TY - JOUR

T1 - Bumepamine, a brain-permeant benzylamine derivative of bumetanide, does not inhibit NKCC1 but is more potent to enhance phenobarbital's anti-seizure efficacy

AU - Brandt, Claudia

AU - Seja, Patricia

AU - Töllner, Kathrin

AU - Römermann, Kerstin

AU - Hampel, Philip

AU - Kalesse, Markus

AU - Kipper, Andi

AU - Feit, Peter W.

AU - Lykke, Kasper

AU - Toft-Bertelsen, Trine Lisberg

AU - Paavilainen, Pauliina

AU - Spoljaric, Inkeri

AU - Puskarjov, Martin

AU - MacAulay, Nanna

AU - Kaila, Kai

AU - Löscher, Wolfgang

N1 - Funding information: We thank Biff Forbush, Yale School of Medicine (CT, USA), for providing the hNKCC1a construct and for extensive discussion during preparation of the manuscript, Søren Nielsen, Aarhus University (Denmark) for providing rat AQP4.M23, Thomas Erker (University of Vienna) for providing an initial sample of bumepamine, and Andrea J. Yool, University of Adelaide (Australia), for providing a sample of AqB013. The experiments were supported by the Deutsche Forschungsgemeinschaft (grant number LO 274/15- 1) (to WL); the ERC Grant 2013-AdG 341116 (to K.K.); and the Academy of Finland (K.K.); and the Danish Medical Research Council (NM). We thank Edith Kaczmarek, Martina Gramer, and Charlotte Goos Iversen for skillful technical assistance. We thank Biff Forbush, Yale School of Medicine (CT, USA), for providing the hNKCC1a construct and for extensive discussion during preparation of the manuscript, Søren Nielsen, Aarhus University (Denmark) for providing rat AQP4.M23, Thomas Erker (University of Vienna) for providing an initial sample of bumepamine, and Andrea J. Yool, University of Adelaide (Australia), for providing a sample of AqB013. The experiments were supported by the Deutsche Forschungsgemeinschaft (grant number LO 274/15-1) (to WL); the ERC Grant 2013-AdG 341116 (to K.K.); and the Academy of Finland (K.K.); and the Danish Medical Research Council (NM). We thank Edith Kaczmarek, Martina Gramer, and Charlotte Goos Iversen for skillful technical assistance.

PY - 2018/12

Y1 - 2018/12

N2 - Based on the potential role of Na-K-Cl cotransporters (NKCCs) in epileptic seizures, the loop diuretic bumetanide, which blocks the NKCC1 isoforms NKCC1 and NKCC2, has been tested as an adjunct with phenobarbital to suppress seizures. However, because of its physicochemical properties, bumetanide only poorly penetrates through the blood-brain barrier. Thus, concentrations needed to inhibit NKCC1 in hippocampal and neocortical neurons are not reached when using doses (0.1–0.5 mg/kg) in the range of those approved for use as a diuretic in humans. This prompted us to search for a bumetanide derivative that more easily penetrates into the brain. Here we show that bumepamine, a lipophilic benzylamine derivative of bumetanide, exhibits much higher brain penetration than bumetanide and is more potent than the parent drug to potentiate phenobarbital's anticonvulsant effect in two rodent models of chronic difficult-to-treat epilepsy, amygdala kindling in rats and the pilocarpine model in mice. However, bumepamine suppressed NKCC1-dependent giant depolarizing potentials (GDPs) in neonatal rat hippocampal slices much less effectively than bumetanide and did not inhibit GABA-induced Ca2+ transients in the slices, indicating that bumepamine does not inhibit NKCC1. This was substantiated by an oocyte assay, in which bumepamine did not block NKCC1a and NKCC1b after either extra- or intracellular application, whereas bumetanide potently blocked both variants of NKCC1. Experiments with equilibrium dialysis showed high unspecific tissue binding of bumetanide in the brain, which, in addition to its poor brain penetration, further reduces functionally relevant brain concentrations of this drug. These data show that CNS effects of bumetanide previously thought to be mediated by NKCC1 inhibition can also be achieved by a close derivative that does not share this mechanism. Bumepamine has several advantages over bumetanide for CNS targeting, including lower diuretic potency, much higher brain permeability, and higher efficacy to potentiate the anti-seizure effect of phenobarbital.

AB - Based on the potential role of Na-K-Cl cotransporters (NKCCs) in epileptic seizures, the loop diuretic bumetanide, which blocks the NKCC1 isoforms NKCC1 and NKCC2, has been tested as an adjunct with phenobarbital to suppress seizures. However, because of its physicochemical properties, bumetanide only poorly penetrates through the blood-brain barrier. Thus, concentrations needed to inhibit NKCC1 in hippocampal and neocortical neurons are not reached when using doses (0.1–0.5 mg/kg) in the range of those approved for use as a diuretic in humans. This prompted us to search for a bumetanide derivative that more easily penetrates into the brain. Here we show that bumepamine, a lipophilic benzylamine derivative of bumetanide, exhibits much higher brain penetration than bumetanide and is more potent than the parent drug to potentiate phenobarbital's anticonvulsant effect in two rodent models of chronic difficult-to-treat epilepsy, amygdala kindling in rats and the pilocarpine model in mice. However, bumepamine suppressed NKCC1-dependent giant depolarizing potentials (GDPs) in neonatal rat hippocampal slices much less effectively than bumetanide and did not inhibit GABA-induced Ca2+ transients in the slices, indicating that bumepamine does not inhibit NKCC1. This was substantiated by an oocyte assay, in which bumepamine did not block NKCC1a and NKCC1b after either extra- or intracellular application, whereas bumetanide potently blocked both variants of NKCC1. Experiments with equilibrium dialysis showed high unspecific tissue binding of bumetanide in the brain, which, in addition to its poor brain penetration, further reduces functionally relevant brain concentrations of this drug. These data show that CNS effects of bumetanide previously thought to be mediated by NKCC1 inhibition can also be achieved by a close derivative that does not share this mechanism. Bumepamine has several advantages over bumetanide for CNS targeting, including lower diuretic potency, much higher brain permeability, and higher efficacy to potentiate the anti-seizure effect of phenobarbital.

KW - Anti-seizure drugs

KW - Epilepsy

KW - GABA

KW - Giant depolarizing potentials

KW - Neonatal seizures

UR - http://www.scopus.com/inward/record.url?scp=85056174866&partnerID=8YFLogxK

U2 - 10.1016/j.neuropharm.2018.09.025

DO - 10.1016/j.neuropharm.2018.09.025

M3 - Article

C2 - 30248303

AN - SCOPUS:85056174866

VL - 143

SP - 186

EP - 204

JO - NEUROPHARMACOLOGY

JF - NEUROPHARMACOLOGY

SN - 0028-3908

ER -