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Objective:To evaluate the effects of propofol on α-amino-3-hydroxy-5-methyl-4-isoxazole-propionate (AMPA) receptor expression in the hippocampus of neonatal rats.Methods:Eighty-four clean-grade healthy Sprague-Dawley rats of either sex, aged 7 days, weighing 14-18 g, were divided into 2 groups ( n=42 each) using a random number table method: control group (group C) and propofol group (group P). Propofol 30 mg/kg was intraperitoneally injected in group P, fat emulsion 3 mg/kg was intraperitoneally injected in group C, 1/2 of the initial dose was given at a 20 min interval, 3 times in total, for 3 consecutive days.The arterial blood samples were taken for blood gas analysis after administration on 1st day.The rats were sacrificed at 3, 7 and 28 days after the last administration of propofol, and the bilateral hippocampus was obtained for detection of the expression of AMPA receptors containing GluR1, GluR2 and GluR3 subunits in total and membrane protein (by Western blot), and the ratio of membrane protein to total protein (M/T) was calculated.The concentrations of free calcium ion were measured.The learning and memory ability was evaluated by Morris water maze test on 28 days after the last administration. Results:Compared with group C, the expression of AMPA receptor containing GluR1 subunit in total and membrane protein was significantly up-regulated, M/T was increased, the expression of AMPA receptor containing GluR2 subunit in total and membrane protein was down-regulated, and M/T was decreased at each time point ( P<0.05), no significant change was found in the expression of AMPA receptor containing GluR3 subunits ( P>0.05), the concentrations of free calcium ion in hippocampal cells were increased, and the escape latency was prolonged, the number of crossing the original platform was decreased, and the time of staying at the target quadrant was shortened at 2-4 days of training in group P ( P<0.05). Conclusion:The mechanism by which propofol reduces cognitive function is related to up-regulation of the expression of AMPA receptors containing GluR1 subunit in the hippocampus and down-regulation of the expression of AMPA receptors containing GluR2 subunits, which increases the concentration of free calcium ions in nerve cells of neonatal rats.
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Objective To evaluate the role of neuroligin 1 (NL-1) in trafficking of GluR1-containing AMPA receptor to cell membrane in spinal cord dorsal horns during remifentanil-induced hyperalgesia in mice with incisional pain.Methods Forty SPF healthy male C57BL/6J mice,aged 8-10 weeks,weighing 18-22 g,were divided into 5 groups (n=8 each) using a random number table method:control group (group C),NL1-shRNA plasmid group (group NL),incisional pain plus remifentanil group (group I+R),incisional pain plus remifentanil plus blank vector group (group I+R+B),and incisional pain plus remifentanil plus NL-1-shRNA plasmid group (group I+R+NL).Negative lentivirus was intrathecally injected in group I+R+B.In NL and I+R+NL groups,10 μl NL-1-shRNA lentivirus at 1×10s IFU/ml was intrathecally injected once a day for 3 consecutive days.Normal saline 10 μl was intrathecally injected at the same time point in C and I+R groups.After transfection was stable,normal saline 0.1 ml was injected via the caudal vein for 4 consecutive times at 15 min intervals in C and NL groups.In I+R,I+R+B and I+R+NL groups,0.1 ml remifentanil 10 μg/kg was injected via the caudal vein for 4 consecutive times at 15 min intervals,and the model of incisional pain was established after the first administration.The mechanical paw withdrawal threshold (MWT) and tail-flick latency (TFL) were measured at 24 h before normal saline or remifentanil administration (T0) and at 3,6,24 and 48 h after the end of administration (T1-4).The animals were sacrificed after measurement of pain threshold at T4,and L4-6 segments of the spinal dorsal horn were then collected for determination of the expression of NL-1 protein and mRNA and AMPA receptors,and the ratio of AMPA receptor expression in the membrane protein to that in the total protein (m/t ratio) was calculated.Results Compared with group C,the MWT was significantly decreased,and TFL was shortened at T1-4,the expression of NL-1 protein and mRNA and GluR1-containing AMPA receptors in membrane and total proteins was up-regulated,and m/t ratio was increased in I+R and I+R+B groups (P＜0.05).Compared with I+R and I+R+B groups,the MWT was significantly increased and TFL was prolonged at T1-4,the expression of NL-1 protein and mRNA and GluR1-containing AMPA receptors in membrane and total proteins was down-regulated,and m/t ratio was decreased in group I+R+NL (P＜0.05).Conclusion NL-1 in spinal cord dorsal horns can promote the trafficking of GluR1-containing AMPA receptors to cell membrane,which is involved in the development and maintenance of remifentanil-induced hyperalgesia in mice with incisional pain.
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Objective@#To evaluate the role of neuroligin 1 (NL-1) in trafficking of GluR1-containing AMPA receptor to cell membrane in spinal cord dorsal horns during remifentanil-induced hyperalgesia in mice with incisional pain.@*Methods@#Forty SPF healthy male C57BL/6J mice, aged 8-10 weeks, weighing 18-22 g, were divided into 5 groups (n=8 each) using a random number table method: control group (group C), NL1-shRNA plasmid group (group NL), incisional pain plus remifentanil group (group I+ R), incisional pain plus remifentanil plus blank vector group (group I+ R+ B), and incisional pain plus remifentanil plus NL-1-shRNA plasmid group (group I+ R+ NL). Negative lentivirus was intrathecally injected in group I+ R+ B.In NL and I+ R+ NL groups, 10 μl NL-1-shRNA lentivirus at 1×108 IFU/ml was intrathecally injected once a day for 3 consecutive days.Normal saline 10 μl was intrathecally injected at the same time point in C and I+ R groups.After transfection was stable, normal saline 0.1 ml was injected via the caudal vein for 4 consecutive times at 15 min intervals in C and NL groups.In I+ R, I+ R+ B and I+ R+ NL groups, 0.1 ml remifentanil 10 μg/kg was injected via the caudal vein for 4 consecutive times at 15 min intervals, and the model of incisional pain was established after the first administration.The mechanical paw withdrawal threshold (MWT) and tail-flick latency (TFL) were measured at 24 h before normal saline or remifentanil administration (T0) and at 3, 6, 24 and 48 h after the end of administration (T1-4). The animals were sacrificed after measurement of pain threshold at T4, and L4-6 segments of the spinal dorsal horn were then collected for determination of the expression of NL-1 protein and mRNA and AMPA receptors, and the ratio of AMPA receptor expression in the membrane protein to that in the total protein (m/t ratio) was calculated.@*Results@#Compared with group C, the MWT was significantly decreased, and TFL was shortened at T1-4, the expression of NL-1 protein and mRNA and GluR1-containing AMPA receptors in membrane and total proteins was up-regulated, and m/t ratio was increased in I+ R and I+ R+ B groups (P<0.05). Compared with I+ R and I+ R+ B groups, the MWT was significantly increased and TFL was prolonged at T1-4, the expression of NL-1 protein and mRNA and GluR1-containing AMPA receptors in membrane and total proteins was down-regulated, and m/t ratio was decreased in group I+ R+ NL (P<0.05).@*Conclusion@#NL-1 in spinal cord dorsal horns can promote the trafficking of GluR1-containing AMPA receptors to cell membrane, which is involved in the development and maintenance of remifentanil-induced hyperalgesia in mice with incisional pain.
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Objective To evaluate the effect of dexmedetomidine on the expression of NR1 subunit-containing NMDA receptors and GIuR2 subunit-containing AMPA receptors during hypoxic injury to rat hippocampal neurons.Methods The hippocampal neurons were isolated from Wistar rats within 24 h after birth and divided into 3 groups (n=24 each) using a random number table:control group (group C),hypoxia group (group H) and dexmedetomidine group (group D).The cells were subjected to hypoxia for 6 h to establish the model of neuronal hypoxic injury in H and D groups.In group D,0.1 μmol/L dexmedetomidine was added at 6 h of hypoxia and neurons were incubated for 3 h,and then the culture medium was replaced with a normal medium and neurons were incubated for 24 h.The neuronal viability was measured by CCK-8 assay,the leakage of LDH was detected,and the leakage rate was calculated.The expression of NR1 subunits-containing NMDA receptors and GluR2 subunits-containing AMPA receptors was detected by Western blot.The concentration of calcium ion in cytoplasm was measured using Fluo-3AM.Results Compared with group C,the neuronal viability was significantly decreased,the LDH leakage rate was increased,the expression of NR1 subunits-containing NMDA receptors in the membrane was up-regulated,the expression of GluR2 subunits-containing AMPA receptors was down-regulated,and the concentration of calcium ion in cytoplasm was increased in H and D groups (P＜0.05).Compared with group H,the neuronal viability was significantly increased,the LDH leakage rate was decreased,the expression of NR1 subunits-containing NMDA receptors in the membrane was down-regulated,the expression of GluR2 subunitscontaining AMPA receptors was up-regulated,and the concentration of calcium ion in cytoplasm was decreased in group D (P＜0.05).Conclusion The mechanism by which dexmedetomidine reduces hypoxic injury to rat hippocampal neurons may be related to inhibiting up-regulation of the expression of NR1 subunits-containing NMDA receptors in the membrane and down-regulation of the expression of GluR2 subunitscontaining AMPA receptors.
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Objective To evaluate the effect of sevoflurane on memory retrieval in mice and the role of hippocampal PSD95 and amino-3-hydroxyl-5-methyl-4-isoxazole-propionate (AMPA) receptor.Methods Sixty-four healthy pathogen-free Kuming mice of both sexes,aged 2-3 months,weighing 30-35 g,were divided into control group (n =32) and sevoflurane group (n =32) in a stratified randomized block design.The ability of memory retrieval was evaluated using dark avoidance test.After setting up the memory of dark avoidance,the mice inhaled 40％ oxygen for 2 h in control group and 3.3％ sevofluane in 40％ oxygen for 2 h in sevoflurane group.Dark avoidance test was performed at 12,24,48 and 72 h after the end of oxygen or sevoflurane inhalation (T1-4),and the test results and development of amnesia were recorded.The animals were sacrificed after behavior test at each time point,and brains were removed and cut into sections which were stained with haematoxylin and eosin for examination of the pathological changes of hippocampal CA1 area (under a light microscope) and for determination of the expression of PSD95 and AMPA receptors in hippocampi (using immunohistochemistry and Western blot).Results Compared with control group,the step-in latency and test results of error times were significantly decreased,the expression of PSD95 and AMPA receptors in hippocampus was down-regulated,and the incidence of amnesia was increased at T1 and T2 in sevoflurane group (P＜0.05).Compared with the basic results,no significant change was found in the step-in latency or test results of error times in control group (P＞0.05),and the step-in latency and test results of error times were significantly decreased at T1 and T2 in sevoflurane group (P＜0.05).Apoptosis in pyramidal cells was not found in sevoflurane group.Conclusion Sevoflurane can inhibit the ability of memory retrieval transiently in mice,and the mechanism is related to inhibiting the expression of hippocampal PSD95 and AMPA receptors.
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Objective To evaluate the changes in the expression of neuroligin1 in excitatory postsynaptic membrane of the spinal dorsal horn in a rat model of inc isional pain.Methods Forty-eight pathogen-free healthy adult male Sprague-Dawley rats,aged 6-8 weeks,weighing 280-320 g,were divided into control group (group C,n =12) and incisional pain group (group Ⅰ,n=36) using a random number table.A1 cm long incision was made in the plantar surface of the right hindpaw in group Ⅰ.Cumulative pain score (CPS) was assessed and mechanical paw withdrawal threshold to von Frey stimuli was measured at 3 hand 1 and 3 days after operation (T1,2,3).The animals were then sacrificed and their lumbar segments (L3-6) of the spinal cord were removed for detection of the expression of neuroligin1,postsynaptic density-95 protein (PSD-95),glutamate receptor 1 (GluR1) and GluR2 in the postsynaptic membrane of spinal dorsal horn (by Western blot) and co-expression of neuroligin1 with PSD-95 in spinal dorsal horn (by co-immuno-precipitation).Results Compared with group C,CPS was significantly increased and mechanical paw withdrawal threshold was decreased at T1-3,and the expression of neuroligin1 and GluR1 in the postsynaptic membrane of spinal dorsal horn at T1,2 and co-expression of neuroligin1 with PSD-95 at T1 were up-regulated in group Ⅰ (P＜0.01 or 0.05).Conclusion The development and maintenance of incisional pain is related to the signaling pathway regulated by neuroligin1 in excitatory postsynaptic membrane of the spinal dorsal horn of rats.
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Objective To evaluate the role of PICK1 in remifentanil-induced miniature excitatory postsynaptic currents (mEPSCs) mediated by AMPA receptors in spinal dorsal horn neurons and in the expression of AMPA receptors in juvenile rats.Methods Thirty-six male Sprague-Dawley rats,aged 14-18 days,weighing 50-60 g,were used in the study.Eighteen rats were randomly selected,their lumbar segments of the spinal cord were immediately removed,and 108 spinal cord slices (400 μm thick,for wholecell patch-clamp recording) aud 108 spinal cord slices (5 μm thick,for immunofluorescence detection) were prepared.The 108 slices with two kinds of thickness were divided into 3 groups (n=36 each) using a random number table:blank control group (group C),remifentanil group (group R) and remifentanil plus PICK inhibitor group (group R+PlCKi).Spinal cord slices were incubated in artificial cerebrospinal fluid (ACSF) for 90 min in group C.Spinal cord slices were incubated for 90 min in ACSF containing remifentanil at the fiual concentration of 4 mnol/L in group R.Spinal cord slices were incubated for 90 min in ACSF containing remifentanil at the final concentration of 4 nmol/L and 50 μmol PICK inhibitor in group R+PICKi.The whole-cell patch-clamp technique was used to record the amplitude and time interval of AMPA receptors-mediated mEPSCs.The method of immunofluorescence was used to detect the expression of AMPA receptors.Results Compared with group C,the amplitude of mEPSCs was significantly increased,and the time interval of mEPSCs was shortened,the expression of GluR1 and GluR3 was up-regulated,and the expression of GluR2 was down-regulated in R and R+PICKi groups (P＜0.05).Compared with group R,the amplitude of mEPSCs was significantly decreased,and the time interval was prolonged,the expression of GluR3 was down-regulated,the expression of GluR2 was up-regulated (P＜0.05),and no significant change was found in GluR1 expression in group R+PICKi (P＞0.05).Conclusion PICK1 is involved in the process of remifentanil-induced mEPSCs mediated by AMPA receptors in spinal dorsal horn neurons and of expression of AMPA receptors in juvenile rats,which may be the mechanism underlying remifentanil-induced hyperalgesia.
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Objective To investigate the effect of isoflurane preconditioning on rat learning and memory in cerebral ischemia-reperfusion injury and its possible mechanism.Methods Thirty-six adult male Sprague-Daw ley rats w ere randomly divided into a sham operation group, a cerebral ischemia-reperfusion group, and an isoflurane preconditioning group (n=12 in each group). A model of middle cerebral artery occlusion and ischemic-reperfusion w as induced by a modified intraluminal suture method. The rats of the isoflurane preconditioning group inhaled 1.5%isoflurane for 1 hour per day for 5 d. At 24 h after the last preconditioning, a model of MCAO w as made. At 24 h after MCAO, the infarct volume w as detected by using 2,3,5 chlorinated diphenyl tetrazolium staining. At day 1, 3, 7, and 14 after MCAO, the modified Neurological Severity Score (mNSS) were performed. At day 9 after MCAO, the Morris w ater maze test w as used to evaluate the learning and memory of rats. At day 14, Western blotting w as used to detect the protein expression level of hippocampal tissue glutamate receptor 1 (GluR1) on the side of ischemia. Results No obvious infarcts w ere observed in the rats of the sham operation group. The infarct volume in the isoflurane preconditioning group w as significantly smal er than that of the cerebral ischemia-reperfusion group (26.383%±3.128%vs.19.107%±1.661%;P<0.05). No neurological deficit w as observed in the sham operation group (score 0). The mNSS scores at day 1, 3, 7, and 14 after MCAO in the isoflurane preconditioning group w ere decreased significantly (day 1:9.000 ±1.195 vs.11.500 ±1.414;day 3:6.6250 ±1.407 vs.6.625 ±1.407vs.6.625 ±1.407; day 7: 5.875 ±0.707 vs.7.375 ±1.407; and day 14:3.375 ±1.187 vs.5.125 ±1.246;al P<0.05). The Morris w ater maze show ed that the escape latencies at day 1-5 after MCAO in the isoflurane preconditioning group w ere al significantly shorter than those in the cerebral ischemia-reperfusion group (day 1: 95.992 ±15.734 s vs.103.008 ±11.654 s; day 2: 70.949 ±14.708 s vs. 94.705 ±14.709 s;day 3:39.660 ±7.413 s vs.65.716 ±10.155 s;day 4:22.692 ±5.778 s vs.35.240 ±8.553 s;day 5: 14.906 ±4.336 s vs.22.890 ±10.381 s; al P<0.05). The numbers of crossing platform (4.556 ± 1.333 vs.2.889 ±1.536 ) and the percentages of time spent in the target quadrant ( 33.014%±5.223%vs. 21.978%±6.697%) in the isoflurane preconditioning group w ere significantly increased than in the cerebral ischemia-reperfusion group (al P<0.01). The levels of hippocampal GluR1 protein on the ischemic sides in the sham operation group, ischemia-reperfusion group, and isoflurane preconditioning group w ere 0.871 ±0.153, 0.456 ±0.130, and 0.689 ±0.126, respectively. There w ere significant differences among the 3 groups ( F=18.329, P<0.001) and the isoflurane preconditioning group w as significantly higher than the ischemia-reperfusion group (P<0.05). Conclusions Isoflurane preconditioning can improve the learning and memory in cerebral ischemia-reperfusion in rats, its mechanism may be associated w ith the uprelagating GluR1 expression in the hippocampus.
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Objective To evaluate the effect of propofol post-conditioning on the expression of GluR1-containing AMPA receptors during cerebral ischemia-reperfusion (I/R) injury in rats.Methods Sixtyfour healthy male Sprague-Dawley rats,aged 7-8 weeks,weighing 250-280 g,were randomly divided into 3 groups (n =18 each) using a random number table:sham operation group (group S),I/R group and propofol group (group P).Focal cerebral ischemia was induced by occlusion of the middle cerebral artery for 1 h followed by reperfusion in I/R and P groups.Propofol was infused at 20 mg · kg-1 · h-1 for 2 h starting from the onset of reperfusion in group P,and the equal volume of normal saline was given in S and I/R groups.Fear conditioning test was performed on 7,14 and 28 days after operation.The time spent freezing induced by context and condition was recorded,and the percentage was calculated.The modified neurological deficit score (NDS) was measured at 24 h of infusion.The cerebral infarct size was measured by TTC.The hippocampal samples were isolated for determination of the expression of protein kinase A (PKA),A-kinase anchoring protein 150 (AKAP150) and AMPA receptor GluR1 subunit and phosphorylation of GluR1 at serine 845 (Ser845) site (pGluR1 Ser845).Results Compared with group S,the NDS and cerebral infarct size were significantly increased,the expression of PKA and pGluR1 Ser845 was down-regulated,and the percentage of time spent freezing induced by context and condition was decreased on 7 and 14 days after operation in I/R and P groups.Compared with group I/R,the NDS and cerebral infarct size were significantly decreased,the expression of PKA and pGluR1 Ser845 was upregulated,and the percentage of time spent freezing induced by context and condition was increased on 7 and 14 days after operation in group P.Conclusion The mechanism by which propofol postconditioning reduces cerebral I/R injury is related to phosphorylation of AMPA receptor GluR1 subunit in rats.
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Objective To investigate the effect of propofol post-conditioning on RNA2 (ADAR2)-α-amino-3-hydroxy-5-methyliso xazole-4-propionic acid (AMPA) receptor subunit glutamate 2 (GluR2) pathway in hippocampal neurons of fetal rats subjected to oxygen-glucose deprivation (OGD).Methods The hippocampal neurons were isolated from the fetal rats obtained from Wistar rats at 16-18 days of gestation and primarily cultured for 7 days.The primarily cultured neurons were randomly divided into 3 groups (n =6 each):control group (group C) ; OGD group (group O) ; propofol post-conditioning group (group P).The cells were subjected to OGD for 1 h followed by restoration of O2-glucose supply in group O.In group P,the cells were subjected to OGD for 1 h followed by restoration of O2-glucose supply and then 1.2 μg/ml propofol was added and the cells were cultured for 2 h and then the culture medium was replaced with plain culture medium.At 24 h of incubation,the cells were collected for assessement of the survival rates of the hippocampal neurons and for determination of the expression of ADAR2 mRNA (by RT-PCR),the total ADAR2 protein (tADAR2) and ADAR2 protein in the nucleus of cells (nADAR2) (by Western bolt).The editing percentage of GluR mRNA at the Q/R site was analyzed by nest RT-PCR and BbV1.Results There was no significant difference in the expression of ADAR2 mRNA and tADAR2 among the three groups.Compared with group C,the survival rates of the hippocampal neurons were significantly decreased,the expression of nADAR2 was down-regulated,the ratio of nADAR2/tADAR2 was decreased,and the editing percentage of GluR mRNA at the Q/R site was decreased in group O.Compared with group O,the survival rates of the hippocampal neurons were significantly increased,the expression of nADAR2 was up-regulated,the ratio of nADAR2/tADAR2 was increased,and the editing percentage of GluR mRNA at the Q/R site was increased in group P.Conclusion Propofol post-conditioning reduces OGD-induced damage to hippocampal neurons of fetal rats through activating ADAR2-AMPA receptor GluR2 pathway.
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Objective To evaluate the role of phosphatidyl-inositol 3 kinase (PI3K)-α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor subunit glutamate receptor 2 (GluR2) pathway in propofol postconditioning-induced reduction of cerebral ischemia-reperfusion (I/R) injury in rats.Methods One hundred and eighty male Sprague-Dawley rats,weighing 250-280 g,were randomly divided into 5 groups with 36 rats in each group:sham operation group (group S),I/R group,propofol postconditioning group (group P),intralipid postconditioning group (group Ⅰ),and PI3K inhibitor wortmannin + propofol postconditioning group (group W +P).The rats were anesthetized with intraperitoneal 10% chloral hydrate 350 mg/kg.Cerebral I/R was produced by 60 min middle cerebral artery occlusion followed by reperfusion.Propofol and 10% intralipid were infused via the femoral vein at a rate of 20 mg· kg-1· h-1 for 2 h starting from the onset of reperfusion in groups P and I,respectively,while the equal volume of normal saline was given instead in groups I/R and S.Wortmannin 0.6 mg/kg was injected intraperitoneally at 30 min before reperfusion in group W + P.The modified Neurological Severity Score (mNSS) was assessed and the cerebral infarct volume was detected at 12 and 24 h after operation.The hippocampi on the ischemic side were obtained at 4,6,12 and 24 h after operation for determination of expression of PI3KGluR2-containing AMPA receptor (by co-immunoprecipitation and Western blot) and activity of PI3K (by ELISA).Results Compared with group S,the mNSSs and infarct volume were significantly increased at 12 and 24 h after operation,and the activity of PI3K was decreased,and the expression of PI3K-GluR2-containing AMPA receptor was down-regulated at 4,6,12 and 24 h after operation in the other four groups (P ＜ 0.05).Compared with group I/R,the mNSSs and infarct volume were significantly decreased at 12 and 24 h after operation,and the activity of PI3K was increased,and the expression of PI3K-GluR2-containing AMPA receptor was up-regulated at 4,6,12 and 24 h after operation in group P (P ＜ 0.05).Compared with group W + P,no significant change was found in the parameters mentioned above in groups I/R and I (P ＞ 0.05),and the mNSSs and infarct volume were significantly decreased at 12 and 24 h after operation,and the activity of PI3K was increased,and the expression of PI3K-GluR2-containing AMPA receptor was up-regulated at 4,6,12 and 24 h after operation in group P (P ＜0.05).Conclusion PI3K-AMPA receptor subunit GluR2 pathway is involved in propofol postconditioning-induced reduction of cerebral I/R injury in rats.
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Objective To investigate the changes in trafficking of GluRl-containing AMPA (GluR1-AMPA) receptor and GluR2-AMPA receptor from cytoplasm to cell membrane in the spinal cord dorsal horn in a rat model of incisional pain.Methods Thirty-two adult male SD rats aged 6-8 weeks weighing 280-300 g were randomly divided into 2 groups:control group (group C,n =8) and incisional pain group (group Ⅰ,n =24).An 1 cm long incision was made in the plautar surface of right hindpaw according to Brennan et al.in group Ⅰ.Cumulative pain score (CPS) and paw-withdrawal threshold to yon Frey stimuli (PWT) were measured at 3 h and day 1 and 3 afar incision ( T1,2,3 ).The animals were sacrificed after pain behavior assessment.Their lumbar segments of the spinal cord (L3-6) were removed.The expression of GluR1 and GluR2 in cell membrane and cytoplasm in spinal cord dorsal horn was determined by Western blot analysis.The co-expression of Stargazing with GluR1 and GluR2 in the spinal cord dorsal horn was examined by co-immuno-precipitation.Results The CPS was increased and PWT decreased; the GluR1 expression in cytoplasm was decreased while the expression of GluR1 in cell membrane and the co-expression of Stargazing with GluR1 were up-regulated in group Ⅰ as compared with group C.There was no significant change in the expression of GluR2 in cytoplasm and cell membrane and the co-expression of Stargazing with GluR2 in group Ⅰ as compared with group C.Conclusion GluR1-AMPA receptor transfers from cytoplasm to cell membrane but GluR2-AMPA receptor does not in rats with incisional pain.
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Objective To investigate the long-term effects of propofol postconditioning on cerebral ischemia-reperfusion (I/R) injury in rats.Methods One hundred and forty-four healthy male SD rats,aged 7-8 weeks,weighing 250-280 g,were equally and randomly divided into 4 groups:sham operation group (group S),I/R group,propofol postconditioning group (group P) and intralipid group (group I).The animals were anesthetized with intraperitoneal 10％ chloral hydrate 300 mg/kg.Focal cerebral ischemia was induced by occlusion of middle cerebral artery for 60 min using a nylon thread with a rounded tip which was inserted into internal carotid artery in groups I/R,P and I.Two hour infusion of propofol was started at 20 mg· kg- 1· h- 1 immediately after the onset of reperfusion in group P,while the equal volume of normal saline was given instead in S and I/R groups,and 10％ intralipid was given instead in group I.Five rats in each group were chosen on day 1,14 and 28 after operation for assessment of neurological behavior and detection of cerebral infarct volume.Six rats in each group were chosen to perform Morris water maze test at day 9 and 23 after operation for 6 consecutive days.Five rats in each group were sacrificed on day 1,14 and 28 after operation and the hippocampal tissues were isolated for determination of the expression of GluR1-containing AMPA (GluR1-AMPA) receptor and GluR1-AMPA receptor in cell membrane.The ratio of GluR1-AMPA receptor in cell membrane/GluR1-AMPA receptor was calculated.Results Compared with group S,neurological behavior scores and the number of animals' swimming across the platform were significantly decreased,cerebral infarct volume was significantly enlarged,escape latency was significantly prolonged,and ratio of GluR1-AMPA receptor in cell membrane/GluR1-AMPA receptor was significantly increased ( P ＜ 0.05),while no significant change in the expression of GluR1-AMPA receptor was found in I/R group ( P ＞0.05).Propofol postconditioning inhibited cerebral I/R-induced changes mentioned above ( P ＜ 0.05).Conclusion The brain protection against focal I/R injury by propofol postconditioning can last for 28 days after operation and the inhibition of trafficking of GluR1-AMPA receptor from cytoplasm to cell membrane may contribute to this long-term brain protection.
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Objective To investigate the effects of propofol on the phosphoryhtion of α-amino-3-hydroxy5-methyl-4-isoxazolepropionate(AMPA)receptor GluR1 subunits at Serine-831 and Serine-845 sites in the spinal cord dorsal horn in a rat model of visceral pain.Methods Thirty male SD rars,aged 6-8 weeks,weighing 200300 g,in which intrathecal catheters were successfully placed without complications,were randomly divided into 3 groups(n-=10 each):sham operation group(group Ⅰ),visceral pain group(group Ⅱ)and propofol group (group Ⅲ).Visceral pain was induced by injection of 10％ capsaicin 50μl via the rectum in groups Ⅱ and Ⅲ.While the equal volume of normal saline was given instead in group Ⅰ.Group Ⅲ received intrathecal injection of propofol 20 tg at 10 min before injection of capsaicin.While the equal volume of dimethyl sulfoxide was given instead of propofol in groups Ⅰ and Ⅱ.The cumulative pain score was recorded during 30 min after capsaicin injection.The rats were then sacrificed,and the lumbar segment(L3-6)of the spinal cord was removed for determination of the expression of GluR1 subunits and phosphorylation of GluR1 subunits at Serine-831 and Serine-845 sites in the spinal cord dorsal horn.Results Compared with group Ⅰ,the cumulative pain score and phosphorylation of GluR1 subunits at Serine-831 sites and Serine-845 in the spinal cord dorsal horn were significantly increased(P ＜0.05 or 0.01),but there were no significant differences in the expression of GluR1 subunits in groups Ⅱ and Ⅲ (P ＞ 0.05).Compared with group Ⅱ,the cumulative pain score and phosphorylation of GluRl subunits at Serine831 and Serine-845 sites in the spinal cord dorsal horn were significantly decreased in group Ⅲ(P ＜ 0.05 or 0.01).Conclusion Propofol can attenuate the visceral pain through the inhibition of the phosphorylation of AMPA receptor GluR1 subunits at Serine-831 and Serine-845 sites in tte rat spinal cord dorsal horn.
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Objective To evaluate the role of hippocampal AMPA receptors in the antidepressant effect of ketamine in rats.Methods Thirty male Wistar rats aged 2 months weighing 180-220 g were randomly divided into 3 groups (n =10 each):control group (group C); ketamine group (group K) and AMPA receptor antagonist NBQX group (group N).The animals were forced to swim for 15 min on the 1st day.On the 2nd day,NBQX 10 mg/kg was injected intrapefitoneally in group N; 30 min later,normal saline was injected intraperitoneally in group C,while ketamine 10 mg/kg was injected intraperitoneally in groups K and N.The forced swimming test was performed again for 5 min at 30 min after administration and the immobility time of the rats was recorded.Then the animals were sacrificed and the hippocampus was removed for determination of the expression of phosphorylated rapamycin (p-mTOR) and phosphorylated glutamate receptor 1 (p-GluR1).Results Compared with group C,the immobility time was significantly shortened and the expression of p-mTOR and p-GluR1 up-regulated in group K,and the immobility time was significantly shortened,the expression of p-mTOR up-regulated and the expression of p-GluR1 down-regulated in group N (P ＜ 0.05).Compared with group K,the immobility time was significantly prolonged and the expression of p-mTOR and p-GluR1 down-regulated in group N (P ＜ 0.05 ).Conclusion AMPA receptors in hippocampus are involved in the antidepressant effect of ketamine in rats and the inhibition of mTOR and GluR1 activities may be involved in the mechanism.
ABSTRACT
Objective To explore the relationship between motor complications of Parkinson's disease(PD)with chronic L-dopa treatment and striatal phosphorylated GluR1Ser845.Methods The hemi-parkinsonian rat model was produced by injecting stereotaxically 6-OHDA to right medial forebrain bundle(MFB).Then the hemi-parkinsonian rat was treated intraperitoneally with L-dopa methylester(25 mg?kg-1?d-1,twice one day)for 22 days and rotational duration and frequency of off period were respectively estimated.After they were sacrificed,their subcellualr distribution of GluR1 and GluR1Ser845 phosphorylation was observed with immunofluorescence and Western blot.Results After the chronic treatment with L-dopa methylester,PD rats displayed shortened rotational duration and increased frequency of off period,which was similar to fluctuations of the symptoms and on-off phenomenon in PD patients.In the lesioned striatum of PD rats,the amount of GluR1 and phosphorylated GluR1Ser845 in striatal membrane was reduced to 73.0%?4.8% and 42.0%?5.6%,respectively,compared with those non-lesioned.After chronic treatment of PD rats with L-dopa,the amount of GluR1 and phosphorylated GluR1Ser845 in striatal membranes were increased to 104.0%?5.5% and 112.0%?3.4%.These unique changes occurred only in parvalbumin-positive interneurons.Conclusions These findings suggest that subcellular distribution and phosphorylation state of GluR1Ser845 in parvalbumin-positive interneurons may play a significant role in the pathogenesis of motor complications of chronic L-dopa treatment for PD.