The relationship between Meckel's cartilage resorption and incisor tooth germ in mice.

Our understanding of the initiation and cellular mechanisms underlying endochondral resorption of Meckel's cartilage (MC) remains limited. Several studies have shown that the resorption site of MC and the mandibular incisor tooth germ are located close to each other. However, whether incisor tooth germ development is involved in MC resorption remains unclear. In this study, we aimed to elucidate the spatio-temporal interaction between the initiation site of MC resorption and the development of incisor tooth germs in an embryonic mouse model. To this effect, we developed a histology-based three-dimensional (3D) reconstruction technique using paraffin-embedded serial sections of various tissues in the jaw. The serial sections were cut in the frontal section and the tissue constituents (e.g., MC, incisor, and mineralized mandible) were studied using conventional and enzyme-based histochemistry. The outline of each component was marked on the frontal sectional images and 3D structures were constructed. To assess the vascular architecture at the site of MC resorption, immunohistochemical staining using anti-laminin, anti-factor VIII, and anti-VEGF antibodies was performed. MC resorption was first observed on the lateral incisor-facing side of the cartilage rods at sites anterior to the mental foramen on E16.0. The 3D analysis suggested that: (a) the posterior region of the clastic cartilage resorption corresponds to the cervical loop of the incisor; (b) the cervical portion of the tooth germ inflates probably due to temporal cellular congestion prior to differentiation into matrix-producing cells; (c) the incisor tooth germ tissue is present in close proximity to MC even in mouse with continuously growing tooth and determines the disappearance of MC as the tooth development.

Effect of local administration of eicosapentaenoic acid on the jaw-opening reflex in rats.

Although eicosapentaenoic acid (EPA) application in vitro inhibits voltage-gated Na+ (Nav) channels in excitable tissues, the acute local effect of EPA on the jaw-opening reflex in vivo remains unknown. The aim of the present study was to determine whether local administration of EPA to adult male Wistar rats could attenuate the excitability of the jaw-opening reflex in vivo, including nociception. The jaw-opening reflex evoked by electrical stimulation of the tongue was recorded by a digastric muscle electromyogram (dEMG) in pentobarbital-anesthetized rats. The amplitude of the dEMG response was significantly increased in proportion to the electrical stimulation intensity (1×-5× threshold). At 3×, local administration of EPA dose-dependently inhibited the dEMG response, lasting 60 min, with maximum inhibition observed within approximately 10 min. The mean magnitude of dEMG signal inhibition by EPA was almost equal to that observed with a local anesthetic, 1% lidocaine, and with a half dose of lidocaine plus a half dose of EPA. These findings suggest that EPA attenuates the jaw-opening reflex, possibly by blocking Nav channels of primary nerve terminals, and strongly support the idea that EPA is a potential therapeutic agent and complementary alternative medicine for the prevention of acute trigeminal nociception.

CXCL12 promotes CCR7 ligand-mediated breast cancer cell invasion and migration toward lymphatic vessels.

Chemokines are a family of cytokines that mediate leukocyte trafficking and are involved in tumor cell migration, growth, and progression. Although there is emerging evidence that multiple chemokines are expressed in tumor tissues and that each chemokine induces receptor-mediated signaling, their collaboration to regulate tumor invasion and lymph node metastasis has not been fully elucidated. In this study, we examined the effect of CXCL12 on the CCR7-dependent signaling in MDA-MB-231 human breast cancer cells to determine the role of CXCL12 and CCR7 ligand chemokines in breast cancer metastasis to lymph nodes. CXCL12 enhanced the CCR7-dependent in vitro chemotaxis and cell invasion into collagen gels at suboptimal concentrations of CCL21. CXCL12 promoted CCR7 homodimer formation, ligand binding, CCR7 accumulation into membrane ruffles, and cell response at lower concentrations of CCL19. Immunohistochemistry of MDA-MB-231-derived xenograft tumors revealed that CXCL12 is primarily located in the pericellular matrix surrounding tumor cells, whereas the CCR7 ligand, CCL21, mainly associates with LYVE-1+ intratumoral and peritumoral lymphatic vessels. In the three-dimensional tumor invasion model with lymph networks, CXCL12 stimulation facilitates breast cancer cell migration to CCL21-reconstituted lymphatic networks. These results indicate that CXCL12/CXCR4 signaling promotes breast cancer cell migration and invasion toward CCR7 ligand-expressing intratumoral lymphatic vessels and supports CCR7 signaling associated with lymph node metastasis.

Local administration of genistein as a local anesthetic agent inhibits the trigeminal nociceptive neuronal activity in rats.

A modulatory role has been reported for the isoflavone, genistein, on voltage-gated Na+ channels in the trigeminal ganglion in vitro. However, the acute effects of genistein in vivo, particularly on nociceptive transmission in the trigeminal system, remain to be determined. The aim of the present study was to examine whether acute local genistein administration to rats attenuates the excitability of wide-dynamic range (WDR) spinal trigeminal nucleus caudalis (SpVc) neurons in response to nociceptive and non-nociceptive mechanical stimulation in vivo. Extracellular single unit recordings were made from SpVc WDR neurons in response to orofacial non-noxious and noxious mechanical stimulation of pentobarbital-anesthetized rats. The effects of local administration of genistein, lidocaine, and lidocaine with genistein to the receptive field on the discharge frequency of SpVc neurons were evaluated. The mean firing frequency of SpVc WDR neurons in response to both non-noxious and noxious mechanical stimuli was significantly and dose-dependently (0.1-10 mM) inhibited by genistein, and maximum inhibition of the discharge frequency of both non-noxious and noxious mechanical stimuli was seen within 10 min. The inhibitory effect of genistein lasted for 20 min and was reversible. No significant difference was seen between the relative magnitude of inhibition by genistein on the SpVc WDR neuronal discharge frequency for noxious and non-noxious stimulation. The mean magnitude of inhibition by genistein (10 mM) on SpVc neuronal discharge frequency was almost equal to that of the local anesthetic, 1 % lidocaine (37 mM). Local injection of half-dose of lidocaine replaced the half-dose of genistein. These results suggest that local injection of genistein into the peripheral receptive field suppresses the excitability of SpVc neurons, possibly via inhibition of voltage-gated Na+ channels in the nociceptive nerve terminals of trigeminal ganglion. Therefore, administration of genistein as a local anesthetic may provide relief from trigeminal nociceptive pain without side effects, thus contributing to the area of complementary and alternative medicines.

Modulatory mechanism underlying how dietary constituents attenuate orofacial pain.

Physiological pain protects the body and its systems from damage, but pathological pain has no obvious biological role. Complementary alternative medicine (CAM) agents are being increasingly studied in the treatment of clinical pain, and some dietary constituents (polyphenol, carotenoids, and fatty acids) and supplements may modify pain pathways. Because these substances modulate neuronal excitability-including the trigeminal pain pathway via various voltage-gated ionic channels and transient receptor potential and ligand-gated channels, dietary constituents could contribute to CAM as therapeutic agents for attenuating orofacial noxious sensory information. This review summarizes the current understanding of the mechanisms by which dietary constituents might attenuate excitability of trigeminal nociceptive neurons implicated in blocking pain, particularly in relation to the authors' recent experimental data, and discusses the development of functional foods and the contribution of dietary constituents in the relief of clinical dental pain without the side effects of nonsteroidal anti-inflammatory drugs.

Effect of resveratrol on the hyperexcitability of nociceptive neurons associated with ectopic hyperalgesia induced by experimental tooth movement.

The present study investigated whether, under in vivo conditions, systemic administration of resveratrol attenuates the experimental tooth movement-induced ectopic hyperalgesia associated with hyperexcitability of nociceptive trigeminal spinal nucleus caudalis (SpVc) neurons. The threshold of escape from mechanical stimulation applied to the ipsilateral whisker pad in rats exposed to experimental tooth movement was significantly lower than seen in control rats from day 1 to 3 following movement of the right maxillary first molar tooth. The lowered mechanical threshold in the rats exposed to experimental tooth movement had almost returned to the level of sham-treated naïve rats at day 3 following administration of resveratrol. The mean mechanical threshold of nociceptive SpVc neurons was significantly lower after experimental tooth movement but the lower threshold could be reversed by administration of resveratrol. The higher discharge frequency of nociceptive SpVc neurons for noxious mechanical stimuli observed in rats exposed to experimental tooth movement was statistically significantly lower following resveratrol administration. These results suggest that resveratrol attenuates experimental tooth movement-induced mechanical ectopic hyperalgesia via suppression of peripheral and/or central sensitization. These findings support the idea that resveratrol, a complementary alternative medicine, is a potential therapeutic agent for the prevention of experimental tooth movement-induced ectopic hyperalgesia.

Resolvin D1 suppresses inflammation-induced hyperexcitability of nociceptive trigeminal neurons associated with mechanical hyperalgesia.

7S,8R,17S-trihydroxy-4Z,9E,11E,13Z,15E,19Z-docosahexaenoic acid (resolvin D1 [RvD1]) is biosynthesized from docosahexaenoic acid (DHA), and belongs to a novel family of lipid mediators showing remarkable anti-inflammatory effects; however, the effect of RvD1 on inflammation-induced hyperexcitability of nociceptive neurons under in vivo conditions remains to be determined. The present study, therefore, investigated whether under in vivo conditions, systemic administration of RvD1 could attenuate the inflammation-induced hyperexcitability of spinal trigeminal nucleus caudalis (SpVc) wide-dynamic range (WDR) neurons associated with hyperalgesia in rats. The threshold of escape from mechanical stimulation applied to the orofacial area in rats with complete Freund's adjuvant-induced inflammation was significantly lower than in naïve rats. The lowered mechanical threshold in rats with inflammation was returned to control levels following administration of RvD1 (3 ng/kg, i.p.) for 3 days. The mean discharge frequency of SpVc WDR neurons in rats with inflammation was significantly decreased after RvD1 administration for both non-noxious and noxious mechanical stimuli. Increased spontaneous discharge of SpVc WDR neurons in rats with inflammation was also significantly decreased after RvD1 administration. Noxious pinch-evoked afterdischarge frequency and occurrence in rats with inflammation was significantly diminished after RvD1 administration. Expansion of the receptive field in rats with inflammation also returned to control levels after RvD1 administration. These results suggest that administration of RvD1 attenuates inflammation-induced hyperexcitability of SpVc WDR neurons associated with inflammatory hyperalgesia. These findings support the idea that RvD1, derived from DHA, as well as DHA itself, are potential complementary or alternative therapeutic agents for the alleviation of inflammatory hyperalgesia.

Resveratrol suppresses nociceptive jaw-opening reflex via 5HT3 receptor-mediated GABAergic inhibition　.

Systemic administration of the dietary constituent, resveratrol, was previously shown to inhibit the nociceptive jaw-opening reflex (JOR) via the endogenous opioid system. The present study investigated whether resveratrol could similarly affect the JOR under in vivo conditions via 5HT3 receptor-mediated GABAergic inhibition. We used electrical stimulation of the tongue in pentobarbital-anesthetized rats to evoke the JOR, which was recorded as the anterior belly of the digastric muscle electromyograms (dEMG). Intravenous administration of resveratrol (2 mg/kg) reduced the dEMG amplitude in response to three times the determined threshold electrical stimulation, with maximum inhibition reached within approximately 10 min. These inhibitory effects on the JOR were reversible to control levels after approximately 20 min. Pretreatment of rats with either 5HT3 receptor antagonist, ondansetron (0.25-1 mg/kg, i.p.), or GABAA receptor antagonist, bicuculline (0.5-1 mg/kg, i.p.), significantly and dose-dependently attenuated the inhibitory effects of resveratrol on dEMG amplitude compared with untreated controls. These findings suggest that resveratrol also attenuates the nociceptive JOR via 5HT3 receptor-mediated GABAergic inhibition. The present study therefore provides new insight into a possible mechanism underlying resveratrol-induced trigeminal antinociception via the descending pain control system and highlights a potential therapeutic agent for complementary alternative medicine.

Dietary constituent genistein inhibits the hyperexcitability of trigeminal nociceptive neurons associated with mechanical hyperalgesia following orofacial inflammation.

OBJECTIVES: Genistein, a dietary constituent, modulates voltage-dependent and ligand-gated ionic channels, suggesting that it could also attenuate inflammatory hyperalgesia. However, the mechanism underlying how genistein affects inflammation-induced hyperexcitability of nociceptive neurons in vivo remains to be determined. The present study therefore investigated whether administration of genistein could attenuate the inflammation-induced hyperexcitability of trigeminal spinal nucleus caudalis (SpVc) neurons associated with mechanical hyperalgesia in vivo. METHODS: Inflammation was induced by injection of complete Freund's adjuvant into the whisker pad. The mechanical thresholds for escape behavior and electrophysiological single-unit recording of SpVc neurons responding to mechanical stimulation were then conducted in naïve rats, inflamed rats, and inflamed rats with genistein administered intraperitoneally. RESULTS: The lowered mechanical threshold in the inflamed rats was returned to control level following administration of genistein for 2 days. The mean number of discharge frequencies of SpVc neurons in inflamed rats was significantly decreased after genistein administration with both non-noxious and noxious mechanical stimuli. The increased spontaneous discharges of SpVc neurons in inflamed rats were significantly decreased after genistein administration. Noxious pinch-evoked after-discharge frequency and occurrence in inflamed rats was also significantly diminished after genistein administration, and expansion of the receptive field was significantly returned to control levels in inflamed rats. CONCLUSION: Herein, we present the first evidence that genistein attenuates hyperexcitability of SpVc neurons associated with inflammatory mechanical hyperalgesia. These findings suggest that genistein could be a potential therapeutic agent in complementary alternative medicine for the prevention of trigeminal inflammatory hyperalgesia.

Effect of lutein on the acute inflammation-induced c-Fos expression of rat trigeminal spinal nucleus caudalis and C1 dorsal horn neurons.

Although lutein is known to inhibit chronic inflammation, its effect on acute inflammation-induced nociceptive processing in the trigeminal system remains to be determined. The aim of the present study was to investigate whether pretreatment with lutein attenuates acute inflammation-induced sensitization of nociceptive processing in rat spinal trigeminal nucleus caudalis (SpVc) and upper cervical (C1) dorsal horn neurons, via c-Fos immunoreactivity. Mustard oil, a transient receptor potential ankyrin-1 channel agonist, was injected into the whisker pads to induce inflammation. Pretreatment of rats with lutein resulted in significant decreases in the inflammation-induced mean times of face grooming and the thickness of inflammation-induced edema in whisker pads relative to those features in inflamed rats (i.e., rats with no lutein pretreatment). In both the ipsilateral superficial and deep laminae of the SpVc and C1 dorsal horn, there were significantly larger numbers of c-Fos-positive neurons in inflamed rats than in naïve rats, and lutein pretreatment significantly decreased that number relative to inflamed rats. These results suggest that systemic administration of lutein attenuates acute inflammation-induced nocifensive behavior and augmented nociceptive processing of SpVc and C1 neurons that send stimulus localization and intensity information to higher pain centers. These findings support lutein as a potential therapeutic agent for use as an alternative, complementary medicine to attenuate, or even prevent, acute inflammatory pain.

Decanoic acid attenuates the excitability of nociceptive trigeminal primary and secondary neurons associated with hypoalgesia.

BACKGROUND: Acute application of decanoic acid (DA) in vivo suppresses the excitability of spinal trigeminal nucleus caudalis (SpVc) wide dynamic range (WDR) neurons associated with the short-term mechanical hypoalgesia via muscarinic M2 receptor signaling; however, the effect of DA on nociceptive trigeminal ganglion (TG) and SpVc nociceptive-specific (NS) neuronal excitability under in vivo conditions remains to be determined. The present study investigated whether this effect could be observed in naive rats. RESULTS: Extracellular single-unit recordings were made from TG and SpVc NS neurons of pentobarbital-anesthetized rats in response to orofacial noxious mechanical stimuli. DA inhibited the mean firing frequency of both TG and SpVc NS neurons, reaching a maximum inhibition of discharge frequency within 1-5 minutes and reversing after approximately 10-minutes; however, this DA-induced suppression of SpVc NS neuronal firing frequency did not occur in rats administered with methoctramine intravenously prior to stimulation. CONCLUSION: This in vivo study indicated that firing of TG and SpVc NS neurons induced by mechanical hypoalgesia through peripheral M2 receptors could be inhibited by acutely administered DA, implicating the potential of DA in the future treatment of trigeminal pain. PERSPECTIVE: This article presents that the acute DA application suppresses the excitability of TG and SpVc NS neurons associated with mechanical hypoalgesia via peripheral M2 receptor signaling, supporting DA as a potential therapeutic agent in complementary and alternative medicine for the attenuation of nociception.

Suppression of hyperexcitability of trigeminal nociceptive neurons associated with inflammatory hyperalgesia following systemic administration of lutein via inhibition of cyclooxygenase-2 cascade signaling.

INTRODUCTION: Lutein is a dietary constituent known to inhibit inflammation; however, its effect on nociceptive neuron-associated hyperalgesia remains to be determined. The present study therefore investigated under in vivo conditions whether administration of lutein attenuates the inflammation-induced hyperexcitability of trigeminal spinal nucleus caudalis (SpVc) neurons that is associated with mechanical hyperalgesia. RESULTS: Complete Freund's adjuvant (CFA) was injected into the whisker pads of rats to induce inflammation, and then mechanical stimulation was applied to the orofacial area to assess the threshold of escape. The mechanical threshold was significantly lower in inflamed rats compared to uninjected naïve rats, and this lowered threshold was returned to control levels by 3 days after administration of lutein (10 mg/Kg, i.p.) Also the lutein administration, inflammation-induced thickness of edema was returned to control levels. The mean increased number of cyclooxygenase-2 (Cox-2)-immunoreactive cells in the whisker pads of inflamed rats was also returned to control levels by administration with lutein. The mean discharge frequency of SpVc wide-dynamic range (WDR) neurons to both nonnoxious and noxious mechanical stimuli in inflamed rats was significantly decreased after lutein administration. In addition, the increased mean spontaneous discharge of SpVc WDR in inflamed rats was significantly decreased after lutein administration. Similarly, lutein significantly diminished noxious pinch-evoked mean after discharge frequency and occurrence in inflamed rats. Finally, lutein restored the expanded mean size of the receptive field in inflamed rats to control levels. CONCLUSION: These results together suggest that administration of lutein attenuates inflammatory hyperalgesia associated with hyperexcitability of nociceptive SpVc WDR neurons via inhibition of the peripheral Cox-2 signaling cascade. These findings support the proposed potential of lutein as a therapeutic agent in complementary alternative medicine strategies for preventing inflammatory mechanical hyperalgesia.

Docosahexaenoic acid attenuates inflammation-induced hyperexcitability of trigeminal spinal nucleus caudalis neurons associated with hyperalgesia in rats.

The present study investigated whether daily systemic administration of docosahexaenoic acid (DHA) in rats could attenuate the hyperexcitability of trigeminal spinal nucleus caudalis (SpVc) neurons associated with hyperalgesia. Inflammation was induced in rats by injecting complete Freund's adjuvant into the whisker pads. The threshold of escape from mechanical stimulation applied to the whisker pads in inflamed rats was significantly lower than that in naïve rats. The lowered mechanical threshold in the inflamed rats was returned to that in naïve rats by 3 d intraperitoneal administration of DHA. The mean discharge frequency of SpVc neurons in inflamed rats was significantly decreased after DHA administration for 3 d with both non-noxious and noxious mechanical stimuli. DHA administration also significantly decreased the increased spontaneous discharges of SpVc neurons in the inflamed rats, while DHA significantly diminished noxious pinch evoked after the discharge frequency and the expanded receptive field in the inflamed rats was returned to control levels. These results suggested that chronic administration of DHA attenuates inflammation-induced mechanical hyperalgesia associated with the suppression of the hyperexcitability of SpVc neurons. These findings support the potential use of DHA as a therapeutic agent in complementary alternative medicine for mitigating trigeminal inflammatory hyperalgesia.

Local anesthetic effect of docosahexaenoic acid on the nociceptive jaw-opening reflex in rats.

Although docosahexaenoic acid (DHA) administration suppresses sodium channels in primary afferent sensory neurons, the acute local effect of DHA on the trigeminal nociceptive reflex remains to be elucidated, in vivo. Therefore, the aim of the present study was to investigate whether local administration of DHA attenuates the nociceptive jaw-opening reflex (JOR) in vivo in the rat. The JOR evoked by electrical stimulation of the tongue was recorded by a digastric muscle electromyogram (dEMG) in pentobarbital-anesthetized rats. The amplitude of the dEMG response was significantly increased in proportion to the electrical stimulation intensity (1-5 x threshold). At 3 x threshold, local administration of DHA (0.1, 10 and 25 mM) dose-dependently inhibited the dEMG response, and lasted 40 min. Maximum inhibition of the dEMG signal amplitude was seen within approximately 10 min. The mean magnitude of inhibition of the dEMG signal amplitude by DHA (25 mM) was almost equal to the local anesthetic, 1% lidocaine (37 mM), a sodium channel blocker. These findings suggest that DHA attenuates the nociceptive JOR via possibly blocking sodium channels, and strongly support the idea that DHA is a potential therapeutic agent and complementary alternative medicine for the prevention of acute trigeminal nociception.

Chemokine ligand 2/chemokine receptor 2 signaling in the trigeminal ganglia contributes to inflammatory hyperalgesia in rats.

This study investigated the functional significance of hyperalgesia in the CCL2/CCR2 signaling system in trigeminal ganglion (TG) neurons following inflammation. Inflammation was induced by injection of complete Freund's adjuvant (CFA) into the whisker pad of rats. The escape threshold from mechanical stimulation applied to the whisker pad 2days later was significantly lower in CFA-treated rats than in naïve rats. Fluorogold (FG) labeling was used to identify the TG neurons innervating the whisker pad. FG-labeled TG neurons were immunoreactive for CCL2/CCR2. The mean number of CCL2/CCR2-immunoreactive small/medium-diameter TG neurons was significantly higher in inflamed rats than in naïve rats. Using whole-cell patch-clamp experiments in small-diameter TG neurons, the threshold current of FG-labeled TG neurons in inflamed rats was significantly decreased compared to naïve rats. The number of spike discharges during current injections by FG-labeled TG neurons in inflamed rats was significantly increased compared to naïve rats. These characteristic effects were abolished by co-application of a CCL2 receptor antagonist. The present study provides evidence that CCL2 enhances the excitability of small-diameter TG neurons following facial skin inflammation via the upregulation of CCR2. These findings suggest that ganglionic CCL2/CCR2 signaling is a therapeutic target for the treatment of trigeminal inflammatory hyperalgesia.

Local subcutaneous injection of chlorogenic acid inhibits the nociceptive trigeminal spinal nucleus caudalis neurons in rats.

Acute administration of chlorogenic acid (CGA) in vitro was recently shown to modulate potassium channel conductance and acid-sensing ion channels (ASICs) in the primary sensory neurons; however, in vivo peripheral effects of CGA on the nociceptive mechanical stimulation of trigeminal neuronal activity remains to be determined. The present study investigated whether local administration of CGA in vivo attenuates mechanical stimulation-induced excitability of trigeminal spinal nucleus caudalis neuronal (SpVc) activity in rats. Extracellular single-unit recordings were made of SpVc wide-dynamic range (WDR) neuronal activity elicited by non-noxious and noxious orofacial mechanical stimulation in pentobarbital anesthetized rats. The mean number of SpVc WDR neuronal firings responding to both non-noxious and noxious mechanical stimuli were significantly and dose-dependently inhibited by local subcutaneous administration of CGA (0.1-10mM), with the maximal inhibition of discharge frequency revealed within 10min and reversed after approximately 30min. The mean frequency of SpVc neuronal discharge inhibition by CGA was comparable to that by a local anesthetic, the sodium channel blocker, 1% lidocaine. These results suggest that local CGA injection into the peripheral receptive field suppresses the excitability of SpVc neurons, possibly via the activation of voltage-gated potassium channels and modulation of ASICs in the nociceptive nerve terminal of trigeminal ganglion neurons. Therefore, local injection of CGA could contribute to local anesthetic agents for the treatment of trigeminal nociceptive pain.

Effect of resveratrol on c-fos expression of rat trigeminal spinal nucleus caudalis and C1 dorsal horn neurons following mustard oil-induced acute inflammation.

The dietary constituent, resveratrol, was recently identified as a transient receptor potential ankyrin 1 (TRPA1) antagonist, voltage-dependent sodium ion (Na+ ) channel, and cyclooxygenase-2 (COX-2) inhibitor. The aim of the present study was to investigate whether pretreatment with resveratrol attenuates acute inflammation-induced sensitization of nociceptive processing in rat spinal trigeminal nucleus caudalis (SpVc) and upper cervical (C1) dorsal horn neurons, via c-fos immunoreactivity. Mustard oil (MO), a TRPA1 channel agonist, was injected into the whisker pads of rats to induce inflammation. Pretreatment with resveratrol significantly decreased the mean thickness of inflammation-induced edema in whisker pads compared with those of untreated, inflamed rats. Ipsilateral of both the superficial and deep laminae of SpVc and C1 dorsal horn, there were significantly more c-fos-immunoreactive SpVc/C1 neurons in inflamed rats compared with naïve rats, and resveratrol pretreatment significantly decreased that number relative to untreated, inflamed rats. These results suggest that systemic administration of resveratrol attenuates acute inflammation-induced augmented nociceptive processing of trigeminal SpVc and C1 neurons. These findings support resveratrol as a potential therapeutic agent for use in alternative, complementary medicine to attenuate, or even prevent, acute trigeminal inflammatory pain.

Dietary constituent, decanoic acid suppresses the excitability of nociceptive trigeminal neuronal activity associated with hypoalgesia via muscarinic M2 receptor signaling.

Background: Although decanoic acid (DA) is thought to act as a muscarinic cholinergic agonist, effect of DA on nociceptive behavioral responses and the excitability of nociceptive neuronal activity under in vivo conditions remain to be determined. The aim of the present study, therefore, was to investigate whether in vivo acute administration of ointment containing DA affects the excitability of nociceptive trigeminal spinal nucleus caudalis (SpVc) neurons associated with hypoalgesia in naïve rats. Results: After local application of DA, the threshold of escape from mechanical stimulation applied to the shaved orofacial skin was significantly higher than before DA application. Vehicle treatment (without DA) had no significant effect on the escape threshold from mechanical stimulation. Extracellular single unit recordings were made from SpVc wide-dynamic range (WDR) neurons in response to orofacial non-noxious and noxious mechanical stimuli of pentobarbital-anesthetized rats. The mean firing frequency of SpVc WDR neurons in response to noxious, but not non-noxious, mechanical stimuli was inhibited by local application of DA, and the maximum inhibition of discharge frequency of both non-noxious and noxious mechanical stimuli was seen within 1­5 min. The DA-induced short-term inhibitory effects were reversed after approximately 10 min. Pretreatment intravenously with the muscarinic-specific M2 receptor antagonist, methoctramine, abolished the DA-induced suppression of firing frequency of SpVc WDR neurons in response to noxious stimulation. Fluorogold (FG) labeling was identified as the trigeminal ganglion (TG) neurons innervating orofacial skin. FG-labeled small-diameter TG neurons expressed M2 receptor immunoreactivity. Conclusion: These results suggest that acute DA application induces short-term mechanical hypoalgesia and this effect was mainly due to suppression of the excitability of SpVc WDR neurons via the peripheral M2 receptor signaling pathway in the trigeminal primary afferents. These findings support the idea that DA is a potential therapeutic agent and complementary alternative medicine for the attenuation of trigeminal nociception in the absence of inflammatory/neuropathic conditions.

The dietary constituent resveratrol suppresses nociceptive neurotransmission via the NMDA receptor.

Background Although we have previously reported that intravenous resveratrol administration inhibits the nociceptive neuronal activity of spinal trigeminal nucleus caudalis neurons, the site of the central effect remains unclear. The aim of the present study was to examine whether acute intravenous resveratrol administration in the rat attenuates central glutamatergic transmission of spinal trigeminal nucleus caudalis neurons responding to nociceptive mechanical stimulation in vivo, using extracellular single-unit recordings and microiontophoretic techniques. Results Extracellular single-unit recordings using multibarrel electrodes were made from the spinal trigeminal nucleus caudalis wide dynamic range neurons responding to orofacial mechanical stimulation in pentobarbital anesthetized rats. These neurons also responded to iontophoretic application of glutamate, and the evoked neuronal discharge frequency was significantly increased in a current-dependent and reversible manner. The mean firing frequency evoked by the iontophoretic application of glutamate (30, 50, and 70 nA) was mimicked by the application of 10 g, 60 g, and noxious pinch mechanical stimulation, respectively. The mean firing frequency of spinal trigeminal nucleus caudalis wide dynamic range neurons responding to iontophoretic application of glutamate and N-methyl-D-aspartate were also significantly inhibited by intravenous administration of resveratrol (2 mg/kg) and the maximal inhibition of discharge frequency was observed within 10 min. These inhibitory effects lasted approximately 20 min. The relative magnitude of inhibition by resveratrol of the glutamate-evoked spinal trigeminal nucleus caudalis wide dynamic range neuronal discharge frequency was similar to that for N-methyl-D-aspartate iontophoretic application. Conclusion These results suggest that resveratrol suppresses glutamatergic neurotransmission of the spinal trigeminal nucleus caudalis neurons responding to nociceptive mechanical stimulation via the N-methyl-D-aspartate receptor in vivo, and resveratrol may be useful as a complementary or alternative therapeutic agent for the treatment of trigeminal nociceptive pain.

Acute intravenous administration of dietary constituent theanine suppresses noxious neuronal transmission of trigeminal spinal nucleus caudalis in rats.

Theanine is a non-dietary amino acid linked to the modulation of synaptic transmission in the central nervous system, although the acute effects of theanine in vivo, particularly on nociceptive transmission in the trigeminal system, remain to be determined. The present study investigated whether acute intravenous theanine administration to rats attenuates the excitability of wide dynamic range (WDR) spinal trigeminal nucleus caudalis (SpVc) neurons in response to nociceptive and non-nociceptive mechanical stimulation in vivo. Extracellular single unit recordings were made from 15 SpVc neurons in response to orofacial mechanical stimulation of pentobarbital-anesthetized rats, and responses to non-noxious and noxious mechanical stimuli were analyzed. The mean firing frequency of SpVc WDR neurons in response to all mechanical stimuli was dose-dependently inhibited by theanine (10, 50, and 100mM, i.v.) with the maximum inhibition of discharge frequency reached within 5min. These inhibitory effects were reversed after approximately 10min. The relative magnitude of theanine's inhibition of SpVc WDR neuronal discharge frequency was significantly greater for noxious than non-noxious stimulation. Iontophoretic application of l-glutamate induced the mean firing frequency of SpVc WDR neuron responding to noxious mechanical stimulation was also inhibited by intravenous administration of 100mM theanine. These results suggest that acute intravenous theanine administration suppresses glutaminergic noxious synaptic transmission in the SpVc, implicating theanine as a potential complementary and alternative therapeutic agent for the treatment of trigeminal nociceptive pain.