Abstracts

Rationale: The development of epilepsy, particulary temporal lobe epilepsy (TLE), can be induced by a variety of brain insults, including status epilepticus (SE). During this process, named epileptogenesis, a variety of molecular and functional changes progress in the brain, but the mechanisms underlying epileptogenesis are still poorly understood. Recent studies indicate that the development of hyperexcitability of neurons and neuronal circuits because of expression changes of cation-chloride-co-transporters might play a crucial role in epileptogenesis. In this context the K+-Cl- co-transporter KCC2 is downregulated, while the Na+-K+-2Cl- co-transporter NKCC1 is upregulated. In consequence, this leads to a GABA-shift from an inhibitory to an excitatory action caused by an accumulation of intracellular chloride. Several studies indicate that NKCC1 is upregulated in models of TLE and that the diuretic drug bumetanide, a selective NKCC1-inhibitor, is a useful tool to counteract the upregulation of NKCC1, and so, may prevent neuronal hyperexcitability during epileptogenesis. The aims of this study are to investigate in the pilocarpine model of TLE whether bumetanide has an effect on alterations of seizure susceptibility developing after a pilocarpine-induced SE, and to determine intracellular chloride levels and GABAergic transmission after sustained seizure activity. Methods: NMRI-mice were used for inducing SE with pilocarpine. Seizure susceptibility was determined by timed i.v. infusion of the GABAA receptor antagonist pentylenetetrazole (PTZ) 24 hours, 48 hours and 1 week after SE. Animals were pretreated with bumetanide or vehicle, respectively, twenty minutes before starting the PTZ infusion. Control animals without SE received bumetanide or vehicle, respectively. In electrophysiological studies, the perforated patch clamp method is used to determine alterations of intracellular chloride levels after SE.Results: Unexpectedly, an increased PTZ threshold was determined after SE. Bumetanide had no effect on the PTZ threshold in control (non-SE) mice. However, bumetanide partly counteracted the increase of PTZ threshold following SE, possibly indicating an involvement of NKCC1 in the alteration in PTZ seizure threshold. Conclusions: Previous studies have shown that NKCC1 is upregulated within 24 hours following a pilocarpine-induced SE in mice and remains increased for at least 2 weeks. In the present study, we determined an increase in seizure threshold to the GABAA receptor antagonist PTZ 24 and 48 hours following SE. This threshold increase could reflect increased GABAergic transmission as a result of sustained seizure activity. However, at least in part, the NKCC1 inhibitor bumetanide counteracted the increase in seizure threshold following SE. Electrophysiological studies are currently under way, to determine intracellular chloride levels and GABAergic transmission in the hippocampus, and to further evaluate bumetanide s effects during epileptogenesis at different time points after SE.