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. It is concluded that latency shifts are reliable indicators of C-fiber activation, being sensitive enough to detect even single spike responses. Furthermore, latency increases may be used as a relative measure of C-fiber activation, e.g., when comparing responses to stimuli of different strength.     

PKC在成年大鼠脊髓背角C-纤维诱发电位长时程增强的诱导和维持中的作用

目的：探讨蛋白激酶C (PKC) 在大鼠脊髓背角C-纤维诱发电位长时程增强（LTP）的诱导和维持中的作用。方法： 细胞外记录技术在脊髓腰膨大部记录背角浅层神经元C-纤维诱发电位。 结果：(1) PKC的选择性抑制剂chelerythrine（200 μmol/L）或G 6983（100 μmol/L）对脊髓背角C-纤维诱发电位的基础电位没有影响，但可完全阻断脊髓背角LTP的诱导。(2) Chelerythrine或G 6983呈时间依赖性翻转脊髓背角LTP。在LTP 诱导后15 min，脊髓局部给予chelerythrine（200 μmol/L）后，LTP逐渐降低，于给药后70 min降至对照水平；而G 6983（100 μmol/L）产生同chelerythrine相似的效应，在用药后110 min，LTP降至对照水平。但同样浓度的chelerythrine或G 6983在LTP 诱导后3 h，均不能翻转业已建立的LTP。结论： PKC参与脊髓背角C-纤维诱发电位LTP的诱导和早期维持，而不影响晚期LTP的维持。     

GABA(A) receptor facilitation of neurokinin release from primary afferent terminals in the rat spinal cord

Our goal was to test the following hypotheses: 1) GABA_A receptors facilitate neurokinin release from primary afferent terminals; 2) they do this by suppressing an inhibitory effect of GABA_B receptors; 3) the activation of these two receptors is controlled by the firing frequency of primary afferents. We evoked neurokinin release by stimulating the dorsal root attached to spinal cord slices, and measured it using neurokinin 1 receptor (NK1R) internalization. Internalization evoked by root stimulation at 1 Hz (but not at 100 Hz) was increased by the GABA_A receptor agonists muscimol (effective concentration of drug for 50% of the increase [EC₅₀] 3 μM) and isoguvacine (EC₅₀ 4.5 μM). Internalization evoked by root stimulation at 100 Hz was inhibited by the GABA_A receptor antagonists bicuculline (effective concentration of drug for 50% of the inhibition [IC₅₀] 2 μM) and picrotoxin (IC₅₀ 243 nM). Internalization evoked by incubating the root with capsaicin (to selectively recruit nociceptive fibers) was increased by isoguvacine and abolished by picrotoxin. Therefore, GABA_A receptors facilitate neurokinin release. Isoguvacine-facilitated neurokinin release was inhibited by picrotoxin, low Cl⁻, low Ca²⁺, Ca²⁺ channel blockers and N-methyl-d-aspartate receptor antagonists. Bumetanide, an inhibitor of the Na⁺-K⁺-2Cl⁻ cotransporter, inhibited isoguvacine-facilitated neurokinin release, but this could be attributed to a direct inhibition of GABA_A receptors. The GABA_B agonist baclofen inhibited NK1R internalization evoked by 100 Hz root stimulation (IC₅₀ 1.5 μM), whereas the GABA_B receptor antagonist (2S)-3-[[(1S)-1-(3,4-dichlorophenyl)ethyl]amino-2-hydroxypropyl](phenylmethyl) phosphinic acid (CGP-55845) increased NK1R internalization evoked by 1 Hz root stimulation (EC₅₀ 21 nM). Importantly, baclofen inhibited isoguvacine-facilitated neurokinin release, and CGP-55845 reversed the inhibition of neurokinin release by bicuculline. In conclusion, 1) GABA_B receptors located presynaptically in primary afferent terminals inhibit neurokinin release; 2) GABA_A receptors located in GABAergic interneurons facilitate neurokinin release by suppressing GABA release onto these GABA_B receptors; 3) high frequency firing of C-fibers stimulates neurokinin release by activating GABA_A receptors and inhibiting GABA_B receptors, whereas low frequency firing inhibits neurokinin release by the converse mechanisms.     

吗啡对外周刺激诱导的大鼠海马CA1区长时程增强(LTP)的抑制作用

目的观察吗啡急性中枢应用,对外周电刺激坐骨神经诱导的海马CA1区LTP及海马痛觉调制作用的影响,并探讨其可能的机制。方法25只雄性SD大鼠200～260g,随机分为5组。单刺激坐骨神经,各组侧脑室给药后给予强直刺激,观察场电位的变化。结果单刺激坐骨神经可在海马CA1区诱导出场电位(21.43±4.54)μV,其潜伏期较长(169.94±14.65)ms及阈值较高(平均15V),据此,推断是C类纤维诱发的;强直刺激后海马CA1区可出现持续时间在2h以上LTP现象;中、大剂量吗啡侧脑室注射可抑制海马CA1区LTP,纳洛酮可阻断这种作用。结论LTP是海马痛觉调制的生物学机制之一,阿片受体在其中起重要作用。吗啡对C-类纤维诱导的海马CA1区LTP的抑制作用可能通过阿片受体实现。     

The role of neuropeptides and capsaicin-sensitive fibres in glutamate-induced nociception and paw oedema in mice

This study sought to establish whether sensory neuropeptides and the capsaicin-sensitive fibres are involved in the nociception and oedema formation caused by intraplantar (i.pl.) injection of glutamate into the mouse paw. The i.pl. co-injection of the selective neurokinin (NK) NK(2) (SR 48968, 0.05-0.5 nmol/paw), and to a lesser extent the selective NK(1) (FK 888, 0.25-1.0 nmol/paw) receptor antagonists, resulted in a significant inhibition of glutamate-induced nociception. The percentages of inhibition were 82 and 37%, respectively. In contrast, the selective NK(3) receptor antagonist (SR 142801, 0.25-1.0 nmol/paw) failed to significantly affect glutamate-induced nociception. SR 48968, but not FK 888 or SR 142801, significantly inhibited (36%) glutamate-induced paw oedema formation. The i.pl. injection of kinin B(1) receptor antagonist des-Arg(9)-[Leu(8)]-BK (0.2-0.8 nmol/paw), but not the B(2) receptor antagonist HOE 140 (1.0-4.0 nmol/paw), together with glutamate, also inhibited glutamate-induced nociception (53%) in a graded manner, without affecting glutamate-induced paw oedema. The i.pl. co-injection of the calcitonin gene-related peptide (CGRP) receptor antagonist CGRP(8-37) (1 nmol/paw) failed to significantly inhibit glutamate-induced nociception or oedema. Finally, neonatal-capsaicin (50 mg/kg, s.c.) treatment inhibited glutamate-induced nociception by 69% and to a lesser extent glutamate-mediated oedema formation (30%). Collectively, the current results indicate that the nociception caused by i.pl. injection of glutamate in mice is clearly mediated by capsaicin-sensitive fibres and by release of neurokinins from sensory neurones that activate NK(2) receptors and to a lesser extent NK(1) receptors. Furthermore, kinins acting at B(1) (but not at B(2)) receptors also largely account for glutamate-mediated nociceptive behaviour response. In contrast, glutamate-induced paw oedema seems to be primarily mediated via activation of NK(2) receptors and stimulation of capsaicin-sensitive C-fibres. CGRP receptors do not seem to be involved in either of the glutamate responses.     

Effect of olvanil on the afferent and efferent function of capsaicin-sensitive C-fibers in guinea pig airways

The aim of the present study was to examine the ability of the nonpungent vanilloid VR1 receptor agonist, olvanil, to activate the afferent and efferent function of capsaicin-sensitive C-fibers in guinea pig airways. We found that while capsaicin (10 nM-10 microM) and resiniferatoxin (0.1 nM-1.0 microM) evoked a robust contraction of the guinea pig trachea in vitro, olvanil (10 nM-10 microM) was a weak spasmogen. In addition, pretreatment with olvanil caused only a minor reduction of subsequent responses to capsaicin or resiniferatoxin. Using single fiber recording from guinea pig airway C-fibers, we found that olvanil (10 microM) did not evoke action potential discharge although these fibers responded vigorously to capsaicin after prolonged treatment with olvanil (10 microM). These findings are indicative of significant differences in the relative sensitivity of vanilloid VR1 receptor-transfected cells and the peripheral terminals of airway C-fibers to pungent and nonpungent vanilloid VR1 receptor agonists.     

Microinjection of a 5-HT3 receptor agonist into the NTS of awake rats inhibits the bradycardic response to activation of the von Bezold-Jarisch reflex

In the present study we investigated the effects of bilateral microinjection into the lateral commissural nucleus tractus solitarius (NTS) of 2-methyl-5-HT, a 5-HT3 receptor agonist, on the bradycardic response of the von Bezold-Jarisch reflex of awake rats. We evaluated mainly the bradycardic response because in previous studies we documented that the hypotensive response of the von-Bezold-Jarisch reflex in awake rats is secondary to the intense bradycardic response. The Bezold-Jarisch reflex was activated by intravenous injection of serotonin (8 microg/kg) in awake rats before and 1, 3, 10, 20 and 60 min after bilateral microinjection of 2-methyl-5-HT (5 nmol/50 nl, n = 8) into the NTS. Microinjections of 2-methyl-5-HT into the NTS produced a significant increase in basal mean arterial pressure [(MAP), 97 +/- 4 vs. 114 +/- 4 mmHg), no changes in basal heart rate and a significant reduction in bradycardic (-78 +/- 19; -94 +/- 24 and -107 +/- 21 bpm) and hypotensive (-16 +/- 4; -10 +/- 5 and -17 +/- 4 mmHg) responses to activation of the von Bezold-Jarisch reflex at 3, 10 and 20 min, respectively, when compared with the control value (-231 +/- 13 bpm and -43 +/- 4 mmHg). The data of the present study suggest that serotonin acting on 5-HT3 receptors in the NTS may play an important inhibitory neuromodulatory role in the bradycardic response to activation of the von Bezold-Jarisch reflex.     

Cholecystokinin and its antagonist lorglumide respectively attenuate and facilitate morphine-induced inhibition of C-fiber evoked discharges of dorsal horn nociceptive neurons

Extracellular single unit recordings were made from dorsal horn nociceptive neurons of intact, urethane-anesthetized rats during controlled electrical stimulation of the hind paw. Neither local superfusion of cholecystokinin octapeptide (CCK; 6.4 pmol to 20 nmol) nor the CCK antagonist lorglumide (LGM; 145 fmol to 145 pmol) significantly altered A- or C-fiber evoked firing or spontaneous activity. Pretreatment with CCK, however, significantly attenuated, whereas LGM enhanced, morphine-induced inhibition of C-evoked firing. These findings provide further evidence that CCK functions as a selective antagonist of opioid-induced analgesia.     

The effects of electrical stimulation of A and C visceral afferent fibres on the excitability of viscerosomatic neurones in the thoracic spinal cord of the cat

Single unit electrical activity has been recorded from viscerosomatic neurons in the lower thoracic spinal cord of decerebrate spinalized cats. The responses of the cells to electrical stimulation of afferent fibres in the splanchnic (SPLN) nerve and the effects of repetitive stimulation of somatic and visceral afferent C-fibres have been studied. Four groups of viscerosomatic neurones could be distinguished according to the type of visceral afferent input of the cells: (1) A-only cells (32.9%), driven only by stimulation of A delta afferent fibres in the SPLN nerve; (2) C-only cells (3%), driven only by stimulation of C afferent fibres in the SPLN nerve; (3) A + C cells (45.7%), driven by both A delta and C afferent fibres in the SPLN nerve; and (4) A + C? cells (18.6%), driven by A delta visceral afferents and showing signs of responsiveness to C-fibres though lacking a distinct response volley to visceral C-fibre activation. Two cells of the A + C group and located in lamina I of the dorsal horn responded to SPLN nerve stimulation in a manner consistent with the afferent fibre composition of the nerve, that is, showed evidence of strong monosynaptic links with SPLN afferent C-fibres and weaker responses to SPLN A delta afferents. Excitability changes of viscerosomatic neurones ('wind up', 'wind down' and changes in background activity) were also observed in the majority of neurones following electrical stimulation of somatic and of visceral afferent C-fibres.(ABSTRACT TRUNCATED AT 250 WORDS)