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Objective:To evaluate the role of transient receptor potential vanillic acid 4 (TRPV4) in dexmedetomidine-induced improvement in cognitive function in mice with mechanical ventilator-caused brain injury.Methods:Ninety clean-grade healthy male C57BL6 mice, weighing 20-25 g, aged 8-12 weeks, were divided into 5 groups ( n=18 each) using a random number table method: control group (group C), mechanical ventilation group (group V), HC-067047 group (group H), dexmedetomidine group (group D), and dexmedetomidine+ GSK1016790A group (group DG). In group C, the animals breathed air spontaneously for 6 h without mechanical ventilation. In group V, the animals were mechanically ventilated for 6 h. In group H, TRPV4 blocker HC-067047 10 mmol was injected into the cerebral ventricle at 3 and 6 h of mechanical ventilation. In D and DG groups, dexmedetomidine 50 μg/kg was intraperitoneally injected at 30 min before mechanical ventilation. In group DG, TRPV4 agonist GSK1016790A 5 μmol was injected into the cerebral ventricle at 60 min before mechanical ventilation. Morris water maze test was performed on 6 mice in each group at 1 day before mechanical ventilation and 3 and 7 days after mechanical ventilation. Six mice in each group were randomly selected and sacrificed at 1 day after mechanical ventilation, and the brain tissue was taken for determination of the neuronal apoptosis in hippocampal CA1 area by TUNEL method, and the apoptosis index was calculated. Six mice in each group were randomly selected and sacrificed at 1 day after mechanical ventilation, and the hippocampal tissues were taken for determination of the expression of TRPV4, serine-threonine protein kinase (Akt), phosphorylated Akt (p-Akt), Bcl-2, Bax and caspase-3 by Western blot. Results:Compared with group C, the escape latency was significantly prolonged and the number of crossing the original platform was reduced at 3 and 7 days after mechanical ventilation, the expression of TRPV4 and caspase-3 was up-regulated, the ratio of Bcl-2/Bax was decreased, and the apoptosis index of neurons was increased in group V and group DG ( P<0.05). Compared with group V, the escape latency was significantly shortened and the number of crossing the original platform was increased at 3 and 7 days after mechanical ventilation, the expression of TRPV4 and caspase-3 was down-regulated, the expression of p-Akt was up-regulated, the ratio of Bcl-2/Bax was increased, and the apoptosis index of neurons was decreased in group D and group H ( P<0.05). Compared with group D, the escape latency was significantly prolonged at 3 and 7 days after mechanical ventilation, the number of crossing the original platform was reduced, the expression of TRPV4 and caspase-3 was up-regulated, the expression of p-Akt was down-regulated, the ratio of Bcl-2/Bax was decreased, and the apoptosis index of neurons was increased in group DG ( P<0.05). Conclusions:TRPV 4 is involved in dexmedetomidine-induced improvement in cognitive function, which is related to up-regulation of p-Akt expression and inhibition of apoptosis in hippocampal neurons in mice with mechanical ventilation-caused brain injury.
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Objectives: As neuropathy predominates vasculopathy, predicting functional deterioration of autonomic neurovascular dysfunction is essential to reduce diabetic foot ulcers. The present study has evaluated the possibility of stimulating the TRPV1 receptors of the small fibres using topical capsaicin to assess diabetic neuropathy in the dorsum of the foot functionally. Materials and Methods: A prospective cross-sectional study was carried out on ten healthy volunteers and 20 diabetic patients after receiving ethical approval. The subjects underwent vascular Doppler analysis after giving written agreement followed by monofilament testing. Then, topical capsaicin was applied to measure the local autonomic neurovascular reaction. With the use of an infrared-based digital instrument that was specially created, the vasodilation and proportional increase in temperature brought on by the application of capsaicin were quantified. Results: The percentage change in the local temperature in the control group varied from 0.478 to 3.315 compared to the diabetic group, which varied from 1.862 to ?3.932. There is a statistically significant difference in the mean of the two groups (P = 0.006) at a 95% confidence interval. Conclusion: This study suggests that TRPV1 receptor stimulation using capsaicin and resultant vasodilation monitored by the increase in local temperature can be used as a quantitative predictor of the early small fibre neuropathy in Distal Symmetric Polyneuropathy before the patient ends up with diabetic foot ulcer.
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Objective:To evaluate the role of transient receptor potential vanilloid receptor 1 (TRPV1)/nuclear factor-κB (NF-κB) signaling pathway in dexmedetomidine-induced alleviation of ventilator-induced lung injury (VILI) in rats.Methods:One hundred clean-grade healthy male Sprague-Dawley rats, weighing 270-320 g, aged 4-5 months, were divided into 5 groups ( n=20 each) using a random number table method: control group (group C), VILI group (group V), AMG9810 group (group A), dexmedetomidine group (group D), and dexmedetomidine + RTX group (group DR). VILI model was prepared by mechanical ventilation with a tidal volume of 40 ml/kg for 4 h. In group A, TRPV1 inhibitor AMG9810 30 mg/kg was intraperitoneally injected at 1 h before mechanical ventilation.Dexmedetomidine 5.0 μg/kg was intravenously infused at 20 min before mechanical ventilation, and dexmedetomidine was intravenously infused at the rate of 5.0 μ g·kg -1·h -1 during ventilation in group D and group DR.In group DR, RTX 70 μ g/kg was intraperitoneally injected for 3 consecutive days before mechanical ventilation.At 4 h of mechanical ventilation, the concentrations of interleukin-1beta (IL-1β), tumor necrosis factor-alpha (TNF-α) and IL-6 in bronchoalveolar lavage fluid (BALF) were detected, oxygenation index (OI) and wet/dry lung weight (W/D) ratio were measured, the histopathological changes of lung tissues were observed, and lung injury was assessed and scored.The expression of TRPV1 and NF-κB in lung tissues was detected by Western blot, and real-time polymerase chain reaction was used to detect the expression of TRPV1 and NF-κB mRNA. Results:Compared with group C, the concentrations of IL-1β, TNF-α and IL-6 in BALF were significantly increased, OI was decreased, the W/D ratio and lung injury scores were increased, and the expression of TRPV1 and NF-κB protein and mRNA was up-regulated in group V ( P<0.05). Compared with group V, the concentrations of IL-1β, TNF-α and IL-6 in BALF were significantly decreased, OI was increased, the W/D ratio and lung injury scores were decreased, and the expression of TRPV1 and NF-κB protein and mRNA was down-regulated in A, D and DR groups ( P<0.05). Compared with group D, the concentrations of IL-1β, TNF-α and IL-6 in BALF were significantly increased, OI was decreased, the W/D ratio and lung injury scores were increased, and the expression of TRPV1 and NF-κB protein and mRNA was up-regulated in group DR ( P<0.05). Conclusions:The mechanism by which dexmedetomidine alleviates VILI is partially related to inhibition of the activation of TRPV1/NF-κB signaling pathway and inhibition of the inflammatory responses in lung tissues of rats.
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Objective To evaluate the role of transient receptor potential vanillic acid subtype 1 (TRPV1)/calcitonin gene-related peptide (CGRP) signaling pathway in lidocaine postconditioning-induced reduction of myocardial ischemia-reperfusion (I/R) injury in rats.Methods Forty healthy male SpragueDawley rats,aged 3 months,weighing 250-300 g,were divided into 5 groups (n=8 each) using a random number table method:sham operation group (group Sham),group I/R,lidocaine postconditioning group (group LP),lidocaine postconditioning plus CGRP8-37 group (group LP+CGRP8-37),and lidocaine postconditioning plus capsazepine group (group LP+Capz).Myocardial ischemia was induced by ligating the anterior descending branch of left coronary artery for 30 min,followed by 120-min repeRFusion in anesthetized rats.Lidocaine 2 mg/kg was injected via the tail vein at 5 min before reperfusion in group LP.In group LP + CGRP8-37,lidocaine was intravenously injected,and CGRP receptor selective antagonist CGRP8-37 2 mg/kg was intravenously injected at the same time.In group LP+Capz,lidocaine was injected intravenously,and TRPV1 blocker capsazepine 3 mg/kg was injected intravenously at the same time.A catheter was retrogradely implanted to the left ventricle,and heart rate (HR),left ventricular systolic pressure (LVSP),left ventricular end-diastolic pressure (LVEDP),and the maximum rate of increase or decrease in left ventricular pressure (±dp/dtmax) were continuously monitored and recorded.Blood samples were collected from the carotid artery at 120 min of reperfusion for determination of cardiac troponin I (cTnI),myoglobin (Myo) and creatine kinase-MB (CK-MB) concentrations in serum.Rats were then sacrificed for determination of myocardial infarct size.Results Compared with Sham group,the serum concentrations of cTnI,Myo and CK-MB were significantly increased,LVSP,+dP/dtmax and HR were decreased,and LVEDP and-dP/dtmax were increased in I/R group and LP group (P＜0.05).Compared with group I/R,the serum concentrations of cTnI,Myo and CK-MB and myocardial infarct size were significantly decreased,LVSP and +dP/dtmax were increased,and LVEDP,-dP/dtmax and HR were decreased in group LP (P＜0.05),and no significant change was found in the parameters mentioned above in group LP+ CGRP8-37 and group LP+Capz (P＞0.05).Compared with group LP,the serum concentrations of cTnI and Myo,myocardial infarct size and LVEDP were significantly increased,and + dP/dtmax was decreased in group LP+CGRP8-37,and the serum concentrations of cTnI and Myo and myocardial infarct size were significantly increased,LVSP and +dP/dtmax were decreased,and LVEDP was increased in group I/R+Lido+Capz (P＜0.05).Conclusion The mechanism by which lidocaine postconditioning mitigates myocardial I/R injury is related to activating TRPV1/CGRP signaling pathway in rats.
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PURPOSE: To evaluate whether mild chemical irritation of the bladder in neonatal rats is associated with persistent vanilloid receptor transient receptor potential vanilloid subfamily 1 (TRPV1) activity in adult rats. METHODS: Female Sprague-Dawley rats were used. Ten-day-old rat pups underwent bladder sensitization via intravesical infusion of 0.2% acetic acid in saline with or without prior bladder desensitization with capsaicin. After 8 weeks, 3 groups of rats (control [group 1], bladder sensitization [group 2], and bladder desensitization [group 3]) underwent cystometry. Inflammation of bladder tissue and the expression of TRPV1 in bladder tissue and dorsal root ganglia (DRG) were also evaluated. RESULTS: The bladder sensitization group showed more frequent voiding contractions. TRPV1 expression in adult bladder tissue was elevated in group 2. TRPV1 mRNA levels in the bladder and DRG were significantly higher in group 2 than in group 1. Moreover, group 2 had significantly more DRG neurons (identified by uptake of the retrograde label Fast Blue) that exhibited TRPV1 immunoreactivity. CONCLUSIONS: We found a significant association between neonatal bladder sensitization and persistent TRPV1 activity in adult rats. This is the first study to focus on the underlying pathogenesis of bladder overactivity from childhood to adulthood. Our findings could lead to the development of new strategies for the treatment and prevention of adult urinary symptoms arising from childhood urinary tract dysfunction.
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Adult , Animals , Female , Humans , Rats , Acetic Acid , Capsaicin , Cystitis, Interstitial , Diagnosis-Related Groups , Ganglia, Spinal , Inflammation , Neurons , Rats, Sprague-Dawley , RNA, Messenger , TRPV Cation Channels , Urinary Bladder , Urinary Tract , Urinary Tract InfectionsABSTRACT
Objective@#To observe the effect of the expressive or functional blockage of TRPV1 on nerve regeneration after sciatic trans-section injury.@*Methods@#AMG-517, a kind of TRPV1 inhibitor, was injected into the surrounding area of the ipsilateral lumbar dorsal root ganglia while unilateral sciatic nerve was transected. A total of 24 healthy male Sprague-Dawley rats were divided into 4 groups: control group, injury only group, injury+ AMG-517 150 μg/kg group, injury+ AMG-517 300 μg/kg group. The injury only group was injected the same volume of medium. The release of CGRP from dorsal-horn of spinal cord, the number of axons at proximal stem of sciatic nerve after transection, and the expression of TRPV1 in dorsal root ganglion were detected using the methods of ELISA, Western blot and semi-thin section (1 μm)- toluidine blue staining 2 weeks after injury.@*Results@#The release of CGRP in lumbar spinal dorsal horn was obviously decreased after AMG-517 treatment, which was the evidence of TRPV1 functional inhibition. CGRP in the control group was 0.15 ng/g, the injury only group 0.17 ng/g, AMG-517 150 μg/kg group 0.09 ng/g, and AMG-517 300 μg/kg group 0.11 ng/g(P<0.01). The number of axons which were myelinated or unmyelinated increased after the TRPV1 was inhibited by AMG-517(P<0.01). In addition, the injection of AMG-517 into surrounding dorsal root ganglion decreased the expression of TRPV1 in dorsal root ganglion(P<0.01).@*Conclusions@#Over expression or activation of TRPV1 after periphery nerve injury has negative effect on nerve regeneration in fact; Inhibiting the over-expression or over-activation of TRPV1 after nerve injury facilitates axonal regeneration and nerve repair.
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Objective To evaluate the changes in the expression of transient receptor potential vanilloid 1 (TRPV1) in dorsal root ganglions (DRGs) during remifentanil-induced hyperalgesia in the rats with incisional pain.Methods Thirty-two SPF healthy male Sprague-Dawley rats,weighing 240-260 g,aged 2-3 months,in which caudal catheters were successfully implanted,were divided into 4 groups (n=8 each) using a random number table:control group (group C),incisional pain group (group Ⅰ),remifentanil group (group R),and incisional pain + remifentanil group (group I+R).A 1 cm longitudinal incision was made through skin,fascia and muscle of the plantar aspect of the left hindpaw to establish the model of incisional pain.In group R,remifentanil was intravenously infused for 60 min at a rate of 1.2 μg · kg-1 · min-1.In group Ⅰ,the model of incisional pain was established,and the equal volume of normal saline was intravenously infused for 60 min at the same time.In group I+R,the model of incisional pain was established,and remifentanil was intravenously infused for 60 min at a rate of 1.2 μg · kg-1 · min-1 at the same time.In group C,the equal volume of normal saline was intravenously infused for 60 min.The mechanical paw withdrawal threshold (MWT) and thermal paw withdrawl latency (TWL) were measured at 24 h before normal saline or remifentanil infusion (To) and 2,6,24 and 48 h after the end of infusion (T1-4).The rats were sacrificed after the last measurement of pain threshold,and the DRGs of the lumbar segment (L4-6) were removed for determination of the expression of TRPV1 protein and mRNA by Western blot and real-time polymerase chain reaction,respectively.Results Compared with group C,the MWT was significantly decreased,and the TWL was shortened at T1-4,and the expression of TRPV1 protein and mRNA was up-regulated in R,I and I+R groups (P＜0.05).Compared with group R or group I,the MWT was significantly decreased,and the TWL was shortened at T1-4,and the expression of TRPV1 protein and mRNA was up-regulated in group I+R (P＜0.05) Conclusion Up-regulated expression of TRPV1 in DRGs may be involved in the mechanism underlying remifentanil-induced hyperalgesia in the rats with incisional pain.
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Objective To investigate the relationship between the expression of trannsient receptor potential vanilloid 1 (TRPV1) and the severity of airway remodeling in elderly patients with chronic obstructive pulmonary disease(COPD).Methods According to airflow obstruction severity,totally 100 cases of elderly patients with COPD were divided into chronic obstructive pulmonary disease Global Initiative(gold) grade 1 in 23 cases,24 cases of grade GOLD2,GOLD3 27 cases,GOLD4 26 cases,respectively.The TRPV1 concentrations in induced sputum supernatant and serum from each level of elderly patients with COPD as well as in 50 cases of healthy old people were analyzed.Results TRPV1 concentrations in serum and induced sputum in the COPD group was significantly increased compared with the healthy elderly group[(9.94±2.91)μg/L vs.(3.68±0.46)μg/L,(3.29± 1.32)μg/L vs.(0.70 ± 0.30)μg/L] (P ＜ 0.01).The serum and induced sputum TRPV1 concentrations in the mutual pairwise comparison between the elderly COPD patients with all levels had statistical difference (P ＜ 0.01).Conclusion The expression of TRPV1 protein become increased with the severity of airway remodeling in elderly patients with chronic obstructive pulmonary disease.
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Objective To investigate the expression of transient receptor potential vanilloid subtype 1 (TRPV1) in uterosacral ligament and its correlation with pain in endometriosis.Methods Total of 54 patients undergoing endometriotic lesions excision in uteroscaral ligament by laparoscopy due to pelvic pain were enrolled in this study.According to visual analogue scale(VAS) scores,27 patients with VAS 5 - 10 were in group A and 27 patients with VAS 0 - 4 were in group B.In the mean time,20 patients with dysmenorrhea without endometriosis (VAS:0 -4 ) were matched as group C.Specimens (including the sacro-ligaments of 20 women without endometriosis) were immunostained with specific antibodies of TRPV1.Western blot and real time PCR were performed to detect TRPV1 expression in endometriosis lesions and control group.Results( 1 ) Immunohistochemnistry:the positive area of TRPV1 was found in endometriotic lesions in uterosacral ligament in group A,B and tissue of uterosacral ligament group C.The semi-qualitification of TRPV1 expression were 3 in group A, 1 in group B and 1 in group C by immunohistochemistry staining.There was significantly different expression between group B and group A ( P =0.005 ) or group C ( P =0.027 ).(2) mRNA expression:the expression of TRPV 1 was 1.84 in group A,0.80 in group B,0.24 in group C,respectively.With higher VAS scores,the expression of TRPV1 exhibited increasing trends.The expression of TRPV1 mRNA was higher in group A than thai in group B ( P =0.022).There was statistically different expression between group B and group C ( P =0.031 ).( 3 ) Western blot:the expression of TRPV1 protein was 0.63 in group A,0.19 in group B,0.02 in group C.There was significant differences between group A and group B ( P =0.022 ),and between group B and group C (P ＜ 0.01 ).Conclusion The expression of TRPV1 was correlated with the degree of pain in patient with endometriosis.
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Objective To investigate the effects of the transient receptor potential V6 (TRPV6) gene silencing on the proliferation and apoptosis of trophoblasts HTR-8/SVneo cells.Methods siRNA sequences targeting the TRPV6 gene were constructed and then transfected into HTR-8/SVneo cells mediated by liposome.The cells were divided three groups,including blank control (add the reagent of transfenction),negative control groups (transfecting nonspecific siRNA) and experimental groups (transfecting TRPV6-siRNA).Those cells in every group were collected at 24,48,72 hours after transfecting.The expression levels of TRPV6 mRNA were detected by reverse transcription (RT) PCR at different times after transfecting.The effects of siRNA on the proliferation and apoptosis of the cells were assayed by methyl thiagolyl tetragolium (MTT) and flow cytometry at different times after transfecting.Results siRNA TRPV6 transfection could inhibit the expression of TRPV6 mRNA in the HTR-8/SVneo cells.The expression was decreased with the extension of time,by 0.72 ± 0.02,0.54 ± 0.02 and 0.29 ± 0.01 after 12,48 and 72 hours of siRNA transfection as compared with the blank control and the negative control groups (P ＜0.01).The rates of proliferation inhibition were (19.29 ± 1.23) ％,(32.12 ± 1.35) ％ and (46.51 ±1.42) ％ at 24,48 and 72 hours respectively when compared with the blank control (2.12 ± 0.03)％,(2.42 ± 0.02) ％,(3.13 ± 0.04) ％ and the negative control groups (2.37 ± 0.01) ％,(2.61 ± 0.05) ％,(2.93 ± 0.03) ％ (P ＜ 0.01).The apoptosis rates of HTR-8/SVneo cells was 16.21％ at 48 hours after transfected with siRNA TRPV6,which were significantly higher than 3.27％ in the blank control and 5.34％ in the negative control groups (P ＜ 0.05).Conclusion Silenceing of TRPV6 genen could inhibit the proliferation and increase the apoptosis of extravillous trophoblas of human placenta.
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Objective To investigate the effects of intrathecal(IT)CX3 CR1 neutralizing antibody(antiFKR)on morphine tolerance in rats with bone cancer pain(BCP)and the unlerlying mechanism.Methods Forty-eight adult female SD rats aged 3 months weighing 180-200 g were randomized into 4 groups(n =12 each):group I sham operation(S); group Ⅱ BCP + normal saline(NS); group Ⅲ BCP + IgG(IgG)and group ⅣBCP + anti-FKR.Bone cancer pain(BCP)was induced by injecting Walker 256 cancer cells 10 μl(400 cells/ μl)into the medullary cavity of right tibia in groups Ⅱ,Ⅲ and Ⅳ.Ten days later morphine 20 μg/kg was administered IT twice a day for 7 consecutive days.Starting from the 8th day NS,IgG and anti-KFR 10 μl was administered IT once a day for 3 consecutive days in groups Ⅱ,Ⅲ and ⅣⅣ respectively.Paw withdrawal threshold to yon Frey filament stimulation(MWT)and paw withdrawal duration(MWD)were determined bcfore(To,baseline)and at 3,6,9 day after intra-tibial cancer cell inoculation(T1.2,3),on the 3rd and 7th day of IT morphine(T4.5)and on the 3rd day of IT NS/lgG/anti-KFR(T6).The animals were killed at T6 after last pain behavior assessment.The lumbar segment(L4-6)of the spinal cord was removed for determination of the expression of CX3 CR1 protein(by Western blot),μ-opioid receptor and TRPV1 receptor(by immuno-histochemistry)in the dorsal horn of spinal cord.Results IT morphine significantly eased BCP at T4,but morphine analgesia was significantly reduced on the 7th day of IT morphine in the 3 groups indicating morphine tolerance which was significantly relieved by anti-KFR in group Ⅳ.IT anti-KFR significantly down-regulated CX3CR1 prolein and TRPVI receptor expression and up-regulated μ-opioid receptor in group Ⅳ as compared with IT NS and lgG in groups Ⅱ and Ⅲ.Conctusion IT anti-KFR can relieve morphine tolerance in the rats with bone cancer pain by up-regulating μ-opioid receptor and down-regulating CX3 CR1 protein and TRPVI receptor expression.
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The transient receptor potential vanilloid 4 (TRPV4) cation channel, a member of the TRP vanilloid subfamily, is expressed in a broad range of tissues where it participates in the generation of Ca2+ signals and/or depolarization of the membrane potential. Regulation of TRPV4 abundance at the cell surface is critical for osmo- and mechanotransduction. Defects in TRPV4 are the cause of several human diseases, including brachyolmia type 3 (MIM:113500) (also known as brachyrachia or spondylometaphyseal dysplasia Kozlowski type [MIM:118452]), and metatropic dysplasia (MIM:156530) (also called metatropic dwarfism or parastremmatic dwarfism [MIM:168400]). These bone dysplasia mutants are characterized by severe dwarfism, kyphoscoliosis, distortion and bowing of the extremities, and contractures of the large joints. These diseases are characterized by a combination of decreased bone density, bowing of the long bones, platyspondyly, and striking irregularities of endochondral ossification with areas of calcific stippling and streaking in radiolucent epiphyses, metaphyses, and apophyses. In this review, we discuss the potential effect of the mutation on the regulation of TRPV4 functions, which are related to human diseases through deviated function. In particular, we emphasize how the constitutive active TRPV4 mutant affects endochondral ossification with a reduced number of hypertrophic chondrocytes and the presence of cartilage islands within the zone of primary mineralization. In addition, we summarize current knowledge about the role of TRPV4 in the pathogenesis of several diseases.
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Humans , Mutation , Osteochondrodysplasias/genetics , Osteogenesis/genetics , TRPV Cation Channels/chemistryABSTRACT
To investigate the changes of peripheral pain threshold after knockout of transient receptor potential vanilloid l (TRPVl) gene. Methods Tail-Flick Analgesia Meterand von-Frey hair were used to determine the peripheral thermal and mechanical thresholds in TRPVl knockout and wild-type female mice, and the results of the two groups were compared. Results The Tail-Flick latency in TRPVl knockout mice was significantly prolonged after hot stimulation compared with that in the wild-type group([3.59±0.65] s vs [2.l9±0.24] s, P0.05). Conclusion It is suggested that TRPVl receptor mediate thermal stimuli response under physiological condition, and has no noticeable influence on mechanical stimuli response.
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Objective To investigate the changes of peripheral pain threshold after knockout of transient receptor potential vanilloid 1(TRPV1) gene. Methods Tail-Flick Analgesia Meterand von-Frey hair were used to determine the peripheral thermal and mechanical thresholds in TRPV1 knockout and wild-type femalemice, and the results of the two groups were compared. Results The Tail-Flick latency in TRPV1 knockout mice was significantly prolonged after hot stimulation compared with that in thewild-type group([3. 59±0. 65] s vs [2. 19±0. 24] s, P0. 05). Conclusion It is suggested that TRPV1 receptor mediate thermal stimuii response under physiological condition, and has no noticeable influence on mechanical stimuii response.
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Mast cells are activated by specific allergens and also by various nonspecific stimuli, which might induce physical urticaria. This study investigated the functional expression of temperature sensitive transient receptor potential vanilloid (TRPV) subfamily in the human mast cell line (HMC-1) using whole-cell patch clamp techniques. The temperature of perfusate was raised from room temperature (RT, 23~25degrees C to a moderately high temperature (MHT, 37~39degrees C to activate TRPV3/4, a high temperature (HT, 44~46degrees C to activate TRPV1, or a very high temperature (VHT, 53~55degrees C to activate TRPV2. The membrane conductance of HMC-1 was increased by MHT and HT in about 50% (21 of 40) of the tested cells, and the I/V curves showed weak outward rectification. VHT-induced current was 10-fold larger than those induced by MHT and HT. The application of the TRPV4 activator 4alpha-phorbol 12,13-didecanoate (4alphaPDD, 1microM) induced weakly outward rectifying currents similar to those induced by MHT. However, the TRPV3 agonist camphor or TRPV1 agonist capsaicin had no effect. RT-PCR analysis of HMC-1 demonstrated the expression of TRPV4 as well as potent expression of TRPV2. The [Ca2+]c of HMC-1 cells was also increased by MHT or by 4alphaPDD. In summary, our present study indicates that HMC-1 cells express Ca2+-permeable TRPV4 channels in addition to the previously reported expression of TRPV2 with a higher threshold of activating temperature.
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Humans , Allergens , Camphor , Capsaicin , Mast Cells , Membranes , Patch-Clamp Techniques , Phorbols , TRPV Cation Channels , UrticariaABSTRACT
Objective To investigate the changes in expression of vanilloid receptor subtype 1 (VR1) in dorsal root ganglion (DRG) and effect of electroacupuncture (EA)on morphine tolerance in rats with inflammatory pain (IP) and morphine tolerance. Methods Thirty 8 month old male SD rats in which intrathecal (IT) catheters were successfully implanted without complication were randomly divided into 6 groups ( n = 5 each): group A IP + normal saline (NS) 10 μl IT twice a day × 7 days;group B intact rats + morphine 10 μg/kg(10 μl )IT twice a day × 7 days;group C IP + morphine 10 μg/kg(10 μl) IT once;group DIP + morphine 10μg/kg(10 μl) IT twice a day × 7 days;group E and F IP + EA (frequency 2/15 Hz) + morphine 10μg/kg(10 μl) IT twice a day × 7 days. IP was induced by injecting complete Freund's adjuvant (CFA) into the anlle joint of the left hindlimb. IT morphine or NS was started on the 4th day after induction of IP. EA of Yanglingquan and Zusanli lasting 30 min was performed once a day after first IT administration of morphine for 7 days. Paw withdrawal latency (PWL) to a thermal nociceptive stimulus was measured before induction of IP (baseline), at 1 day before and 1-7days of consecutive IT administration. The animals were sacrificed after last PWL measurement. The DRGs of the lumbar segment (L4-6) were removed for determination of VR1 expression in total and membrane protein using Western blot analysis. Results There was no significant difference in the baseline PWL measured before induction of IP among the 6 groups. Morphine tolerance developed in group B and D but did not develop in group E and F.The expression of VR1 in total and membrane protein of DRG was highest in group D and was significantly lower in group E than in group F. Conclusion VR1 in DRG is involved in the development of morphine tolerance. EA can inhibit morphine tolerance by down-regulating the expression of VR1.
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Objective To investigate the change in the phosphorylation of vanilloid receptor 1 (VR1) in dorsal root ganglion (DRG) in rats with inflammatory pain (IP)-morphine tolerance. Methods Twenty 8-12 months old male SD rats in which intrathecal (IT) catheters were successfully implanted without complication were randomly divided into 4 groups (n =5 each): group NS, IP + normal saline (NS) 10 μl IT twice a day ×7 days; group M0 ,intact rats + morphine 10 μg/kg ( 10 μl) IT twice a day × 7 days; group M1 , IP + morphine 10 μg/kg (10 μl) IT once; group M2 ,IP + morphine 10 μg/kg(10 μl) IT twice a day × 7 days. IP was induced by injecting complete Freund's adjuvant (CFA) into the ankle joint of the left hindlimb. IT morphine or NS was started on the 3rd day after induction of IP. Paw withdrawal threshold (PWT) and paw withdrawal latency (PWL)were measured after catheterization, before and at 1-7 days of IT consecutive administration. The rats were sacrificed after last pain threshold measurement. The phosphorylated VR1 (p-VR1) protein expression in DRG was determined by Western blot. Results There was no significant difference in the baseline PWL measured before induction of IP among the 4 groups. Morphine tolerance developed in group M0 and M2. The expression of p-VR1 in DRG was highest in group M2. Conclusion The phosphorylation of VR1 in DRG in rats with IP tolerance is increased, which is involved in the development of morphine tolerance.
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Plasma calcium concentration is maintained within a narrow range (8.5-10.5 mg/dL) by the coordinated action of parathyroid hormone (PTH), 1,25(OH)2D3, calcitonin, and ionized calcium (iCa2+) itself. The kidney plays a key role in this process by the fine regulation of calcium excretion. More than 95% of filtered calcium is reabsorbed along the renal tubules. In the proximal tubules, 60% of filtered calcium is reabsorbed by passive mechanisms. In the thick ascending limb, 15% of calcium is reabsorbed by paracellular diffusion through paracellin-1 (claudin-16). The calcium sensing receptor (CaSR) in the basolateral membrane of the thick ascending limb senses the change in iCa2+ and inhibits calcium reabsorption independent to PTH and 1,25(OH)2D3. The fine regulation of calcium excretion occurs in the distal convoluted tubules and connecting tubules despite the fact that only 10-15% of filtered calcium is reabsorbed there. Transient receptor potential vanilloid 5 (TRPV5) and 6 (TRPV6) in the apical membrane act as the main portal of entry, calbindin-D28K delivers Ca2+ in the cytoplasm, and then Na2+/Ca2+ exchanger (NCX1) and plasma membrane Ca2+-ATPase in the basolateral membrane serve as an exit. In the cortical collecting duct, TRPV6 is expressed, but the role might be negligible. In addition to PTH and 1,25(OH)2D3, acid-base disturbance, diuretics, and estrogen affect on these calcium channels. Recently, klotho and fibroblast growth factor 23 (FGF23) are suggested as new players in the calcium metabolism. Klotho is exclusively expressed in the kidney and co-localized with TRPV5, NCX1, and calbindin-D28K. Klotho increases calcium reabsorption through trafficking of TRPV5 to the plasma membrane, and also converts FGF receptor to the specific FGF23 receptor. FGF23:klotho complex bound to FGF receptor inhibits 1alpha- hydroxylase of vitamin D, and contributes to calcium reabsorption and phosphate excretion in the kidney.
Subject(s)
Calcitonin , Calcium , Calcium Channels , S100 Calcium Binding Protein G , Cell Membrane , Cytoplasm , Diffusion , Diuretics , Estrogens , Extremities , Fibroblast Growth Factors , Homeostasis , Kidney , Membranes , Parathyroid Hormone , Plasma , Receptors, Calcium-Sensing , Receptors, Fibroblast Growth Factor , TRPV Cation Channels , Vitamin DABSTRACT
PURPOSE: Hypercalciuria is a risk factor of renal calcium stone and proper initial management is dietary salt restriction. But the molecular mechanism responsible for this sodium calcium relationship remains unclear. The present study investigates the relationship between different amount of sodium intake and the expression level of transporters involved in the active calcium transport. METHODS: Sprague-Dawley rats were randomized into three groups: a normal salt group, a low salt group, and a high salt group. Expression of mRNA and protein of transient receptor potential vanilloid (TRPV) 5 and calbindin-D28K was determined by real-time quantitative PCR and western blots, respectively. RESULTS: Hematocrit and body weight showed no difference among the three groups. High salt diet led to significant increase in the amount of urinary calcium excretion and decreased mRNA expression of calbindin-D28K and TRPV5. Protein abundance of calbindin-D28K and TRPV5 was decreased but the result was statistically insignificant. Low salt diet decreased the amount of urinary calcium excretion without significant difference. Messenger RNA expression and protein abundance of calbindin-D28K and TRPV5 showed no difference, compared to those of normal salt group. CONCLUSION: These data suggest that high sodium intake increases urinary calcium excretion, which is accompanied by a decreased expression of calbindin-D28K mRNA.