughter branches were unmyelinated. These results demonstrate that collision techniques can be used to study the functional anatomy of physiologically identified nociceptive afferent terminals. Furthermore these results indicate that some nociceptive afferents branch quite proximal to their peripheral receptive field. Occlusion of action potential activity can occur in these long branches such that the shorter branches dominate in the response to natural stimuli.

Mechanical hyperalgesia after spinal nerve ligation in rat is not reversed by intraplantar or systemic administration of adrenergic antagonists.

The development of alpha-adrenergic sensitivity in cutaneous nociceptors has been postulated as a mechanism for sympathetically maintained pain (SMP). In order to characterize the adrenergic receptors involved, we investigated the effects of intraplantar administration of alpha1-(prazosin) and alpha2-(yohimbine) adrenergic antagonists and systemic injection of phentolamine, a non-specific alpha-adrenergic blocker, on allodynic/hyperalgesic behavior in an animal model thought to mimic SMP in humans. Peripheral neuropathy in rats was induced by tight ligation of the L5/L6 spinal nerves. Mechanical hyperalgesia was quantified with von Frey hairs applied either for 3 s or repetitively to the plantar surface of the hindpaw. Responses to the 3 s duration stimulus were used to determine the paw withdrawal threshold with the up-down paradigm and repetitive stimuli were used to determine the response incidence of paw withdrawal to a given von Frey hair. Mechanical thresholds on the ipsilateral paw decreased significantly after ligation and were stable over the following 3 weeks. Intradermal administration of yohimbine or prazosin did not significantly alleviate mechanical hyperalgesia in L5/L6 ligated animals. Also systemic administration of phentolamine (1 and 5 mg/kg) did not alleviate the increased incidence of paw withdrawal in L5/L6 spinal nerve ligated animals. These results suggest that an alpha adrenergic interaction between sympathetic efferent and somatic afferent fibers does not play a critical role for the maintenance of mechanical hyperalgesia in this model for neuropathic pain.

Lumbar sympathectomy failed to reverse mechanical allodynia- and hyperalgesia-like behavior in rats with L5 spinal nerve injury.

The L5 spinal nerve ligation model of neuropathic pain in rats has been proposed as a model for sympathetically maintained pain (SMP) based on the effects of surgical or chemical sympathectomy on nerve injury induced behavior. In an attempt to confirm that the lesion produces an animal model of SMP, surgical sympathectomies were independently conducted in two different laboratories (Johns Hopkins and University Kiel) using male Sprague-Dawley (n = 30) or Wistar rats (n = 14). The L5 spinal nerve was ligated or cut and ligated. Using von Frey hairs, paw withdrawal threshold and incidence of paw withdrawal were tested concurrently before and after the sympathectomy. The sympathectomy was either verified by (a) glyoxylic acid staining of peripheral blood vessels of the hindpaw or (b) skin temperature measurements of the hindpaws. To blind the experimenter, surgeries and behavioral tests were performed by two different investigators and a sham sympathectomy was performed at Johns Hopkins. Decreased paw withdrawal thresholds and increased frequencies of paw withdrawal on the lesioned side were observed after the L5 lesion. Thus, the L5 spinal nerve ligation resulted in behavioral signs of allodynia and hyperalgesia to mechanical stimuli. Lumbar surgical sympathectomy 1-3 weeks after the lesion or prior to lesion with bilateral removal of the sympathetic ganglia L2-L4, however, did not reverse or prevent the behavioral changes induced by the nerve injury. The lack of effect of the sympathectomies was independent of the testing paradigm used. Experiments in Wistar and Sprague-Dawley rats yielded the same results. Potential reasons for the discrepancies between the present study and earlier reports are discussed. These results indicate that an L5 spinal nerve injury rat model is not a reliable model for SMP.

Uninjured C-fiber nociceptors develop spontaneous activity and alpha-adrenergic sensitivity following L6 spinal nerve ligation in monkey.

We investigated whether uninjured cutaneous C-fiber nociceptors in primates develop abnormal responses after partial denervation of the skin. Partial denervation was induced by tightly ligating spinal nerve L6 that innervates the dorsum of the foot. Using an in vitro skin-nerve preparation, we recorded from uninjured single afferent nerve fibers in the superficial peroneal nerve. Recordings were made from 32 C-fiber nociceptors 2-3 wk after ligation and from 29 C-fiber nociceptors in control animals. Phenylephrine, a selective alpha1-adrenergic agonist, and UK14304 (UK), a selective alpha2-adrenergic agonist, were applied to the receptive field for 5 min in increasing concentrations from 0.1 to 100 microM. Nociceptors from in vitro control experiments were not significantly different from nociceptors recorded by us previously in in vivo experiments. In comparison to in vitro control animals, the afferents found in lesioned animals had 1) a significantly higher incidence of spontaneous activity, 2) a significantly higher incidence of response to phenylephrine, and 3) a higher incidence of response to UK. In lesioned animals, the peak response to phenylephrine was significantly greater than to UK, and the mechanical threshold of phenylephrine-sensitive afferents was significantly lower than for phenylephrine-insensitive afferents. Staining with protein gene product 9.5 revealed an approximately 55% reduction in the number of unmyelinated terminals in the epidermis of the lesioned limb compared with the contralateral limb. Thus uninjured cutaneous C-fiber nociceptors that innervate skin partially denervated by ligation of a spinal nerve acquire two abnormal properties: spontaneous activity and alpha-adrenergic sensitivity. These abnormalities in nociceptor function may contribute to neuropathic pain.

Sympathectomy decreases formalin-induced nociceptive responses independent of changes in peripheral blood flow.

The present experiments studied the effect of surgical sympathectomy on mechanical paw withdrawal thresholds and formalin-induced pain behaviors (FIPB) and tested if the effect of sympathectomy on FIPB is dependent on changes in peripheral blood flow. Compared to sham-operated animals, surgical sympathectomy decreased second phase FIPB but did not affect paw withdrawal thresholds to mechanical stimuli. In sympathectomized and sham-operated animals, subcutaneous intraplantar injection of the nonadrenergic vasoconstrictor angiotensin II (20 microl, 1 mM) or the nonadrenergic vasodilator hydralazine (20 microl, 1 mM) prior to formalin injection decreased and increased peripheral blood flow, respectively. In sympathectomized animals, pretreatment with angiotensin II inhibited the increase of peripheral blood flow following formalin injection when compared to animals pretreated with saline or hydralazine. In sham-operated animals, pretreatment with hydralazine or angiotensin II did not alter vasodilation after formalin injection when compared to pretreatment with saline. Pretreatment with vasoactive drugs had no effect on the sympathectomy-induced decrease in FIPB. It is concluded that sympathectomy decreases FIPB by mechanisms that are independent of sympathectomy-induced changes in peripheral blood flow. The formalin test is an easy, reliable model that can be used to study the mechanisms by which the sympathetic nervous system modulates pain processing.

Innervation territories of mechanically activated C nociceptor units in human skin.

Innervation territories of mechanically activated C nociceptor units in human skin. J. Neurophysiol. 78: 2641-2648, 1997. Innervation territories of single mechanically activated C nociceptors in the skin of the leg and foot were explored in normal human subjects. Microneurographic recordings were obtained in the peroneal nerve from 70 mechano-heat responsive (CMH) and 7 mechano-(but not heat) responsive (CM) units. Units were identified by their constant long-latency response to intracutaneous electrical stimulation of their terminals. Responsiveness to mechanical, heat, or transcutaneous electrical stimuli was verified by transient slowing of conduction velocity after activation by such stimuli. We determined their thresholds to mechanical stimuli (mean 33.7 mN, median 30 mN, range 3-750 mN) and heat (mean 42.5 degrees C, median 42.5 degrees C, range 37-49 degrees C). Most mechano-receptive fields (mRFs) were found on the foot dorsum (60 units) and some on the lower leg (14 units) and toes (3 units). Most units had one continuous mRF, but 10 units had more complex fields. Areas of mRFs mapped with a von Frey filament (750 mN) ranged from 10 to 363 mm2 (mean, 106 mm2). The mRFs were oval or irregularly shaped with greatest diameters ranging from 3 to 45 mm. Mean areas of mRFs were largest on the lower leg (198 mm2), smaller on the foot dorsum (88 mm2), and smallest on the toes (35 mm2). Forty-nine of the 77 units had identical mRFs and electro-receptive fields (eRFs). Twenty-six units had larger eRFs than mRFs, whereas the opposite was found for two units only. Areas of eRFs ranged from 16 to 511 mm2 (mean 121 mm2). An estimate of the innervation density based on the present data and the presumed number of C fibers in cutaneous fascicles of the peroneal nerve suggests a considerable overlap of nociceptive endings in the skin. Such overlapping nociceptor innervation in the skin allows for substantial spatial summation in response to punctate noxious stimuli, which may be a prerequisite for high accuracy in localizing painful events from a C-fiber input. The reduction in size of innervation territories distally allows for finer discrimination of spatial dimensions of noxious stimuli distally as compared with proximal regions of the extremities. Mean maximal diameters of the mechano-receptive fields of CMH and CM units on the lower leg (22.3 mm) and foot (15.3 mm) are of similar size as the radius of axon reflex flares evoked by noxious mechanical stimuli in these regions.

Recordings from brain stem neurons responding to chemical stimulation of the subarachnoid space.

The subarachnoid space at the base of the skull was perfused continuously with artificial cerebrospinal fluid in anesthetized rats. A combination of inflammatory mediators consisting of histamine, bradykinin, serotonin, and prostaglandin E2 (10(-5) M) at pH of 6.1 was introduced into the flow for defined periods to stimulate meningeal primary afferents. Secondary neurons in the caudal nucleus of the trigeminal brain stem were searched by electrical stimulation of the cornea. Of the units receiving oligosynaptic input from the cornea, 44% were excited by stimulation of the meninges with inflammatory mediators. Most of these units had small receptive fields including cornea and the periorbital region, and their responsiveness was restricted to stimuli of noxious intensity. Three types of responses to stimulation of the meninges with algogenic agents were encountered: responses that did not outlast the stimulus period, responses outlasting the stimulus period for several minutes, and oscillating response patterns containing periods of enhanced and suppressed activity. The response pattern of a unit was reproducible, however, upon repetitive stimulation at 20-min intervals; the response magnitude showed tachyphylaxis upon stimulus repetition. The preparation presented mimics pathophysiolocial states normally accompanied by headache, e.g., subarachnoidal bleeding. Responsiveness of neurons in the caudal nucleus of the trigeminal brain stem to inflammatory mediators may play a role in the generation and maintenance of headache, e.g., migraine.

Intracutaneous injections of platelets cause acute pain and protracted hyperalgesia.

Suspensions of autologous, washed platelets were intracutaneously injected at the volar forearms of healthy volunteers. Injections of serum and vehicle served as control. Subjects and experimenter were blind with respect to the sequence of injections. In contrast to serum and solvent solution, platelets induced graded burning pain lasting several minutes. Platelet but not serum or vehicle injections dose-dependently caused large axon-reflex flares. At the site of platelet injections an induration developed and in parallel delayed mechanical and heat hyperalgesia was observed. Hyperalgesia to pressure and impact stimulation reached a maximum after 6 h and subsided during the following 48 h. Also, the threshold to heat stimuli decreased moderately by about 1 degree C, on average, after 24 h. Neither indurations nor hyperalgesia could be detected at the injection sites of serum or vehicle. The pathophysiological significance of this new inflammatory model for the research of posttraumatic hyperalgesia is discussed.

Hemodynamic and sympathetic nerve responses to painful stimuli in normotensive and borderline hypertensive subjects.

Observations in animals and humans show that pain sensitivity might be lower (and pain tolerance higher) in hypertensive as compared to normotensive subjects. One hypothesis, derived from experimental studies, assumes that enhanced activation of baroreceptors leads to an enhanced central inhibition. A central hypothesis assumes changes in the central (endogenous) control of the nociceptive system. To investigate these two hypotheses we quantitatively assessed the minute-by-minute changes in mean arterial pressure (MAP), central venous pressure (CVP) heart rate (HR), muscle sympathetic nerve activity (MSNA), and individual pain ratings during noxious mechanostimulation in 10 normotensive (NT) and 13 borderline hypertensive (BH) subjects. Linear regression analysis indicated a close negative correlation for the overall data between resting levels of MAP and pain ratings (r = -0.57, P < 0.0001). The BH group exhibited a lower pain sensitivity compared to the NT group (P < 0.001). The extent of baroreceptor activation during the application of pain was not different between the two groups (P = NS) as assessed by almost identical increases in MAP (+8 +/- 1 vs. +9 +/- 1 mmHg NT vs. BH group), CVP (+0.7 +/- 0.1 vs. +0.5 +/- 0.1 mmHg), HR (+2 +/- 1 vs. +2 +/- 1 beats/min), and MSNA (+5 +/- 1 +4 +/- 1 bursts/min). The NT subjects exhibited significant correlations between the pain ratings and the increases in MAP (r = +0.52; P < 0.05) and MSNA (r = +0.49; P < 0.05) whereas the BH subjects did not show such a relationship. Thus, the increased pain tolerance in human hypertension cannot be explained by hemodynamically mediated differences in the activation of baroreceptors or by an altered baroreflex sensitivity during the application of pain. We conclude, that the reduced pain sensitivity in hypertensive humans is more likely related to central changes.

Limitation of sensitization to injured parts of receptive fields in human skin C-nociceptors.

Unmyelinated cutaneous mechano-heat fibers (CMH) in the peroneal nerve of healthy human volunteers were studied by means of a "marking" technique which allows stable recordings from identified single units over extended periods. Mechanoreceptive field sizes were 105 +/- 13 mm2 in 25 units. These large receptive fields indicate extensive terminal branching of C fibers in the skin of foot and lower leg. Sensitization of CMHs was tested by assessment of thresholds for mechanical (von Frey hair) and heat stimuli before and after topical application of mustard oil (allyl isothiocyanate) and capsaicin (8-methyl-N-vanillyl-6-noneamide). While in a group of 14 CMHs the entire receptive field was treated with these irritant substances, in another group of 11 CMH units only parts of the receptive field were treated to check for signs of spreading sensitization through axon collaterals. Mustard oil application did not change mechanical thresholds, regardless of whether parts of or complete receptive fields were treated. However, mean heat thresholds dropped by 5.6 degrees C to 36.5 +/- 1.5 degrees C in completely treated receptive fields and by 5.7 degrees C to 37.3 +/- 3.4 degrees C in treated parts of receptive fields ("primary sensitization"). In contrast, heat thresholds in the nontreated parts did not change significantly (42.1 +/- 3.4 degrees C vs 41.2 +/- 3.9 degrees C), i.e. "secondary sensitization" to heat was lacking. The absence of primary sensitization to probing with von Frey hairs indicates that sensitization of insensitive C fibers and recruitment of insensitive axon collaterals may be more important for mechanical hyperalgesia than sensitization of conventional CMH units-apart from the contribution of central mechanisms. The lack of spread of sensitization to untreated parts of the receptive fields o CMHs ("secondary sensitization") indicates that this fiber group is probably not involved in any form of secondary hyperalgesia to heating.

Novel classes of responsive and unresponsive C nociceptors in human skin.

One hundred ninety-four cutaneous C-fibers were recorded from the human peroneal nerve; 118 units were found by mechanical stimuli and 76 units were detected by electrical stimulation through a surface electrode. Needle electrodes were then inserted for electrical intradermal stimulation in the innervation territory of the units. Afferent and efferent sympathetic C-fibers were identified by slowing of conduction velocity after activation either by physical or chemical skin stimuli, or by arousal maneuvers eliciting sympathetic reflexes. In addition to mechano-heat-responsive C units (CMH) also found in previous studies, we here report on novel classes of C nociceptors in human skin, namely, units responding only to mechanical stimuli (CM), units responding only to heating (CH), and units that were insensitive to mechanical and heating stimuli and also to sympathetic provocation tests (CMiHi). With the electrical search technique we found 45% CMH, 13% CM, 6% CH, 24% CMiHi, and 12% sympathetic units. Excitation by topically applied mustard oil occurred in 58% of CMH units, and in one-third of CM and CMiHi units, respectively. Some CM, CH, and CMiHi units were sensitized to heating and/or to mechanical stimuli after topical application of mustard oil or capsaicin. These units then acquired responsiveness to a stimulus modality to which they previously were insensitive. Such recruitment of previously silent nociceptors implies spatial summation to the nociceptive barrage at central levels, and may contribute both to primary hyperalgesia to heat and pressure after chemical irritation, and to secondary hyperalgesia as a consequence of central sensitization.

Delayed responses to electrical stimuli reflect C-fiber responsiveness in human microneurography.

The slowing of impulse conduction during the relative refractory period has often been used to assess activation of C-fibers, in particular, in human microneurography. This study aimed to evaluate the sensitivity of this method and the factors affecting it. Thirty cutaneous C-fibers were recorded from the peroneal nerves of healthy human subjects. Intracutaneous electrical stimulation in the receptive field at 4 s intervals, after some minutes of adaptation, induced spike discharges at constant latency. One or more conditioning stimulus pulses were interpolated at different intervals and the increase in latency after the subsequent regular pulse was assessed. The latency shift was found to depend on the number of interposed pulses, on the time interval between conditioning and conditioned stimulus, and on the conduction velocity of the C-unit. The increase in latency was larger with greater distance between stimulating and recording electrodes, indicating a contribution of the conductile membrane over its whole length. On the other hand, slowing was more pronounced, on average, in slower conducting C-units and conduction velocities were slower when recordings were performed more distally. These findings indicate that the slower terminal nerve branches contribute most to the latency increases. Even a single additional spike in between two regular pulses caused a reliable latency shift of 1.2 +/- 0.2 ms (mean +/- SEM) and additional pulses lead to an approximately linear latency increase (2 pulses: 2.3 +/- 0.3 ms; 4 pulses: 5.9 +/- 0.7 ms). In contrast to the number of interposed stimuli, different intervals between interposed and regular stimuli had only a minor impact on the latency shifts.(ABSTRACT TRUNCATED AT 250 WORDS)

Sensitization of insensitive branches of C nociceptors in human skin.

1. Eighteen cutaneous mechanosensitive C nociceptors were recorded from the peroneal nerves of healthy human subjects. Their identity was continuously monitored by intracutaneous electrical stimulation, and their activation by mechanical or transcutaneous electrical stimulation was detected by slowing of conduction velocity during the relative refractory period. 2. Mechanoreceptive fields (mRFs) mapped with suprathreshold von Frey hair stimuli covered an area of 99 +/- 21 mm2 (mean +/- S.E.M.). Two of the units had separate mRFs, with borders about 0.5-1.5 cm apart from each other and the largest of these units had a maximal diameter of 4.5 cm. 3. Successive topical application of mustard oil and capsaicin induced expansions of mRFs by 57 +/- 14 mm2 in eight of fifteen units. 4. In twelve units transcutaneous electrical stimulation delivered through a pointed electrode was used for mapping the electroreceptive fields (eRFs). The borders of the eRFs and the mRFs were identical for two of twelve units only. In the other ten units additional mechano-insensitive areas (55 +/- 22 mm2) were detected from which transcutaneous electrical stimuli could activate the respective unit. 5. Application of mustard oil and capsaicin to these mechano-insensitive areas sensitized five of eight units to mechanical stimuli. In these cases the mRF after sensitization exactly corresponded to the eRF. 6. It is concluded that there are insensitive branches in human mechanosensitive cutaneous C nociceptors that can be detected by transcutaneous electrical stimulation and sensitized by topical application of chemical irritants. Activation of those branches in the course of inflammatory processes may contribute to spatial summation at central synapses and hence to hyperalgesia.