(R)-[11C]verapamil PET studies to assess changes in P-glycoprotein expression and functionality in rat blood-brain barrier after exposure to kainate-induced status epilepticus

Abstract

Background: Increased functionality of efflux transporters at the blood-brain barrier may contribute to decreased drug concentrations at the target site in CNS diseases like epilepsy. In the rat, pharmacoresistant epilepsy can be mimicked by inducing status epilepticus by intraperitoneal injection of kainate, which leads to development of spontaneous seizures after 3 weeks to 3 months. The aim of this study was to investigate potential changes in P-glycoprotein (P-gp) expression and functionality at an early stage after induction of status epilepticus by kainate.

Methods: (R)-[11C]verapamil, which is currently the most frequently used positron emission tomography (PET) ligand for determining P-gp functionality at the blood-brain barrier, was used in kainate and saline (control) treated rats, at 7 days after treatment. To investigate the effect of P-gp on (R)-[11C]verapamil brain distribution, both groups were studied without or with co-administration of the P-gp inhibitor tariquidar. P-gp expression was determined using immunohistochemistry in post mortem brains. (R)-[11C]verapamil kinetics were analyzed with approaches common in PET research (Logan analysis, and compartmental modelling of individual profiles) as well as by population mixed effects modelling (NONMEM).

Results: All data analysis approaches indicated only modest differences in brain distribution of (R)-[11C]verapamil between saline and kainate treated rats, while tariquidar treatment in both groups resulted in a more than 10-fold increase. NONMEM provided most precise parameter estimates. P-gp expression was found to be similar for kainate and saline treated rats.

Conclusions: P-gp expression and functionality does not seem to change at early stage after induction of anticipated pharmacoresistant epilepsy by kainate.

Figure 1

Mean whole brain uptake of (R)-[¹¹C]verapamil, expressed as SUV (radioactivity normalized to injected dose and rat weight), as function of time in the four rat groups. Vertical bars represent standard deviation. No difference was found in the profiles between vehicle co-administered saline and kainate treated animals. Somewhat slower kinetics was observed in the kainate treated compared to the saline treated rats that were also co-administered with tariquidar.

Figure 2

Logan (left), 1T2k (middle) and 2T4k (right) fits for a vehicle (upper panel) and a tariquidar (lower panel) co-administered saline treated rat. Circles represent measured concentrations, solid lines best fits. The 2T4k model resulted in better fits than the 1T2k model for most of the rats.

Figure 3

The final population model. V_c, V_p1, and V_p2 are pharmacological volumes of distribution in central and two peripheral plasma compartments, respectively. V_br1 and V_br2 are volumes of distribution in central and peripheral brain compartment, respectively. CL, Q₁, Q₂, Q_in, Q_out and Q_br are total body clearance, bidirectional clearance between plasma and peripheral compartment 1, bidirectional clearance between plasma and peripheral compartment 2, clearance into and out of the brain, and bidirectional clearance between central and peripheral brain compartments respectively. The plus-sign indicates an increase in affected parameter estimate, i.e. tariquidar co-administration increased peripheral plasma volume of distribution (V_p1), influx clearance to the brain (Q_in) and brain volume of distribution (V_br1), animal weight increased systemic clearance (CL), and the kainate treatment increased V_br1.

Figure 4

Diagnostic plots for the final population model including both brain and plasma (R)-[¹¹C]verapamil concentrations. All dots represent individual data points and the lines (upper panels) identity lines. Observed versus population and individual predictions are shown in the upper panels, respectively. Most of the data points are randomly distributed around the line of identity which indicates that the model describes the concentrations adequately. Absolute individual weighted residuals versus individual predictions and weighted residuals versus time are shown in the lower panels. Except for some outliers at early time points, most residuals are clustered around zero.

Figure 5

Staining of P-glycoprotein in the brain capillaries in the region of hippocampus. The arrows indicate the stained capillaries in a kainate treated animal.

Figure 6

Mean P-gp expression measured as P-gp labelled area (upper panel) and optical density (lower panel) at 7 days after saline (n = 20) or kainate treatment (n = 22). Vertical bars represent standard deviation. There was no significant difference between the two rat groups, but the P-gp labelled area tended to be somewhat larger for kainate treated rats.