Pharmacokinetic Stuidies of 5-HT3 Receptor Antagonists in Rats: Microdialysis Application

• Main Authors: Chien-Tsai Huang, 黃建才
• Other Authors: Chieh-Fu Chen
• Format: Others
• Language: zh-TW
• Published: 2000
• Online Access: http://ndltd.ncl.edu.tw/handle/25014790108048606303

博士 === 國立陽明大學 === 藥理學研究所 === 88 === 5-HT3 receptor antagonists (granisetron, ondansetron, tropisetron), are potent and highly selective antagonists to 5-HT3 receptors in the peripheral vagus nerve and vomiting center. They are particularly effective for the relief of nausea and vomiting induced by for cancer therapy. In this study, we used an in vivo microdialysis technique to investigate in the rat the peripheral disposition and the distribution in the central nervous system of 5-HT3 receptor antagonists, the drug interaction with P-glycoprotein inhibitors, the pharmacokinetics differences in the anesthetized with awake rats, and the enterohepatic circulation of the drugs. We implanted microdialysis probes into the jugular vein, cerebral frontal cortex and bile duct of the rat, and then 5-HT3 receptor antagonists alone or with P-glycoprotein inhibitors were intravenously administered via the femoral vein. Unbound-drug sample were collected and each sample 20μl of dialysate was assayed using the high-performance liquid chromatographic system. Pharmacokinetics parameters were calculated from the corrected dialysate concentration of 5-HT3 receptor antagonists versus time data. 1. Pharmacokinetic study of drug in the rat blood This microdialysis sampling method facilitates pharmacokinetic studies by reducing the effects of blood volume changes as compared with conventional blood withdrawing assays. Tropisetron (5 or 10 mg/kg, i.v.) was administered and then a microdialysis system coupled with liquid chromatogeaphy was applied for the measurement of unbound tropisetron in the rat blood. Rapid distribution followed by a slower elimination phase was observed from the blood concentration-time curve. The disposition of tropisetron at each dose fitted well to a two-compartment model. Microdialysis is an in vivo sampling technique that allows continuously determination of drug concentration from protein unbound and extracellular space of most tissue and to provide near-time analysis of tropisetron in blood dialysate samples, and therefore minimal strain on hemodyanamics. 2. Pharmacokinetic study of drug in the rat blood and brain Simultaneous microdialysis probes in the blood and brain with sensitive high-performance liquid chromatography-fluoresence detection were used to examine the granisetron concentration in the blood and frontal cortex of rats after granisetron (6mg/kg, i.v.) administration. Our results showed that the average brain/blood ratio of granisetron deduced from the areas under the concentration curves continued to increase between 12 to 144 min after drug administration. This enhanced ratio suggested that granisetron penetrated the blood-brain barrier. The microdialysis technique provides protein-free drug samples from blood and brain, which can be injected directly into a chromatographic system for continuous in vivo monitoring. Furthermore, compared with other in vivo method for pharmacokinetic study, this microdialysis sampling method causes less tissue damage in both the peripheral circulation system and the central nervous system. 3. Pharmacokinetic study of drug in the rat bile This study was designed to evaluate the biliary excretion of granisetron employing a specially designed microdialysis shunt probe to monitor biliary drug concentrations without fluid losses. To avoid obstruction of the bile duct or bile salts waste, the shunt linear probe connected the bile duct between liver and duodenum. The utility of this microdialysis design was demonstrated by studying the biliary excretion of granisetron in the rats. Following intravenous administration (3 mg/kg), granisetron was measurable in the bile microdialysate sample with chromatographic analysis. It was concluded that the in vivo microdialysis technique yielded useful data on the biliary excretion of granisetron. 4. Pharmacokinetic study of drug in the rat blood and bile The present study utilized the minimally invasive sampling technique of microdialysis in pharmacokinetics and biliary excretion studies. We used dual-site in vivo microdialysis sampling to estimate the biliary excretion of unbound tropisetron concentration during the steady state in the rats. Microdialysis probes were implanted into the jugular vein and bile duct of the male Sprague-Dawley rat. Tropisetron ( 20 mg/kg, i.v., n=5) was then administered via a femoral vein of rat. Unbound tropisetron were identified using a mobile phase containing acetonitrile -100 mM sodium phosphate (pH 5.0, 22-24:76-78, v/v) with ultraviolet detection. Parmacokinetics parameters were calculated from the corrected microdialysate concentrations of tropisetron versus time data. The areas under the concentration curves (AUCs) for blood and bile were 556.67±97.51 and 194.69±13.12 for 20mg/kg, respectively, indicate the disposition of tropisetron in bile system. This biliary excretion of the tropisetron might be undergoing enterohepatic circulation. These studies provide evidence for the biliary excretion of unbound tropisetron in rats. 5. Pharmacokinetic study of drug in the rat blood, brain and bile Studies were conducted to simultaneous measurement the unbound ondansetron in the rat blood, brain and bile duct using a microdialysis system coupled with a high-performance liquid chromatography. We implanted microdialysis probes into the jugular vein, cerebral frontal cortex, and bile duct of the male Sprague-Dawley rat. Ondansetron (10 or 20 mg/kg, i.v., n=6) was then administered via a femoral vein of rat. Samples were eluted with a mobile phase containing acetonitrile -25 mM sodium acetate (pH 4.8, 28:72, v/v). Parmacokinetics parameters were calculated from the corrected microdialysate concentrations of ondansetron versus time data. The brain/blood ratios of ondansetron were 0.12±0.02 and 0.11±0.03 for 10 or 20mg/kg, respectively, indicating the penetration of ondansetron of the central nervous system. The bile/blood ratios of ondansetron were 0.36±0.01 and 0.36±0.03 for 10 or 20mg/kg, respectively, indicating that biliary excretion of ondansetron was dose-independent. The results suggested that simultaneous multiple-site microdialysis in blood, brain and bile can be usefully applied to study the pharmacokinetics of ondansetron in the periphery and the central nervous system. 6. Pharmacokinetic study of drug interaction on the 5-HT3 antagonists with P-glycoprotein inhibitor Cyclosporin A (Sandimmune) is a very potent and specific drug-transporting P-glycoprotein inhibitor. Previous study data suggest that antiemetic 5-HT3 antagonists, ondansetron and granisetron, across the blood-brain barrier of the mice may be augmented by P-glycoprotein inhibition. To test this, we used the simultaneous microdialysis method for evaluated the effect of cyclosporin A on the pharmacokinetics of the unbound ondansetron and granisetron in the rat blood, bile duct and brain. Male Sprague-Dawley rats received either ondansetron (10 or 20 mg/kg, i.v., n=6) or granisetron (10 mg/kg, i.v.) with cyclosporin A (20 mg/kg i.v.) or ondansetron, granisetron alone. Unbound ondansetron and granisetron concentrations were monitored by a simultaneous microdialysis system coupled with a high-performance liquid chromatography. We found that the area under of the concentration-time curve for unbound ondansetron and granisetron in brain was significantly increased in the cyclosporin A-treated rats compared with control rats, but in blood and bile were not altered significantly. The mean-residence time for unbound ondansetron and granisetron in the brain was significantly increase in the cyclosporin A-treated rats compared with control rats and a decrease in ondansetron clearance, but not in the blood and bile. The present results suggest that P-glycoprotein inhibitors might be useful in specifically enhancing the penetration of blood-brain barrier of ondansetron and granisetron into the central nervous system. 7. Pharmacokinetic study of drug in anesthetized and awake, freely-moving animals Previously reports suggest that general anesthetics is extremely perturbing to the pharmacokinetics of drugs, due may be to interfer with metabolic enzyme activity or the cardiovascular system. Microdialysis probe was implanted into the jugular vein of the male Sprague-Dawley rats. Ondansetron (10 and 20 mg/kg, i.v., n=6) was then administered via the femoral vein. We found that the area under of the concentration-time curve for unbound ondansetron in the anesthetized rats was significantly higher than in the awake, freely-moving rats, and clearance was also profoundly faster, and indicating dose-dependence. This finding suggests that using a conscious, freely-moving rats provides near real physiological conditions for pharmacokinetics studies. 8. Pharmacokinetic study of bioavailability of unbound drugs Drugs given orally are normally absorbed in gastrointestinal tract and transported via the mesenteric vessels to the hepatic portal vein and then to the liver prior to the systemic circulation. First-pass effects may be suspected when there is a lack of parent drug in the systemic circulation after oral administration. In such a case, the bioavailability for a drug given orally is less than the bioavailability for the same dose of drug given intravenously. Microdialysis probe was implanted into the jugular vein of the male Sprague-Dawley rat. Ondansetron (10 and 20 mg/kg, i.v., n=6) and then administered via a femoral vein and orally in awake, freely-moving animals. Estimate for the bioavailability of unbound ondansetron after oral administration of ondansetron was 20.36%. It has been previously suggested that ondansetron is secreted into the bile and excreted into the duodenum via the common bile duct. Subsequently, the parent ondansetron might be reabsorbed from intestine and become systemically circulation available, through enterohepatic circulation. 9. Pharmacokinetic study of the enterohepatic circulation of drugs Certain drugs that undergo biliary excretion may be reabsorbed in the gastrointestinal tract again through enterohepatic circulation. The biliary excretion of a drug that might be undergo the enterohepatic circulation and may significantly increase the area under blood drug concentration curve and pharmacological effects. If enterohepatic circulation is interrupted then there might be a decrease in blood drug concentration and therapeutic effects. A paired rat model consisting of a hepato-duodenal shunt, in which the bile of a drug-treated donor rat was transported to the duodenum of an untreated recipient rat via a bile cannula, was used to assess the effects of enterohepatic circulation on the pharmacokinetics of ondansetron. Results indicated that a minor extent of the parent ondansetron was excreted into the bile, and reabsorbed from intestine, to undergo enterohepatic circulation. and pharmacological effects. If enterohepatic circulation interruption, might be decrease in blood drug concentration and therapeutic effects. A paired rat model consisting of a hepato-duodenal shunt in which the bile of a drug- Conclusions: (1). In vivo pharmacokinetics of 5-HT3 antagonists were studied in the rat blood, brain and bile by simultaneous multiple-site microdialysis, demonstrating that it was feasible to simultaneously monitor the disposition of drugs, (2) the results of these studies suggest granisetron and ondansetron of the 5-HT3 antagonists rapidly penetrated the blood-brain barrier, (3) the results of these studies suggest that granisetron, ondansetron and tropisetron are excreted through the bile duct to the duodenum, (4) the P-glycoprotein inhibitor cyclosporin A significantly elevated the AUC of unbound granisetron and ondansetron in the cerebral frontal cortex, (5) the microdialysis sampling method coupled with HPLC and ultraviolet or fluorescence detection has provide sufficient sensitivity, accuracy and reproducibility is suitable for pharmacokinetics studies.