Abstracts

Rationale: Temporal lobe epilepsy (TLE) is a common form of adult epilepsy involving the limbic structures of the temporal lobe. Layer II neurons of the medial entorhinal cortex (mEC) are spared and hyper-excitable in TLE. Recent studies have suggested a role for T-type Ca2+ channels in facilitating increases in neuronal activity associated with TLE. We sought to determine if T-type Ca2+ channels are involved in maintaining neuronal hyperexcitability of layer II stellate and non-stellate neurons in TLE. Methods: TLE in rats was induced by electrical stimulation of the hippocampus for 90 mins to induce status epilepticus (SE). Rats having two or more spontaneous seizures per day by EEG, 3 months after SE were used in the study. Brain slices were prepared and membrane properties recorded from visually identified mEC layer II neurons. To establish a role for T-type Ca2+ channels we used TTA-P2 at 1 μM, a selective T-type Ca2+ channel antagonist. Results: Action potentials (APs) were evoked by a series of depolarizing current injection steps under whole cell current clamp conditions. Both mEC layer II stellate and non-stellate neurons from TLE rat brain slices had higher AP firing frequencies than control. At a current injection step of 470 pA, stellate frequencies were increased (P<0.05) from 28.0 ± 1.4 Hz (n = 15) in control to 32.5 ± 2.6 Hz (n = 8) in TLE. Bath application of TTA-P2 significantly (P<0.01) reduced firing frequencies in TLE neurons by 30% (to 22.6 ± 2.9 Hz; n=8). In contrast, TTA-P2 had little effect on firing rates in control neurons (24.6 ± 0.9 Hz; n=15). Non-stellate neurons were also hyperexcitable in TLE (21.9 ± 0.7 Hz; n = 7 in control to 33.3 ± 1.6 Hz; n = 13 in TLE, P<0.001). In contrast to stellate neurons, TTA-P2 had little effect on non-stellate firing rates reducing them to 15.9 ± 2.4 Hz (n = 7) in control and 28.3 ± 1.6 Hz (n = 2) in TLE. Stimulation of mEC layer III evoked AP bursts in TLE mEC stellate and non-stellate neurons, but single APs in control neurons. Application of TTA-P2 reduced evoked AP burst firing in TLE stellate and non-stellate neurons. TTA-P2 had little effect on evoked AP's in both types of control neurons. Rebound APs evoked by increasing amplitudes of hyperpolarizing steps evoked single APs in control stellate neurons, but no APs in non-stellate neurons. In TLE neurons hyperpolarizing steps evoked AP bursts in both subtypes of neurons. TTA-P2 had a more pronounced effect on APs in both subtypes of TLE neurons than control neurons, delaying and inhibiting rebound AP bursts. Quantitative RT-PCR (qPCR) of mEC layer II tissue revealed an almost 3-fold increase in Cav3.1 mRNA in TLE tissue as compared to controls. Cav3.2 or Cav3.3 levels were unchanged. Conclusions: These findings suggest that T-type Ca2+ channels, in part, contribute to the increased hyper-excitability of mEC layer II neurons in TLE. Increased expression of the Cav3.1 isoform, in particular, may be important for facilitating and maintaining neuronal hyper-excitability of mEC layer II neurons in TLE.