Differences between the Lewis and Sprague-Dawley rats in chronic inflammation induced norepinephrine sensitivity of cutaneous C-fiber nociceptors

We investigated whether there are any differences between the Lewis and Sprague-Dawley (SD) rats in chronic inflammation-induced norepinephrine (NE) sensitivity of nociceptors. Activities of C-fiber nociceptors innervating rat hairy hindpaw skin were recorded in an in vitro skin-nerve preparation. Sixty-five percent of C-fibers from inflamed Lewis rats were excited by NE (10 microM), against only 38% of C-fibers from inflamed SD rats. The average of the total impulses evoked in response to NE was also significantly higher in Lewis rats. The alpha2-adrenoceptor antagonist CH 38083 (10 microM) and yohimbine (10 microM) consistently blocked the NE-excitation of both strains. These results show that after chronic inflammation, C-fiber nociceptors of Lewis strain rats have a stronger sensitivity to NE, and that alpha2-adrenoceptors are predominately involved in the NE-sensitivity of inflamed rats in both strains.     

Comparison of heat and mechanical receptive fields of cutaneous C-fiber nociceptors in monkey

1. Receptive-field properties were investigated in cutaneous C-fiber nociceptive afferents (CMH) responsive to mechanical and heat stimuli. Teased-fiber techniques were used to record from 28 CMHs that innervated the hairy skin of upper or lower limb in anesthetized monkeys. 2. The response to mechanical stimuli was studied with the use of calibrated von Frey probes. The response to heat stimuli was studied with the use of a laser thermal stimulator that provided stepped increases in skin temperature with rise times to the desired temperature near 100 ms. The size of the receptive field (RF) for mechanical stimuli was determined by use of a suprathreshold stimulus that consisted of a 0.5-mm-diam probe that exerted a 200-mN force (10 bar). The size of the heat RF was determined by use of a 49 degrees C stimulus applied to a 7.5-mm-diam area for 1 s. 3. Heat thresholds were determined with an ascending series of stimulus intensities and were found to be stable over many hours: they ranged from 37 to 46 degrees C (mean, 41.1 degrees C). Mechanical thresholds ranged from 1.3 to 7.3 bar (mean, 3.3 bar). There was no correlation between mechanical and heat thresholds. Both thresholds extended well below the corresponding psychophysical pain thresholds in the literature. This suggests that spatial and/or temporal summation of C-fiber input are important for pain induced by either stimulus modality. 4. Mechanical RF diameters ranged from 3.3 to 9.6 mm (mean, 4.7 mm); heat RF diameters ranged from punctate (less than 1 mm) to 9.5 mm (mean, 4.3 mm). There was a significant linear correlation between mechanical and heat RF sizes with a slope of one. The distance between the center of the mechanical RF and the center of the heat RF along one axis ranged from 0 to 1.1 mm (mean, 0.4 mm). These data indicate that the heat RFs coincided with the mechanical RFs. 5. Within the mechanical RF determined with the suprathreshold stimuli, all CMHs had one or more punctate areas of maximal mechanical sensitivity where mechanical threshold was lowest. Heat excitability extended greater than 2 mm beyond these mechanically sensitive spots. Because lateral transmission of the heat stimulus is small, this indicates that heat transduction occurs outside the regions of maximal mechanical sensitivity. 6. Both the threshold to heat and the response magnitude at suprathreshold intensities depended on the percentage of the RF area overlapped by the heat stimulus. This indicates that multiple transducer sites probably contribute to the total evoked response.(ABSTRACT TRUNCATED AT 400 WORDS)     

Adrenergic sensitivity of uninjured C-fiber nociceptors in neuropathic rats

We investigated the adrenergic sensitivity of afferent fibers in the L4 dorsal roots of rats with a unilateral ligation of the L5-L6 spinal nerves. About 12% of nociceptive fibers on the affected side were excited by sympathetic stimulation or by intra-arterial injection of norepinephrine which did not affect A beta-fiber activity. Sympathetic excitation of nociceptive fibers was suppressed by alpha 1-antagonist prazosin, while it was unaffected by alpha 2-antagonist yohimbine. Most of these fibers were excited by intra-arterial injection of alpha 1-agonist phenylephrine, without being affected by an injection of alpha 2-agonist clonidine. Sympathetic excitation was blocked by lidocaine applied near the receptive fields of recorded fibers. The results suggested that some nociceptors remaining intact after partial nerve injury become sensitive to sympathetic activity by the mediation of alpha 1-adrenoceptors in the peripheral endings.     

Response of cutaneous A- and C-fiber nociceptors in the monkey to controlled-force stimuli

The goal of this study was to determine the capacity of primary afferent nociceptive fibers (nociceptors) to encode information about noxious mechanical stimuli in primates. Teased-fiber techniques were used to record from 14 A-fiber nociceptors and 18 C-fiber nociceptors that innervated the hairy skin. Stimulus-response functions were examined with an ascending series of force-controlled stimuli. Stimulus-interaction effects were examined with use of a series of paired stimuli in which the interval between the stimulus pairs was varied systematically. Both A-fiber and C-fiber nociceptors exhibited a slowly adapting response to the stepped force stimuli. The response of the A fibers increased monotonically with increasing force, whereas the response of the C fibers reached a plateau at low force levels. The slope of the stimulus-response function for the A fibers was significantly steeper than that for the C fibers, and the total response was greater. The A fibers also provided more discriminative information regarding stimulus intensity. The C fibers demonstrated a significant fatigue in response when the interstimulus interval between the paired stimuli was 相似文献   

Mechanical and heat sensitization of cutaneous nociceptors in rats with experimental peripheral neuropathy

This study examined whether or not the properties of cutaneous nociceptive fibers are altered in the neuropathic state by comparing lumbars 5 and 6 spinal nerve ligation (SNL) rats with sham-operated controls. The rats with the unilateral SNL developed mechanical allodynia in the ipsilateral hind limb, whereas the sham group did not. Two to 5 weeks after the neuropathic or sham surgery, rats were subjected to single fiber-recording experiments to examine the properties of afferent fibers in the sural and plantar nerves. A total of 224 afferents in the C- and Adelta-ranges were characterized in the neuropathic and sham groups. Spontaneous activity was observed in 16 of 155 fibers in the neuropathic group and one of 69 fibers in the sham group. The response threshold of both the C- and Adelta-fibers to mechanical stimuli was lower in the neuropathic group than the sham group. The afferent fibers responsive to heat stimuli were all C-fibers, and none were Adelta-fibers. The response threshold of the C-fibers to the heat stimuli was lower in the neuropathic group than the sham group. The magnitude of the responses of both C- and Adelta-fibers to the suprathreshold intensity of the mechanical stimulus was greater in the neuropathic group than the sham group. However, the magnitude of the responses of C-fibers to the suprathreshold intensity of the heat stimulus in the neuropathic group was not different from that in the sham group. These results suggest that after a partial peripheral nerve injury, the nociceptors on the skin supplied by an uninjured nerve become sensitized to both mechanical and heat stimuli. This nociceptor sensitization can contribute to neuropathic pain.     

NOS inhibitor antagonism of PGE2-induced mechanical sensitization of cutaneous C-fiber nociceptors in the rat

Prostaglandins, metabolites of arachidonic acid, released during tissue injury and inflammation sensitize primary afferent nociceptors. While it has been suggested that this effect on nociceptors is mediated mainly via the cAMP second messenger system, recent evidence suggests that nitric oxide (NO) is also involved in peripheral pain mechanisms. To test the hypothesis that NO contributes to the sensitization of nociceptors to mechanical stimuli induced by hyperalgesic prostaglandins, we compared von Frey hair mechanical threshold as well as the response evoked by 10-s sustained threshold mechanical stimulation before and after injection of prostaglandin E2 (PGE2) alone, and NOS inhibitor NG-methyl-L-arginine (L-NMA) or its inactive stereoisomer NG-methyl-D-arginine (D-NMA) plus PGE2, adjacent to the receptive field of C-fiber nociceptors. The reduction of mechanical threshold and increase in number of action potentials to sustained mechanical stimulation induced by intradermal application of PGE2 was blocked by L-NMA, but not D-NMA. It is suggested that NO contributes to nociceptor sensitization induced by hyperalgesic prostaglandins.     

B2 receptor-mediated enhanced bradykinin sensitivity of rat cutaneous C-fiber nociceptors during persistent inflammation.

Bradykinin (BK), which has potent algesic and sensitizing effect on nociceptors, is of current interest in understanding the mechanisms of chronic pain. BK response is mediated by B2 receptor in normal conditions; however, findings that B1 receptor blockade alleviated hyperalgesia in inflammation have been highlighting the role of B1 receptor in pathological conditions. It has not yet been clear whether nociceptor activities are modified by B1 receptor agonists or antagonists during inflammation. In addition, previous studies reported the change in BK sensitivity of nociceptors during short-lasting inflammation, and data in persistent inflammation are lacking. Therefore we investigated whether an experimentally induced persistent inflammatory state modulates the BK sensitivity of nociceptors and which receptor subtype plays a more important role in this condition. Complete Freund's adjuvant was injected into the rat-tail and after 2-3 wk, persistent inflammation developed, which was prominent in the ankle joint. Using an in vitro skin-saphenous nerve preparation, single-fiber recordings were made from mechano-heat sensitive C-fiber nociceptors innervating rat hairy hindpaw skin, and their responses were compared with those obtained from C-fibers tested similarly in normal animals. BK at 10(-8) M excited none of the 10 C-fibers in normal animals while it excited 5 of 11 (45%) C-fibers of inflamed animals, and at 10(-6) M BK excited all of the 11 inflamed C-fibers (or 94% of 36 tested C-fibers) but only 4 of 10 (or 45% of 58 tested C-fibers) in normal animals. Thus the concentration-response curves based on the incidence of BK induced excitation, and the total number of impulses evoked in response to BK were significantly shifted to the left. Moreover, an increased percentage of the inflamed C-fibers responded to 10(-6) M BK with bursting or high-frequency discharges. Thirty-percent of inflamed C-fibers had spontaneous activity, and these fibers showed comparatively less tachyphylaxis to consecutive second and third 10(-6) M BK stimulation. A B2 receptor antagonist (D-Arg-[Hyp3, Thi5,8,D-phe7]-BK) completely eliminated BK responses in inflamed rats, while B1 receptor antagonists (B 9958 and Des-Arg9-[Leu8]-BK) had no effect. Selective B1 receptor agonist (Des-Arg10-Kallidin) excited 46% (n = 13) of inflamed C-fibers at 10(-5) M concentration, which is 1,000 times higher than that of BK needed to excite the same percentage of inflamed C-fibers. We conclude that in chronically inflamed tissue, sensitivity of C-fiber nociceptors to BK, which is B2 receptor mediated, is strongly increased and that B1 receptor may not be important to a persistent inflammatory state, at least at the primary afferent level.     

The tetrodotoxin-resistant Na+ channel Nav1.8 reduces the potency of local anesthetics in blocking C-fiber nociceptors

The generation of action potentials in nociceptive neurons is accomplished by the tetrodotoxin-resistant (TTXr) Na⁺ channel Na_v1.8. Following nerve injury, a redistribution of Na_v1.8 from dorsal root ganglion (DRG) neurons into peripheral axons contributes to hyperexcitability and possibly to neuropathic pain. Na_v1.8 has been reported to display a lower sensitivity to block by Na⁺ channel blockers as compared to TTX-sensitive (TTXs) Na_v subunits. Furthermore, the antinociceptive efficacy of lidocaine is increased in Na_v1.8-knockout mice. Here, we asked if Na_v1.8 expression can reduce the susceptibility of sensory neurons to block by lidocaine. Employing wild-type and Na_v1.8-knockout mice, we examined C-fibers in the skin-nerve preparation and Na⁺ currents in DRG neurons by patch-clamp recordings. Deletion of Na_v1.8 resulted in an enhanced tonic block of Na⁺ currents in DRG neurons held at ?80 mV but not at ?140 mV. Accordingly, lower concentrations of lidocaine were required for a conduction block of C-fibers from Na_v1.8-knockout as compared to wild-type mice. The efficacy of lidocaine on neurons lacking Na_v1.8 was further increased by cold temperatures, due to a synergistic hyperpolarizing shift of the slow inactivation of TTXs Na⁺ channels by lidocaine and cooling. Finally, the ～90% reduction of TTXr Na⁺ currents in injured neurons from mice with a peripheral nerve injury was accompanied with an enhanced tonic block by lidocaine. In conclusion, our data demonstrate that the expression of Na_v1.8 in sensory neurons can confine the antinociceptive efficacy of lidocaine and other Na⁺ channel blockers employed for pain treatment.     

Different mechanical transduction mechanisms for the immediate and delayed responses of rat C-fiber nociceptors.

1. In this electrophysiological study, action potentials from single C-fibers were recorded in fine filaments teased from the rat saphenous nerve. We evaluated the effect of pharmacological agents on the responses of C-fiber mechanoheat nociceptors (C-MH; n = 53) after sustained suprathreshold and subthreshold stimuli. 2. Sustained suprathreshold mechanical stimuli elicit an immediate burst of activity that quickly adapts to a low-level firing that is maintained during the stimulus. Sustained subthreshold stimuli activate C-MHs after a delay and elicit a constant, low-level firing. 3. Gentamicin, a known suppressor of mechanosensory cell activity, blocked the initial rapid burst response to suprathreshold stimuli (n = 11) but had no effect on the adaptive low-level firing. The latency of the delayed activation of C-MHs induced by sustained subthreshold stimuli was not affected by gentamicin. 4. Sphingosine, a protein kinase inhibitor, increased the latency of the delayed activation of C-MHs (n = 7) to sustained subthreshold stimuli; phorbol 12-myristate 13-acetate (TPA), a protein kinase C activator, decreased the latency of the delayed activation of C-MHs (n = 9); and 4 alpha-phorbol, an inactive isomer of TPA, had no effect on the latency of the delayed activation (n = 7). Sphingosine, TPA, and 4 alpha-phorbol had no affect on the initial burst response induced by suprathreshold stimuli. 5. K+ channel blockers, 4-aminopyridine (n = 9) and noxiustoxin (n = 5), decreased the latency of the delayed activation of C-MHs to sustained subthreshold stimuli but had no effect on the initial burst response of C-MHs to suprathreshold stimuli.(ABSTRACT TRUNCATED AT 250 WORDS)     

Buprenorphine reduces central sensitization after repetitive C-fiber stimulation in rats

The partial micro-opioid receptor agonist buprenorphine produces antinociception through mechanisms different from those of classical opioids. In this study, we compared the effect of buprenorphine and morphine on C-fiber conditioning stimulation (CS)-induced facilitation of the flexor reflex, a model of central sensitization in decerebrate, spinalized unanesthetized rats. Intraperitoneal morphine and buprenorphine moderately depressed the baseline flexor reflex to a similar extent at doses of 1-2 and 0.03-0.1 mg/kg, respectively. Buprenorphine significantly reduced C-fiber CS-induced reflex facilitation whereas morphine at 1 or 2 mg/kg had no effect. Thus, some of the atypical antinociceptive effects of buprenorphine may be derived from its effect on central sensitization.     

Mechanical and heat sensitization of cutaneous nociceptors after peripheral inflammation in the rat

Tissue injuries commonly cause an increase in pain sensitivity, so that normally painful stimuli become more painful (hyperalgesia), and those usually associated with nonnoxious sensations evoke pain (allodynia). The neural bases for these sensory phenomena have been explored most extensively using heat injuries and experimental arthritis as models. Heat sensitization of cutaneous nociceptors is observed after burns, and sensitization of articular afferents to limb movements occurs after knee joint inflammation. These are likely to be peripheral mechanisms of hyperalgesia. Others, using different models of peripheral inflammation, have only rarely found mechanical sensitization of cutaneous nociceptors. In general these studies have failed to evaluate suprathreshold mechanical sensitivity, which has led to the concept of enhanced spinal cord processing ("central sensitization") serving as the neural substrate for mechanical hyperalgesia. In the current experiments, the mechanical and heat responses of cutaneous nociceptors supplying the glabrous skin of the rat hindpaw were studied 16-24 h after induction of acute inflammation with complete Freund's adjuvant. Single-fiber recordings were made from nociceptors in the sciatic nerve of barbiturate-anesthetized animals, and their responses compared with those obtained from nociceptors tested identically in normal animals. Nociceptors were characterized by the following: 1) graded mechanical stimuli (5-90 g) delivered with probes of tip area of 1 and 0.1 mm(2), 2) their adaptive responses to 2-min mechanical stimuli at three intensities, and 3) their responses to graded heat stimuli (40-50 degrees C). Forty-three nociceptors were studied in the inflamed state; 20 were A fibers, and the remainder were C fibers. Mechanical thresholds, determined with calibrated monofilaments, were not significantly different from controls. Sensitization to suprathreshold mechanical stimuli was observed for both A- and C-fiber nociceptors, although it was greater for the A fibers. Similarly, sensitization during testing of adaptive properties of A- and C-fiber nociceptors was seen, although it was limited to the dynamic (initial) and not the static (plateau) phase of the response. Heat sensitization was observed in 25% of A-fiber nociceptors, but the responses of C fibers to heat were depressed. Other indicators of neuronal sensitization, such as spontaneous activity and expanded receptive fields, were also observed. It was concluded that the mechanical hyperalgesia caused by peripheral inflammation could be explained by nociceptor sensitization. Central mechanisms cannot be completely ruled out as contributing to such hyperalgesia, although their role may be much smaller than previously envisaged.     

Transient receptor potential channel A1 and noxious cold responses in rat cutaneous nociceptors

The role of transient receptor potential channel A1 (TRPA1) in noxious cold sensation remains unclear. Some data support the hypothesis that TRPA1 is a transducer of noxious cold whilst other data contest it. In this study we investigated the role of TRPA1 in cold detection in cutaneous nociceptors in vivo using complementary experimental approaches. We used noxious withdrawal reflex electromyography, and single fibre recordings in vivo, to test the hypothesis that TRPA1-expressing primary afferents mediate noxious cold responses in anaesthetised rats. TRPV1 and TRPM8 agonists sensitise their cognate receptors to heat and cold stimuli respectively. Herein we show that the TRPA1 agonist cinnamaldehyde applied to the skin in anaesthetised rats did not sensitise noxious cold evoked hind limb withdrawal. In contrast, cinnamaldehyde did sensitise the C fibre-mediated noxious heat withdrawal, indicated by a significant drop in the withdrawal temperature. TRPA1 agonist thus sensitised the noxious reflex withdrawal to heat, but not cold. Thermal stimuli also sensitise transient receptor potential (TRP) channels to agonist. Activity evoked by capsaicin in teased primary afferent fibres showed a significant positive correlation with receptive field temperature, in both normal and Freund's complete adjuvant-induced cutaneous inflammation. Altering the temperature of the receptive field did not modulate TRPA1 agonist evoked-activity in cutaneous primary afferents, in either normal or inflamed skin. In addition, block of the TRPA1 channel with Ruthenium Red did not inhibit cold evoked activity in either cinnamaldehyde sensitive or insensitive cold responsive nociceptors. In cinnamaldehyde-sensitive–cold-sensitive afferents, although TRPA1 agonist-evoked activity was totally abolished by Ruthenium Red, cold evoked activity was unaffected by channel blockade. We conclude that these results do not support the hypothesis that TRPA1-expressing cutaneous afferents play an important role in noxious cold responses.     

Responses of C-fiber low threshold mechanoreceptors and nociceptors to cold were facilitated in rats persistently inflamed and hypersensitive to cold

Cold allodynia is an annoying symptom in conditions of chronic inflammation such as rheumatoid arthritis. To examine whether primary afferent nerve activities are changed in association with hypersensitivity to cold, we recorded single nerve activities from the sural nerve in persistently inflamed rats in vivo. Inflammation was induced by an injection of complete Freund's adjuvant (CFA) solution into the tibio-tarsal joint. Inflamed rats showed an increased number of paw shakes to paw immersion in 25 degrees C water (pre-inflammation: 1.15+/-0.58, 2-week inflammation: 4.70+/-1.15). We also recorded cutaneous C-fiber activities under pentobarbital anesthesia and studied their responses to thermal and mechanical stimuli. The response of C-low threshold mechanoreceptors to cooling (total discharges between 27 and 23 degrees C) increased 1.8-fold (control group: 5.17+/-1.04 impulses, inflamed group: 9.38+/-1.47 impulses). In addition, the proportion of C-nociceptor units responding to cold down to 2 degrees C was significantly greater in the inflamed group (9 out of 18 units; threshold: 10.0+/-2.6 degrees C) than in the intact group (1 out of 14 units; threshold: 4.0 degrees C). These results suggest that the facilitated responses of these primary afferents are associated with cold hypersensitivity in chronically inflamed conditions.     

Differential effects of TRPV channel block on polymodal activation of rat cutaneous nociceptors in vitro

The capsaicin receptor TRPV1 is a polymodal sensory transducer molecule in the pain pathway. TRPV1 integrates noxious heat, tissue acidosis and chemical stimuli which are all known to cause pain. Studies on TRPV1-deficient mice suggest that TRPV1 is essential for acid sensing by nociceptors and for thermal hyperalgesia in inflammation of the skin, but not for transducing noxious heat. After TRPV1, other TRPV channels were cloned with polymodal properties and sensitivity to noxious heat, named TRPV2, TRPV3 and TRPV4. While TRPV3 and TRPV4 are predominantly warm sensors, TRPV2’s threshold is in the noxious range (>52°C). However, mice deficient of TRPV2 and TRPV1 or TRPV3 or TRPV4 show no major impairment of noxious heat sensing. Ruthenium red, a water soluble polycationic dye, was found to block the pore of the capsaicin-operated cation channel TRPV1 thus interfering with all polymodal ways of TRPV1 activation. Antagonistic effects of the dye were subsequently described on many other TRP-channels, especially on the heat-sensitive ones of the vanilloid family, TRPV2, TRPV3 and TRPV4. In this study, we used the rat skin-nerve preparation to define the possible actions of ruthenium red on the proton, capsaicin and noxious heat activation of native polymodal nociceptors. Ruthenium red was found to suppress only the capsaicin-induced excitation and desensitization of these nerve endings. On the contrary, the proton and heat-induced discharge responses of the single fibres were not influenced. Additionally, we found that the dye concentration dependently increases the excitability of the neurons resulting in ongoing activity and burstlike discharge. These differential results are discussed in the light of recent findings from transgenic mouse models, and they point once more to major (pharmacological) differences between cellular models of nociception, including spinal ganglion neuron and transfected cell lines, and the real native nerve endings.     

Hyper-responsivity in a subset of C-fiber nociceptors in a model of painful diabetic neuropathy in the rat

While clinical characteristics of diabetic painful neuropathy are well described, the underlying electrophysiological basis of the exaggerated painful response to stimuli, as well as the presence of spontaneous pain, are poorly understood. In order to elucidate peripheral contributions to painful diabetic neuropathy, we quantitatively evaluated the function of C-fibers in a rat model of painful diabetic neuropathy, diabetes induced by the pancreatic beta-cell toxin streptozotocin. While there was no significant effect of diabetes on conduction velocity, mechanical threshold or spontaneous activity, the number of action potentials in response to sustained threshold and suprathreshold mechanical stimuli was significantly increased in the diabetic rats. Moreover, there was a clustering of responses of C-fibers in diabetic rats; while two-thirds of C-fibers fired at the same mean frequency as C-fibers in control rats, one-third of C-fibers in diabetic rats were markedly hyper-responsive, demonstrating a threefold increase in firing frequency. The high-firing-frequency C-fibers in rats with diabetes also had faster conduction velocity than the low-firing-frequency C-fibers in rats with diabetes or in C-fibers in control rats. The hyper-responsiveness was characterized by a selective increase of the shortest interspike intervals (<100ms) in the burst component (first 10s) of the response to a sustained suprathreshold stimulus; in the plateau phase (last 50s) of the response to a 60-s suprathreshold stimulus, we found a selective increase of interspike intervals between 100 and 300ms in hyper-responsive C-fibers in rats with diabetes. The hyper-responsiveness did not correlate with mechanical threshold, presence of spontaneous activity or location of the fiber's receptive field. In summary, in an established model of painful diabetic neuropathy in the rat, a subset of C-fibers demonstrated a marked hyper-responsiveness to mechanical stimuli. The subset was also found to have a greater mean conduction velocity than the fibers not demonstrating this hyper-responsivity. The present findings suggest that study of individual neurons in vitro may allow elucidation of the ionic basis of enhanced nociception in diabetic neuropathy.