Ketamine (KTM), a N-methyl-D-aspartate (NMDA) receptor antagonist, was found to has an anti-inflammatory effect, but some patients suffered from exacerbated pro-inflammatory reactions after anesthesia with KTM. The present study was aimed to examine the underlying mechanism of pro-inflammatory effects of KTM. In this study, RAW264.7 cells were exposed to KTM and NMDA alone or combined for 30 min before lipopolysaccharide (LPS) stimulation. The expression levels of IL-6 and TNF-α were detected by RT-PCR and ELISA, and those of NMDA receptors by RT-PCR in RAW264.7 cells. Additionally, the TLR4 expression was determined by RT-PCR and flow cytometry, respectively. The results showed that in RAW264.7 cells, KTM alone promoted the TLR4 expression, but did not increase the expression of IL-6 or TNF-α. In the presence of LPS, KTM caused a significantly higher expression of IL-6 and TNF-α than LPS alone. NMDA could neither alter the IL-6 and TNF-α mRNA expression, nor reverse the enhanced expression of IL-6 and TNF-α mRNA by KTM in LPS-challenged cells. After TLR4-siRNA transfection, RAW264.7 cells pretreated with KTM no longer promoted the IL-6 and TNF-α expression in the presence of LPS. In conclusion, KTM accelerated LPS-induced inflammation in RAW264.7 cells by promoting TLR4 expression, independent of NMDA receptor.
Anesthetics, Dissociative ; pharmacology ; Animals ; Cell Survival ; drug effects ; Gene Expression Regulation ; Inflammation Mediators ; pharmacology ; Interleukin-6 ; genetics ; Ketamine ; pharmacology ; Lipopolysaccharides ; pharmacology ; Macrophages ; drug effects ; metabolism ; Male ; Mice ; N-Methylaspartate ; pharmacology ; RAW 264.7 Cells ; Signal Transduction ; drug effects ; Toll-Like Receptor 4 ; genetics ; metabolism ; Tumor Necrosis Factor-alpha ; genetics

OBJECTIVE: To select specific DNA aptamer for determining ketamine by FluMag-SELEX. METHODS: Based on magnetic beads with tosyl surface modification as solid carrier and ketamine as target, a random ssDNA library with total length of 78 bp in vitro was compounded. After 13 rounds screening, DNA cloning and sequencing were done. Primary and secondary, structures were analyzed. The affinity, specificity and Kd values of selected aptamer were measured by monitoring the fluorescence intensity. RESULTS: Two ssDNA aptamers (Apt#4 and Apt#8) were successfully selected with high and specific abilities to bind ketamine as target with Kd value of 0.59 and 0.66 μmol/L. The prediction of secondary structure was main stem-loop and G-tetramer. The stem was the basis of stability of aptamer's structure. And loop and G-tetramer was the key of specific binding of ketamine. CONCLUSION FluMag-SELEX can greatly improve the selection efficiency of the aptamer, obtain the ketamine-binding DNA aptamer, and develop a new method for rapid detection of ketamine.
Aptamers, Nucleotide/metabolism* ; DNA ; DNA, Single-Stranded/genetics* ; In Vitro Techniques ; Ketamine/metabolism* ; Oligonucleotides ; SELEX Aptamer Technique/methods*

The deficiency of aquaporin-4 (AQP4) has been reported to alter release of neurotransmitters in the mouse brain. However, the functional relevance of AQP4 in mediating essential components of the general anaesthetic state is unknown. The aim of the present study was to investigate the role of AQP4 in general anaesthesia in mice lacking AQP4. The hypnotic effects of propofol, ketamine, and pentobarbital in AQP4 knockout (KO) and CD1 control mice were evaluated using the behavioural endpoint of loss of righting reflex (LORR). The effects of propofol on extracellular levels of amino acids in prefrontal cortex of freely moving mice were investigated using microdialysis coupled to high performance liquid chromatography with fluorescent detection. The result showed that, after receiving ketamine or pentobarbital, LORR occurred at earlier time in KO mice than that in control animals. Intraperitoneal injection of ketamine or pentobarbital increased the duration of LORR. After the administration of propofol, the duration of LORR was significantly reduced in KO mice compared with that in controls. Propofol increased the extracellular levels of aspartate, glutamate, and GABA, but not taurine, in prefrontal cortex. There were significant differences of increase patterns of the three kinds of neurotransmitters between KO and WT mice. Notably, the duration of GABA level increase correlated with the duration of LORR in two genotypes of mice. These results provide in vivo evidence of different responses in time-dependent release of excitatory and inhibitory neurotransmitters in prefrontal cortex of the two genotypes of mice. It is suggested that changes in anaesthetic reactions in mice with AQP4 loss may be related to neurotransmitter regulation, and that normal functioning of AQP4 plays an important role in the maintenance of anaesthetic hypnosis.
Anesthetics, Intravenous ; pharmacology ; Animals ; Aquaporin 4 ; deficiency ; genetics ; Hypnotics and Sedatives ; pharmacology ; Ketamine ; pharmacology ; Mice ; Mice, Knockout ; Neurotransmitter Agents ; metabolism ; Pentobarbital ; pharmacology ; Prefrontal Cortex ; drug effects ; metabolism ; Propofol ; pharmacology

OBJECTIVETo investigate the protective effect of low-dose ketamine against intestinal ischemia reperfusion injury following pneumoperitoneum with carbon dioxide in rats.

METHODSThirty healthy male adult SD rats (body weight 280-320 g) were randomized into sham-operated group, model group and ketamine group and subjected to pneumoperitoneum for 120 min with carbon dioxide (not in sham-operated group). The rats in ketamine group received an intraperitoneal injection of 10 mg/kg ketamine 10 min before pneumoperitoneum, and those in the other two groups received saline injection. Fifteen minutes after pneumoperitoneum or sham operation, the small intestines were sampled to detect the content of malondialdehyde (MDA) and fore pathological testing. ELISA was used to detect the serum levels of I-FABP, TNF-α IL-6 and IL-8.

RESULTSPneumoperitoneum caused a significant increase in intestinal MDA content (P<0.05), which was lowered by ketamine pretreatment (P<0.05). Serum I-FABP, TNF-α, IL-6 and IL-8 levels all significantly increased following pneumoperitoneum (P<0.05) and were obviously lowered by ketamine pretreatment (P<0.05). Pneumoperitoneum also caused obvious pathologies in intestinal mucosa, which were ameliorated by ketamine pretreatment.

CONCLUSIONLow-dose ketamine preconditioning can reduce the inflammatory reaction and lessen oxidative damage in the intestinal mucosa following pneumoperitoneum in rats.

Animals ; Carbon Dioxide ; Dose-Response Relationship, Drug ; Fatty Acid-Binding Proteins ; blood ; Interleukin-6 ; blood ; Interleukin-8 ; blood ; Intestine, Small ; blood supply ; metabolism ; pathology ; Ketamine ; administration & dosage ; therapeutic use ; Male ; Malondialdehyde ; metabolism ; Pneumoperitoneum ; chemically induced ; complications ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Reperfusion Injury ; blood ; etiology ; metabolism ; pathology ; prevention & control ; Tumor Necrosis Factor-alpha ; blood

OBJECTIVE: To study the effect of chronic poisoning of ketamine on brain cell apoptosis in adult mouse under different duration and doses. METHODS: The mouse model of chronic poisoning of ketamine was established on adult mouse by tail vein injection of ketamine twice every week with different doses (4, 10, 20 and 30 mg/kg). The mice were sacrificed after continuous injection of ketamine of 1, 2, 4, 8 and 12 weeks. The qualitative assessment of apoptosis was made by transmission electron microscope and the quantitative assessment was made by Caspase-3 immumofluorescence staining method and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) to estimate the time point of apoptosis. All the experimental results were statistically analyzed. RESULTS: The neuron apoptosis was observed in hippocampus and corpus striatum by transmission electron microscope one week after administration, and continued for eight weeks. High level of Caspase-3 expression was observed one week after administration, but with a low level expression after 4 weeks. The number of TUNEL positive cells obviously increased one week after administration and maintained in a high number at 4 weeks. CONCLUSION Ketamine by tail vein injection could induce neuron apoptosis in adult mouse.
Animals ; Apoptosis/drug effects* ; Behavior, Animal/drug effects* ; Brain/pathology* ; Caspase 3/metabolism* ; Disease Models, Animal ; Dose-Response Relationship, Drug ; Female ; Forensic Pathology/methods* ; Hippocampus/pathology* ; In Situ Nick-End Labeling ; Injections, Intravenous ; Ketamine/poisoning* ; Male ; Mice ; Neurons/pathology* ; Time Factors

OBJECTIVE: To study the effects of ketamine and alcohol on learning and memory in mice and its possible mechanism. METHODS: Forty mice were divided into 4 groups: normal control group, ketamine group, alcohol group, and alcohol plus ketamine group. Ketamine and alcohol were given by intraperitoneal injection and intragastric administration, respectively, 1 time per day, for 14 days. The ability of learning and memory in mice was tested by the method of step-down and Morris water maze. Acetylcholine (ACh) and 5-hydroxy tryptamine(5-HT) in mice brain tissue were analyzed for the possible mechanism. RESULTS: (1) Step-down: The treatment groups lessened the latency and added wrong times (P < 0.05). The number of errors in the combined treatment group significantly increased comparing with the single drug treatment group (P < 0.05). (2) Morris water-maze: The treatment groups prolonged the latency (P < 0.05), reduced the target quadrant activity time significantly (P < 0.05), and decreased the numbers of crossing the former platform significantly (P < 0.05). (3) Biochemical index determination: The concentrations of ACh and 5-HT in treatment groups decreased significantly (P < 0.05), showed a more decreasement comparing with the single drug treatment group. CONCLUSION Ketamine has a synergistic effect with alcohol on learning and memory impairment in mice, which may be related to the common inhibitive effect on the ACh and 5-HT.
Acetylcholine/metabolism* ; Alcohols/pharmacology* ; Animals ; Brain/physiopathology* ; Drug Synergism ; Ketamine/pharmacology* ; Male ; Maze Learning/drug effects* ; Memory/drug effects* ; Memory Disorders/physiopathology* ; Mice ; Mice, Inbred ICR ; Serotonin/metabolism* ; Spatial Behavior/drug effects*

OBJECTIVETo investigate the expression of protein kinase C (PKC) in the spinal dorsal horn of rats with formalin-induced pain and the effect of intrathecal ketamine on PKC expression.

METHODSThirty-two SD rats were randomly divided into 4 equal groups, namely the control group, intrathecal saline group (NS), 50 µg ketamine group (K1) and 100 µg ketamine group (K2). The rats were anesthetized with 10% chloral hydrate, and a microspinal catheter was inserted intrathecally into the lumbar region. Five days later, the rats in groups, K1 and K2 were subjected to intrathecal administration of 50 and 100 µg ketamine (10 µl), respectively, followed by 10 µl saline, and those in NS group received 20 µl saline only. Thirty minutes later, 5% formalin (50 µl) was subcutaneously injected into the left hindpaw. The pain intensity score (PIS) was utilized to assess antinociceptive behavior within 1 h after formalin injection. Twenty-four hours later, the left hindpaw thickness was measured and the expression of PKC in the spinal dorsal horn in the L5 segment was assayed using immunohistochemistry.

RESULTSCompared to group NS, groups K1 and K2 showed significantly decreased PIS (P<0.01) in the second phase of formalin-induced pain; 24 h later, the left hindpaw thickness of group NS increased obviously in comparison with that in the control group (P<0.01), whereas the thickness was significantly reduced in group K1 and K2 as compared to that in group NS (P<0.05). The number of immunoreactive cells and the immunohistochemical score of PKC in the spinal dorsal horn were significantly higher in group NS than in group C (P<0.01), but significantly lower in groups K1 and K2 than in group NS (P<0.05).

CONCLUSIONIntrathecal ketamine produces obvious antinociception against formalin-induced pain in rats and inhibits the enhanced PKC expression in the spinal dorsal horn in response to formalin-induced pain, suggesting the important role of PKC in nociceptive signal transmission and modulation in the spinal cord.

Animals ; Formaldehyde ; adverse effects ; Injections, Spinal ; Ketamine ; administration & dosage ; pharmacology ; Male ; Pain ; chemically induced ; metabolism ; Pain Measurement ; Posterior Horn Cells ; metabolism ; Protein Kinase C ; metabolism ; Rats ; Rats, Sprague-Dawley ; Spinal Cord ; drug effects ; metabolism

OBJECTIVETo study the effects of electroacupuncture (EA) on the changes of behavior after ketamine anesthesia, and changes of serum antibodies against beta-amyloid (Abeta) and Abeta protein in the hippocampus of aged rats, thus exploring the effects of EA on the cognitive dysfunction.

METHODSThirty 14-month old SD rats were randomly divided into 3 groups, i. e. , the control group (Group A), the ketamine anesthesia group (Group B), and the EA+ketamine anesthesia group (Group C), 10 in each group. 50 mg/kg katemine was intraperitoneally injected to rats in Group B and Group C, once daily for 7 successive days. EA was performed to rats in Group C from the 1st day of the experiment after rats awoke completely from anesthesia, twice daily for 7 successive days. Changes of the ratio of the swim time in the original platform quadrant to the total swim time and the escape latency phase were observed by Morris water maze. The peripheral blood was withdrawn by the end of the experiment. Serum anti-Abeta antibody contents were detected using enzyme-linked immunosorbent assay (ELISA). The expressions of Abeta in the hippocampus were detected using Westen blot.

RESULTSLong-term application of ketamine could lower aged rats' cognitive function. In the navigation test, the escape latency phase of rats in Group B was significantly prolonged ( P < 0.01) . On the 7th day of the experiment, the serum level of anti-Abeta antibodies was lower in Group B than in Group A (P < 0.05), while the serum level of anti-Abeta antibodies was significantly higher in Group C than in Group B (P < 0.01). On the 7th day of the experiment, the expression of Abeta in the hippocampus was higher in Group B than in Group A (P < 0.05).

CONCLUSIONEA could increase the contents of anti-Abeta antibodies in aged rats with ketamine anesthesia, decrease the expression of Abeta in the hippocampus, alleviate the deposition of Abeta, thus improving rats' cognitive dysfunction.

Amyloid beta-Peptides ; immunology ; Anesthesia ; adverse effects ; Animals ; Antibodies ; blood ; Cognitive Dysfunction ; therapy ; Electroacupuncture ; Female ; Hippocampus ; metabolism ; Ketamine ; adverse effects ; Male ; Maze Learning ; Rats ; Rats, Sprague-Dawley

OBJECTIVE: To explore the effect of ketamine on adrenal pheochromocytoma (PC12) cell proliferation inhibition and induction of apoptosis and its mechanism. METHODS: PC12 cells of rats were models for dopaminergic neuron. PC12 cells were cultured with ketamine at concentrations of 0.9, 1.2, 1.5, 1.8 and 2.1 mmol/L, respectively. The cell viability was measured by MTT method after incubation at 12, 24, 48 and 72h. Hoechst stain was used to observe the morphological changes of apoptosis. PC12 cells cultured after 48 h with different concentrations of ketamine were selected to detect apoptotic rate using flow cytometry and detect the expression of bax and bcl-2 proteins using Western blotting. RESULTS: For different concentrations of ketamine, vitality of PC12 cells significantly decreased with increase of the incubation time. Apoptosis was obviously observed using Hoechst staining. Flow cytometry showed that apoptosis rates significantly increased with increasing ketamine concentrations. CONCLUSION Ketamine can inhibit the proliferation of PC12 cell by inducing apoptosis of the PC12 cell in a concentrations-dependent manner. The underlying mechanism may be related to promoting the expression of bax and inhibiting the expression of bcl-2 in the cells.
Anesthetics, Dissociative/pharmacology* ; Animals ; Apoptosis/drug effects* ; Blotting, Western ; Cell Proliferation/drug effects* ; Dose-Response Relationship, Drug ; Flow Cytometry ; Gene Expression Regulation/drug effects* ; Ketamine/pharmacology* ; PC12 Cells ; Proto-Oncogene Proteins c-bcl-2/metabolism* ; Rats ; Time Factors ; bcl-2-Associated X Protein/metabolism*

OBJECTIVE: To explore the correlation between signs similar to schizophrenia in mice after ketamine administration and the expressions of NRG1 and ErbB4 mRNA in order to explain the possible pathogenesis of schizophrenia. METHODS: Fifty KM mice were randomly divided into 5 groups which were administered intraperitoneally with saline, clozapine and different dosages ketamine. The ketamine groups were administered intraperitoneally with low dosage (25 mg/kg), middle dosage (50 mg/kg) and high dosage (100 mg/kg) one time every day for 7 days. After administration of 100 mg/kg ketamine for 7 days, the clozapine group was introgastrically administered 20 mg/kg with clozapine one time every day for 7 days. The pathological changes of hippocampus neurons were observed by HE stain. The expressions of the NRG1 and ErbB4 mRNA in hippocampus were detected by reverse transcriptase polymerase chain reaction (RT-PCR). RESULTS: In the group with high dosage of ketamine, the levels of NRG1 and ErbB4 mRNA were significantly lower than that of the group with saline. CONCLUSION Ketamine may induce signs similar to schizophrenia in KM mice. The mechanism may be involved in the reduction of NRG1 and ErbB4 mRNA expression.
Animals ; Clozapine/therapeutic use* ; Disease Models, Animal ; Dose-Response Relationship, Drug ; ErbB Receptors/metabolism* ; Hippocampus/pathology* ; Ketamine/adverse effects* ; Male ; Mice ; Neuregulin-1/metabolism* ; Neurons/metabolism* ; RNA, Messenger/metabolism* ; Random Allocation ; Receptor, ErbB-4 ; Reverse Transcriptase Polymerase Chain Reaction ; Schizophrenia/genetics*