RESUMEN
Cocaine- and amphetamine-regulated transcript (CART) peptides are involved in several physiological and pathological processes, but their mechanism of action is unrevealed due to the lack of identified receptor(s). We provided evidence for the antihyperalgesic effect of CART(55-102) by inhibiting dipeptidyl-peptidase 4 (DPP4) in astrocytes and consequently reducing neuroinflammation in the rat spinal dorsal horn in a carrageenan-evoked inflammation model. Both naturally occurring CART(55-102) and CART(62-102) peptides are present in the spinal cord. CART(55-102) is not involved in acute nociception but regulates spinal pain transmission during peripheral inflammation. While the full-length peptide with a globular motif contributes to hyperalgesia, its N-terminal inhibits this process. Although the anti-hyperalgesic effects of CART(55-102), CART(55-76), and CART(62-76) are blocked by opioid receptor antagonists in our inflammatory models, but not in neuropathic Seltzer model, none of them bind to any opioid or G-protein coupled receptors. DPP4 interacts with Toll-like receptor 4 (TLR4) signalling in spinal astrocytes and enhances the TLR4-induced expression of interleukin-6 and tumour necrosis factor alpha contributing to inflammatory pain. Depending on the state of inflammation, CART(55-102) is processed in the spinal cord, resulting in the generation of biologically active isoleucine-proline-isoleucine (IPI) tripeptide, which inhibits DPP4, leading to significantly decreased glia-derived cytokine production and hyperalgesia.
Asunto(s)
Hiperalgesia , Receptor Toll-Like 4 , Ratas , Animales , Hiperalgesia/metabolismo , Dipeptidil Peptidasa 4 , Isoleucina , Nocicepción , Dolor/metabolismo , Fragmentos de Péptidos/farmacología , Médula Espinal/metabolismo , Inflamación/metabolismoRESUMEN
Complex Regional Pain Syndrome (CRPS) represents severe chronic pain, hypersensitivity, and inflammation induced by sensory-immune-vascular interactions after a small injury. Since the therapy is unsatisfactory, there is a great need to identify novel drug targets. Unbiased transcriptomic analysis of the dorsal root ganglia (DRG) was performed in a passive transfer-trauma mouse model, and the predicted pathways were confirmed by pharmacological interventions. In the unilateral L3-5 DRGs 125 genes were differentially expressed in response to plantar incision and injecting IgG of CRPS patients. These are related to inflammatory and immune responses, cytokines, chemokines and neuropeptides. Pathway analysis revealed the involvement of Tumor Necrosis Factor (TNF) and Janus kinase (JAK-STAT) signaling. The relevance of these pathways was proven by abolished CRPS IgG-induced hyperalgesia and reduced microglia and astrocyte markers in pain-associated central nervous system regions after treatment with the soluble TNF alpha receptor etanercept or JAK inhibitor tofacitinib. These results provide the first evidence for CRPS-related neuroinflammation and abnormal cytokine signaling at the level of the primary sensory neurons in a translational mouse model and suggest that etanercept and tofacitinib might have drug repositioning potentials for CRPS-related pain.
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Dolor Crónico , Síndromes de Dolor Regional Complejo , Animales , Síndromes de Dolor Regional Complejo/tratamiento farmacológico , Síndromes de Dolor Regional Complejo/patología , Modelos Animales de Enfermedad , Etanercept/farmacología , Etanercept/uso terapéutico , Ganglios Espinales/patología , Inmunoglobulina G , Quinasas Janus , Ratones , Factores de Transcripción STAT , Transducción de Señal , Transcriptoma , Factor de Necrosis Tumoral alfaRESUMEN
Neuroimmune interactions may contribute to severe pain and regional inflammatory and autonomic signs in complex regional pain syndrome (CRPS), a posttraumatic pain disorder. Here, we investigated peripheral and central immune mechanisms in a translational passive transfer trauma mouse model of CRPS. Small plantar skin-muscle incision was performed in female C57BL/6 mice treated daily with purified serum immunoglobulin G (IgG) from patients with longstanding CRPS or healthy volunteers followed by assessment of paw edema, hyperalgesia, inflammation, and central glial activation. CRPS IgG significantly increased and prolonged swelling and induced stable hyperalgesia of the incised paw compared with IgG from healthy controls. After a short-lasting paw inflammatory response in all groups, CRPS IgG-injected mice displayed sustained, profound microglia and astrocyte activation in the dorsal horn of the spinal cord and pain-related brain regions, indicating central sensitization. Genetic deletion of interleukin-1 (IL-1) using IL-1αß knockout (KO) mice and perioperative IL-1 receptor type 1 (IL-1R1) blockade with the drug anakinra, but not treatment with the glucocorticoid prednisolone, prevented these changes. Anakinra treatment also reversed the established sensitization phenotype when initiated 8 days after incision. Furthermore, with the generation of an IL-1ß floxed(fl/fl) mouse line, we demonstrated that CRPS IgG-induced changes are in part mediated by microglia-derived IL-1ß, suggesting that both peripheral and central inflammatory mechanisms contribute to the transferred disease phenotype. These results indicate that persistent CRPS is often contributed to by autoantibodies and highlight a potential therapeutic use for clinically licensed antagonists, such as anakinra, to prevent or treat CRPS via blocking IL-1 actions.
Asunto(s)
Autoanticuerpos/inmunología , Síndromes de Dolor Regional Complejo/inmunología , Inmunoglobulina G/inmunología , Interleucina-1alfa/inmunología , Interleucina-1beta/inmunología , Adulto , Animales , Autoanticuerpos/administración & dosificación , Autoanticuerpos/sangre , Síndromes de Dolor Regional Complejo/sangre , Síndromes de Dolor Regional Complejo/diagnóstico , Síndromes de Dolor Regional Complejo/tratamiento farmacológico , Modelos Animales de Enfermedad , Femenino , Humanos , Inmunoglobulina G/administración & dosificación , Inmunoglobulina G/sangre , Proteína Antagonista del Receptor de Interleucina 1/administración & dosificación , Interleucina-1alfa/genética , Interleucina-1alfa/metabolismo , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Extremidad Inferior/lesiones , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Microglía/inmunología , Microglía/patología , Persona de Mediana Edad , Dimensión del Dolor , Receptores Tipo I de Interleucina-1/antagonistas & inhibidores , Receptores Tipo I de Interleucina-1/inmunología , Receptores Tipo I de Interleucina-1/metabolismo , Asta Dorsal de la Médula Espinal/inmunología , Asta Dorsal de la Médula Espinal/patologíaRESUMEN
Pharmacotherapy of neuropathic pain is still challenging. Our earlier work indicated an analgesic effect of dimethyl trisulfide (DMTS), which was mediated by somatostatin released from nociceptor nerve endings acting on SST4 receptors. Somatostatin release occurred due to TRPA1 ion channel activation. In the present study, we investigated the effect of DMTS in neuropathic pain evoked by partial ligation of the sciatic nerve in mice. Expression of the mRNA of Trpa1 in murine dorsal-root-ganglion neurons was detected by RNAscope. Involvement of TRPA1 ion channels and SST4 receptors was tested with gene-deleted animals. Macrophage activity at the site of the nerve lesion was determined by lucigenin bioluminescence. Density and activation of microglia in the spinal cord dorsal horn was verified by immunohistochemistry and image analysis. Trpa1 mRNA is expressed in peptidergic and non-peptidergic neurons in the dorsal root ganglion. DMTS ameliorated neuropathic pain in Trpa1 and Sstr4 WT mice, but not in KO ones. DMTS had no effect on macrophage activity around the damaged nerve. Microglial density in the dorsal horn was reduced by DMTS independently from TRPA1. No effect on microglial activation was detected. DMTS might offer a novel therapeutic opportunity in the complementary treatment of neuropathic pain.
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Neuralgia/tratamiento farmacológico , Sulfuros/farmacología , Canal Catiónico TRPA1/metabolismo , Animales , Ganglios Espinales/metabolismo , Hiperalgesia , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Microglía/metabolismo , Microscopía Confocal , ARN Mensajero/metabolismo , Nervio Ciático/patología , Somatostatina/metabolismoRESUMEN
Agonists at the δ opioid receptor are known to be potent antihyperalgesics in chronic pain models and effective in models of anxiety and depression. However, some δ opioid agonists have proconvulsant properties while tolerance to the therapeutic effects can develop. Previous evidence indicates that different agonists acting at the δ opioid receptor differentially engage signaling and regulatory pathways with significant effects on behavioral outcomes. As such, interest is now growing in the development of biased agonists as a potential means to target specific signaling pathways and potentially improve the therapeutic profile of δ opioid agonists. Here, we report on PN6047 (3-[[4-(dimethylcarbamoyl)phenyl]-[1-(thiazol-5-ylmethyl)-4-piperidylidene]methyl]benzamide), a novel G protein-biased and selective δ opioid agonist. In cell-based assays, PN6047 fully engages G protein signaling but is a partial agonist in both the arrestin recruitment and internalization assays. PN6047 is effective in rodent models of chronic pain but shows no detectable analgesic tolerance following prolonged treatment. In addition, PN6047 exhibited antidepressant-like activity in the forced swim test, and importantly, the drug had no effect on chemically induced seizures. PN6047 did not exhibit reward-like properties in the conditioned place preference test or induce respiratory depression. Thus, δ opioid ligands with limited arrestin signaling such as PN6047 may be therapeutically beneficial in the treatment of chronic pain states. SIGNIFICANCE STATEMENT: PN6047 (3-[[4-(dimethylcarbamoyl)phenyl]-[1-(thiazol-5-ylmethyl)-4-piperidylidene]methyl]benzamide) is a selective, G protein-biased δ opioid agonist with efficacy in preclinical models of chronic pain. No analgesic tolerance was observed after prolonged treatment, and PN6047 does not display proconvulsant activity or other opioid-mediated adverse effects. Our data suggest that δ opioid ligands with limited arrestin signaling will be beneficial in the treatment of chronic pain.
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Analgésicos Opioides/metabolismo , Antidepresivos/química , Benzamidas/química , Benzamidas/farmacocinética , Dolor Crónico/tratamiento farmacológico , Proteínas de Unión al GTP/metabolismo , Receptores Opioides delta/metabolismo , Animales , Antidepresivos/administración & dosificación , Antidepresivos/efectos adversos , Antidepresivos/farmacocinética , Arrestina/metabolismo , Benzamidas/administración & dosificación , Benzamidas/efectos adversos , Tolerancia a Medicamentos , Células HEK293 , Humanos , Masculino , Ratones , Modelos Animales , Ratas Wistar , Resultado del TratamientoRESUMEN
Semicarbazide-sensitive amine oxidase (SSAO) produces tissue irritants by deamination of primary amines, which activate transient receptor potential ankyrin 1 (TRPA1) and vanilloid 1 (TRPV1) receptors expressed predominantly on nociceptors. Since there are no data about its functions in pain, we studied the effects and mechanisms of action of our novel SSAO inhibitor and dual TRPA1/TRPV1 antagonist multi-target drug SZV 1287 in different pain models. Acute chemonociception was induced by TRPV1 and TRPA1 activation (resiniferatoxin and formalin, respectively), chronic arthritis by K/BxN serum transfer, traumatic mononeuropathy by sciatic nerve ligation. SZV 1287 (20â¯mg/kg i.p.) was investigated in C57BL/6J wildtype (WT), TRPA1- (TRPA1-/-) and TRPV1-deficient (TRPV1-/-) mice. Paw mechanonociception was measured by aesthesiometry, thermonociception by hot plate, nocifensive behavior by licking duration, volume by plethysmometry, myeloperoxidase activity by luminescence and plasma extravasation by fluorescence imaging, glia activation in pain-related brain regions by immunohistochemistry. SZV 1287 significantly inhibited both TRPA1 and TRPV1 activation-induced acute chemonociception and hyperalgesia. In K/BxN arthritis, daily SZV 1287 injections significantly decreased hyperalgesia, L4-L6 spinal dorsal horn microgliosis, edema and myeloperoxidase activity. SZV 1287-evoked antihyperalgesic and anti-edema effects were absent in TRPV1-/-, and remarkably reduced in TRPA1-/- mice. In contrast, myeloperoxidase-inhibitory effect was absent in TRPA1-/-, but not in TRPV1-/- animals. Acute SZV 1287 administration resulted in approximately 50% significant reduction of neuropathic hyperalgesia 7â¯days after nerve ligation, which was not observed in either TRPA1-/- or TRPV1-/- mice. SZV 1287 inhibits chronic inflammatory and neuropathic pain via TRPV1 and TRPA1/TRPV1 activation, respectively, highlighting its drug developmental potential.
Asunto(s)
Amina Oxidasa (conteniendo Cobre)/antagonistas & inhibidores , Analgésicos/uso terapéutico , Dolor Crónico/tratamiento farmacológico , Inhibidores Enzimáticos/uso terapéutico , Oxazoles/uso terapéutico , Oximas/uso terapéutico , Canal Catiónico TRPA1/metabolismo , Canales Catiónicos TRPV/metabolismo , Amina Oxidasa (conteniendo Cobre)/metabolismo , Analgésicos/farmacología , Animales , Dolor Crónico/genética , Dolor Crónico/metabolismo , Modelos Animales de Enfermedad , Inhibidores Enzimáticos/farmacología , Eliminación de Gen , Masculino , Ratones Endogámicos C57BL , Neuralgia/tratamiento farmacológico , Neuralgia/genética , Neuralgia/metabolismo , Oxazoles/farmacología , Oximas/farmacología , Canal Catiónico TRPA1/genética , Canales Catiónicos TRPV/genéticaRESUMEN
The tachykinin NK1 receptor was suggested to be involved in psychiatric disorders, but its antagonists have failed to be effective as antidepressants in clinical trials. Hemokinin-1 (HK-1), the newest tachykinin, is present in several brain regions and activates the NK1 receptor similarly to substance P (SP), but acts also through other mechanisms. Therefore, we investigated the roles of the Tac4 gene-derived HK-1 in comparison with SP and neurokinin A (NKA) encoded by the Tac1 gene, as well as the NK1 receptor in anxiety and depression-like behaviors in mice. Mice lacking SP/NKA, HK-1 or the NK1 receptor (Tac1-/-, Tac4-/-, Tacr1-/-, respectively) compared to C57Bl/6 wildtypes (WT), and treatment with the NK1 antagonist CP99994 were used in the experiments. Anxiety was evaluated in the light-dark box (LDB) and the elevated plus maze (EPM), locomotor activity in the open field (OFT) tests. Hedonic behavior was assessed in the sucrose preference test (SPT), depression-like behavior in the tail suspension (TST) and forced swim (FST) tests. FST-induced neuronal responsiveness was evaluated with Fos immunohistochemistry in several stress-related brain regions. In the LDB, Tac4-/- mice spent significantly less, while Tacr1-/- and CP99994-treated mice spent significantly more time in the lit compartment. In the EPM only Tac4-/- showed reduced time in the open arms, but no difference was observed in any other groups. In the OFT Tac4-/- mice showed significantly reduced, while Tac1-/- and Tacr1-/- animals increased motility than the WTs, but CP99994 had no effect. NK1-/- consumed markedly more, while Tac4-/- less sucrose solution compared to WTs. In the TST and FST, Tac4-/- mice showed significantly increased immobility. However, depression-like behavior was decreased both in cases of genetic deletion and pharmacological blockade of the NK1 receptor. FST-induced neuronal activation in different nuclei involved in behavioral and neuroendocrine stress responses was significantly reduced in the brain of Tac4 -/- mice. Our results provide the first evidence for an anxiolytic and anti-depressant-like actions of HK-1 through a presently unknown target-mediated mechanism. Identification of its receptor and/or signaling pathways might open new perspectives for anxiolytic and anti-depressant therapies.
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Ansiolíticos/farmacología , Ansiedad/genética , Depresión/genética , Precursores de Proteínas/genética , Precursores de Proteínas/fisiología , Taquicininas/genética , Taquicininas/fisiología , Anhedonia , Animales , Ansiedad/psicología , Depresión/psicología , Preferencias Alimentarias , Genes fos , Suspensión Trasera , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Actividad Motora , Receptores de Neuroquinina-1/genética , Sustancia P/genéticaRESUMEN
The rodent transient receptor potential ankyrin-1 (TRPA1) channel has been hypothesized to serve as a temperature sensor for thermoregulation in the cold. We tested this hypothesis by using deletion of the Trpa1 gene in mice and pharmacological blockade of the TRPA1 channel in rats. In both Trpa1(-/-) and Trpa1(+/+) mice, severe cold exposure (8°C) resulted in decreases of skin and deep body temperatures to â¼8°C and 13°C, respectively, both temperatures being below the reported 17°C threshold temperature for TRPA1 activation. Under these conditions, Trpa1(-/-) mice had the same dynamics of body temperature as Trpa1(+/+) mice and showed no weakness in the tail skin vasoconstriction response or thermogenic response to cold. In rats, the effects of pharmacological blockade were studied by using two chemically unrelated TRPA1 antagonists: the highly potent and selective compound A967079, which had been characterized earlier, and the relatively new compound 43 ((4R)-1,2,3,4-tetrahydro-4-[3-(3-methoxypropoxy)phenyl]-2-thioxo-5H-indeno[1,2-d]pyrimidin-5-one), which we further characterized in the present study and found to be highly potent (IC50 against cold of â¼8 nm) and selective. Intragastric administration of either antagonist at 30 mg/kg before severe (3°C) cold exposure did not affect the thermoregulatory responses (deep body and tail skin temperatures) of rats, even though plasma concentrations of both antagonists well exceeded their IC50 value at the end of the experiment. In the same experimental setup, blocking the melastatin-8 (TRPM8) channel with AMG2850 (30 mg/kg) attenuated cold-defense mechanisms and led to hypothermia. We conclude that TRPA1 channels do not drive autonomic thermoregulatory responses to cold in rodents.
Asunto(s)
Sistema Nervioso Autónomo/fisiología , Regulación de la Temperatura Corporal/genética , Péptidos y Proteínas de Señalización Intracelular/fisiología , Sensación Térmica/genética , Animales , Regulación de la Temperatura Corporal/efectos de los fármacos , Células CHO , Frío , Cricetulus , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Femenino , Regulación de la Expresión Génica/efectos de los fármacos , Regulación de la Expresión Génica/genética , Proteínas HSP90 de Choque Térmico , Péptidos y Proteínas de Señalización Intracelular/antagonistas & inhibidores , Péptidos y Proteínas de Señalización Intracelular/genética , Masculino , Ratones , Ratones Transgénicos , Oximas/sangre , Oximas/farmacología , Dolor/inducido químicamente , Dolor/tratamiento farmacológico , Ratas , Ratas Sprague-Dawley , Ratas Wistar , Temperatura Cutánea/efectos de los fármacos , Canales Catiónicos TRPM/antagonistas & inhibidores , Sensación Térmica/efectos de los fármacosRESUMEN
A 2-day closed workshop was held in Liverpool, United Kingdom, to discuss the results of research concerning symptom-based disorders (SBDs) caused by autoantibodies, share technical knowledge, and consider future plans. Twenty-two speakers and 14 additional participants attended. This workshop set out to consolidate knowledge about the contribution of autoantibodies to SBDs. Persuasive evidence for a causative role of autoantibodies in disease often derives from experimental "passive transfer" approaches, as first established in neurological research. Here, serum immunoglobulin (IgM or IgG) is purified from donated blood and transferred to rodents, either systemically or intrathecally. Rodents are then assessed for the expression of phenotypes resembling the human condition; successful phenotype transfer is considered supportive of or proof for autoimmune pathology. Workshop participants discussed passive transfer models and wider evidence for autoantibody contribution to a range of SBDs. Clinical trials testing autoantibody reduction were presented. Cornerstones of both experimental approaches and clinical trial parameters in this field were distilled and presented in this article. Mounting evidence suggests that immunoglobulin transfer from patient donors often induces the respective SBD phenotype in rodents. Understanding antibody binding epitopes and downstream mechanisms will require substantial research efforts, but treatments to reduce antibody titres can already now be evaluated.
RESUMEN
Hydrogen sulfide (H2S) has been shown in previous studies to cause hypothermia and hypometabolism in mice, and its thermoregulatory effects were subsequently investigated. However, the molecular target through which H2S triggers its effects on deep body temperature has remained unknown. We investigated the thermoregulatory response to fast-(Na2S) and slow-releasing (GYY4137) H2S donors in C57BL/6 mice, and then tested whether their effects depend on the transient receptor potential ankyrin-1 (TRPA1) channel in Trpa1 knockout (Trpa1-/-) and wild-type (Trpa1+/+) mice. Intracerebroventricular administration of Na2S (0.5-1 mg/kg) caused hypothermia in C57BL/6 mice, which was mediated by cutaneous vasodilation and decreased thermogenesis. In contrast, intraperitoneal administration of Na2S (5 mg/kg) did not cause any thermoregulatory effect. Central administration of GYY4137 (3 mg/kg) also caused hypothermia and hypometabolism. The hypothermic response to both H2S donors was significantly (p < 0.001) attenuated in Trpa1-/- mice compared to their Trpa1+/+ littermates. Trpa1 mRNA transcripts could be detected with RNAscope in hypothalamic and other brain neurons within the autonomic thermoeffector pathways. In conclusion, slow- and fast-releasing H2S donors induce hypothermia through hypometabolism and cutaneous vasodilation in mice that is mediated by TRPA1 channels located in the brain, presumably in hypothalamic neurons within the autonomic thermoeffector pathways.
RESUMEN
SZV 1287 (3-(4,5-diphenyl-1,3-oxazol-2-yl)propanal oxime) is a novel multi-target candidate under preclinical development for neuropathic pain. It inhibits amine oxidase copper containing 3, transient receptor potential ankyrin 1 and vanilloid 1 (TRPV1) receptors. Mainly under acidic conditions, it is transformed to the cyclooxygenase inhibitor oxaprozin, which is ineffective for neuropathy. Therefore, an enterosolvent capsule is suggested for oral formulation, which we investigated for nociception, basic kinetics, and thermoregulatory safety in mice. The antihyperalgesic effect of SZV 1287 (10, 20, 50, and 200 mg/kg, p.o.) was determined in partial sciatic nerve ligation-induced traumatic neuropathy by aesthesiometry, brain and plasma concentrations by HPLC, and deep body temperature by thermometry. Its effect on proton-induced TRPV1 activation involved in thermoregulation was assessed by microfluorimetry in cultured trigeminal neurons. The three higher SZV 1287 doses significantly, but not dose-dependently, reduced neuropathic hyperalgesia by 50% of its maximal effect. It was quickly absorbed; plasma concentration was stable for 2 h, and it entered into the brain. Although SZV 1287 significantly decreased the proton-induced TRPV1-mediated calcium-influx potentially leading to hyperthermia, it did not alter deep body temperature. Oral SZV 1287 inhibited neuropathic hyperalgesia and, despite TRPV1 antagonistic action and brain penetration, it did not influence thermoregulation, which makes it a promising analgesic candidate.
RESUMEN
Diabetes mellitus is a common metabolic disease leading to hyperglycemia due to insufficient pancreatic insulin production or effect. Amine oxidase copper containing 3 (AOC3) is an enzyme that belongs to the semicarbazide-sensitive amine oxidase family, which may be a novel therapeutic target to treat diabetic complications. We aimed to explore the effects of AOC3 inhibition and to test the actions of our novel AOC3 inhibitor multi-target drug candidate, SZV 1287, compared to a selective reference compound, LJP 1207, in an 8-week long insulin-controlled streptozotocin (STZ)-induced (60â¯mg/kg i.p.) rat diabetes model. Both AOC3 inhibitors (20â¯mg/kg, daily s.c. injections) were protective against STZ-induced pancreatic beta cell damage determined by insulin immunohistochemistry and radioimmunoassay, neuropathic cold hypersensitivity measured by paw withdrawal latency decrease from 0⯰C water, and retinal dysfunction detected by electroretinography. SZV 1287 showed greater inhibitory effects on beta cell damage, and reduced retinal apoptosis shown by histochemistry. Mechanical hypersensitivity measured by aesthesiometry, cardiac dysfunction and nitrosative stress determined by echocardiography and immunohistochemistry/Western blot, respectively, serum Na+, K+, fructosamine, and urine microalbumin, creatinine, total protein/creatinine ratio alterations did not develop in response to diabetes. None of these parameters were influenced by the treatments except for SZV 1287 reducing serum fructosamine and LJP 1207 increasing urine creatinine. We provide the first evidence for protective effects of AOC3 inhibition on STZ-induced pancreatic beta cell damage, neuropathic cold hypersensitivity and diabetic retinal dysfunction. Long-term treatment with our novel multi-target analgesic candidate, SZV 1287, is safe and effective also under diabetic conditions.
Asunto(s)
Complicaciones de la Diabetes/tratamiento farmacológico , Diabetes Mellitus Experimental/tratamiento farmacológico , Inhibidores de la Monoaminooxidasa/farmacología , Oxazoles/farmacología , Oximas/farmacología , Amina Oxidasa (conteniendo Cobre)/metabolismo , Analgésicos/farmacología , Animales , Moléculas de Adhesión Celular/metabolismo , Complicaciones de la Diabetes/prevención & control , Diabetes Mellitus Experimental/prevención & control , Nefropatías Diabéticas/tratamiento farmacológico , Retinopatía Diabética/tratamiento farmacológico , Humanos , Hidrazinas/farmacología , Insulina/metabolismo , Células Secretoras de Insulina , Masculino , Ratas , Ratas Sprague-Dawley , Estreptozocina/efectos adversosRESUMEN
The inhibitory neuropeptide somatostatin regulates several functions in the nervous system including memory. Its concentrations decrease by age leading to functional alterations, but there are little known about the receptorial mechanism. We discovered that somatostatin receptor 4 (sst4) mediates analgesic, anti-depressant, and anti-inflammatory effects without endocrine actions, and it is a unique target for drug development. We investigated the exploratory and locomotor activities and learning and memory functions of male and female sst4gene-deficient mice compared with their wild-types (WT) at ages of 3, 12, 17 months in the Y-maze test, open field test (OFT), radial-arm maze (RAM) test and novel object recognition (NOR) test. Young sst4 gene-deficient females visited, repeated, and missed significantly less arms than the WTs in the RAM; males showed decreased exploration in the NOR. Young mice moved significantly more, spend longer time in OFT center, and visited more arms in the Y-maze than older ones. Young WT females spend significantly longer time in the OFT center, visited, missed and repeated more arms of the RAM than males. Old males found more rewards than females. Young males explored longer the novel object than young females and older males in the NOR; the recognition index was smaller in females. We conclude that aging and sex are important factors of behavioral parameters that should be focused on in such studies. Sst4 is likely to influence locomotion and exploratory behavior only in young mice, but not during normal aging, which is a beneficial feature of a good drug target focusing on the elderly.
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Envejecimiento/fisiología , Conducta Exploratoria/fisiología , Locomoción/fisiología , Receptores de Somatostatina/fisiología , Envejecimiento/genética , Animales , Femenino , Aprendizaje/fisiología , Masculino , Aprendizaje por Laberinto , Memoria a Corto Plazo/fisiología , Ratones Noqueados , Receptores de Somatostatina/genética , Factores Sexuales , Memoria Espacial/fisiologíaRESUMEN
Somatostatin released from activated capsaicin-sensitive afferents of the lung inhibits inflammation and related bronchial hyperreactivity presumably via somatostatin 4 receptors (sst(4)). The aim of this study was to examine the effects of TT-232, a heptapeptide sst(4)/sst(1) receptor agonist and J-2156, a high affinity sst(4) receptor-selective peptidomimetic agonist in airway inflammation models. Acute pneumonitis was evoked by intranasal lipopolysaccharide 24 h before measurement. Chronic inflammation was induced by ovalbumin inhalation on days 28, 29 and 30 after i.p. sensitization on days 1 and 14. Semiquantitative histopathological scoring was based on perivascular/peribronchial oedema, neutrophil/macrophage infiltration, goblet cell hyperplasia in the acute model and eosinophil infiltration, mucosal oedema, mucus production and epithelial cell damage in chronic inflammation. Myeloperoxidase activity of the lung was measured spectrophotometrically to quantify granulocyte accumulation and the broncoalveolar lavage fluid was analysed by flow cytometry. Carbachol-induced bronchoconstriction was assessed by unrestrained whole body plethysmography and its calculated indicator, enhanced pause (Penh) was determined. TT-232 and J-2156 induced similar inhibition on granulocyte recruitment and histopathological changes in both models, although macrophage infiltration in LPS-induced inflammation was unaltered by either compounds. Both agonists diminished inflammatory airway hyperresponsiveness. Since their single administration after the development of the inflammatory reactions also inhibited carbachol-induced bronchoconstriction, somatostatin sst(4) receptor activation on bronchial smooth muscle cells is likely to be involved in their anti-hyperreactivity effect. These results suggest that stable, somatostatin sst(4) receptor-selective agonists could be potential candidates for the development of a completely novel group of anti-inflammatory drugs for the treatment of airway inflammation and hyperresponsiveness.
Asunto(s)
Antiinflamatorios/farmacología , Asma/tratamiento farmacológico , Hiperreactividad Bronquial/prevención & control , Butanos/farmacología , Pulmón/efectos de los fármacos , Proteínas de la Membrana/agonistas , Naftalenos/farmacología , Péptidos Cíclicos/farmacología , Neumonía Bacteriana/tratamiento farmacológico , Receptores de Somatostatina/agonistas , Fármacos del Sistema Respiratorio/farmacología , Sulfonas/farmacología , Animales , Antiinflamatorios/uso terapéutico , Asma/inducido químicamente , Asma/metabolismo , Asma/patología , Asma/fisiopatología , Hiperreactividad Bronquial/metabolismo , Hiperreactividad Bronquial/patología , Hiperreactividad Bronquial/fisiopatología , Pruebas de Provocación Bronquial , Líquido del Lavado Bronquioalveolar/citología , Broncoconstrictores/administración & dosificación , Butanos/uso terapéutico , Carbacol/administración & dosificación , Células Cultivadas , Femenino , Interleucina-1beta/metabolismo , Lipopolisacáridos , Pulmón/metabolismo , Pulmón/patología , Pulmón/fisiopatología , Macrófagos Peritoneales/efectos de los fármacos , Macrófagos Peritoneales/metabolismo , Proteínas de la Membrana/metabolismo , Ratones , Ratones Endogámicos BALB C , Naftalenos/uso terapéutico , Ovalbúmina , Péptidos Cíclicos/uso terapéutico , Neumonía Bacteriana/inducido químicamente , Neumonía Bacteriana/metabolismo , Neumonía Bacteriana/patología , Neumonía Bacteriana/fisiopatología , Receptores de Somatostatina/metabolismo , Fármacos del Sistema Respiratorio/uso terapéutico , Somatostatina/análogos & derivados , Sulfonas/uso terapéutico , Acetato de Tetradecanoilforbol/farmacología , Factores de TiempoRESUMEN
Neurokinin (NK) signaling is involved in various inflammatory processes. A common manifestation of systemic inflammation is fever, which is usually induced in animal models with the administration of bacterial lipopolysaccharide (LPS). A role for the NK1 receptor was shown in LPS-induced fever, but the underlying mechanisms of how the NK1 receptor contributes to febrile response, especially in the early phase, have remained unknown. We administered LPS (120 µg/kg, intraperitoneally) to mice with the Tacr1 gene, i.e., the gene encoding the NK1 receptor, either present (Tacr1+/+ ) or absent (Tacr1-/- ) and measured their thermoregulatory responses, serum cytokine levels, tissue cyclooxygenase-2 (COX-2) expression, and prostaglandin (PG) E2 concentration. We found that the LPS-induced febrile response was attenuated in Tacr1-/- compared to their Tacr1+/+ littermates starting from 40 min postinfusion. The febrigenic effect of intracerebroventricularly administered PGE2 was not suppressed in the Tacr1-/- mice. Serum concentration of pyrogenic cytokines did not differ between Tacr1-/- and Tacr1+/+ at 40 min post-LPS infusion. Administration of LPS resulted in amplification of COX-2 mRNA expression in the lungs, liver, and brain of the mice, which was statistically indistinguishable between the genotypes. In contrast, the LPS-induced augmentation of COX-2 protein expression was attenuated in the lungs and tended to be suppressed in the liver of Tacr1-/- mice compared with Tacr1+/+ mice. The Tacr1+/+ mice responded to LPS with a significant surge of PGE2 production in the lungs, whereas Tacr1-/- mice did not. In conclusion, the NK1 receptor is necessary for normal fever genesis. Our results suggest that the NK1 receptor contributes to the early phase of LPS-induced fever by enhancing COX-2 protein expression in the periphery. These findings advance the understanding of the crosstalk between NK signaling and the "cytokine-COX-2-prostaglandin E2" axis in systemic inflammation, thereby open up the possibilities for new therapeutic approaches.
Asunto(s)
Ciclooxigenasa 2/metabolismo , Fiebre/inmunología , Lipopolisacáridos/efectos adversos , Receptores de Neuroquinina-1/metabolismo , Animales , Ciclooxigenasa 2/genética , Citocinas/sangre , Dinoprostona/sangre , Femenino , Fiebre/inducido químicamente , Inflamación/sangre , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Receptores de Neuroquinina-1/genética , Transducción de SeñalRESUMEN
Altered pain sensations such as hyperalgesia and allodynia are characteristic features of various pain states, and remain difficult to treat. We have shown previously that spinal application of dipeptidyl peptidase 4 (DPP4) inhibitors induces strong antihyperalgesic effect during inflammatory pain. In this study we observed low level of DPP4 mRNA in the rat spinal dorsal horn in physiological conditions, which did not change significantly either in carrageenan-induced inflammatory or partial nerve ligation-generated neuropathic states. In naïve animals, microglia and astrocytes expressed DPP4 protein with one and two orders of magnitude higher than neurons, respectively. DPP4 significantly increased in astrocytes during inflammation and in microglia in neuropathy. Intrathecal application of two DPP4 inhibitors tripeptide isoleucin-prolin-isoleucin (IPI) and the antidiabetic drug vildagliptin resulted in robust opioid-dependent antihyperalgesic effect during inflammation, and milder but significant opioid-independent antihyperalgesic action in the neuropathic model. The opioid-mediated antihyperalgesic effect of IPI was exclusively related to mu-opioid receptors, while vildagliptin affected mainly delta-receptor activity, although mu- and kappa-receptors were also involved. None of the inhibitors influenced allodynia. Our results suggest pathology and glia-type specific changes of DPP4 activity in the spinal cord, which contribute to the development and maintenance of hyperalgesia and interact with endogenous opioid systems.
Asunto(s)
Dipeptidil Peptidasa 4/genética , Hiperalgesia/tratamiento farmacológico , Inflamación/tratamiento farmacológico , Neuralgia/tratamiento farmacológico , Analgésicos Opioides/administración & dosificación , Animales , Astrocitos/efectos de los fármacos , Linaje de la Célula/genética , Inhibidores de la Dipeptidil-Peptidasa IV/administración & dosificación , Hiperalgesia/genética , Hiperalgesia/patología , Inflamación/genética , Inflamación/patología , Masculino , Neuralgia/genética , Neuralgia/patología , Neuroglía/efectos de los fármacos , Ratas , Ratas Sprague-Dawley , Ratas Wistar , Receptores Opioides kappa/genética , Receptores Opioides mu , Médula Espinal/efectos de los fármacos , Médula Espinal/patologíaRESUMEN
The antinociceptive action of botulinum toxin type A (BoNT/A) has been demonstrated in behavioral animal studies and clinical settings. It was shown that this effect is associated with toxin activity in CNS, however, the mechanism is not fully understood. Substance P (SP) is one of the dominant neurotransmitters in primary afferent neurons transmitting pain and itch. Thus, here we examined association of SP-mediated transmission and BoNT/A antinociceptive action by employing gene knockouts. Antinociceptive activity of intraplantarly (i.pl.) injected BoNT/A was examined in mice lacking the gene encoding for SP/neurokinin A (tac1-/-) or SP-preferred receptor neurokinin 1 (tac1r-/-), compared to control C57Bl/6J wild type animals. BoNT/A action was assessed in inflammatory pain induced by formalin and CFA, and neuropathic pain induced by partial sciatic nerve ligation. BoNT/A activity in CNS was examined by c-Fos and BoNT/A-cleaved SNAP-25 immunohistochemistry. In wild type mice, acute (formalin-evoked) and chronic pain (neuropathic and inflammatory) was reduced by peripherally injected BoNT/A. In tac1-/- and tac1r-/- knockout mice, BoNT/A exerted no analgesic effect. In control animals BoNT/A reduced the formalin-evoked c-Fos expression in lumbar dorsal horn, while in knockout mice the c-Fos expression was not reduced. After peripheral toxin injection, cleaved SNAP-25 occurred in lumbar dorsal horn in all animal genotypes. BoNT/A antinociceptive activity is absent in animals lacking the SP and neurokinin 1 receptor encoding genes, in spite of presence of toxin's enzymatic activity in central sensory regions. Thus, we conclude that the integrity of SP-ergic system is necessary for the antinociceptive activity of BoNT/A.
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Toxinas Botulínicas Tipo A/uso terapéutico , Inflamación/tratamiento farmacológico , Neurotoxinas/uso terapéutico , Dolor/tratamiento farmacológico , Receptores de Neuroquinina-1/deficiencia , Sustancia P/metabolismo , Animales , Modelos Animales de Enfermedad , Formaldehído/toxicidad , Adyuvante de Freund/toxicidad , Lateralidad Funcional , Regulación de la Expresión Génica/genética , Hiperalgesia/tratamiento farmacológico , Hiperalgesia/etiología , Inflamación/inducido químicamente , Inflamación/complicaciones , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Dolor/inducido químicamente , Dolor/complicaciones , Fosfopiruvato Hidratasa/metabolismo , Receptores de Neuroquinina-1/genética , Sustancia P/genética , Proteína 25 Asociada a Sinaptosomas/metabolismo , Factores de TiempoRESUMEN
Semicarbazide-sensitive amine oxidase (SSAO) catalyses oxidative deamination of primary amines. Since there is no data about its function in pain and arthritis mechanisms, we investigated the effects of our novel SSAO inhibitor SzV-1287 in chronic mouse models of joint inflammation. Effects of SzV-1287 (20 mg/kg i.p./day) were investigated in the K/BxN serum-transfer and complete Freund's adjuvant (CFA)-evoked active immunization models compared to the reference SSAO inhibitor LJP-1207. Mechanonociception was assessed by aesthesiometry, oedema by plethysmometry, clinical severity by scoring, joint function by grid test, myeloperoxidase activity by luminescence, vascular leakage by fluorescence in vivo imaging, histopathological changes by semiquantitative evaluation, and cytokines by Luminex assay. SzV-1287 significantly inhibited hyperalgesia and oedema in both models. Plasma leakage and keratinocyte chemoattractant production in the tibiotarsal joint, but not myeloperoxidase activity was significantly reduced by SzV-1287 in K/BxN-arthritis. SzV-1287 did not influence vascular and cellular mechanisms in CFA-arthritis, but significantly decreased histopathological alterations. There was no difference in the anti-hyperalgesic and anti-inflammatory actions of SzV-1287 and LJP-1207, but only SzV-1287 decreased CFA-induced tissue damage. Unlike SzV-1287, LJP-1207 induced cartilage destruction, which was confirmed in vitro. SzV-1287 exerts potent analgesic and anti-inflammatory actions in chronic arthritis models of distinct mechanisms, without inducing cartilage damage.
Asunto(s)
Amina Oxidasa (conteniendo Cobre)/antagonistas & inhibidores , Analgésicos no Narcóticos/uso terapéutico , Antiinflamatorios/uso terapéutico , Artritis/tratamiento farmacológico , Hidrazinas/uso terapéutico , Articulaciones/patología , Oxazoles/uso terapéutico , Oximas/uso terapéutico , Animales , Células Cultivadas , Enfermedad Crónica , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Adyuvante de Freund/inmunología , Humanos , Hidrazinas/farmacología , Articulaciones/efectos de los fármacos , Ratones , Ratones Endogámicos , Ratones Transgénicos , Oxazoles/farmacología , Oximas/farmacologíaRESUMEN
Somatostatin regulates stress-related behavior and its expression is altered in mood disorders. However, little is known about the underlying mechanisms, especially about the importance of its receptors (sst1-sst5) in anxiety and depression-like behavior. Here we analyzed the potential role of sst4 receptor in these processes, since sst4 is present in stress-related brain regions, but there are no data about its functional relevance. Genetic deletion of sst4 (Sstr4(-/-)) and its pharmacological activation with the newly developed selective non-peptide agonist J-2156 were used. Anxiety was examined in the elevated plus maze (EPM) and depression-like behavior in the forced swim (FST) and tail suspension tests (TST). Neuronal activation during the TST was monitored by Fos immunohistochemistry, receptor expression was identified by sst4(LacZ) immunostaining in several brain regions. Sstr4(-/-) mice showed increased anxiety in the EPM and enhanced depression-like behavior in the FST. J-2156 (100 µg/kg i.p.) exhibited anxiolytic effect in the EPM and decreased immobility in the TST. J-2156 alone did not influence Fos immunoreactivity in intact mice, but significantly increased the stress-induced Fos response in the dorsal raphe nucleus, central projecting Edinger-Westphal nucleus, periaqueductal gray matter, the magnocellular, but not the parvocellular part of the hypothalamic paraventricular nucleus, lateral septum, bed nucleus of the stria terminalis and the amygdala. Notably, sst4(LacZ) immunoreactivity occurred in the central and basolateral amygdala. Together, these studies reveal that sst4 mediates anxiolytic and antidepressant-like effects by enhancing the stress-responsiveness of several brain regions with special emphasis on the amygdala.
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Antidepresivos/uso terapéutico , Ansiedad/metabolismo , Butanos/uso terapéutico , Depresión/metabolismo , Naftalenos/uso terapéutico , Receptores de Somatostatina/metabolismo , Sulfonas/uso terapéutico , Análisis de Varianza , Animales , Antidepresivos/farmacología , Ansiedad/tratamiento farmacológico , Ansiedad/genética , Ansiedad/patología , Encéfalo/efectos de los fármacos , Encéfalo/metabolismo , Butanos/farmacología , Depresión/tratamiento farmacológico , Depresión/genética , Depresión/patología , Modelos Animales de Enfermedad , Conducta Exploratoria/efectos de los fármacos , Conducta Exploratoria/fisiología , Regulación de la Expresión Génica/efectos de los fármacos , Regulación de la Expresión Génica/genética , Suspensión Trasera , Aprendizaje por Laberinto/efectos de los fármacos , Aprendizaje por Laberinto/fisiología , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Naftalenos/farmacología , Proteínas Oncogénicas v-fos/metabolismo , Receptores de Somatostatina/genética , Sulfonas/farmacología , Natación/psicologíaRESUMEN
AIMS: To investigate the roles of TRPV1 and TRPA1 channels in baseline and allyl isothiocyanate (AITC)-evoked nociceptive responses by comparing wild-type and gene-deficient mice. MAIN METHODS: In contrast to conventional methods of thermonociception measuring reflex latencies, we used our novel methods to determine the noxious heat threshold. KEY FINDINGS: It was revealed that the heat threshold of the tail measured by an increasing-temperature water bath is significantly higher in TRPV1(-/-), but not TRPA1(-/-), mice compared to respective wild-types. There was no difference between the noxious heat thresholds of the hind paw as measured by an increasing-temperature hot plate in TRPV1(-/-), TRPA1(-/-) and the corresponding wild-type mice. The withdrawal latency of the tail from 0°C water was prolonged in TRPA1(-/-), but not TRPV1(-/-), mice compared to respective wild-types. In wild-type animals, dipping the tail or paw into 1% AITC induced an 8-14°C drop of the noxious heat threshold (heat allodynia) of both the tail and paw, and 40-50% drop of the mechanonociceptive threshold (mechanical allodynia) of the paw measured by dynamic plantar esthesiometry. These AITC-evoked responses were diminished in TRPV1(-/-), but not TRPA1(-/-), mice. Tail withdrawal latency to 1% AITC was significantly prolonged in both gene-deleted strains. SIGNIFICANCE: Different heat sensors determine the noxious heat threshold in distinct areas: a pivotal role for TRPV1 on the tail is contrasted with no involvement of either TRPV1 or TRPA1 on the hind paw. Noxious heat threshold measurement appears appropriate for preclinical screening of TRP channel ligands as novel analgesics.