Shiori Murase

Bradykinin and Nerve Growth Factor Play Pivotal Roles in Muscular Mechanical Hyperalgesia after Exercise (Delayed-Onset Muscle Soreness)

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 ∼1 d. Using B2 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. B1 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 B2 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.

Cutaneous C-fiber nociceptor responses and nociceptive behaviors in aged Sprague-Dawley rats.

&NA; The change with age in pain perception in humans and the nociceptive behaviors in animals elicited by noxious stimuli to the skin are not well understood, and little is known about the peripheral neural mechanisms of cutaneous nociception in the aged. We systematically examined cutaneous nociceptor responses and nociceptive behaviors in young (9–14 w) and in aged (127–138 w) Sprague–Dawley rats. C‐fiber nociceptors in the skin were identified by mechanical and electrical stimulation, and extracellularly recorded from hind paw skin‐saphenous nerve preparations in vitro. In the aged rats, the proportions of mechano‐responsive and/or heat‐responsive C‐nociceptors were significantly lower. The proportion of mechano‐ and thermo‐insensitive units, on the other hand, was significantly increased. In addition, the response threshold to mechanical stimulus tended to be higher and the magnitude of the response tended to be smaller. There were no differences between the two age groups in the response magnitudes of mechano‐responsive C‐nociceptors to bradykinin, cold or heat. Repetitive electrical stimulation of afferent fibers revealed exaggerated slowing of conduction velocity in mechano‐responsive C‐fibers in the aged. This showed for the first time that not only receptive properties of afferent terminals but also membrane properties of conducting axons are changed in aged rats. Nociceptive behaviors in response to noxious levels of cold (cold plate test) and heat (Hargreaves’ radiant heat test) were facilitated in aged animals, while mechanical sensitivity measured by von Frey hairs remained unchanged. These discrepancies between the changes in peripheral afferents and the behavioral outcomes might be explained by facilitatory changes in the central nervous system.

TRPV1 and TRPV4 Play Pivotal Roles in Delayed Onset Muscle Soreness

Unaccustomed strenuous exercise that includes lengthening contraction (LC) often causes tenderness and movement related pain after some delay (delayed-onset muscle soreness, DOMS). We previously demonstrated that nerve growth factor (NGF) and glial cell line-derived neurotrophic factor (GDNF) are up-regulated in exercised muscle through up-regulation of cyclooxygenase (COX)-2, and they sensitized nociceptors resulting in mechanical hyperalgesia. There is also a study showing that transient receptor potential (TRP) ion channels are involved in DOMS. Here we examined whether and how TRPV1 and/or TRPV4 are involved in DOMS. We firstly evaluated a method to measure the mechanical withdrawal threshold of the deep tissues in wild-type (WT) mice with a modified Randall-Selitto apparatus. WT, TRPV1−/− and TRPV4−/− mice were then subjected to LC. Another group of mice received injection of murine NGF-2.5S or GDNF to the lateral gastrocnemius (LGC) muscle. Before and after these treatments the mechanical withdrawal threshold of LGC was evaluated. The change in expression of NGF, GDNF and COX-2 mRNA in the muscle was examined using real-time RT-PCR. In WT mice, mechanical hyperalgesia was observed 6–24 h after LC and 1–24 h after NGF and GDNF injection. LC induced mechanical hyperalgesia neither in TRPV1−/− nor in TRPV4−/− mice. NGF injection induced mechanical hyperalgesia in WT and TRPV4−/− mice but not in TRPV1−/− mice. GDNF injection induced mechanical hyperalgesia in WT but neither in TRPV1−/− nor in TRPV4−/− mice. Expression of NGF and COX-2 mRNA was significantly increased 3 h after LC in all genotypes. However, GDNF mRNA did not increase in TRPV4−/− mice. These results suggest that TRPV1 contributes to DOMS downstream (possibly at nociceptors) of NGF and GDNF, while TRPV4 is located downstream of GDNF and possibly also in the process of GDNF up-regulation.

Upregulated glial cell line‐derived neurotrophic factor through cyclooxygenase‐2 activation in the muscle is required for mechanical hyperalgesia after exercise in rats

•  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.

A novel Caspr mutation causes the shambling mouse phenotype by disrupting axoglial interactions of myelinated nerves.

The neurological mouse mutation shambling (shm) exhibits ataxia and hindlimb paresis. Positional cloning of shm showed that it encodes contactin-associated protein (Caspr), which is required for formation of the paranodal junction in myelinated nerves. The shm mutation is a TT insertion in the Caspr gene that results in a frame shift and a premature stop codon at the COOH-terminus. The truncated Caspr protein that is generated lacks the transmembrane and cytoplasmic domains. Here, we found that the nodal/paranodal axoplasm of shm mice lack paranodal junctions and contain large mitochondria and abnormal accumulations of cytoplasmic organelles that indicate altered axonal transport. Immunohistochemical analysis of mutant mice showed reduced expression of Caspr, contactin, and neurofascin 155, which are thought to form a protein complex in the paranodal region; protein 4.1B, however, was normally distributed. The mutant mice had aberrant localization of voltage-gated ion channels on the axolemma of nodal/paranodal regions. Electrophysiological analysis demonstrated that the velocityof saltatory conduction was reduced in sciatic nerves and that thevisual response was attenuated in the primary visual cortex. These abnormalities likely contribute to the neurological phenotype of the mutant mice.

Decreased nerve growth factor upregulation is a mechanism for reduced mechanical hyperalgesia after the second bout of exercise in rats.

Delayed onset muscle soreness (DOMS) is reduced when the same exercise is repeated after a certain interval. However, the mechanism for this adaptation, called a repeated bout effect, is still not well understood. Recently, we showed that upregulated nerve growth factor (NGF) triggered by B2 bradykinin receptor (B2R) activation in exercised muscle was responsible for DOMS. In this study, we investigated whether NGF upregulation was reduced after repeated bouts of exercise in rats, and if so, whether this change occurred upstream of B2R. A bout of 500 lengthening contractions (LC) was applied on day 0 and again 5 days later. DOMS was evaluated by the mechanical withdrawal threshold of the exercised extensor digitorum longus (EDL) muscle. Mechanical hyperalgesia and NGF mRNA upregulation in EDL were observed after the first LC, but not after the second LC. We then injected HOE140, a B2R antagonist with effects lasting only several hours, once before the first LC. This blocked the development of mechanical hyperalgesia and NGF mRNA upregulation not only after the first LC but also after the second LC. This suggests that adaptation occurred upstream of B2R, as the influence of the first LC was limited to that area by HOE140.

Absence of mechanical hyperalgesia after exercise (delayed onset muscle soreness) in neonatally capsaicin-treated rats.

Delayed onset muscle soreness (DOMS) appears with some delay after unaccustomed, strenuous exercise, especially after lengthening contraction (LC). It is characterized by tenderness and movement related pain, namely muscular mechanical hyperalgesia. To clarify the involvement of C-fibers in this mechanical hyperalgesia, we examined whether DOMS could be induced in rats treated neonatally with capsaicin. We confirmed that a large portion of unmyelinated afferent fibers were lost in capsaicin treated rats. In these animals, LC failed to induce muscular mechanical hyperalgesia. mRNA of nerve growth factor (NGF) in the muscle, which plays a pivotal role in maintaining mechanical hyperalgesia, was upregulated in the capsaicin treated animals similar to the vehicle treated animals. These results demonstrate that C-fiber afferents are essential in transmitting the nociceptive information from exercised muscle in DOMS.

Glial cell line‐derived neurotrophic factor sensitized the mechanical response of muscular thin‐fibre afferents in rats

The role of glial cell line‐derived neurotrophic factor (GDNF) in pain and muscular nociceptor activities is not well understood. We examined pain‐related behaviour and mechanical response of muscular thin‐fibre afferents after intramuscular injection of GDNF in rats.

Animal Models of Myofascial Trigger Points

ABSTRACT Objective: One of the typical symptoms of myofascial pain syndrome is trigger points [TrPs] in a taut band. It has been reported that hyperalgesic muscle [delayed onset muscle soreness or DOMS] after lengthening contraction [LC] has similarities with clinical TrPs, such as mechanically sensitive spots in hardening of the muscle, and electromyographic activities. Using this model we studied the mechanism by which DOMS is generated and maintained. Findings: We found that bradykinin released during exercise triggers nerve growth factor [NGF] upregulation in the muscle through the activation of B2 bradykinin receptors. NGF upregulation started 12 hours after LC, and lasted for one to two days, a compatible time course for muscle mechanical hyperalgesia. Injection of anti-NGF antibody into the exercised muscle two days after LC reversed the mechanical hyperalgesia. NGF sensitized muscle thin-fiber receptors recorded in vitro to mechanical stimulation after a short latency of 10 to 20 minutes. Conclusions: The sensitization of muscle nociceptors to mechanical stimulation by NGF upregulated in the muscle after LC is considered to be a mechanism for mechanical hyperalgesia after exercise. Determining whether there is any difference in expression of NGF or sensitivity of muscle nociceptors in the TrP and in other areas will be an important key for clarifying the mechanism of TrPs.

EP2 receptor plays pivotal roles in generating mechanical hyperalgesia after lengthening contractions

We previously demonstrated that nerve growth factor (NGF) and glial cell line‐derived neurotrophic factor (GDNF) were upregulated after lengthening contractions (LC) in exercised muscle through B2 bradykinin receptor activation and cyclooxygenase (COX)‐2 upregulation, respectively, and that these trophic factors sensitized nociceptors resulting in mechanical hyperalgesia (delayed‐onset muscle soreness, DOMS). Here, we examined the prostaglandin receptor subtype involved in DOMS. The mechanical withdrawal threshold of the exercised muscle was measured before and after LC in rats administered prostaglandin E2 receptor (EP) antagonists before LC, or in wild‐type (WT), EP2 knockout (EP2−/−), and IP knockout (IP−/−) mice. The change in expression of NGF, GDNF, or COX‐2 mRNA was examined using real‐time RT‐PCR in the muscle in EP2−/− and WT mice. None of the antagonists to EP1, EP3, and EP4 receptors (ONO‐8713, ONO‐AE5‐599, and ONO‐AE3‐208, respectively) induced a significant difference in DOMS compared with controls in rats. WT and IP−/− mice developed mechanical hyperalgesia after LC, but EP2−/− mice did not. Upregulation of NGF, GDNF, and COX‐2 mRNA was observed after LC in WT mice but not in EP2−/− mice. Injecting an EP2 agonist (ONO‐AE1‐259‐01) into the mouse muscle increased expression of COX‐2 mRNA. These results suggest that EP2 contributes to generating mechanical hyperalgesia through positive feedback upregulation of COX‐2 expression in muscle after LC.