Unaccustomed strenuous exercise that includes lengthening contraction (LC) often causes delayed-onset muscle soreness (DOMS), a kind of muscular mechanical hyperalgesia. The substances that induce this phenomenon are largely unknown. Peculiarly, DOMS is not perceived during and shortly after exercise, but rather is first perceived after ϳ1 d. Using B 2 bradykinin receptor antagonist HOE 140, we show here that bradykinin released during exercise plays a pivotal role in triggering the process that leads to muscular mechanical hyperalgesia. HOE 140 completely suppressed the development of muscular mechanical hyperalgesia when injected before LC, but when injected 2 d after LC failed to reverse mechanical hyperalgesia that had already developed. B 1 antagonist was ineffective, regardless of the timing of its injection. Upregulation of nerve growth factor (NGF) mRNA and protein occurred in exercised muscle over a comparable time course (12 h to 2 d after LC) for muscle mechanical hyperalgesia. Antibodies to NGF injected intramuscularly 2 d after exercise reversed muscle mechanical hyperalgesia. HOE 140 inhibited the upregulation of NGF. In contrast, shortening contraction or stretching induced neither mechanical hyperalgesia nor NGF upregulation. Bradykinin together with shortening contraction, but not bradykinin alone, reproduced lasting mechanical hyperalgesia. We also showed that rat NGF sensitized thin-fiber afferents to mechanical stimulation in the periphery after 10 -20 min. Thus, NGF upregulation through activation of B 2 bradykinin receptors is essential (though not satisfactory) to mechanical hyperalgesia after exercise. The present observations explain why DOMS occurs with a delay, and why lengthening contraction but not shortening contraction induces DOMS.

The roles of ion channels in sensory neurons were examined in experimental models of muscle pain in the rat. Rats were injected with 50 microl of 4% carrageenan or subjected to an eccentric exercise (ECC) of the gastrocnemius muscle (GM). The Randall-Selitto and von Frey tests were performed on the calves to evaluate mechanical hyperalgesia of the muscle. The changes in expression of four genes and proteins of ion channels in dorsal root ganglia were examined using quantitative PCR and immunohistochemistry, respectively. Effects of antagonists to transient receptor potential (TRP) channels and acid sensing ion channels (ASICs) on the mechanical hyperalgesia induced by carrageenan injection or ECC were evaluated. The mechanical hyperalgesia was observed 6-24h after carrageenan injection and 1-3 days after ECC in the Randall-Selitto test. Infiltrations of the inflammatory cells in the GM were seen in carrageenan-injected animals but not in those subjected to ECC. Expressions of genes and proteins in sensory neurons showed no changes. Intramuscular injection of antagonists to TRPV1 showed an almost complete suppressive effect on ECC-induced muscle hyperalgesia but not a carrageenan-induced one. Antagonists to TRP channels and ASICs showed suppressive effects for both carrageenan- and ECC-induced muscle hyperalgesia. The carrageenan injection and ECC models are useful models of acute inflammatory pain and delayed onset muscle soreness (DOMS), respectively, and the time course and underlying etiology might be different. TRP channels and ASICs are closely related to the development of muscle mechanical hyperalgesia, and TRPV1 is involved in ECC-induced DOMS.

Taguchi, Toru, Jun Sato, and Kazue Mizumura. Augmented mechanical response of muscle thin-fiber sensory receptors recorded from rat muscle-nerve preparations in vitro after eccentric contraction. J Neurophysiol 94: 2822Neurophysiol 94: -2831Neurophysiol 94: , 2005 doi:10.1152/jn.00470.2005. Unaccustomed strenuous exercise, especially that from eccentric muscular work, often causes muscle tenderness, which is a kind of mechanical hyperalgesia. We developed an animal model of delayed-onset muscle soreness (DOMS) from eccentric muscular contraction (ECC) in rats and demonstrated the existence of muscle tenderness by means of behavioral pain tests and c-Fos protein expression in the spinal dorsal horn. The purpose of the present study was to examine whether the sensitivities of muscle thin-fiber sensory receptors to mechanical, chemical, and thermal stimuli were altered after repetitive ECC in a rat model of DOMS. ECC was caused in the animals by electrical stimulation of the common peroneal nerve innervating the extensor digitorum longus muscle (EDL) while the muscle was being stretched. Activities of single thin-fiber receptors (sensitive to pressure but insensitive to stretch, with conduction velocity slower than 2.0 m/s) were recorded from muscle (EDL)-nerve preparations in vitro 2 days after ECC when mechanical hyperalgesia was at its peak. The mechanical threshold of thin-fiber receptors was found to be very much lower in the ECC preparations than in the nontreated control (CTR) [median 65.4 mN (interquartile range [IQR]; 46.6 -122.0 mN) in the CTR preparation vs. 38.2 mN (IQR; 26.8 -55.8 mN) in the ECC, P Ͻ 0.001]. In addition, the total number of evoked discharges during a ramp mechanical stimulus, taken as an index of the magnitude of the mechanical response, nearly doubled in the ECC preparations compared with the CTR [24.7 spikes (IQR; 14.2-37.1 spikes) in the CTR preparation vs. 54.2 spikes (IQR; 24.3-89.0 spikes) in the ECC, P Ͻ 0.001]. In contrast, the numbers of discharges induced by chemical (pH 5.5, lactic acid, adenosine triphosphate, and bradykinin) and thermal (cold and heat) stimuli were not different between the two preparations. These results suggest that augmentation of the mechanical response in muscle thin-fiber sensory receptors might be related to the muscle tenderness in DOMS after ECC.

Key points• Unaccustomed strenuous exercise that includes lengthening contraction often causes delayed onset muscle soreness (DOMS), characterised as muscular mechanical hyperalgesia.• It has been reported that bradykinin triggers upregulation of nerve growth factor in exercised muscle, sensitizing nociceptors and resulting in DOMS, but additional mechanism(s) may be involved.• We showed that pretreatment with cyclooxygenase (COX)-2 inhibitors completely suppressed the development of DOMS, but treatment 2 days after lengthening contraction failed to reverse existing mechanical hyperalgesia.• We demonstrated that COX-2 induced upregulation of glial cell line-derived neurotrophic factor (GDNF) and that intramuscularly injected anti-GDNF antibody reduced muscle mechanical hyperalgesia after exercise.• These results suggest that upregulation of GDNF through COX-2 activation is essential to mechanical hyperalgesia after exercise, and is another pathway alongside the bradykinin-nerve growth factor pathway that is involved in DOMS development.Abstract Unaccustomed strenuous exercise that includes lengthening contraction (LC) often causes delayed onset muscle soreness (DOMS), characterised as muscular mechanical hyperalgesia. Previously we reported that a bradykinin-like substance released from the muscle during exercise plays a pivotal role in triggering the process of muscular mechanical hyperalgesia by upregulating nerve growth factor (NGF) in exercised muscle of rats. We show here that cyclooxygenase (COX)-2 and glial cell line-derived neurotrophic factor (GDNF) are also involved in DOMS. COX-2 inhibitors but not COX-1 inhibitors given orally before LC completely suppressed the development of DOMS, but when given 2 days after LC they failed to reverse the mechanical hyperalgesia. COX-2 mRNA and protein in exercised muscle increased six-to 13-fold in mRNA and 1.7-2-fold in protein 0-12 h after LC. COX-2 inhibitors did not suppress NGF upregulation after LC. Instead, we found GDNF mRNA was upregulated seven-to eight-fold in the exercised muscle 12 h-1 day after LC and blocked by pretreatment of COX-2 inhibitors. In situ hybridisationThe authors E. Terazawa and K. Hirate passed away from illness during the course of this experiment (on 13 studies revealed that both COX-2 and GDNF mRNA signals increased at the periphery of skeletal muscle cells 12 h after LC. The accumulation of COX-2 mRNA signals was also observed in small blood vessels. Intramuscular injection of anti-GDNF antibody 2 days after LC partly reversed DOMS. Based on these findings, we conclude that GDNF upregulation through COX-2 activation is essential to mechanical hyperalgesia after exercise.

Delayed onset muscle soreness (DOMS) is quite common, but the mechanism for this phenomenon is still not understood; even the existence of muscle tenderness (mechanical hyperalgesia) has not been demonstrated in experimental models. We developed an animal model of DOMS by inducing eccentric contraction (lengthening contraction, ECC) to the extensor digitorum longus muscle (EDL), and investigated the existence of mechanical hyperalgesia in the EDL by means of behavioural pain tests (Randall-Selitto test and von Frey hair test, applied to/through the skin on the EDL muscle) and c-Fos expression in the spinal dorsal horn. We found that the mechanical withdrawal threshold measured with the Randall-Selitto apparatus decreased significantly between 1 and 3 days after ECC, while that measured by von Frey hairs did not. The group that underwent stretching of the muscle only (SHAM group) showed no change in mechanical pain threshold in either test. These results demonstrated that the pain threshold of deep tissues (possibly of the muscle) decreased after ECC. c-Fos immunoreactivity in the dorsal horn (examined 2 days after ECC/SHAM exercise) was not changed by either ECC or compression (1568 mN) to the EDL muscle by itself, but it was significantly increased by applying compression to the EDL muscle 2 days after ECC. This increase was observed in the superficial dorsal horn of the L4 segment of the ipsilateral side, and was clearly suppressed by morphine treatment (10 mg kg −1 , I.P.). These results demonstrated the existence of mechanical hyperalgesia in the muscle subjected to ECC. This model could be used for future study of the neural mechanism of muscle soreness.

Delayed-onset muscle soreness (DOMS) is quite a common consequence of unaccustomed strenuous exercise, especially exercise containing eccentric contraction (lengthening contraction, LC). Its typical sign is mechanical hyperalgesia (tenderness and movement related pain). Its cause has been commonly believed to be micro-damage of the muscle and subsequent inflammation. Here we present a brief historical overview of the damage-inflammation theory followed by a discussion of our new findings. Different from previous observations, we have observed mechanical hyperalgesia in rats 1-3 days after LC without any apparent microscopic damage of the muscle or signs of inflammation. With our model we have found that two pathways are involved in inducing mechanical hyperalgesia after LC: activation of the B2 bradykinin receptor-nerve growth factor (NGF) pathway and activation of the COX-2-glial cell line-derived neurotrophic factor (GDNF) pathway. These neurotrophic factors were produced by muscle fibers and/or satellite cells. This means that muscle fiber damage is not essential, although it is sufficient, for induction of DOMS, instead, NGF and GDNF produced by muscle fibers/satellite cells play crucial roles in DOMS.

The level of wakefulness is one of the major factors affecting nociception and pain. Stress-induced analgesia supports an animal’s survival via prompt defensive responses against predators or competitors. Previous studies have shown the pharmacological effects of orexin peptides on analgesia. However, orexin neurons contain not only orexin but also other co-transmitters such as dynorphin, neurotensin and glutamate. Thus, the physiological importance of orexin neuronal activity in nociception is unknown. Here we show that adult-stage selective ablation of orexin neurons enhances pain-related behaviors, while pharmacogenetic activation of orexin neurons induces analgesia. Additionally, we found correlative activation of orexin neurons during nociception using fiber photometry recordings of orexin neurons in conscious animals. These findings suggest an integrative role for orexin neurons in nociceptive perception and pain regulation.

Mechanisms and structures which are involved in eccentric exercise-induced delayed onset muscle soreness (DOMS) are not yet clarified. Tissue and site specificity may be important considerations in afferent sensitisation following eccentric exercise. This study investigated the nociceptive response to hypertonic sodium solution applied to fascial/epimysium tissue and mechanically sensitised sites in muscle by assessing (1) afferent recordings in animals and (2) psychophysical assessment in humans. Seventeen male rats underwent eccentric contraction of extensor digitorum longus muscle, while 11 rats served as an unexercised naïve group. Two days post-exercise, group IV afferent fibre activity was recorded in response to superfusion of hypertonic Krebs solution on the mechanically sensitised muscle/epimysium site. Mechanical sensitisation was confirmed with significant increases in afferent response and decreases in threshold to mechanical stimulation in the eccentrically exercised rats compared to naïve rats. There was no difference in afferent response magnitude to hypertonic Krebs solution between exercise and naïve groups. In the human study, 13 volunteers participated. After bilateral assessment of pressure pain thresholds (PPT) along the tibialis anterior muscles, eccentric exercise was performed to induce DOMS in m. tibialis anterior of one leg. Site of maximal mechanical sensitivity was identified 24 h later and injected with hypertonic saline at fascial and deep muscle levels. The corresponding site on the opposite unexercised leg served as a control. Fascial injection of the exercised muscle caused significantly higher pain intensity compared to all other injections. Response to deep muscle stimulation was not different between sides. This suggests that fascia rather than muscle tissue is important in DOMS associated sensitisation.