Résumé
Objective:To establish an ultra-performance liquid chromatography-mass spectrometry （UPLC-MS/MS） for determining the plasma concentrations of 10 active ingredients in Wujiwan at different time points after oral administration， and to compare the pharmacokinetic characteristics between normal rats and rats with chronic visceral hypersensitive irritable bowel syndrome （CVH-IBS）. Method:CVH-IBS rat model was prepared by the neonatal rat colon percutaneous transluminal coronary angioplasty （PTCA） balloon stimulation method. After intragastric administration of Wujiwan （0.245 g·kg<sup>-1</sup>）， blood was collected from the jugular vein at different time points， and the plasma concentrations of 10 active ingredients （berberine hydrochloride， palmatine hydrochloride， coptisine hydrochloride， jatrorrhizine hydrochloride， epiberberine， dihydroberberine， evodiamine， evodine， paeoniflorin， albiflorin） in Wujiwan was detected simultaneously by UPLC-MS/MS， the pharmacokinetic parameters of each component in normal rats and CVH-IBS rats were calculated. Result:The established UPLC-MS/MS could sensitively and accurately detect the plasma concentrations of 10 active ingredients of Wujiwan in rats. Compared with the normal group， the absorption rates of these 10 active ingredients of Wujiwan in the blood of CVH-IBS rats all decreased to a certain extent， and the peak time （<italic>t</italic><sub>max</sub>） was prolonged. Among them， the <italic>t</italic><sub>max</sub> of berberine hydrochloride and jatrorrhizine hydrochloride were significantly prolonged from 54 minute and 39 minute to 90 minute， respectively （<italic>P</italic><0.05， <italic>P</italic><0.01）. Area under the plasma concentration-time curve （AUC<sub>0-</sub><italic><sub>t</sub></italic>） of each component increased， and evodiamine and paeoniflorin were significantly different （<italic>P</italic><0.05，<italic> P</italic><0.01）. The clearance rates （CL/<italic>F</italic>） of these 10 active ingredients were all decreased， among which berberine hydrochloride， palmatine hydrochloride and evodiamine had significant differences （<italic>P</italic><0.05， <italic>P</italic><0.01）. Conclusion:There are significant differences in the pharmacokinetic behavior of the active ingredients in Wujiwan between normal rats and CVH-IBS rats， which may be related to the destruction of microstructure of intestinal epithelial cells and the change of activity of liver enzymes under the pathological state of IBS.
Résumé
Objective@#To investigate the effects of glutamate (Glu) injected into hypothalamic paraventricular nucleus (PVN) on visceral pain of chronic visceral hypersensitivity (CVH) rats and its possible mechanism.@*Methods@#Newborn SD rats were given CVH rat model by colorectal distension (CRD) on the 8th, 10th and 12th day after birth. Thirty rats with successful CVH model were randomly divided into CVH model group (CVH group), CVH + injection of saline into PVN group (NS group), CVH+ injection of Glu into PVN (3, 6, and 12 μg Glu, namely G3, G6, and G12, respectively), 6 rats in each group, and 6 SD rats with matching body mass were taken as sham operation group (Sham group). The pain behavior of the rats was evaluated by pain threshold, abdominal withdrawal reflex (AWR) score, and abdominal external oblique muscle electromyography (EMG). The expression of arginine vasopressin (AVP) and the proliferation of colon tissue were detected by immunohistochemical staining. The apoptosis of colon tissue was detected by TUNEL.@*Results@#Compared with the NS group, the pain thresholds of the G3, G6 and G12 groups increased, and the AWR scores and EMG amplitudes decreased. The differences were statistically significant(Pain threshold: t=7.65, 16.31, 24.78, both P<0.05; AWR scores: t=-2.98, -4.77, -7.29, both P<0.05; EMG amplitudes: t=-3.06, -5.75, -8.92, both P<0.05). Compared with the Sham group, the expression of AVP in PVN of the CVH group and NS group decreased ((42.63±5.20) %, (18.67±2.94) %, (17.53±2.47) %; t=6.95, t=7.56, both P<0.05). The expression of AVP was increased after different doses of Glu into PVN, and the AVP level in G12 group ((18.15±6.49)%) was higher than that of NS group, the difference was statistically significant (t=-4.21, P<0.05). Compared with the Sham group, the expression of PCNA in colonic mucosal cells of the CVH group and NS group decreased ((65.48±1.68) %, (18.39±1.67) %, (17.69±1.68) %; t=34.35, t=34.80, both P<0.05). The expression of PCNA was increased after different doses of Glu injected into PVN, and the PCNA level in G12 group ((59.91±5.63)%) was higher than that of NS group, the difference was statistically significant (t=-12.44, P<0.05). Compared with the Sham group, the expression of apoptotic cells in colonic mucosal cells of the CVH group and NS group increased ((23.38±11.40)%, (83.79±3.57)%, (80.91±2.47)%; t=-8.77, t=-8.54, both P<0.05). The expression of apoptotic cells was decreased after different doses of Glu into PVN, and the G12 group was ((18.15±6.49) %). Compared with NS group, the difference was statistically significant (t=15.65, P<0.05).@*Conclusion@#Injection of Glu into hypothalamic PVN can alleviate the visceral pain behaviors in CVH rats, and its mechanism may be related to arginine vasopressin.
Résumé
Objective To investigate the effects of glutamate (Glu) injected into hypothalamic pa-raventricular nucleus (PVN) on visceral pain of chronic visceral hypersensitivity (CVH) rats and its possi-ble mechanism. Methods Newborn SD rats were given CVH rat model by colorectal distension (CRD) on the 8th,10th and 12th day after birth. Thirty rats with successful CVH model were randomly divided into CVH model group (CVH group),CVH + injection of saline into PVN group (NS group),CVH+ injection of Glu into PVN (3,6,and 12 μg Glu,namely G3,G6,and G12,respectively),6 rats in each group,and 6 SD rats with matching body mass were taken as sham operation group (Sham group). The pain behavior of the rats was evaluated by pain threshold,abdominal withdrawal reflex (AWR) score,and abdominal external ob-lique muscle electromyography (EMG). The expression of arginine vasopressin (AVP) and the proliferation of colon tissue were detected by immunohistochemical staining. The apoptosis of colon tissue was detected by TUNEL. Results Compared with the NS group, the pain thresholds of the G3, G6 and G12 groups in-creased,and the AWR scores and EMG amplitudes decreased. The differences were statistically significant (Pain threshold:t=7. 65,16. 31,24. 78,both P<0. 05;AWR scores:t=-2. 98,-4. 77,-7. 29,both P<0. 05;EMG amplitudes:t=-3. 06,-5. 75,-8. 92,both P<0. 05). Compared with the Sham group,the expression of AVP in PVN of the CVH group and NS group decreased ((42. 63±5. 20) %,(18. 67±2. 94) %,(17. 53± 2. 47) %; t=6. 95,t=7. 56,both P<0. 05). The expression of AVP was increased after different doses of Glu into PVN,and the AVP level in G12 group ((18. 15±6. 49)%) was higher than that of NS group,the difference was statistically significant (t=-4. 21,P<0. 05). Compared with the Sham group,the expression of PCNA in colonic mucosal cells of the CVH group and NS group decreased ((65. 48±1. 68) %,(18. 39± 1. 67) %,(17. 69±1. 68) %;t=34. 35,t=34. 80,both P<0. 05). The expression of PCNA was increased after different doses of Glu injected into PVN,and the PCNA level in G12 group ((59. 91±5. 63)%) was higher than that of NS group,the difference was statistically significant (t=-12. 44,P<0. 05). Compared with the Sham group,the expression of apoptotic cells in colonic mucosal cells of the CVH group and NS group increased ((23. 38±11. 40)%,(83. 79± 3. 57)%,(80. 91± 2. 47)%;t=-8. 77,t=-8. 54,both P<0. 05). The expression of apoptotic cells was decreased after different doses of Glu into PVN,and the G12 group was ((18. 15±6. 49) %). Compared with NS group,the difference was statistically significant ( t=15. 65,P<0. 05). Conclusion Injection of Glu into hypothalamic PVN can alleviate the visceral pain be-haviors in CVH rats,and its mechanism may be related to arginine vasopressin.