Effects of administration of α2 adrenergic receptor agonist into psoas major muscle on inflammatory pain induced by injection of complete Freund's adjuvant in rats.

α2 adrenergic agonists are widely used in clinical anesthesia and ICU sedation owing to their effective sedative and analgesic effects. Lumbago and leg pain is the most common clinical pain disease. Studies have reported that lumbago and leg pain is associated with dysregulation of paravertebral muscles, especially psoas major muscles. In the present study, a unilateral lower extremity chronic inflammation and pain model was established by subcutaneous administration of low-dose complete Fredrin's adjuvant (CFA) into the posterior paw of rats. α2 adrenalin receptor agonist was then injected into the psoas major muscle. Behavioral tests were conducted for 21 days. Psoas major muscle tissue was harvested for evaluation of biochemical indexes related to pain. The effect of α2 adrenergic receptor agonist injected into psoas major muscle on chronic inflammatory pain of lower extremities in rats was explored. The results showed that injection of α2 adrenergic receptor agonist into the psoas major muscle relieved CFA-induced mechanical hyperalgesia. Administration of α2 adrenergic receptor antagonist yohimbine reversed the analgesic effect of α2 adrenergic receptor agonists. Administration of dexmedetomidine into psoas major muscle downregulated the levels of norepinephrine, interleukin-6 and tumor necrosis factor-α in tissues. The findings of the present study show that administration of α2 adrenoceptor agonists into the psoas major muscle relieves chronic inflammatory pain induced by CFA. Local injection of dexmedetomidine also exerted anti-inflammatory and anti-sympathetic effect by activating α2-adrenoceptor in the psoas major muscle.

The Protective Effects of Hydrogen Sulfide New Donor Methyl S-(4-Fluorobenzyl)-N-(3,4,5-Trimethoxybenzoyl)-l-Cysteinate on the Ischemic Stroke.

In this paper, we report the design, synthesis and biological evaluation of a novel S-allyl-l-cysteine (SAC) and gallic acid conjugate S-(4-fluorobenzyl)-N-(3,4,5-trimethoxybenzoyl)-l-cysteinate (MTC). We evaluate the effects on ischemia-reperfusion-induced PC12 cells, primary neurons in neonatal rats, and cerebral ischemic neuronal damage in rats, and the results showed that MTC increased SOD, CAT, GPx activity and decreased LDH release. PI3K and p-AKT protein levels were significantly increased by activating PI3K/AKT pathway. Mitochondrial pro-apoptotic proteins Bax and Bim levels were reduced while anti-apoptotic protein Bcl-2 levels were increased. The levels of cleaved caspase-9 and cleaved caspase-3 were also reduced in the plasma. The endoplasmic reticulum stress (ERS) was decreased, which in turns the survival rate of nerve cells was increased, so that the ischemic injury of neurons was protected accordingly. MTC activated the MEK-ERK signaling pathway and promoted axonal regeneration in primary neurons of the neonatal rat. The pretreatment of MEK-ERK pathway inhibitor PD98059 and PI3K/AKT pathway inhibitor LY294002 partially attenuated the protective effect of MTC. Using a MCAO rat model indicated that MTC could reduce cerebral ischemia-reperfusion injury and decrease the expression of proinflammatory factors. The neuroprotective effect of MTC may be due to inhibition of the over-activation of the TREK-1 channel and reduction of the current density of the TREK1 channel. These results suggested that MTC has a protective effect on neuronal injury induced by ischemia reperfusion, so it may have the potential to become a new type of neuro-ischemic drug candidate.

Effect of reactive oxygen species of the psoas major muscle in complete Freund's adjuvant-induced inflammatory pain in rats.

Lower limb pain is a common clinical disease that affects millions of people worldwide. It is found in previous studies that reactive oxygen species is closely related to neuropathic, cancer, chemotherapy, and inflammatory pain, which can be relieved by reactive oxygen species scavengers. Furthermore, acupuncture or electroacupuncture on the psoas major muscle has a great effect on adjuvant-induced arthritis and lower back pain. In our study, we investigated the function of reactive oxygen species scavengers locally injecting into the ipsilateral psoas major muscle on complete Freund's adjuvant-induced inflammatory pain. Our results demonstrated that in the development of complete Freund's adjuvant-induced inflammatory pain, early local continuous application of N-tert-Butyl-α-phenylnitrone (PBN, 1 and 5 mg/kg/0.2 ml) on the ipsilateral psoas major muscle effectively reduced mechanical and cold hyperalgesia. However, intraperitoneal injection of PBN (1 and 5 mg/kg) or local injection of PBN (1 and 5 mg/kg/0.2 ml) into contralateral psoas major muscle, ipsilateral quadratus lumborum, and ipsilateral erector spinae showed limited effect. In the developed inflammatory pain model, local injection of PBN into the ipsilateral psoas major muscle also alleviated pain and paw edema. In addition, reactive oxygen species level increased in ipsilateral psoas major muscle at seven days after complete Freund's adjuvant injection. In general, PBN reduces complete Freund's adjuvant-evoked inflammatory pain by inhibiting reactive oxygen species in the psoas major muscle.

The protective effects of hydrogen sulfide on the myocardial ischemia via regulating Bmal1.

BACKGROUND: To investigate the effect of hydrogen peroxide (H2S) on myocardial clock gene Bmal1 in ischemic cardiomyocytes. MATERIALS & METHODS: Quantitative PCR (qPCR) was used to detect the expression of Bmal1 at the mRNA level in H9C2 rat cardiomyocytes. The protein expressions of Bax and Bcl-2, PI3K/Akt, caspase-3 were measured by western blotting. The levels of reactive oxygen species (ROS) were determined by ELISA. RESULTS: The expression level of clock gene Bmal1 demonstrated a clock rhythm of periodic oscillation within 24 h. Compared with the control group, H2S treatment maintained the rhythm of the clock gene in ischemic cardiomyocytes and increased the transcription and expression levels of Bmal1. H2S increased cell survival by activating PI3K/Akt signaling pathway, inhibiting mitochondrial apoptosis signaling, and reducing intracellular oxidative stress. PI3K/Akt and Bmal1 were demonstrated to be involved in H2S protection of cardiomyocyte ischemia. Knockout of Bmal1 gene affects the degree of phosphorylation of Akt and Erk proteins, and the level of ROS production, resulting in a decrease in the protective effects of H2S. CONCLUSION: The expression level of Bmal1 has effects on the function of cardiomyocytes such as ROS production. The potential mechanism by which H2S regulates clock genes may be related to the effect of clock genes on protein phosphorylation levels in ischemic cardiomyocytes.

Recyclable fluorescent chemodosimeters based on 8-hydroxyquinoline derivatives for highly sensitive and selective detection of mercury(II) in aqueous media and test strips.

Four novel highly selective 8-hydroxyquinoline-based fluorescent chemodosimeters (1-4) were synthesized for the rapid analysis of Hg2+ in aqueous solution and on paper strips, which probably attributed to the excited state intramolecular proton transfer (ESIPT) process. Chemodosimeter 1 was evaluated as a Hg2+-ratiometric fluorescent sensor while others (2, 3 and 4) displayed fluorescence turn-on response for Hg2+ among the various survey metal ions. We demonstrated that chemodosimeters (1-4) could recognized Hg2+ ions based on a 1:1 stoichiometric binding event with fast detection time. More importantly, the detection limits for Hg2+ could reach at 10-9 M level except chemodosimeter 1 (4.05 × 10-8 M). In addition, it was found that chemodosimeters (1-4) were recycled efficiently because the Hg2+ induced emission spectra were reversed after adding NaBH4. Finally, these four sensors were successfully applied for fabrication of simple device test strips for rapid and on-site detection of Hg2+ ions.

Role of ERK1/2 activation on itch sensation induced by bradykinin B1 activation in inflamed skin.

It has previously been demonstrated that bradykinin receptor B1 (B1R) agonists evoke an itch-related scratching response in inflamed skin via the B1 receptor; however, the mechanisms responsible for this abnormal itch sensation remain unclear. Therefore, the present study utilized a complete Freund's adjuvant (CFA)-induced mouse model of inflammation to elucidate the mechanisms responsible. Over a period of 30 min, scratching behavior was quantified by the number of hind limb scratches of the area surrounding the drug injection site on the neck. Furthermore, western blot analysis was used to investigate the potential role of extracellular signal-regulated kinase (ERK) 1/2 signaling as a mediator of itch in CFA-treated mice. The results demonstrated that CFA-induced inflammation at the back of the neck is associated with sustained enhancement of ERK1/2 activation in the spinal cord. Moreover, B1R agonist treatment resulted in increased expression of phosphorylated ERK1/2 in the spinal cord, which peaked at 45 min. Consistent with these findings, inhibition of either mitogen-activated protein/ERK kinase or ERK1/2, as well as inhibition of ERK1/2 activation following inflammation, attenuated B1 receptor-mediated scratching responses to a greater extent, as compared with control mice. Collectively, the results of the present study indicated that enhanced and persistent ERK1/2 activation in the spinal cord may be required to induce a scratching response to B1R agonists following CFA-induced inflammation.

The effect of kinin B1 receptor on chronic itching sensitization.

BACKGROUND: Altered kallikrein-related peptidase activity and bradykinin are associated with skin disorders in humans and mice under chronic inflammation conditions. The bradykinin B1 receptor (B1R), also known as one of the G-protein-coupled receptor family and usually absent in intact tissues and upregulated during tissue injury, is responsible for vasodilation, capillary permeability, nociceptor sensitization, and pain; it is indispensable for physiopathological progress in chronic inflammation conditions, but its roles and effectors in the itching sensation of the allergic contact dermatitis model are poorly defined. RESULTS: We focused on incurable itching in a diphenylcyclopropenone (DCP) chronic inflammation experimental model. Preventive treatment with the B1R antagonist R892 significantly suppressed spontaneous scratching, while the B2R selective antagonist did not. B1R expression in the skin tissues of this model was detected using a quantitative, real-time polymerase chain reaction, Western blotting, and immunohistochemistry; B1R mRNA and protein levels were increased compared with a sham-treated control group. A higher B1R IHC staining signal was observed in the keratinocytes in DCP-treated mice compared with a vehicle-treated group, so we studied the B1R function when superimposed on a protease-activated receptor 2 (PAR2) background, establishing B1R as a pivotal mediator of PAR2 function in HaCaT cell lines. CONCLUSION: Our data provide evidence that B1R facilitates the chronic itching sensation related to keratinocytes in a DCP-treated chronic inflammation experimental model.

The kinin B1 receptor mediates alloknesis in a murine model of inflammation.

Noxious stimuli and non-noxious mechanical stimuli elicit itch (alloknesis) instead of pain on skin lesions of patients with atopic dermatitis. We previously found that bradykinin evokes an itch-related scratching response through activation of kinin B1 receptor in skin inflamed using complete Freund's adjuvant. In this study we investigated whether alloknesis is evoked in CFA-inflamed skin and the involvement of kinin receptors. In our results, alloknesis was elicited four days after CFA-inflammation. Furthermore, pretreatment with a B1 receptor antagonist or µ-opioid receptor antagonist significantly reduced alloknesis. In contrast, treatment with a B2 receptor antagonist significantly increased alloknesis. These results suggest that the alloknesis response is mediated by the activation of kinin B1 receptor but antagonized by the B2 receptor in CFA-inflamed mice.

Paradoxic effects of propofol on visceral pain induced by various TRPV1 agonists.

Intraperitoneal injection of propofol inhibits subsequent acetic acid-induced writhing response in mice. Propofol increases the sensitivity of dorsal root ganglion neurons to capsaicin through transient receptor potential ankyrin subtype-1 (TRPA1) and protein kinase Cε (PKCε)-mediated phosphorylation of transient receptor potential vanilloid subtype-1 (TRPV1). Intraperitoneal co-injection of propofol may increase visceral nociception induced by TRPV1 agonists via sensitization of TRPV1. Therefore, we investigated the effects of intraperitoneal co-injection of propofol on nociception induced by acetic acid and capsaicin. The number of writhing movements induced by acetic acid or nociception time by capsaicin with or without propofol were counted. Neonatal capsaicin-treated mice were also used to demonstrate the role of TRPV1 in the effects of propofol on nociception, induced by TRPV1 agonists. Co-injection of propofol resulted in a pronociceptive effect on the writhing response induced by acetic acid, while the same dose of propofol ameliorated the response to capsaicin. The writhing response to intraperitoneal acetic acid was sharply inhibited following neonatal treatment with capsaicin. Co-injection with propofol reduced the number of writhing movements induced by acetic acid in neonatal capsaicin-treated mice. These results suggest that propofol binds to TRPV1 at the capsaicin-binding pocket.

Endothelin B receptors exert antipruritic effects via peripheral κ-opioid receptors.

Endothelin B receptor agonists exert antipruritic effects on itching induced via endothelin-1 (ET-1) and compound 48/80. Peripheral µ- and κ-opioid receptors (MORs and KORs, respectively) are reported to be involved in the anti-nociceptive properties triggered by ET(B) agonists. Therefore, we investigated the role of peripheral opioid receptors in the scratching response induced by ET-1. ET(A) and ET(B) antagonists and non-selective and selective opioid receptor antagonists were co-injected with ET-1 in the neck of mice and the number of scratching bouts was counted. Pretreatment with systemically administered naloxone significantly reduced the number of scratches, while co-injection of naloxone substantially augmented the effect of ET-1. Co-injection of nor-Binaltorphimine (nor-BNI), a KOR antagonist, significantly increased the number of scratches induced by ET-1. However, CTOP (a MOR antagonist) and naltrindole [a δ-opioid receptor (DOR) antagonist] did not alter the scratching response elicited by ET-1. These results indicate that peripheral KORs mediate the antipruritic effect of endothelin B receptor activation.

Bradykinin-evoked scratching responses in complete Freund's adjuvant-inflamed skin through activation of B1 receptor.

Capsaicin, a potent algogen, induces an itch-related behavior in the presence of inflammation. In this study, we tested whether bradykinin (BK) can evoke a similar response and investigated the potential mechanisms involved in this process. Local inflammation was induced by intradermal injection of complete Freund's adjuvant (CFA) into the back of the neck, left hind foot or left cheek of male C57BL/6J mice. BK was then injected intradermally into the same area on indicated days. Four days after CFA inflammation, BK treatment evoked scratching responses in a time- and dose-dependent manner. For BK receptor antagonist treatment, inflamed-mice were either given an intraperitoneal injection of B(1) receptor (B(1)R) or B(2) receptor (B(2)R) antagonist 30 min prior to BK administration, or an intradermal co-injection of antagonist and BK into the inflamed area. Our results indicate that B(1)R and B(2)R act in an opposite fashion during this process, as pretreatment with B(1)R antagonist by intraperitoneal injection significantly reduced BK-induced scratching behavior, whereas B(2)R antagonist treatment dramatically increased scratching behavior. Moreover, combined injection of BK and B(2)R antagonist enhanced BK-induced scratching activity in CFA-inflamed mice. In addition, pretreatment or co-injection with B(2)R antagonist dramatically reduced the pain-related licking behavior induced by BK injection. The data suggest that BK-induced scratching responses in CFA-inflamed mouse skin occur via activation of B(1)R. Furthermore, B(1) and B(2) receptors play different roles in modulating BK-induced itch-related behavior in CFA-inflamed mice.

Effect of the cation-chloride cotransporter inhibitor furosemide in a rat model of postoperative pain.

OBJECTIVE: Recently, evidence has accumulated to show that cation-chloride cotransporters (CCCs) participate in the modulation of pain transmission at the spinal cord level. To investigate whether CCC inhibitors might affect surgical pain, we examined the effect of furosemide in a rat incisional pain model. DESIGN: We examined pain thresholds using von Frey filaments in intact and incision model Sprague-Dawley rats before and after intrathecal furosemide administration (100 µg/20 µL). Twenty-four rats were divided into four groups (N=6), groups A and B were intact rats, which received furosemide and its solvent, respectively. Groups C and D were incision rats, which received furosemide and its solvent, respectively. Non-parametric tests were used to calculate pain thresholds, and P<0.05 was considered significant. RESULTS: Furosemide decreased the pain threshold (vs solvent) in intact rats for 2 hours after administration and caused excitatory behavior. However, furosemide increased the pain threshold in incision model rats ([1] at the incision point: at 20 minutes, 2 hours, 3 hours, 4 hours, and on the second to fifth days after incision; [2] at the remote point: at 20 minutes, 3-5 hours, and from the 3rd to the 7th day after incision) and improved wound recovery. CONCLUSIONS: Intrathecal administration of the CCC inhibitor furosemide had antinociceptive effects in rats with incisional pain. Furosemide may be a novel treatment for postoperative pain.

Capsaicin induces reflex scratching in inflamed skin.

We investigated whether capsaicin induces itching in skin with existing inflammation. We induced skin inflammation by intradermal injection of complete Freund's adjuvant (CFA) in the neck of mice. Four days later, we injected capsaicin in the same area and counted the number of scratching bouts for 30 min. We examined potential effects on pain in parallel experiments in which CFA and capsaicin were intradermally injected into hind paws. We used the time spent licking the hind paws during the 15 min after capsaicin injection as an estimate of pain. Capsaicin injection into the skin pretreated with CFA, but not into healthy skin, induced scratching. The scratching behavior was reduced by pretreatment with naloxone or capsazepine, selective antagonists for transient receptor potential vanilloid receptor-1 (TRPV1), but not morphine or mepyramine, selective antagonists for histamine 1 receptor. In animals injected with capsaicin into the hind paws, licking behavior was significantly inhibited via a µ-receptor-dependent mechanism. Our results show that TRPV1 activation, which normally induces pain, evokes an itch-related response in the presence of inflammation. This model may be interesting for future studies to explore the mechanism of a painful stimuli-induced itch observed under pathological conditions.

Role of transient receptor potential ankyrin subfamily member 1 in pruritus induced by endothelin-1.

Noxious cold reduces pruritus and transient receptor potential ankyrin subfamily member 1 (TRPA1), a non-selective cation channel, is known as a noxious cold-activated ion channel. Recent findings implicated the involvement of TRPA1 in pain induced by endothelin-1 (ET-1). Therefore, we evaluated its potential role in pruritus induced by ET-1. We found that ruthenium red (RR; a nonselective TRP inhibitor) and AP18 (a TRPA1 antagonist) significantly increased scratching bouts caused by ET-1, while capsazepine (a TRPV1 antagonist) and morphine showed no effects in the ET-1-induced scratching response. However, RR and capsazepine significantly reduced scratching bouts caused by histamine. Our results suggested that activation of TRPA1 could suppress itch induced by ET-1 and this is not related to pain induced by ET-1.

Molecular signaling of pruritus induced by endothelin-1 in mice.

Endothelin-1 (ET-1) has recently been identified to evoke pruritus/itching sensation in both humans and animals. It is most likely that the signaling is through the specific G-protein-coupled ET(A) and ET(B) receptors, but the downstream signaling mediators for ET-1 remain elusive. In the present study, we examined the potential involvement of several distinct signaling molecules in ET-1-induced pruritus in a murine model. We applied an in vivo pruritus model in C57BL/6J mice by injecting ET-1 intradermally into the scruff, and recording the number of scratching bouts within 30 min after injection. Then specific antagonists/inhibitors for distinct signaling molecules, including cell-surface ET(A) and ET(B) receptors, histamine receptor type 1 (H1 receptor), protein kinases A (PKA) and C (PKC), phospholipase C (PLC) or adenylyl cyclase (AC), were co-injected with ET-1. The results showed that ET-1 induced a vigorous scratching response in mice in a dose-dependent manner. This response was further enhanced by a specific antagonist for ET(B) receptor, BQ-788, reduced by a specific antagonist for ET(A) receptor, BQ-123, and not affected by mepyramine, the specific inhibitor for H1 receptor. In addition, the scratching response was significantly reduced by inhibitors for PKC and AC, but was significantly enhanced by PLC inhibitor, while PKA inhibitors showed no effects in the ET-1-induced scratching response. Our data suggested that ET-1 may signal through the ET(A) receptor, AC and PKC pathway to induce pruritus sensation, while ET(B) receptor and PLC may antagonize the pruritus evoked by ET-1. These results may provide a basis for the future development of antipruritic therapy.

Involvement of TRPA1 in ET-1-induced pain-like behavior in mice.

Transient receptor potential ankyrin subfamily member 1 (TRPA1) is a nonselective cation channel known as a noxious cold-activated ion channel. Recent findings implicated its involvement in acute and chronic cold nociception processes. Here, we investigated whether TRPA1 is involved in endothelin-1 (ET-1)-induced spontaneous pain-like behavior in C57BL/6J mice. We found that TRPA1 antagonists, HC-030031 and AP18, significantly reduced the pain-like behavior caused by ET-1. AP18 also significantly reduced the pain caused by cinnamaldehyde, an agonist of TRPA-1. However, AP18 did not alleviate the pain caused by capsaicin. The pain-like behavior caused by ET-1 was inhibited by phospholipase C inhibitor, but not by protein kinase C inhibitor. Low dose of ET-1 could potentiate cinnamaldehyde-induced nociception. Our results suggested that TRPA1 is involved in ET-1-induced spontaneous pain-like behavior in mice.

Effects of static magnetic fields on Escherichia coli.

This paper represents the study of the biological effects of static magnetic fields (SMFs) on Escherichia coli (E. coli). The bacterial strain E. coli was exposed to SMFs in order to test its viability (evaluated by the number of colony-forming units (CFU)). In this study, we measured the dependence of CFU on the duration of exposure, on the treatment temperature T and on the value of the magnetic field induction B. The results showed that the number of CFU decreased with longer exposure time and higher treatment temperature (from 25 degrees C to 40 degrees C), whereas multiple extreme values of number of CFU were obtained when the induction B changed. In order to explain the results, transmission electron microscopy (TEM) and scanning electron microscopy (SEM) were used to investigate the morphology of the cells. We found obvious cell surface damage when the cells were exposed to SMFs.

Involvement of transient receptor potential vanilloid subfamily 1 in endothelin-1-induced pain-like behavior.

Although local administration of endothelin-1 (ET-1) is known to evoke spontaneous pain, the mechanism of ET-1-induced pain has not been elucidated. We investigated the involvement of protein kinase C (PKC) and transient receptor potential vanilloid subfamily 1 (TRPV1) in ET-1-induced pain-like behavior. Intraplantar ET-1 evoked pain-like behaviors, including licking, flinching, and biting, in a dose-dependent manner in wild-type mice. ET-1-induced pain-like behavior was attenuated by an endothelin type A receptor antagonist but not by PKC inhibitors and was also attenuated in TRPV1-deficient (KO) mice. In addition, we found a significant reduction of spinal Fos expression caused by the same dose of ET-1 in KO mice compared with that in wild-type mice. This study showed that endothelin type A receptor and TRPV1 are involved in ET-1-induced pain-like behaviors but failed to reveal the contribution of PKC.

[Role of vanilloid receptors in thermal hyperalgesia induced by intraplantar endothelin-1 administration].

OBJECTIVE: To assess the role of vanilloid receptor (VR) in thermal hyperalgesia induced by intraplantar endothelin-1 (ET-1) injection. METHODS: VR gene-knockout mice (KO group) and wild type C57BL/6J mice (WT group) in 3 subgroups were subjected to intraplantar administration of ET-1 at the doses of 3, 10, 30 and 100 pmol (dissolved in 10 microl of PBS, pH 7.4, n=6 in each group), respectively. The latency time of paw withdrawal (PWT) from heat irradiation stimulation was recorded before injection and 15, 30, 45 and 60 min after injection. RESULTS: ET-1 induced thermal hyperalgesia in both groups. The mice in WT group showed a more sharply shortened PWT than those in KO group. ET-1 decreased PWT as the dose administered increased in WT group, which was different from the responses of the KO mice. At the dose of 100 pmol of ET-1, no further decrement of latency time was observed in WT group, whereas such response occurred at 30 pmol in KO group. CONCLUSION: Intraplantar injection of ET-1 induces thermal hyperalgesia mediated partially by VR.