Formalin-induced Nociceptive Responses Research Articles

Ovariectomy induces hyperalgesia accompanied by upregulated estrogen receptor α and protein kinase B in the rat spinal cord

Hormone supplementation is one of the common therapies for menopause-related disorders. Among different tools, the ovariectomy (OVX) rodents are widely accepted as an appropriate menopausal pain model. Our previous study has showed that OVX produces a significant pain facilitation in both acute pain and tonic pain, however, the underlying mechanisms remain unclear. In this study, we examined the effects of OVX treatment and estradiol (E2) supplementation on formalin-induced nociceptive responses, and explored the associated spinal mechanisms. Female Sprague-Dawley rats underwent bilateral OVX, and E2 supplementation was given subcutaneously from the 5th week after surgery (30 μg/day for 7 days). Our results showed that formalin-induced nociceptive behaviors did not differ between diestrus and proestrus stages of the estrous in intact rats. However, OVX exacerbated formalin-evoked inflammatory pain, especially in the late phase at 4–5 weeks but not 2 weeks post-surgery. E2 supplementation significantly reversed the OVX-triggered hyperalgesia. Double immunofluorescence staining revealed that both ERα and ERβ in the spinal dorsal horn were co-labeled with the neuronal markers, but not with markers of astrocytes or microglia. The spinal ERα (but not ERβ) expression significantly increased in the OVX group, which was reversed by E2 supplementation. Moreover, the OVX individuals showed an increased protein kinase B (AKT) level in lumbar spinal cord, and E2 supplementation diminished the AKT expression in OVX rats. Finally, intrathecal injection Wortmannin, an inhibitor for AKT signaling, effectively reduced the nociceptive behaviors in the late phase and the number of c-fos positive cells. Together, our findings indicate that E2 supplementation alleviates the OVX-induced hyperalgesia, which might be involved in spinal ERα and AKT mechanisms.

Rhodojaponin III-Loaded Chitosan Derivatives-Modified Solid Lipid Nanoparticles for Multimodal Antinociceptive Effects in vivo.

BackgroundRhodojaponin III (RJ-III) is a bioactive diterpenoid, which is mainly found in Rhododendron molle G. Don (Ericaceae), a potent analgesia in traditional Chinese medicine with several years of clinical applications in the country. However, its clinical use is limited by its acute toxicity and poor pharmacokinetic profiles. To reduce such limitations, the current study incorporated RJ-III into the colloidal drug delivery system of hydroxypropyl trimethyl ammonium chloride chitosan (HACC)-modified solid lipid nanoparticles (SLNs) to improve its sustained release and antinociceptive effects in vivo for oral delivery.ResultsThe optimized RJ-III@HACC-SLNs were close to spherical, approximately 134 nm in size, and with a positive zeta potential. In vitro experiments showed that RJ-III@HACC-SLNs were stable in the simulated gastric fluid and had a prolonged release in PBS (pH = 6.8). Pharmacokinetic results showed that after intragastric administration in mice, the relative bioavailability of RJ-III@HACC-SLNs was 87.9%. Further, it was evident that the peak time, half-time, and mean retention time of RJ-III@HACC-SLNs were improved than RJ-III after the administration. In addition, pharmacodynamic studies revealed that RJ-III@HACC-SLNs markedly reduced the acetic acid, hot, and formalin-induced nociceptive responses in mice (P < 0.001), and notably increased the analgesic time (P < 0.01). Moreover, RJ-III@HACC-SLNs not only showed good biocompatibility with Caco-2 cells in vitro but its LD50 value was also increased by 1.8-fold as compared with that of RJ-III in vivo.ConclusionThese results demonstrated that RJ-III@HACC-SLNs improved the pharmacokinetic characteristics of the RJ-III, thereby exhibiting toxicity-attenuating potential and antinociceptive enhancing properties. Consequently, HACC-SLNs loaded with RJ-III could become a promising oral formulation for pain management that deserves further investigation in the future.

Translational Value of the Transdermal Administration of Bergamot Essential Oil and of Its Fractions.

The essential oil of bergamot (BEO) has consistently proven antinociceptive and antiallodynic properties. Accordingly, the analgesic efficacy of the decolored essential oil (DEC), with higher levels of limonene, and the deterpenated (DET) fraction, with higher levels of linalool and linalyl acetate, was investigated using a formalin test after inhalation. The present study was aimed at characterizing the effects of BEO, its components with the highest pharmacological activity (represented by linalool, limonene, and linalyl acetate), and its DEC and DET fractions on the formalin test after transdermal administration relevant to clinical translation through topical application. To this aim, the schedule of intervention involved administration immediately after formalin injection or as a 5 min pretreatment followed by washout in ddY-strain mice. This study demonstrates, for the first time, the significant analgesic effect of all three constituents in the first and second phases, accounting for the efficacy of the essential oil in the formalin test. While all fractions revealed equal activity toward the phytocomplex in the early phase, the reduction in time of licking/biting during the late phase was more markedly induced by DEC. Moreover, pretreatment with BEO and its fractions followed by washout did not produce a significant reduction in licking/biting time in both phases of formalin-induced nociceptive response.

Cuminum cyminum L.-Mediated Synthesis of Silver Nanoparticles: Their Characterization and Effect on Formalin-Induced Nociceptive Response in Male Rats.

In this study, a simple, low-cost, rapid, and eco-friendly approach for the biosynthesis of silver nanoparticles (AgNPs) using the aqueous extract of Cuminum cyminum L. (cumin) seed (CcAgNPs) was developed. Also, the anti-nociceptive properties of these synthesized AgNPs were evaluated in vivo. The CcAgNPs characterized using Ultraviolet-visible (UV-Vis) spectrophotometer, X-ray diffraction analysis (XRD), Fourier transform infrared (FTIR) spectroscopy, atomic force microscopy (AFM), and transmission electron microscopy (TEM). The analysis of phytochemical components in the aqueous extract of cumin seeds showed high concentrations of total phenols and ascorbic acid and low concentrations of total flavonoids. The analysis of phytochemical components and FTIR spectroscopy confirmed the presence of functional groups responsible for the bioreduction of Ag+ to AgNPs. The UV-Vis absorbance spectrum of CcAgNPs showed a maximum wavelength at 442nm. The analysis of TEM images showed a spherical shape with a size of less than 50nm, while XRD spectra revealed the crystallinity of CcAgNPs. The analysis of anti-nociceptive properties of CcAgNPs showed that the first phase of formalin-induced pain was significantly reduced in the groups receiving 200, 500, and 1000mg/kg CcAgNPs compared with the controls and the group receiving 300mg/kg of sodium salicylate (SS300). The second phase of formalin pain was also significantly reduced in the groups receiving 200 and 500mg/kg CcAgNPs compared to the controls and SS300 group. Overall, we introduced a new AgNPs synthesized from cumin seeds (CcAgNPs) and showed their anti-nociceptive properties in the formalin-induced pain.

Contribution of spathulenol to the anti-nociceptive effects of Psidium guineense

ABSTRACT Objectives: Araçá-verdadeiro is the popular name of Psidium guineense (Myrtaceae), whose fruits and leaves are used in Brazilian folk medicine for treatment of inflammation and pain. The focus of the present research was an investigation of the anti-nociceptive, and anti-inflammatory effects of the essential oil from P. guineense (EOPG) leaves, and of spathulenol. The anxiolytic and antidepressive effects associated with chronic pain were also investigated in models of acute or persistent nociception or/and inflammatory pain. Methods and Results: Oral treatment with EOPG (10–100 mg/kg) or spathulenol (10 mg/kg) significantly inhibited formalin-induced nociceptive responses, both sensitivity to cold and edema. Oral treatment with EOPG (10 mg/kg) and spathulenol (10 mg/kg) did not reduce locomotor activity (open field test). Local administration of spathulenol (1000 µg/paw) significantly prevented formalin-induced nociceptive sensitivity to cold and paw edema, and carrageenan-induced mechanical hyperalgesia, paw edema and sensitivity to cold. In the Freund's complete adjuvant (CFA) model, oral treatment with EOPG (10 mg/kg) or spathulenol (10 mg/kg) for 21 days significantly inhibited all analyzed parameters. The percentage maximal inhibition by spathulenol was 76.00% (mechanical hyperalgesia), 71.90% (cold response), 85.00% (edema), 77.16% (myeloperoxidase activity), 97.72% (time in the closed arms in the elevated plus maze), and 49.00% (immobility time in the tail suspension test), in the CFA model. Models employed male Swiss mice, except for the CFA test, which employed C57bL6 male mice (n=6 /group). Conclusion: This study demonstrates that EOPG is an anti-nociceptive and anti-hyperalgesic agent, in acute and continuous treatment, and an anxiolytic and antidepressive agent when tested with the chronic pain experimental state.

Pain impairs consolidation, but not acquisition or retrieval of a declarative memory.

Among the physical conditions that impair memory performance, pain is one of the most prevalent. However, the mechanisms by which pain impairs memory are largely unknown. In this study, we asked whether pain affects memory acquisition, consolidation and retrieval as well as whether memory impairment depends on pain intensity. Wistar rats received a hind paw injection of formalin (1%) at different phases of object recognition test. The injection of formalin after training but not before training or testing impaired object recognition memory. We concluded that pain impairs the consolidation but not acquisition or retrieval of object recognition memory, which is a subtype of declarative memory. Morphine, at a dose that did not affect object recognition memory in control rats, drastically reduced formalin-induced nociceptive behavior without reversing memory impairment. A lower dose of formalin (0.25%) induced less nociceptive behavior, but similar memory impairment. There is no statistical correlation between the intensity of nociceptive response and the performance in object recognition test. However, when formalin-induced nociceptive response was blocked by a local anesthetic, memory impairment was prevented. These findings suggest that pain-induced impairment in the consolidation of object recognition memory does not directly depend on the intensity of nociceptive activity.

Oral Spirulina Platensis Attenuates Hyperglycemia and Exhibits Antinociceptive Effect in Streptozotocin-Induced Diabetic Neuropathy Rat Model.

IntroductionDiabetic neuropathy is a common consequence of diabetes. Hyperalgesia is one of the main symptoms of diabetic neuropathy. In response to noxious stimuli, streptozotocin (STZ)-induced diabetic rats show exaggerated hyperalgesic behavior, while Spirulina platensis has anti-inflammatory, antioxidant, and insulin-like effects. To assess the antinociceptive effect of oral Spirulina platensis (SP) powder on formalin-induced nociceptive responses in STZ-induced diabetic rats.MethodsSixty mature male albino rats were randomly allocated into six equal groups (10 in each group). Group 1 (control non-diabetic group) received 0.9% saline; group 2 was given oral pure SP powder-treated as a non-diabetic control group, group 3 was sodium salicylate-treated rats and used as a positive non-diabetic control group, group 4 managed as vehicle-treated diabetic rats, group 5 considered as SP-treated-diabetic group, and sodium salicylate-treated-diabetic rats used as a diabetic positive control group (group 6). STZ-diabetic rats were orally given SP in a dose of 500 mg kg/day for 1 month. The formalin test was implemented in two phases: the early phase in the first 10-min post-formalin injection, and the late phase was considered in the 15–60 min post-formalin injection time interval.ResultsPain scores were increased in the diabetic groups during both phases of the experiment. Blood glucose was significantly reduced in diabetic rats that received oral SP, P < 0.01. Besides, SP-treated rats had lower pain scores during both phases of the experiment than untreated diabetic ones. However, in the sodium salicylate group, the pain score was reduced only during the second phase. An exaggerated nociceptive response occurred in diabetic rats after the formalin test. A significant antinociceptive effect appeared in SP-treated control and diabetic rats.DiscussionThe findings suggest that oral Spirulina platensis could have a potential therapeutic role for managing induced painful diabetic neuropathy in rats.

Behavioral Effects of Continuously Administered Bergamot Essential Oil on Mice With Partial Sciatic Nerve Ligation.

Neuropathic pain is an intractable chronic pain condition that is mainly caused by allodynia. We had previously reported that intra-plantar administration of bergamot essential oil (BEO) containing an aromatic compound significantly suppressed partial sciatic nerve ligation (PSNL)-induced mechanical allodynia via opioid mu receptors in mice. However, it has also been reported that the inhalation of BEO reduced formalin-induced nociceptive responses. Therefore, we aimed to elucidate whether the analgesic action of BEO is mediated by olfactory stimulation through volatile components. In the current study, BEO was continuously administered with an osmotic pump during PSNL surgery, and the effects on mice behavior were examined pharmacologically using a double activity monitoring system, which can detect two-dimensional planar motion in a cage with an infrared beam sensor as well as active motion with a running wheel. Here, we report that the two-dimensional planar activity significantly increased in mice with PSNL in the light phase (from 8 o’clock to 20 o’clock) but not in the dark phase (from 20 o’clock to 8 o’clock) from the second day after surgery. However, this increase was not observed when BEO was continuously administered. The effect of BEO on the two-dimensional planar counts in mice with PSNL was antagonized by naloxone hydrochloride. Regarding the running wheel activity, the number of rotations decreased by PSNL in the dark phase from the 8th day after surgery. However, this was not apparent with BEO use. The effect of BEO on the number of rotations was also antagonized by naloxone hydrochloride. Furthermore, inhalation of BEO in PSNL mice did not affect mechanical allodynia or the two-dimensional planar motion or running wheel activities. These findings indicate that BEO exhibits an analgesic action, which is mediated by opioid receptors and not by the olfactory system.

Mesaconitine plays the major role in the antinociceptive and anti-inflammatory activities of Radix Aconiti Carmichaeli (Chuan Wu)

This study investigated the antinociceptive and anti-inflammatory activities of the aqueous extracts of Radix Aconiti Carmichaeli (Chuan Wu) by oral gavage in mice. Carrageenan-induced paw edema served as an acute inflammation model. The acetic acid-induced writhing test and formalin-induced nociceptive responses in the early and late phases were used to assess the analgesic activity. The contents of Aconitum alkaloids were analyzed by high-performance liquid chromatography. The order of concentration of aconitine-type alkaloids in Radix Aconiti Carmichaeli was mesaconitine > aconitine > hypaconitine. Radix Aconiti Carmichaeli water extract at the dosage of 60 mg/kg not only effectively inhibited carrageenan-induced paw edema, but also significantly attenuated the abdominal constriction induced by an intraperitoneal injection of acetic acid, as well as the late phase of pain response caused by a hindpaw injection of formalin. Mesaconitine and aconitine are effective in decreasing the pain sensation elicited by acetic acid. The aqueous extracts of Radix Aconiti Carmichaeli exhibits antinociceptive activity and anti-inflammatory effect probably due to the presence of high content of mesaconitine.

Effect of spinal angiotensin-converting enzyme 2 activation on the formalin-induced nociceptive response in mice

We have previously demonstrated that the phosphorylation of p38 MAPK, through spinal AT1 receptor activation, is involved in formalin-induced nociception and follows accompanied by the increase in spinal angiotensin (Ang) II levels. We have also found that Ang (1–7), an N-terminal fragment of Ang II generated by ACE2, prevents the Ang II-induced nociceptive behavior via spinal MAS1 and the inhibition of p38 MAPK phosphorylation. Here, we examined whether the ACE2 activator diminazene aceturate (DIZE) can prevent the formalin-induced nociception in mice. The i.t. administration of DIZE attenuated the second, but not the first phase of formalin-induced nociceptive response. An increase in the activity of spinal ACE2 was measured following DIZE administration. The inhibitory effect of DIZE on nociception was abolished by the i.t. co-administration of the MAS1 antagonist A779. The i.t. administration of Ang (1–7) showed a similar effect on the second phase of the response which was also attenuated by A779. Furthermore, DIZE and Ang (1–7) each inhibited the formalin-induced phosphorylation of p38 MAPK on the dorsal lumbar spinal cord. This inhibition was again prevented by A779. ACE2 was expressed in neurons and microglia but absent from astrocytes in the superficial dorsal horn. Our data show that the i.t.-administered DIZE attenuates the second phase of the formalin-induced nociception which is accompanied by the inhibition of p38 MAPK phosphorylation. They also suggest the involvement of MAS1 activation on spinal neurons and microglia in response to the increase in Ang (1–7) following ACE2 activation.

Zinc oxide nanoparticles synthesised from the Vernonia amygdalina shows the anti-inflammatory and antinociceptive activities in the mice model

In this study, we synthesised the zinc oxide nanoparticles from Vernonia amygdalina and evaluated its anti-inflammatory and antinociceptive potentials against the different inflammation and pain induced mice model. The synthesised zinc oxide nanoparticles were characterised by UV, SEM, XRD and FTIR techniques. The anti-nociceptive effects of V. amygdalina were examined by different stimuli e.g. acetic acid, glutamate, capsaicin, and formalin-induced nociception in mice. The anti-inflammatory effects of synthesised zinc oxide nanoparticles were assessed by air sack assessment and the level of inflammatory cytokines were studied. The muscle tension of animals were studied through open field assessment. The present study exhibited proficient antinociceptive and anti-inflammatory actions of the synthesised Zinc oxide nanoparticles from V. amygdalina. The sormulated zinc oxide nanoparticles were appreciably reduced the acetic acid, glutamate, capsaicin, and formalin-induced nociceptive responses in mice. Further the zinc nanoparticles were exhibited the potent anti-inflammatory actions via reducing the inflammatory response and pro-inflammatory cytokines level in the mice. In conclusion, the findings of this study proved the beneficial effects of zinc oxide nanoparticles from V. amygdalina against the different pain and inflammation-induced mice. Hence, it was clear that the zinc nanoparticles from V. amygdalina could be promising antinociceptive and anti-inflammatory agent in the future.

Expressions of spinal microglia activation, BDNF, and DREAM proteins correlated with formalin-induced nociceptive responses in painful and painless diabetic neuropathy rats

The complications of diabetic polyneuropathy (DN) determines its level of severity. It may occur with distinctive clinical symptoms (painful DN) or appears undetected (painless DN). This study aimed to investigate microglia activation and signalling molecules brain-derived neurotrophic factor (BDNF) and downstream regulatory element antagonist modulator (DREAM) proteins in spinal cord of streptozotocin-induced diabetic neuropathy rats. Thirty male Sprague-Dawley rats (200-230 g) were randomly assigned into three groups: (1) control, (2) painful DN and (3) painless DN. The rats were induced with diabetes by single intraperitoneal injection of streptozotocin (60 mg/kg) whilst control rats received citrate buffer as a vehicle. Four weeks post-diabetic induction, the rats were induced with chronic inflammatory pain by intraplantar injection of 5% formalin and pain behaviour responses were recorded and assessed. Three days later, the rats were sacrificed and lumbar enlargement region of spinal cord was collected. The tissue was immunoreacted against OX-42 (microglia), BDNF and DREAM proteins, which was also quantified by western blotting. The results demonstrated that painful DN rats exhibited increased pain behaviour score peripherally and centrally with marked increase of spinal activated microglia, BDNF and DREAM proteins expressions compared to control group. In contrast, painless DN group demonstrated a significant reduction of pain behaviour score peripherally and centrally with significant reduction of spinal activated microglia, BDNF and DREAM proteins expressions. In conclusions, the spinal microglia activation, BDNF and DREAM proteins correlate with the pain behaviour responses between the variants of DN.

Formalin-induced nociceptive response is enhanced by serum exosomes isolated from partial sciatic nerve ligation (PSL) mice

Formalin-induced nociceptive response is enhanced by serum exosomes isolated from partial sciatic nerve ligation (PSL) mice

The novel small-molecule antagonist of PAC1 receptor attenuates formalin-induced inflammatory pain behaviors in mice

We recently developed PA-8, a novel small-molecule antagonist of PACAP type 1 (PAC1) receptor. In the present study, we examined whether PA-8 was effective against formalin-induced inflammatory pain in mice. Both intrathecal and oral administration of PA-8 resulted in the dose-dependent attenuation of the second phase of formalin-induced nociceptive responses. PA-8 also inhibited c-fos upregulation in the ipsilateral dorsal horn of the spinal cord. The results suggested that PACAP-PAC1 receptor signaling system in the spinal cord were primarily involved in the transmission of inflammatory pain, and PA-8 could be useful for the development of novel analgesics for treating inflammatory pain.

Antinociceptive effect of inhalation of the essential oil of bergamot in mice

Bergamot essential oil (BEO) has proven wide evidence of pharmacological antinociceptive effectiveness both in nociceptive and in neuropathic pain models. The antinociceptive properties of BEO for inhalation have not been investigated. The purpose of this study is to evaluate the effects of the inhalation of BEO on formalin-induced nociceptive response in mice.Male ddY-strain mice (Japan SLC, Hamamatsu, Japan) of 23–25 g of weight at the time the experiments underwent the formalin test. Twenty μl of formalin (2% in saline) were administered into the plantar surface of the mice hindpaw and the time of licking/biting was observed and recorded at intervals of 5 min. The device for BEO inhalatory delivery consisted in a filter paper disc soaked with known volume of BEO placed on the edge of the cage.Inhalation of BEO exerted antinociceptive activity. In particular, it reduced the formalin-induced licking/biting behaviour in a manner that was dependent on the volume of BEO used in the device for its release and on the time of exposure to the phytocomplex.The results support the use of BEO in aromatherapy for complementary management of chronic pain relief in a stepwise therapeutic programme.

Antinociceptive effects of novel epibatidine analogs through activation of α4β2 nicotinic receptors.

The study of α4β2 nicotinic receptors has provided new indications in the treatment of pain. Efforts have been made to explore new α4β2 nicotinic receptor agonists, including TC-2559, as antinociceptive drugs. In this study, we discovered a set of novel epibatidine analogs with strong binding affinities to the α4β2 nicotinic receptors. Among these compounds, C-159, C-163, and C-9515 attenuated formalin-induced nociceptive responses in mice; C-9515 caused the most potent analgesic effect, which was blocked by mecamylamine, a non-selective nicotinic receptor antagonist. Furthermore, C-9515 potently inhibited chronic constriction injury (CCI)-induced neuropathic pain in rats, which was sensitive to DHβE, a selective α4β2 subtype antagonist, indicating that its analgesic effect was mediated by the activation of the α4β2 nicotinic receptors. In conclusion, the epibatidine analog C-9515 was found to be a potent α4β2 nicotinic receptor agonist with potent analgesic function, which demonstrated potential for the further exploration of its druggability.

Mouse mast cells and mast cell proteases do not play a significant role in acute tissue injury pain induced by formalin.

Subcutaneous formalin injections are used as a model for tissue injury-induced pain where formalin induces pain and inflammation indirectly by crosslinking proteins and directly through activation of the transient receptor potential A1 receptor on primary afferents. Activation of primary afferents leads to both central and peripheral release of neurotransmitters. Mast cells are found in close proximity to peripheral sensory nerve endings and express receptors for neurotransmitters released by the primary afferents, contributing to the neuro/immune interface. Mast cell proteases are found in large quantities within mast cell granules and are released continuously in small amounts and upon mast cell activation. They have a wide repertoire of proposed substrates, including Substance P and calcitonin gene-related peptide, but knowledge of their in vivo function is limited. We evaluated the role of mouse mast cell proteases (mMCPs) in tissue injury pain responses induced by formalin, using transgenic mice lacking either mMCP4, mMCP6, or carboxypeptidase A3 (CPA3), or mast cells in their entirety. Further, we investigated the role of mast cells in heat hypersensitivity following a nerve growth factor injection. No statistical difference was observed between the respective mast cell protease knockout lines and wild-type controls in the formalin test. Mast cell deficiency did not have an effect on formalin-induced nociceptive responses nor nerve growth factor-induced heat hypersensitivity. Our data thus show that mMCP4, mMCP6, and CPA3 as well as mast cells as a whole, do not play a significant role in the pain responses associated with acute tissue injury and inflammation in the formalin test. Our data also indicate that mast cells are not essential to heat hypersensitivity induced by nerve growth factor.

Spinal Sigma-1 Receptor-mediated Dephosphorylation of Astrocytic Aromatase Plays a Key Role in Formalin-induced Inflammatory Nociception

Aromatase is a key enzyme responsible for the biosynthesis of estrogen from testosterone. Although recent evidence indicates that spinal cord aromatase participates in nociceptive processing, the mechanisms underlying its regulation and its involvement in nociception remain unclear. The present study focuses on the potential role of astrocyte aromatase in formalin-induced acute pain and begins to uncover one mechanism by which spinal aromatase activation is controlled. Following intraplantar formalin injection, nociceptive responses were quantified and immunohistochemistry/co-immunoprecipitation assays were used to investigate the changes in spinal Fos expression and the phospho-serine levels of spinal aromatase. Intrathecal (i.t.) injection of letrozole (an aromatase inhibitor) mitigated both the late phase formalin-induced nociceptive responses and formalin-induced spinal Fos expression. Furthermore, formalin-injected mice showed significantly reduced phospho-serine levels of aromatase, which is associated with the rapid activation of this enzyme. However, sigma-1 receptor inhibition with i.t. BD1047 blocked the dephosphorylation of aromatase and potentiated the pharmacological effect of letrozole on formalin-induced nociceptive responses. In addition, i.t. administration of a sub-effective dose of BD1047 potentiated the pharmacological effect of cyclosporin A (a calcineurin inhibitor) on both the formalin-induced reduction in phospho-serine levels of aromatase and nociceptive behavior. These results suggest that dephosphorylation is an important regulatory mechanism involved in the rapid activation of aromatase and that spinal sigma-1 receptors mediate this dephosphorylation of aromatase through an intrinsic calcineurin pathway.

Orai1 Plays a Crucial Role in Central Sensitization by Modulating Neuronal Excitability.

Pathological pain is a common and debilitating condition that is often poorly managed. Central sensitization is an important mechanism underlying pathological pain. However, candidate molecules involved in central sensitization remain unclear. Store-operated calcium channels (SOCs) mediate important calcium signals in nonexcitable and excitable cells. SOCs have been implicated in a wide variety of human pathophysiological conditions, including immunodeficiency, occlusive vascular diseases, and cancer. However, the role of SOCs in CNS disorders has been relatively unexplored. Orai1, a key component of SOCs, is expressed in the human and rodent spinal cord dorsal horn, but its functional significance in dorsal horn neurons is poorly understood. Here we sought to explore a potential role of Orai1 in the modulation of neuronal excitability and A-type potassium channels involved in pain plasticity. Using both male and female Orai1 knock-out mice, we found that activation of Orai1 increased neuronal excitability and reduced A-type potassium channels via the protein kinase C-extracellular signal-regulated protein kinase (PKC-ERK) pathway in dorsal horn neurons. Orai1 deficiency significantly decreased acute pain induced by noxious stimuli, nearly eliminated the second phase of formalin-induced nociceptive response, markedly attenuated carrageenan-induced ipsilateral pain hypersensitivity and abolished carrageenan-induced contralateral mechanical allodynia. Consistently, carrageenan-induced increase in neuronal excitability was abolished in the dorsal horn from Orai1 mutant mice. These findings uncover a novel signaling pathway involved in the pain process and central sensitization. Our study also reveals a novel link among Orai1, ERK, A-type potassium channels, and neuronal excitability.SIGNIFICANCE STATEMENT Orai1 is a key component of store-operated calcium channels (SOCs) in many cell types. It has been implicated in such pathological conditions as immunodeficiency, autoimmunity, and cancer. However, the role of Orai1 in CNS disorders remains poorly understood. The functional significance of Orai1 in neurons is elusive. Here we demonstrate that activation of Orai1 modulates neuronal excitability and Kv4-containing A-type potassium channels via the protein kinase C-extracellular signal-regulated protein kinase (PKC-ERK) pathway. Genetic knock-out of Orai1 nearly eliminates the second phase of formalin-induced pain and markedly attenuates carrageenan-induced pain hypersensitivity and neuronal excitability. These findings reveal a novel link between Orai1 and neuronal excitability and advance our understanding of central sensitization.

Role of orexin-2 receptors in the nucleus accumbens in antinociception induced by carbachol stimulation of the lateral hypothalamus in formalin test.

Orexins, which are mainly produced by orexin-expressing neurons in the lateral hypothalamus (LH), play an important role in pain modulation. Previously, it has been established that the nucleus accumbens (NAc) is involved in the modulation of formalin-induced nociceptive responses, a model of tonic pain. In this study, the role of intra-accumbal orexin-2 receptors (OX2rs) in the mediation of formalin-induced pain was investigated. A volume of 0.5 μl of 10, 20, and 40 nmol/l solutions of TCS OX2 29, an OX2r antagonist, were unilaterally microinjected into the NAc 5 min before an intra-LH carbachol microinjection (0.5 μl of 250 nmol/l solution). After 5 min, animals received a subcutaneous injection of formalin 2.5% (50 μl) into the hind paw. Pain-related behaviors were assessed at 5 min intervals during a 60-min test period. The findings showed that TCS OX2 29 administration dose dependently blocked carbachol-induced antinociception during both phases of formalin-induced pain. The antianalgesic effect of TCS OX2 29 was greater during the late phase compared with the early phase. These observations suggest that the NAc, as a part of a descending pain-modulatory circuitry, partially mediates LH-induced analgesia in the formalin test through recruitment of OX2rs. This makes the orexinergic system a good potential therapeutic target in the control of persistent inflammatory pain.