The purpose of this study was to investigate whether unilateral inflammation of masseter muscle can cause a bilateral allodynia in rats and if so, whether TRPV1 channels may contribute to the bilateral allodynia. We took advantage of the CFA models of inflammation that have been used in studies investigating nociception in the orofacial region [16, 37, 39]. Our findings show the nocifensive behavioral response in both inflamed and contralateral non-inflamed masseter muscle. The mechanism of bilateral nocifensive behavioral response is not fully understood although bilateral allodynia after unilateral inflammation of deep craniofacial tissues has also been shown in recent studies [11, 36]. Characteristics of contralateral events after unilateral injury that follows the same pattern as at ipsilateral side but usually smaller in magnitude with briefer time course are well documented . Unilateral injection of CFA in masseter muscle complex is capable of inducing bilateral and also widespread hyperalgesia involving different parts of body in rats , and unilateral injection of hypertonic saline in masseter or overlying skin evokes bilateral enhancement of signal intensity in the brain in humans .
In this study we found bilateral nocifensive behavioral response 1 day and 4 days after unilateral injection of CFA. This result confirms induction of bilateral allodynia by unilateral deep tissue inflammation in orofacial region. Mechanical hypersensitivity developed almost immediately at site of CFA injection and the highest hypersensitivity reached at day 4. Contralateral behavioral response to mechanical stimuli follows the same pattern as the ipsilateral inflamed side, although not as immediately nor as robustly as the injection side. Head withdrawal threshold reached lowest and highly significant difference in both ipsilateral and contralateral masseter 4 days after CFA injection. Similar to this study in rats, clinical studies report bilateral pain in patients with myofascial temperomandibular disorder [41, 42]. Besides bilateral hypersensitivity, overlaps of symptoms especially in hypersensitivity disorders associated with nociceptive impairment like fibromyalgia and whiplash injury suggest that alterations in nociceptive processing may also play role in etiopathogenesis of orofacial pain [43, 44].
Previous studies from ours and others have reported the involvement of TRPV1 in orofacial pain, and its importance in muscle nociception and mechanical hyperalgesia in animal models [16, 18]. In this study, real time RT PCR was performed to detect alterations in TRPV1 mRNA expression in bilateral TRG after unilateral masseter inflammation. Up-regulation of TRPV1 mRNA expression in the trigeminal ganglion ipsilateral to inflammation found in this study confirms the importance of TRPV1 in inflammatory orofacial pain conditions [24, 45]. No changes in expression level of TRPV1 in contralateral TRG suggest that changes in ipsilateral TRG are specific to inflamed tissues. However, it is noteworthy to mention that TRPV1 is expressed not only in neurons but also in mast cells as TRPV1 expression can be induced in fibroblasts following inflammation [46–48]. Therefore, the upregulation TRPV1 observed in this study cannot exclude non-neuronal type of cells in TRG, although the great majority of expression is in neurons. The finding of increased calcitonin gene-related peptide (CGRP) mRNA level just in ipsilateral TRG in animals with bilateral allodynia also supports the view that changes in TRG are specific to inflammation and nerves innervating inflamed tissue . An increased mechanical sensitivity of the contralateral side observed in this and previous studies suggests development of secondary mechanical allodynia after masseter inflammation [11, 36]. The role of central mechanisms in contralateral pain response of masseter muscle has previously been suggested by results of bilateral c-fos expression in the brainstem after unilateral inflammation of masseter muscle . Beside TRPV1 are expressed in the sensory nerves of peripheral nervous system, including masseter muscle afferents in the TRG , a significant increase in TRPV1 expression is found in hippocampus after TMJ inflammation . Since the effect of TMJ inflammation on TRPV1 expression in the hippocampus is specific but in other brain regions (16), we decided to test if blocking the TRPV1 in CA1 region of hippocampus can attenuate bilateral masseter muscle pain response as well. In this study, the involvement of central mechanism in masseter pain conditions was supported by results of reversed head withdrawal threshold in both ipsilateral and in contralateral masseter after hippocampal injection of TRPV1 antagonist, suggesting that blocking the function of hippocampal TRPV1 can attenuate bilateral masseter muscle allodynia in rats. While ipsilateral nociception seems to depend on alterations in both peripheral and central neurons, contralateral response seems to depend mainly on alterations in higher neural centers. Although it was reasonable to expect changes in nocifensive behavioral response at the inflamed muscle site, the contralateral allodynia suggests that inflammation of masticatory muscles is likely a peripheral trigger of CNS alterations that lead to nociception of both inflamed and non inflamed tissue.
In conclusion, the results of this study support the possibility of bilateral changes in pain response after unilateral inflammation of masseter muscle in rats, and suggest the involvement of TRPV1 channels in masseter muscle pain conditions.