| ObjectiveBased on the organic cation transporters (Octs) and multidrug and toxin extrusion protein 1 (Matel) to study the effects of atenolol/metoprolol on the pharmacokinetics of metformin.Methods1 To study the effects of atenolol on pharmacokinetics of metformin1.1 For the oral administration study, rats were divided into two groups (n=8 per group). Rats in drug co-administration group were orally treated with metformin (100 mg/kg) plus atenolol (20 mg/kg), while rats in metformin group were treated with metformin (100 mg/kg). After the single or 7 days drug administration, serial blood samples (0.25 mL) were collected through the right side of the femoral artery with heparinized syringes at 10,20,40,60,90,120,180,240,360,480, and 720 min, respectively. The plasma concentration of metformin was determined by HPLC, and the pharmacokinetic parameters were calculated using two-compartment model.1.2 For the intravenous administration study, the group setting was as same as section "1.1". Rats in drug co-administration group were orally treated with metformin (100 mg/kg) plus atenolol (20 mg/kg) once daily for 6 days, while rats in metformin group were treated with metformin (100 mg/kg) once daily for 6 days. On day 7, rats in drug co-administration group were orally given atenolol (20 mg/kg) and then intravenously given metformin (25 mg/kg), while rats in metformin group were intravenously given metformin (25 mg/kg). Blood samples were collected at 2,5,10, 20,30,60,90,120,240, and 480 min post metformin, respectively.1.3 To investigate the expression of Oct2 and Matel in rat kidney, the group setting and drug administration was as same as section "1.1". After 7 days drug administration, kidney samples were dissected. Western blotting and immunohistochemistry technique were used to investigate the protein expression of Oct2 and Matel in the kidney of rats.1.4 To test the effect of atenolol on glomerular filtration rate and renal blood flow, the group setting and drug administration was as same as section "1.1". After 7 days drug administration, blood samples were collected from orbital at 2 h post metformin. The plasma concentration of creatinine was determined by LC-MS/MS. In addition, renal blood flow and glomerular filtration rate were also determined using Single-Photon Emission Computed Tomography (SPE-CT).1.5 To investigate the effect of long-term atenolol administration on urinary excretion of metformin, the group setting and drug administration was as same as section "1.1". On day 1,3,7,15,30, and 60, urine samples were collected during the interval of 0-2,2-4,4-6,6-8,8-10,10-12, and 12-24 h post metformin.1.6 To investigate the effect of long-term atenolol administration on the plasma and tissue concentration of metformin as well as biochemical parameters, the group setting and drug administration was as same as section "1.1". On day 1,3,7,15,30, and 60, blood samples were collected from orbital at 2 h post metformin in order to monitor plasma metformin concentration. On day 60, rats were sacrificed at 2 h post metformin, and blood samples were collected from abdominal aorta in order to determinate the plasma level of lactate (LCA), glucose (GLU), aspartate aminotransferase (AST) and alanine aminotransferase (ALT). The residual blood in the body was removed by saline cardiac perfusion, and then tissues, i.e., heart, liver, brain, lung, kidney and small intestine, were dissected, weighed and homogenized in saline for metformin determination. Another part of liver and kidney tissues were also removed and fixed in 10% neutral buffered formalin for histological evaluation.2 To study the effects of metoprolol on pharmacokinetics of metformin2.1 For single or 7 days drug administration, rats were divided into two groups (n=8 per group). Rats in drug co-administration group were orally treated with metformin (100 mg/kg) plus metoprolol (20 mg/kg), while rats in metformin group were treated with metformin (100 mg/kg). After single or 7 days drug administration, serial blood samples (0.25 mL) were collected through the right side of the femoral artery with heparinized syringes at 10,20,40,60,90,120,180,240,360,480, and 720 min post metformin, respectively. The plasma concentration of metformin was determined by HPLC, and the pharmacokinetic parameters were calculated using two-compartment model.2.2 To investigate the effect of metoprolol on uptake of metformin in liver, kidney and thigh muscle, the group setting and drug administration was as same as section "2.1". After single or 7 days drug administration, blood, liver, kidney and thigh muscle were collected at 2 h post metformin in order to determinate the tissue uptake ratio.2.3 To investigate the expression of Octs and Matel in rat liver and kidney, the groups setting and drug administration was as same as section "2.1". After single or 7 days drug administration, liver and kidney samples were dissected. Western blotting was used to investigate the protein expression of Octs and Matel in liver and kidney of rats.2.4 To investigate the effect of long-term metoprolol administration on urinary excretion of metformin, the group setting and drug administration was as same as section "2.1". On day 1,3,7,15,30, and 60, urine samples were collected during the interval of 0-2,2-4,4-6,6-8,8-10,10-12, and 12-24 h post metformin.2.5 To investigate the effect of long-term metoprolol administration on the tissue distribution of metformin as well as biochemical parameters, the groups setting and drug administration was as same as section "2.1". After 60 days drug administration, rats were sacrificed at 2 h post metformin, and blood samples were collected from abdominal aorta in order to determinate the plasma level of LCA, GLU, AST, ALT, uric acid and urea. The residual blood in the body was removed by saline cardiac perfusion, and then tissues, i.e., heart, liver, brain, lung, kidney, thigh muscle and small intestine, were dissected, weighed and homogenized in saline to determinate of metformin concentration. Another part of liver and kidney samples were removed and fixed in 10% neutral buffered formalin for histological evaluation.Results1 Effects of atenolol on pharmacokinetics of metforminAfter single drug administration, there was no significant difference in the plasma concentrations and urinary excretion of metformin between metformin group and drug co-administration group. After 7 days drug administration, the plasma concentrations and parameter AUC0â†'t of metformin in drug co-administration group were significantly increased compared to that of metformin group. However, the cumulative urinary excretions, CL/F and ke of metformin in drug co-administration group were significantly decreased compared to that of metformin group (p<0.01). In addition, the expression of Matel (p<0.01), but not Oct2 (p>0.05), was significantly decreased in drug co-administration group when compared to that of metformin group.The metformin concentrations of kidney and liver in drug co-administration group were significantly increased compared to that of the metformin group after 60 days drug administration. In addition, the LCA levels in drug co-administration group were significantly elevated compared to that of metformin group after 60 days drug administration (p<0.01). Long-term atenolol and metformin administration did not show any effect on the histological structure of kidney and liver tissue.2 Effects of metoprolol on pharmacokinetics of metforminAfter single or 7 days drug administration, plasma concentrations and AUC0â†'t of metformin in drug co-administration group were significantly decreased compared to that of metformin group (p<0.01). However, the V/F of metformin in drug co-administration group was markedly increased compared to that of metformin group (p<0.05). The Kp of metformin in liver, kidney and thigh muscle were significantly increased in drug co-administration group when compared to that of metformin group (p<0.01). After 7 days drug administration, the renal expression of Matel in drug co-administration group was significantly decreased compared to that of metformin group. On the other hand, there were no significant differences in Octs levels between drug co-administration group and metformin group (p<0.05).The cumulative urinary excretion of metformin in drug co-administration group was significantly increased when compared to that of metformin group after single drug administration. However, the cumulative urinary excretion of metformin in drug co-administration group was significantly higher than that of metformin group on day 3,7,15,30, and 60, respectively (p<0.05). After 60 days drug administration, the metformin concentration of liver, kidney and thigh muscle in drug co-administration group were significantly elevated compared to that of metformin group (p<0.01). In addition, LCA and uric acid levels in drug co-administration group were significantly higher than that of metformin group after 60 days drug administration (p<0.01). The results also found that long-term metoprolol and metformin administration did not show any effect on the histological structure of kidney and liver.Conclusions1 After 7 days drug administration, atenolol inhibited the urinary excretion of metformin and increased plasma concentration of metformin via down-regulating the expression of Matel, but not Oct2, in the kidney of rats. On the other hand, long-term atenolol administration increased the metformin concentration of liver and kidney as well as plasma LCA levels.2 After single or 7 days drug administration, metoprolol decreased the plasma concentration of metformin as a result of increasing the uptake of metformin in liver, kidney and thigh muscle. After 7 days drug administration, metoprolol inhibited the urinary excretion of metformin via down-regulating the expression of Matel, but not Octl and Oct2, in the kidney of rats. In addition, long-term metoprolol administration elevated metformin concentration of liver, kidney and thigh muscle as well as the level of LCA and uric acid. |
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