ABSTRACT
Objective:To study the effect of modified Xiao Chaihutang on the expressions of excitatory amino acid transporters（EAATs） and vesicle glutamate transporters（VGLUTs）in hippocampus of rats with chronic depression， in order to explore the anti-depressant mechanism of modified Xiao Chaihutang based on glutamate transport. Method:A total of 120 SD rats were randomly divided into normal group， model group， and low， middle and high-dose modified Xiaochaihutang groups （6.5， 13， 26 g·kg-1） and riluzole group， with 20 rats in each group．Except normal group， the depression model of rats was prepared through Chronic restraint stress（CRS）. The normal group and the model group were intragastrically （ig） given normal saline. The modified Xiao Chaihutang groups were intragastrically given corresponding herbal drugs （6.5， 13， 26 g·kg-1）， and the Riluzole group was given Riluzole 20 mg·kg-1 through intraoeritoneal injection for 21 days， once a day. Then the depressive behaviors of rats were observed by forced swimming test （FST） and tail suspension test （TST）. The level of glutamic acid （Glu） in rats hippocampus was determined by high performance liquid chromatography （HPLC）. The mRNA expressions of EAAT1， EAAT2 and EAAT3 in hippocampus were detected by Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR）method. Western blot was used to detect the protein expressions of EAAT1， EAAT2， EAAT3， VGLUT1 and VGLUT2 in rat hippocampus tissue. Nissl staining was used to observe the morphology of hippocampal neurons in rats. Immunohistochemical（IHC）S-P method were used to detect the location expressions of EAAT1， EAAT2 and NeuN proteins in rat hippocampal CA1 region tissue. Result:The immobility times in FST and TST were increased significantly（P<0.01）， the mRNA and protein expressions of EAAT1，EAAT2，EAAT3 were decreased significantly （P<0.01）， and as well as the expressions of VGLUT1 and NeuN were decreased significantly（P<0.01）， while the level of Glutamate and the expression of VGLUT2 were increased significantly（P<0.01） in model group， compared with normal group. Compared with model group，the immobility times in FST and TST were decreased significantly（P<0.05， P<0.01）， mRNA and protein expressions of EAAT1，EAAT2，EAAT3 were increased significantly（P<0.01）， and expressions of VGLUT1 and NeuN were increased significantly（P<0.01）. However， the level of Glutamate and the expression of VGLUT2 were decreased significantly（P<0.01）， and the damage of hippocampal neurons in rats was mild in middle and high-dose modified Xiao Chaihutang groups. Conclusion:Modified Xiao Chaihutang has an anti-depressive effect. Its mechanism may be related to its up-regulation of expressions of EAAT1， EAAT2， EAAT3 genes and VGLUT1 protein in the hippocampus of depression model rats.
ABSTRACT
Glutamate serves as a major excitatory neurotransmitter in the mammalian central nervous system and is stored in synaptic deft by an uptake system that is dependent on the high-affinity glutamate transporters (ETTAs),which locate in the plasma membrane of glial cells and neurons.ETTAs can rapidly terminate the action of glutamate and maintain its normal physiological functions.If the content or function of glutamate transporters is abnormal,it can result in many physiological dysfunctions.Studies have demonstrated that high-affinity glutamate transporters play an important role in the development of chronic pain,which might be a new therapeutic target for the pain.
ABSTRACT
AIM:To investigate the role of minocycline on glutamate uptake in the periventricular zone and its putative mechanism after hypoxic exposure in neonatal rats.METHODS: A model of hypoxic-ischemic brain damage (HIBD) was developed by putting postnatal 1 d rat pups in 5%O2 for 3.5 h.The glutamate level in periventricular zone was measured by liquid chromatography coupled with tandem mass spectrometry assay ( LC-MS/MS) after hypoxic exposure for 4 h and 1 d.The dynamic changes of glutamate transporters EAAT1, and EAAT2 during developmental period in periventricular zone were determined by Western blot.Moreover, the expression of EAAT1, EAAT2, Iba-1, IL-1β, TNF-αand TGF-β1 was also detected by Western blot after hypoxic exposure for 4 h and 1 d in that region.The effects of mino-cycline on all parameters mentioned above were tested after minocycline treatment at the same time points and in the same region.RESULTS:After hypoxic exposure, glutamate level was increased, but it was decreased after minocycline treat-ment.EAAT1 and EAAT2 kept a low expression level at the first postnatal week, but a predominant elevation was found at the end of the second postnatal week.The expression of EAAT1, EAAT2, Iba-1, IL-1βand TGF-β1 was increased at 1 d after hypoxic exposure.EAAT1 and TNF-αexpression was significantly up-regulated, while EAAT2 was down-regulated af-ter minocycline treatment.CONCLUSION: Minocycline inhibits the increase in the glutamate level after hypoxia in periventricular region of the neonatal rats.The mechanism may relate to the selective regulation of glutamate transporters, rather than the inhibition of neuroinflammation in periventricular zone.
ABSTRACT
Objective To investigate the effects of inhibiting vesicular glutamate transporters (VGLUT) on pain behaviors in animals. Methods The latency in hot plate test, number of writhes in acetic acid and licking time in formeldehyde solution (formalin) tests were recorded to determine the analgesic effect of Chicago sky blue 6B (CSB6B), a selective VGLUT inhibitor. Results Intraperitoneal administration of CSB6B did not affect the acute thermal pain responses in hot plate test. In acetic acid writhing, compared with control group (26.50±2.97), CSB6B (2.5 mg/kg, ip) significantly attenuated the acetic acid-induced writhing (8.22±1.90) 30 min after administration (P<0.01); CSB6B (2.5 mg/kg, ip) significantly reduced the acetic acid-induced writhing (9.60±1.84) 60 min after administration (P<0.01). In Formalin test, compared with control group(139.40±21.02), CSB6B 0.5 mg/kg, ip) significantly reduced the licking time 75.10±19.45) 30 min after administration P<0.05) during the second phase, but not during the first phases. CSB6B(p) did not affect the licking time 2 h after administration during the first and second phases. Conclusion Inhibition of VGLUTs activity is sufficient to attenuate the inflammatory pain and this finding suggests that VGLUT participate in regulation of inflammatory pain and be a novel therapeutic strategy for treatment of pain.
ABSTRACT
Objective To investigate the effects of inhibiting vesicular glutamate transporters (VGLUT) on pain behaviors in animals. Methods The latency in hot plate test, number of writhes in acetic acid and licking time in formeldehyde solution (formalin) tests were recorded to determine the analgesic effect of Chicago sky blue 6B (CSB6B), a selective VGLUT inhibitor. Results Intraperitoneal administration of CSB6B did not affect the acute thermal pain responses in hot plate test. In acetic acid writhing, compared with control group (26.50 ±2.97), CSB6B (2.5 mg/kg, ip) significantly attenuated the acetic acid-induced writhing (8.22±1.90) 30 min after administration (P<0.01);CSB6B (2.5 mg/kg, ip) significantly reduced the acetic acid-induced writhing(9.60±1.84) 60 min after administration(P<0.01). In Formalin test, compared with control group(139.40±21.02), CSB6B (0.5 mg/kg, ip) significantly reduced the licking time(75.10±19.45) 30 min after administration(P<0.05) during the second phase, but not during the first phases. CSB6B(ip) did not affect the licking time 2 h after administration during the first and second phases. Conclusion Inhibition of VGLUTs activity is sufficient to attenuate the inflammatory pain and this finding suggests that VGLUT participate in regulation of inflammatory pain and be a novel therapeutic strategy for treatment of pain.
ABSTRACT
Objective To investigate the effect of recombinant human erythropoietin(rh‐EPO ) on acute cerebral injury and expression of GLT‐1 and GLAST in rat .Methods Sixty SD rats were randomly divided into three groups by weight :control group (n=18) ,acute cerebral injury group(n=22) and rh‐EPO conditioning group(n=20) .Acute cerebral injury models were made by modified Feeney′s method .rh‐EPO was injected in abdominal cavity 15 min after acute cerebral injury in rh‐EPO conditioning group .Rats′brain were removed 48 h after experiments .Rat GLT‐1 and GLAST mRNA expression were determined by RT‐PCR , GLT‐1 and GLAST protein expression were determined by Western blot .Results GLT‐1/GLAST mRNA and protein expression decreased significantly after acute cerebral injury(all P<0 .01) ,but increased significantly in rh‐EPO preconditioning group com‐pared with acute cerebral injury group(all P<0 .01) .Conclusion rh‐EPO preconditioning may protect against acute cerebral injury by up regulating the expression of GLT‐1/GLAST .
ABSTRACT
Manganese (Mn) is an essential element that is required in trace amount for normal growth, development as well maintenance of proper function and regulation of numerous cellular and biochemical reactions. Yet, excessive Mn brain accumulation upon chronic exposure to occupational or environmental sources of this metal may lead to a neurodegenerative disorder known as manganism, which shares similar symptoms with idiopathic Parkinson's disease (PD). In recent years, Mn exposure has gained public health interest for two primary reasons: continuous increased usage of Mn in various industries, and experimental findings on its toxicity, linking it to a number of neurological disorders. Since the first report on manganism nearly two centuries ago, there have been substantial advances in the understanding of mechanisms associated with Mn-induced neurotoxicity. This review will briefly highlight various aspects of Mn neurotoxicity with a focus on the role of astrocytic glutamate transporters in triggering its pathophysiology.
Subject(s)
Astrocytes , Brain , Glutamic Acid , Manganese , Nervous System Diseases , Neurodegenerative Diseases , Parkinson Disease , Public HealthABSTRACT
BACKGROUND: Clonidine has been shown to be a potent neuroprotectant by acting at alpha2 receptors on glutamatergic neurons to inhibit the release of glutamate. The aim of this study is to investigate the effects of clonidine on the activity of EAAT3 that can regulate extracellular glutamate. METHODS: EAAT3 was expressed in the Xenopus oocytes. Using a two-electrode voltage clamp, membrane currents were recorded after application of 30 microM L-glutamate both in the presence and absence of various concentrations of clonidine. To determine the effects of clonidine on the Km and Vmax of EAAT3 and the reversibility of clonidine effects, membrane currents were recorded after the application of various concentrations of L-glutamate both in the presence and absence of 1.50 x 10(-7) M clonidine. RESULTS: Clonidine reduced the EAAT3 responses to L-glutamate in a concentration-dependent manner. This inhibition was statistically significant at higher concentrations than at the clinically relevant range. Clonidine at 1.50 x 10(-7) M reduced the Vmax, but did not affect the Km of EAAT3 for L-glutamate. CONCLUSIONS: These results suggest that the direct inhibition of EAAT3 activity is not related to the sedation effect of clonidine and that the clonidine-induced reduction of EAAT3 activity provides additional data for the possible involvement of glutamatergic hyperactivity in the proconvulsant effect of clonidine.
Subject(s)
Animals , Rats , Amino Acid Transport System X-AG , Clonidine , Glutamic Acid , Membranes , Neurons , Oocytes , XenopusABSTRACT
In the present study, we examined the distribution and amount of two important glutamate transporters, GLT-1 and GLAST in the cerebellum of young and aged rats. Sprague-Dawley rats were used at the age of three months for young control (n=3) and 24 months for aged group (n=4). After transcardial perfusion with 4% paraformaldehyde, brain sections were immunostained for GLT-1, and GLAST. We found that GLT-1- and GLAST-immunoreactive materials were diffusely distributed throughout the gray matter of the cerebellum. Pre-embedding immunoelectron microscopic study demonstrated that the two glutamate transporters in the cerebellum were restricted to glial cells with astrocytic features. The intensity of GLT-1-immunostaining in the cerebellum appeared to be higher in aged rats than in young rats whereas GLAST-immunostaining decreased with aging. Western blot results were also consistent with the immunohistochemical observations. Conclusively, GLT-1 and GLAST expression in the rat cerebellum was changed with aging, i.e, increase of GLT-1 and decrease of GLAST expression with aging, which suggests that the two glutamate transporters might be regulated by different underlying mechanisms with aging.
Subject(s)
Aged , Animals , Humans , Rats , Aging , Blotting, Western , Brain , Cerebellum , Formaldehyde , Glutamic Acid , Neuroglia , Perfusion , Polymers , Rats, Sprague-DawleyABSTRACT
The effects of glutamate transporters on synaptic plasticity in rat models of pilocarpine-induced status epilepticus were investigated. Male Wista rats ((304.06?13.79) g) were randomly divided into 5 groups, short-term seizures (SE) and its control (SC), long-term seizures (LE) and its control(LC), normal control (Sham) groups. Epilepsy rat models were induced by injection of pilocarpine(25 mg/kg, i.d.). Glutamate transporter inhibitor, DL-threo-benzyloxyaspartate (TBOA, 7.5 nmol,1 ?l) was microinjected into right side of hippocampus after 14 days of initial status epilepticus in SE and LE groups. The same volumes of artificial cerebrospinal fluid were injected into same side of hippocampus in SC and LC groups. Electroencephalographys (EEG) were detected in SE and SC groups after 2 h of drug injection. Long term potential (LTP) at perforant pathway and dentate gyrus(PP-DG) and EEG were recorded in LE and LC groups after two weeks of drug injection. Example of Fluoro-Jade-B staining in the rat brain was made at the end of electrophysiological experiment. The results showed that there was a significant decrease in theta band power of EEG in SE group compared with that of SC group (P 0.05). The slope of excitatory postsynaptic potential (EPSP) was significantly increased in LE group compared with that of LC group (P
ABSTRACT
AIM: To demonstrate the effect of mitochondrial ATP-sensitive potassium channel opener,Diazoxide,on glutamate uptake activity of astrocyte. METHODS: Primary astrocytic cultures were prepared from cerebral cortices of rats within 3 days old.Glutamate uptake activity of astrocytes was measured using isotopic techniques to detect intracellular -labeled D,L-glutamate concentration.RESULTS: Diazoxide increased -glutamate uptake in a concentration-dependent manner,and could significantly reverse MPP~+-induced glutamate uptake inhibition.The above effects of Diazoxide were blocked by 5-hydroxydecanoate,a selective mitochondrial ATP-sensitive potassium channel blocker.CONCLUSION: Diazoxide increase the activity of glutamate transporters in astrocytes by opening mitochondrial ATP-sensitive potassium channel.
ABSTRACT
Objective To study the effect of Ceftriaxone on glutamate-induced neurocyte injury. Methods Cultured rat cerebral cortical neurocytes were divided into Ceftriaxone (A)group, glutamate(B) group ,Ceftriaxone-pretreated (C) group and control (D) group. Observe morphous and MTT colorimetry were used to detect cytoactive. Laser scanning confocal microscope was used to measure the change of Ca2+ in rest state and after brief glutamate-induced Ca2+ loading. RT-PCR was used to determine the change of GLT1 mRNA expression.Results Compared with B group, cellular morphous was closer to normal in C group. Absorptance values of groups A,B,C,D were 0.795?0.010, 0.624?0.028, 0.738?0.021 and 0.813?0.023, respectively, which there were lower in groups B and C compared with group D, but absorptance value was higher in group C than that in group B(allP
ABSTRACT
Vesicular glutamate transporters(VGLUTs)package specifically glutamate into synaptic vesicles in the presynaptic terminal for subsequent release into the synaptic cleft.VGLUT1 and VGLUT2 together label all glutamatergic neurons,are highly specific markers of glutamatergic neurons and their axon terminals.VGLUT1 and VGLUT2 are respectively the neurochemical marker of cortico-cortical projection and thalamo-cortical projection.VGLUT3 is also expressed in cholinergic interneurons,serotoninergic neurons,subsets of GABAergic interneurons in the hippocampus and cerebral cortex.The disfunction of VGLUTs can lead to the abnormal excitatory neurotransmitter glutamate,resulting in many nervous system disease.In order to give a clue for prevention and therapy of these diseases,this paper reviews the disfunction of VGLUTs effects on Alzheimer's disease(AD),Parkinson's disease(PD),schizophrenia,depressive disorder,epilepsy and deafness.