Reconstructions of Kp brain ratios (P-glycoprotein function at the blood-brain barrier) and Kp brain values of P-glycoprotein substrate verapamil in control, pentylenetetrazole, phenytoin, and spontaneous model of epilepsy mice

Reconstructed brain-to-plasma concentration ratios (Kp brain) were calculated from the reported in vitro P-glycoprotein (P-gp) efflux ratio of verapamil (Uchida et al., 2011a), the reported protein expression level of P-gp/mdr1a in P-gp/mdr1a–expressing LLC-PK1 cell monolayer (Uchida et al., 2011a), and the protein expression level of P-gp/mdr1a in brain capillaries isolated from control, PTZ, EL, and PHT mice, using eq. 2 described in Materials and Methods. Unbound fractions of verapamil in plasma (fu,plasma) and brain (fu,brain) were determined by the equilibrium dialysis method and brain slice uptake method, respectively. Plasma was spiked with 500 nM verapamil and dialyzed against phosphate-buffered saline buffer (pH 7.4) at 37°C for 6 hours (n = 3–4). Brain slices (500 μm) were incubated in extracellular fluid buffer (pH 7.4) containing 50 nM verapamil at 37°C for 6 hours (n = 3–4). The reconstructed Kp brain values were calculated from the reconstructed Kp brain ratios and the fu,plasma and fu,brain values using eq. 3 described in Materials and Methods. Each value represents the mean ± S.E.M. The S.E.M. was calculated according to the law of propagation of error.

Reconstructed Kp brain ratiofu,plasmafu,brainReconstructed Kp brain
(ml/g brain)
Control10.3 ± 0.80.125 ± 0.0050.0246 ± 0.00140.493 ± 0.034
PTZ13.0 ± 1.10.109 ± 0.0070.0237 ± 0.00090.353 ± 0.026
EL14.3 ± 1.30.128 ± 0.0050.0250 ± 0.00150.358 ± 0.028
PHT18.9 ± 1.50.121 ± 0.0040.0231 ± 0.00050.277 ± 0.020