Proceedings of The Physiological Society

Puerto de la Cruz, Tenerife (2003) J Physiol 548P, P181

Poster Communications

Increased mRNA expression of a potassium-chloride co-transporter in chronically epileptic rats

Begonia Morales-Aza, Gavin L. Woodhall, Roland S.G. Jones and Lucy F. Donaldson

Department of Physiology, School of Medical Sciences, University of Bristol, Bristol BS8 1TD, UK

One factor which may determine the strength of GABAergic inhibition in cortical neurones is the expression of the electro-neutral neuronal-specific potassium-chloride co-transporter, KCC2 (Payne et al. 1996). KCC2 helps to maintain the transmembrane Cl- concentration providing a driving force for GABAA receptor-mediated Cl- currents, which are inward in adult cortical neurones. In states of altered neuronal excitability such as experimental epilepsy, numerous aspects of GABAergic function may be altered. In the present study we examined the hypothesis that altered KCC2 expression/function could contribute to epileptogenesis in specific neuronal areas.

An acute state of status epilepticus was induced in male Wistar rats by repeated (30 min intervals) low doses (5 mg kg-1) of pilocarpine 24 h following administration of 320 meq kg-1 LiSO4 (see Glien et al. 2001). Four to six weeks following this acute epileptic episode rats developed recurrent spontaneous seizures. Following a period of 4-6 weeks of chronic seizure activity the expression of KCC2 mRNA was determined using in situ hybridisation. Animals (2 epileptic, 2 age-matched controls) were anaesthetized with ketamine (120 mg kg-1 I.M.), decapitated and the brains removed. Brains were rapidly frozen, sectioned horizontally at 10 µm and subject to in situ hybridisation as previously described (Payne et al. 1996). mRNA levels were determined by silver grain counting of 12-15 fields from each brain area, in each of three sections from each animal. Regions studied were the superficial and deep layers of the lateral and medial entorhinal cortices (EC), hippocampal areas CA1, CA2, CA3 and the dentate gyrus.

KCC2 mRNA expression was high in both hippocampus and entorhinal cortex. There was no apparent change in expression in epileptic animals for lateral EC (control 27915 ± 4680 pixels/field (P/F), epileptic 33206 ± 6574 P/F; mean ± S.E.M.) or CA1-3 regions of the hippocampus (104-120 % control values). In contrast, KCC2 mRNA expression was slightly increased (control 28715 ± 4189 P/F vs. epileptic 36519 ± 5793 P/F; 127% control) in the deep layers of the medial EC and substantially increased in the superficial layers (control 29027 ± 4776 P/F vs. epileptic 49283 ± 8743 P/F; 170% control). In addition, KCC2 mRNA expression was also significantly increased in the dentate gyrus (control 86640 ± 11286 P/F vs. epileptic 122514 ± 11606 P/F; 141% control) of epileptic rats.

Assuming that KCC2 is localised to postsynaptic sites, these data could suggest that GABAA-receptor mediated responses may be strengthened by an increased inward Cl- gradient in the EC and dentate gyrus. The implications of this for epileptogenesis remain to be determined.

Where applicable, experiments conform with Society ethical requirements