The possible role of P-glycoprotein in the distribution of clarithromycin to the middle ear

Distribution of clarithromycin into the middle ear was investigated in this study with an emphasis on the possible involvement of P-glycoprotein (P-gp). Clarithromycin was given to chinchillas at two dose levels, 30 mg/kg and 50 mg/kg as short intravenous (IV) infusions. In vivo microdialysis with on-line HPLC-MS-MS analysis was used to quantify clarithromycin concentrations in the middle ear fluid (MEF). Clarithromycin showed good penetration into the chinchilla MEF following IV administration. Unbound drug levels in the MEF surpassed those in the plasma about 2 hours after dosing. The unbound drug concentration ratios of MEF to plasma (mean +/- SD) increased from 1.9 +/- 1.0 at 2 hours to 7.1 +/- 3.4 at 12 hours after the end of infusion. The AUC ratio of unbound drug in MEF to plasma at the two dose levels was determined to be 1.58 +/- 0.58 (n=16), which is significantly greater than unity. The effect of a selective P-gP inhibitor, quinidine on clarithromycin clearance out of the middle ear was examined. Clanthromycin (10 mug/ml) alone or with quinidine (200 muM or 400 muM) was given as intrabulla doses and clarithromycin clearance from the middle ear was compared. The P-gp inhibition study with quinidine did not support that P-gp was involved in the distribution of clarithromycin into chinchilla MEF because adding of quinidine changed the unbound clarithromycin concentration in dosing solutions. An RT-PCR method and an immunohistochemistry (IHC) method were used to investigate the existence of P-gP mRNA and the corresponding protein in the middle ear epithelial cells (MEEC). RT-PCR products were detected from rat MEEC and cultured rat MEEC. Sequencing of these products showed that they match the rat P-gp DNA sequence. The RT-PCR method did not detect P-pg mRNA in the chinchilla MEEC tissue due to the lack of chinchilla P-pg mRNA sequence information. The IHC results showed the existence of P-gp protein in the rat cultured MEEC but not the rat MEEC tissue. IHC result in the chinchilla tissue was inconclusive. Nevertheless, these data showed that clarithromycin accumulates in the middle ear.