| Objective:Through observing the study of the action of different doses of CAP on gastric acid secretion and gastrin (GAS) connected with the specific immunohistochemical distribution of VR1, CGRP and SP during the effects of these compounds, and mechanism of different doses of capsaicin(CAP) on gastric acid secretion in visceral hypersensitivity rats and the effects of intragastric administration of CAP on gastric mucosal histology in rats. Methods:(1) Experimental animals grouping:Take 60 SD rats (sex is not limited) which weigh 180g～200g. The rats are grouped into group A (control group), group B (model group), group C1 (CAP 1 group), group C2 (CAP2 group) and group C3 (CAP3 group) randomly with 12 rats in each group. (2) The model of visceral hypersensitivity rats:The rats adapt to diet about 2 weeks before the initiation of experimentation. Group B, C1, C2 and C3 were sensitized by intraperitoneal injection of chicken egg albumin, while group A were intraperitoneal injection with normal saline. The responses of viscerosensitive neurons were recorded by the abdominal withdrawal reflex (AWR) to investigate visceral nociceptive transmission on the repeated colon distention 2 weeks later. (3) The dose and use of CAP:After the success of model, set up the experiment by intragastric administration. In Group A and B, normal saline were divided into 2 times to use for 10mL/kg/d in the successive 2 weeks under ungiven free access to food and water. In Group C1, C2 and C3, use the same volume of the CAP solution which were divided into 2 times for 10mL/kg/d with 1mg/kg,5mg/kg and 20mg/kg respectively and observe the rats's common states in the successive 2 weeks under ungiven free access to food and water. From the 14th day of intragastric administration on, all the rats were deprived of food, but not prohibited from drugs and ungiven free access to water after the last drugs of the next morning. (4) Indicators and methods of detection:①Observe the rats's common states in the course of model and intragastric administration.②Gastric acid secration:The rats were injected with pentobarbital sodium (40mg/kg) by intraperitoneal after pyloric ligation respectively. Open the abdominal cavity, their stomachs were quickly ligated at the esophagus-gastric junction and antral and then removed. The stomachs were incised and their contents were washed with normal saline, the washed liquid was collected for detecting gastric acid output.by the titration of collected gastric juice. The titration of was carried out with 0.01 M NaOH in presence of 0.5%Phenolphthalein until pH 7.0. The expressions of VR1,CGRPandSP in each rat's stomach sections:Took a full-thickness organization of each rat's fundic gland (about 0.5cm×0.5cm) and dipped into 4% poly formaldehyde and fixed fully,then Paraffin-embedded, did a pathological section with the thickness of 0.5μm.The expression of VR1, CGRP and SP were detected with immunohistoche-mical method. Finally, Observed under the microscope and photoed.④plasma GAS:Collect the blood sample (2ml) from heart. Take blood sample centrifuged under 4℃condition, and then supernatant extracted was used to measure the changes of GAS by the radioimmunoassay (RIA) method.⑤The change of gastric mucosal barrier:The stomachs which were washed were dipped into 4% poly formaldehyde and fixed fully, gastric mucosa was observed through naked eyes and its injury was evaluated by Guth standard-modulated scores.Take out some gastric mucosa to do the histopathologic examination. Use the light microscope to observe gastric mucous membrane's injuring degree after the HE dyeing. The related data was checked with statistic method. Results:(1) General states:The rats in each group were in good state with gloomy color pattern, acted smartly and their stools were basically normal. The rats which were intraperitoneal injection of chicken egg albumin had emergence of aggressive, mutual bite sometime, but all the rats were no died. In the process of intragastric administration, there were some rats choking in group C1, C2 and C3 and no difficulty in breathing and death and with the CAP treatment extended, the symptoms of aggressive, mutual bite had reduced obviously. While the symptoms of group B did not significantly reduced.(2)The results of AWR scores:The levels of AWR scores in model group were 0.53±0.52 scores,2.53±0.52 scores,3.40±0.63 scores respectively, and in control group were 0.67±0.50 scores,1.44±0.53 scores,2.56±0.53 scores respectively. With the increase in airbag volume, there has incremental in AWR score of each group. There has a marked increase of AWR score in 3ml and 5ml airbag distension experimental group than in control group (P<0.05) (3) The change of gastric acid secration:The levels of gastric acid secration in Group A,B,C1,C2 and C3 were 48.17±5.75 mmol/L,45.67±3.05 mmol/L,45.25±2.93 mmol/L,34.42±2.28 mmol/L, 30.50±4.25 mmol/L respectively. The gastric acid secration of group B was lower than group A, but no significant difference (P>0.05), after the treatment of CAP, group C1, C2 and C3 were significant lower than group B (P<0.05), and also lower than group A. The gastric acid secration of group C2 and C3 were significant lower than group C1 (P<0.05), and group C3 was significant lower than group C2 (P<0.05). (4) The expressions of VR1 and CGRP and SP in each rat's fundic gland. The level of VR1 of group A,B,C1,C2 and C3 were 0.58±0.51 scores,1.25±0.62 scores,2.00±0.60 scores,2.58±0.52 scores,3.25±0.75 scores, The expressions of VR1 group B were significant higher than group A (P<0.05).After the treatment of CAP, The expressions of VR1 in group Cl, C2 and C3 were significant higher than group B (P<0.05), and also higher than group A (P<0.05). The expressions of VR1 in group C2 and group C3 were significant higher than group C1(P<0.05),and group C3 was significant higher than group C2(P<0.05). The level of CGRP of group A,B,C1,C2 and C3 were 1.00±0.43 scores,1.08±0.52 scores,1.58±0. 52 scores,2.17±0.58 scores,2.67±0.49 scores, There was no significant difference between group A and group B (P>0.05).After the treatment of CAP, The expressions of CGRP in group C1, C2 and C3 were significant higher than group B (P<0.05), and also higher than group A (P<0.05). The expressions of CGRP in group C2 and group C3 were significant higher than group C1(P< 0.05),and group C3 was significant higher than group C2(P< 0.05).The expressions of SP of group A,B,C1,C2 and C3 were 0.50±0.52scores,0.83±0.58 scores,0.75±0.45 scores,0.75±0.45 scores,为0.58±0.52 scores, respectively. The expressions of SP in group B were no significant higher than group A (P>0.05), after the treatment of CAP, group C1, C2 and C3 were no significant change(P>0.05).(5) The change of plasma GAS:That of group A,B,C1,C2 and C3 were A组49.50±7.51 pg/ml, B组63.25±5.99 pg/ml, C1组44.33±2.90 pg/ml, C2组37.42±3.55 pg/ml, C3组33.00±2.37pg/ml respectively. The plasma GAS of group B was higher than group A (P<0.05), after the treatment of CAP, group C1, C2 were significant lower than group B (P<0.05), and lower than group A. The plasma GAS of group C2 was significant lower than group C1 (P<0.05). While the plasma GAS of group C3 were significant lower than group C1 and group C2(P<0.05).(6)Gastric mucosal barrier:Group A, B, C1,C2 and C3 had no significant difference (Guth standard-modulated scores,p>0.05.To observe the histopathology changes in the gastric mucosa under the light microscope, we could see that gastric mucosa was smooth and cellular epithelialis defluvium are not observed, the rats' mucosa has no hyperemia in group A,B,C1 and C2,the rats' mucosa has slightly epithelialis defluvium, hyperemia edema and inflammatory cell infiltration, focal necrosis are not observe in group C3, levels of mucosa pathohistology scores in Group A,B, C1, C2 and C3 were 0.25±0.45 scores,0.17±0.39 scores,0.32±0.45 scores,0.75±0.75 scores,0.33±0.47 scores,0.83±0.72 scores respectively. The scores of group C3 were significant higher than the other groups (P<0.05). Conclusion:(1) It was successfully to set up visceral hypersensitivity rats model by intraperitoneal injection of chicken egg albumin. (2)Successive intragastric administration of lmg/kg/d 5mg/kg/d and 20mg/kg/d CAP for 2 weeks can restrain gastric acid secration in visceral hypersensitivity rats, and effects of restrain gastric acid secration can get more and more significant with the add of doses of CAP. (3) The gastric acid secration of visceral hypersensitivity rat which impacted by CAP may be effected by regulating the expression of antral myenteric plexus VR1,CGRP. (4) CAP impacted gastric acid secration in visceral hypersensitivity rats by way of possible correlation with CGRP, no correlation with SP. (5) CAP impacted gastric acid secration in visceral hypersensitivity rats by way of possible correlation with GAS. (6) Successive intragastric administration of lmg/kg/d and 5mg/kg/d CAP for 2 weeks have no injury to gastric mucosa in visceral hypersensitivity rats, but 20mg/kg/d CAP can cause slightly injury. |
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