Objective To investigate the effect of propofol on the response of spinal cord to pain stimulation induced by intraplantar injection of formalin Methods Thirty SD rats of both sexes weighing 200 250g were randomly divided into six groups Pain stimulation was produced by subcutaneous injection of formalin (2 5%100?l) into the plantar region of unilateral front paw Group F received intraplantar injection of formalin only (n=6);group FP received intraperitoneal propofol 100?g?kg -1 10min after formalin injection (n=6); group PF received formalin injection 10min after intraperitoneal propofol 100?g?kg -1 (n=6); group P received intraperitoneal propofol 100?g?kg -1 only; group FS received intraperitoneal normal saline 10ml?kg -1 10min after formalin injection; and group S received intraperitoneal normal saline 10ml?kg -1 only 1 h after last injection (intraperitoneal or intraplantar) the animals were anesthetized and cervical spinal cord (where sensory nerves from front paw enter) was removed and sliced and examined for fos expression in the spinal cord using fos immunohistochemistry technique Results After formalin injection large numbers of fos like immunoreactive neurons (FLINs) were found in the ipsilateral dorsal horn Most of FLINs were confined to the medial part of outer area of laminal I and II Intraperitoneal propofol injected either before or after formalin stimulation significantly suppressed fos expression in all laminal (P

BACKGROUND: It is indistinct that whether ketamine can exert antinociceptive effect througb influencing the transmission of nocuous information in spinal cord; Nitric oxide (NO) in spinal cord participates mainly in the formation and development of hyperalgesia, and it can also induce Fos protein expression. It is still controversal whether it contributes to the transmission and mediation of ketamine to pain signal.OBJECTIVE: To observe the response to formalin stimulation in spinal cord of the rats and the effect of ketamine.DESIGN: Balanced randomized animal trial.SETTING: Department of Anesthesiology, Affiliated Hospital of Xuzhou Medical College; Jiangsu Provincial Key Laboratory of Anesthesiology.MATERIALS: This trial was carried out in the Jiangsu Provincial Key Laboratory of Anesthesiology, Xuzhou Medical College from January to March 2000. Totally 30 Sprague-Dawley rats were chosen and balanced randomized into 6 groups: formalin group (n=6), formalin + ketamine group (n=6), ketamine +formalin group (n=6), ketamine group (n=6), formalin+normal saline group (n=3) and normal saline group (n=3). The gender ratio was the same in each group.METHODS: Formalin group:The rats were stimulated for one hour by subcutaneous injection of 0.05 volume fraction of 200 μL in the center of palm of unilateral fore-claw. Formalin +ketamine group: The rats were stimulated for 10 minutes by formalin, then for one hour by intraperitoneal injection of 100 rg/kg ketamine. Ketamine + formalin group: The rats were injected with ketamine for 10 minutes, then with formalin for one hour. Ketamine group: the same dosage of ketamine was intraperitoneally injected into the rats for one hour. Formalin + normal saline group: The rats were stimulated for 10 minutes by formalin, then intraperitoneally given 10 mL/kg normal saline for one hour. Normal saline group: the same volume of normal saline was intraperitoneally injected into the rats for one hour.MAIN OUTCOME MEASURES: ① Behavioral performance of the rats in each group. ② Spinal sections were chosen, and stained with c-fos genetic immunohistochemical and NADPH-d histochemical methods. The changes of the number of Fos-like immuno-positive neurons (FLI) and FLI/nitric oxide synthase (NOS) double-labeled neurons in the 4-layer sections (layer Ⅰ -Ⅱ ,layer Ⅲ-Ⅳ ,layerⅤ-Ⅵ ,layer Ⅶ-X )of spinal dorsal horn of the rats were observed.RESULTS: All the thirty rats entered the stage of result analysis. ① Behavioral changes: The rats of formalin group and formalin+ normal salinegroup had apparent pain response; Several minutes after injection with ketamine, righting reflex disappeared and did not recover at perfusion period.Prolonged sleep was found without obvious pain response performance. ② FLI neuron expression: A lot of FLI positive neurons were found in the spinal dorsal horn of injec tion side of the rats in the formalin group and formalin+ normal saline group, and they distributed principally in the layer Ⅰ - Ⅱ of spinal dorsal horn.The distribution in the ketamine + formalin group and formalin + ketamine group was basically similar to that in the formalin group and formalin + normal saline group, but positive neuron counts were significantly reduced (P ＜ 0.01). ③ The expression of FLI/NOS double-labeled neurons: The number of double-labeled neurons in the spinal dorsal horn layer Ⅰ - Ⅱ of the rats in the ketamine+ formalin group and formalin+ ketamine group were significantly less than that in the formalin group and formalin+normal saline group [(1±1), (1±1), (7±3), (8±3),P ＜ 0.01].CONCLUSION: Some neurons of ipsilateral corresponding spinal segments participate in the transmission and mediation of pain signal. Ketamine can suppress the activities of these neurons and exert antinociceptive effect. The antinococeptive function of ketamine may be caused by the activity depression of the NOS-positive neurons in spinal cord.

Aim To observe the neuron nitric oxide synthase(nNOS) expression in the distal cerebrospinal fluid contacting neurons(CSF-CNs) of rat brain parenchyma, and investigate the role of CSF-CNs in the development of morphine dependence and withdrawal.Methods Male adult Sprague-Dawley rats, weighed 260?20 g,were experimented with A 3 ?l volume of 30% cholera toxin subunit B with horseradish peroxidase(CB-HRP) was injected into one of the rats′lateral ventricles to trace and locate the distal CSF-CNs of rat brain parenchyma 48 hours before the animals were killed. All animals were perfused and the relative tissue of rats′brain was removed.Frozen serial coronal sections (40 ?m) were cut. Then TMB-ST reaction procedure was used to stain the CB-HRP positive neurons,followed by immunohistochemistry double-labeling of the nNOS with CB-HRP positive neurons. The withdrawal symptoms were observed and scored. The numbers of the CB-HRP, and CB-HRP/nNOS positive neurons on the same segmental brain sections were counted.Results The withdrawal symptoms of the withdrawal group were significant, scores of all signs were significantly higher than those of the dependence groups and control group(P

Aim To investigate the effects of intrathecal levobupivacaine(LB)on substance P(SP)expression in spinal dorsal horn(L4~5)and the distal cerebrospinal fluid contacting neuron(dCSF-CN)in a rat model of formalin induced inflammatory pain.Methods Twelve male Sprague-Dawley rats(290~310 g,Grade SPF)were randomly divided into 2 groups with 6 in each.Rats were anesthetized with sodium pentobarbital(40 mg?kg-1,ip)and then fixed in the stereotaxic instrument.A 3 ?l volume of 30% CB-HRP to trace dCSF-CN was injected into one of the rats'lateral ventricles.The animals kept alive,forty eight hours later artificial cerebrospinal fluid(ACSF)10 ?l and 0.5% LB 10 ?l(diluted with ACSF)were intrathecally injected in group ACSF and group LB respectively.Inflammatory pain was induced by formalin injection into left hind paw.Then nociception of the animals was assessed behaviorally with the formalin test.Two hours after formalin administration,rats were sacrificed with deeply anesthesia and transcardially perfused.L4~5 segments of spinal cord were removed for immunohistochemical SP-labeling.Brainstem segments containing dCSF-CN were removed for SP/CB-HRP dual-labeling with immunohistochemical procedures and colloidal gold immunoelectronmicroscopy.Results LB significantly reduced paw licking time in the formalin test(P 0.05vsbaseline,P

Objective The ultrastructure of the distal CSF-contacting neurons in the dorsal raphe and the relationships with their surrounding tissues were observed and a new morphological evidence for their functional significance was offered. Methods We combined CB-HRP tracing with electron microscopic techniques and observed the ultrastructure of the distal CSF-contacting neurons in the dorsal raphe and the relationships with their surrounding tissues. Results 1.The CSF-contacting neurons have the general cytological structure of neurons; 2.CB-HRP mainly is localized in the trans face of the Golgi apparatus and lysosomes;3.There are two kinds of contrary direction synaptic relationships between the distal CSF-contacting neurons and the non-CSF-contacting neurons.Conclusions 1.There are no obvious differences between the CSF-contacting neurons and other neurons in ultrastructure;2.The Golgi apparatus and the lysosomes may exist the special function in take and transport the material from CSF;3.The CSF-contacting neurons can not only transport the material or deliver the information from the dorsal raphe to CSF,but also from CSF to the dorsal raphe.

AIMTo investigate the effect of shor tly inhaled isoflurane or isoflurane on c-fos gene expression of limbic system in rat. METHODS18 SD male rats weighing 200～250 g were randomi zed into three groups, control group, isoflurane group, enflurane group.The anim als in enflurane group or isoflurane group breathed 2% enflurane or 2% isofluran e till righing reflex nearly disappear.The animals in control group underwent th e same experimental steps except inhalation of anesthesia.The effect of shortly inhaled isoflurane or enflurane on c-fos gene in limbic system was observed wit h Fos immunohistochemical staining technique. RESULTNumber of FL I positive neurons of the 10 nucleus was increased significantly in limbic syst em by isoflurane, but enflurane only influenced FLI positive neurons expression of 8 out of the 10 nucleus. CONCLUSIONNuclei with significant ch ange of c-fos expression might be related to isoflurane or enflurane anesthes ia induction.

Objective To explore prevention of cyclosporine A (CsA) combined with Cobalt protoporphyrin (CoPP) against murine graft versus-host disease (GVHD) after allogeneic hematopoietic stem cell transplantation (allo-HSCT).Methods C57BL/6 (H-2Kb) mice were used as donors and BALB/c (H-2Kd) mice as recipients,which were randomly divided into 4 groups.The mice in total body irradiation group (TBI group) were lethally irradiated and injected intravenously with PBS; The mice in Allo-HSCT group (BS group) were lethally irradiated and injected intravenously with bone marrow cells and spleen cells; The mice in CsA intervention group (CsA group) were injected with CsA intraperitoneally after allo-HSCT; The mice in CsA combine with CoPP intervention group (combination group) received both CsA and CoPP intraperitoneally after alloHSCT.Recipients were monitored for condition,survival rate and weight.The liver,small intestine and skin in the recipients were gained and pathological changes of GVHD were assessed.The kidney was stained with Masson staining dye to observe the tissue fibrosis.The expression levels of renal HO-1 mRNA in the recipients were detected.Results In contrast to BS and CsA groups,GVHD degree in combination group was mild,with less reduction and quick recovery of weight.On the day 30 after HSCT,survival rate in BS group was 36.8％,and that in combination group and CsA group was 69.6％ and 53.5％ respectively (P＜0.05).In comparison with BS and CsA groups,pathological changes in combination group were mild,cellular edema and degeneration degree of the liver,small intestine and skin were slight,and few necrosis and infiltrated inflammatory cells were observed.Tubulointerstitial fibrosis hardly occurred in combination group,but it occurred in CsA group abundantly.As compared with BS group,the expression levels of HO-1 mRNA was increased in combination group,while decreased in CsA group (P＜0.05).Conclusion CsA combined with CoPP enhanced the protective effect of CsA against GVHD,moreover,CoPP could alleviate the side effects of CsA,which might be related with up-regulation of the expression levels of HO-1.

Objective To (e)xplore inhibition of endothelial progenitor cells (EPCs) against hepatic vein thrombosis after allogeneic bone marrow transplantation (BMT).Methods Balb/c mice were randomly divided into three groups: (1) BMT group [Balb/c mice were injected intravenously with 5 × 106 bone marrow cells after total body irradiation (TBI)]; (2) EPCs co-transfusion with bone marrow cells group: 5 × 105 EPCs were infused into recipient mice simultaneously; (3) Normal control group.Liver index was detected on the day 0,5,10,15 and 20 after transplantation.Hepatic vein thrombosis,hepatic cells and vascular endothelial damage were observed under the light microscopy after H&E staining.The injury of liver cells,liver veins,hepatic sinusoidal endothelial cells (SECs)and platelet adhesion conditions were observed under a transmission electron microscope (TEM).The proportion of activated platelets and TNF-α concentration in peripheral blood were detected by using flow cytometry.Results On the day 0,5,10,15 and 20 after transplantation,the proportion of activated platelets,liver index and TNF-α concentrations in BMT group and EPCs co-transfusion group showed an upward trend,peaked on the 15th day,and then decreased.However,they were still significantly higher than those in normal control group (P＜0.05).The above parameters in EPCs co-transfusion group at each time point were significantly lower than those in BMT group (P＜0.05).As compared with BMT group,platelet adhesion decreased,hepatic vein thromboses were reduced,hepatocyte swelling and necrosis were alleviated,and liver damage repaired rapidly in EPCs co-transfusion group.Conclusion EPCs co-transfusion with bone marrow cells could inhibit the hepatic veins thrombosis and ameliorate liver damage significantly.

Objective To explore the growth and developmental parameters and behavioral characteristics of rhesus monkeys during the first year of birth and to establish the background data.Methods A total of 18 (♂=11,♀=7 ) infant rhesus monkeys born from individually caged mothers and with known genetic background and postnatal days were monitored monthly for body weight, body height, head circumference, chest circumference, forelimb length, hind limb length, crown-rump length, tail length and anal-genital distance from postnatal day ( PND) 1 to 360, while hematology, blood chemistry and lymphocyte subsets were examined on PND 28, 175 and 360, and finger maze test was carried out on PND 208.Results The body weight showed linear growth with no significant difference between genders (P>0.05). Except for the anal-genital distance of male infants was significantly greater than that of female infants ( P<0.01 ) , no significant differences were observed between sexes in other morphological parameters.No significant differences of hematology were seen between genders (P >0.05).Compared with that at PND28, TP and BUN were significantly increased (P<0.01) while ALP decreased with no significant difference (P>0.05) at PND 175 and 360.Compared with that at PND28, CD4 +and CD4 +/CD8 +were significantly decreased ( P<0.01) while CD8+significantly increased ( P<0.01) at PND175 and 360.The number of sessions to solve task 2 in learning test was significantly greater than other tasks with females significantly less than males ( P<0.05) .The females had higher correct rate than males in the 2-day random memory test (P<0.05).Conclusions Body weight and morphological parameters show a linear growth.The PND.The RBC, HGB, LYMPH, TP, BUN, ALP, CD4 +, CD8 +and CD4 +/CD8+in hematology, blood chemistry and lymphocyte subsets show relevant changes to the growth and development of organs and systems in infants, which should be highly concerned in drug evaluation.The finger maze test indicates that female infants have better reversal learning and long-term memory than male infants.Background data and behavioral characteristics of infant rhesus monkeys during the first 12 months of birth are established in this study, which provide useful reference and support the evaluation of developmental and reproductive toxicity of drugs in rhesus monkeys.

Objective To explore a proper dose of endothelial progenitor cells (EPCs)administration that can achieve optimal hematopoietic improving effectiveness in a murine allogeneic hernatopoietic stem cell transplantation (allo-HSCT) model.Methods Female Balb/c mice were lethally irradiated with 60Co source,and then were injected intravenously with 5 106 bone marrow cells from C57BL/6 mice (bone marrow transplantation group).In co-transfer experiments,5 × 104,1 ×105,5 × 105 or 1 × 106 donor EPCs (EPCs treated groups) were injected simultaneously with bone marrow cells.The recipients were monitored for survival,peripheral white blood cells,hematopoietic stem cells (HSCs) and bone marrow histology.Results Compared with bone marrow transplantation group,all EPCs treated groups had accelerated recovery of peripheral white blood cells (P＜0.05),platelets (P＜0.05) and HSCs (P＜0.05).When infused with less than 5 × 105 EPCs,these effective hernatopoietic improving phenomena showed a positive correlation with the administrated doses of EPCs.However,when infused with 1 × 106 EPCs,the mice showed lower survival rate (P＜0.05)and slower recovery of peripheral white blood cells (P＜0.05),platelets (P＜0.05) and HSCs (P＜0.05) than 5 × 105 EPCs treated grpup.Bone marrow histopathology analysis confirmed the above findings.Conclusion Co-transfer with donor EPCs can improve survival rate and hematopoietic reconstitution of recipient mice in allo-HSCT,and 5 × 105 EPCs should be a proper dose to achieve the best effectiveness.