Pilocarpine-Induced Convulsive Activity Is Limited by Multidrug Transporters at the Rodent Blood-Brain Barrier

Abstract

As a result of the growing availability of genetically engineered mouse lines, the pilocarpine post-status epilepticus (SE) model of temporal lobe epilepsy is increasingly used in mice. A discrepancy in pilocarpine sensitivity in FVB/N wild-type versus P-glycoprotein (PGP)-deficient mice precipitated the investigation of the interaction between pilocarpine and two major multidrug transporters at the blood-brain barrier. Doses of pilocarpine necessary for SE induction were determined in male and female wild-type and PGP-deficient mice. Brain and plasma concentrations were measured following low (30-50 mg×kg 21 i.p.) and/or high (200 mg×kg 21 i.p.) doses of pilocarpine in wild-type mice, and mice lacking PGP, breast cancer resistance protein (BCRP), or both transporters, as well as in rats with or without pretreatment with lithium chloride or tariquidar. Concentration equilibrium transport assays (CETA) were performed using cells overexpressing murine PGP or BCRP. Lower pilocarpine doses were necessary for SE induction in PGP-deficient mice. Brain-plasma ratios were higher in mice lacking PGP or PGP and BCRP, which was also observed after pretreatment with tariquidar in mice and in rats. Lithium chloride did not change brain penetration of pilocarpine. CETA confirmed transport of pilocarpine by PGP and BCRP. Pilocarpine is a substrate of PGP and BCRP at the rodent blood-brain barrier, which restricts its convulsive action. Future studies to reveal whether strain differences in pilocarpine sensitivity derive from differences in multidrug transporter expression levels are warranted.

A face-to-face comparison of claudin-5 transduced human brain endothelial (hCMEC/D3) cells with porcine brain endothelial cells as blood–brain barrier models for drug transport studies

“…5a, significant Pgp-mediated transport of the selective Pgp substrate dLop [37] was determined in pBCECs. As shown previously for various other Pgp substrates in the CETA assay [27,[38][39][40][41][42], asymmetrical (basolateral to apical) transport of dLop was indicated by the significant increase of drug concentration in the apical chamber and simultaneous decrease of drug concentration in the basolateral chamber, demonstrating basolateral-to-apical drug transport across the pBCEC monolayer in the Transwells. This…”

Functional Neuroplasticity in the Nucleus Tractus Solitarius and Increased Risk of Sudden Death in Mice with Acquired Temporal Lobe Epilepsy

“…A large proportion of the mice that survived pilocarpine-induced SE died suddenly and unexpectedly at time points corresponding with the development of spontaneous seizures ( Shibley and Smith, 2002 ; Winokur et al, 2004 ; Bhaskaran and Smith, 2010b ), promoting this mouse as a reasonable model of SUDEP in TLE. Pilocarpine plasma and brain levels peak in the minutes after injection and fall to almost zero by 2 h postinjection; it is therefore doubtful that the single exposure to pilocarpine itself is responsible for our findings, which were measured days to months after injection ( Römermann et al, 2015 ). Additionally, microinjection of muscarinic receptor agonists in the NTS alters function for 4–6 min after application without sustained changes ( Sundaram et al, 1988 ).…”

The evolution of the pilocarpine animal model of status epilepticus

“…One possibility for the observed pilocarpine resistance is the presence of the multidrug transporter P-glycoprotein (PGP), which is a transporter that removes pilocarpine from the BBB, serving a protective role. Studies have shown that PGP knockout mice require a lower dose of pilocarpine for SE induction than their wild-type counterparts ( Römermann et al., 2015 ). Studies have revealed that PGP knockout mice injected with a low dose of pilocarpine do not show any increase in brain uptake of the drug, but those injected with a higher dose show significantly increased brain uptake ( Sahin et al., 2003 ; van Vliet et al., 2007 ; Li et al., 2013 ).…”

“…In addition, pre-treatment of wild-type mice with the PGP inhibitor tariquidar causes an increase in pilocarpine brain uptake similar to that observed in PGP knockout mice, thus emphasizing that pilocarpine is carried by the PGP multidrug transporter ( Bankstahl et al., 2013 ). Female wild-type mice reportedly require a higher pilocarpine dose for SE induction than males ( Römermann et al., 2015 ). The absence of this phenomenon in PGP knockout mice could therefore be explained by a PGP-dependent mechanism ( Römermann et al., 2015 ).…”

“…Female wild-type mice reportedly require a higher pilocarpine dose for SE induction than males ( Römermann et al., 2015 ). The absence of this phenomenon in PGP knockout mice could therefore be explained by a PGP-dependent mechanism ( Römermann et al., 2015 ). Verapamil, a common PGP substrate, showed lower brain uptake in female mice than male mice, suggesting higher PGP expression levels in female mice ( Dagenais et al., 2001 ).…”