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Voltage-gated Na(+) channels control neuronal excitability and are the primary target for the majority of anti-epileptic drugs. This study investigates the (sub)cellular expression patterns of three important brain-associated Na(+) channel α subunits: NaV1.1, NaV1.2 and NaV1.6 during epileptogenesis (induced by kainic acid) using time points that cover the period from induction to the chronic phase of epilepsy. NaV1.1 immunoreactivity was persistently reduced at 1 day, 3 weeks and 2 months after SE in CA1 and CA3. About 50% of the NaV1.1-positive interneurons was lost at one day after SE in all regions investigated. In the hilus a similar reduction in NeuN-positive neurons was found, while in the CA1 and CA3 region the loss in NeuN-positive neurons only reached 15% in the chronic phase of epilepsy. This implies a stronger shift in the balance between excitation and inhibition toward excitation in the CA1 and CA3 region than in the hilus. NaV1.2 immunoreactivity in the inner molecular layer of the dentate gyrus was lower than control at 1 day after SE. It increased at 3 weeks and 2 months after SE in the inner molecular layer and overlapped with sprouted mossy fibers. NaV1.6 immunoreactivity in the dendritic region of CA1 and CA3 was persistently reduced at all time-points during epileptogenesis. Some astrocytes expressed NaV1.1 and NaV1.6 at 3 weeks after SE. Expression data alone are not sufficient to explain changes in network stability, or infer causality in epileptogenesis. These results demonstrate that hippocampal sub-regional expression of NaV1.1, NaV1.2 and NaV1.6 Na(+) channel α subunits is altered during epileptogenesis in a time and location specific way. This implies that understanding epileptogenesis has to take into account several distinct and type-specific changes in sodium channel expression., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Aim: To study whether the functional properties of sodium channels, and subsequently the channel modulation by carbamazepine (CBZ) in hippocampal CA1 neurons can be changed after epileptic seizures., Methods: We used the acutely dissociated hippocampal CA1 pyramidal cells from epilepsy model rats 3 weeks and 3 months respectively after kainate injection, and whole-cell voltage-clamp techniques., Results: After long-term epileptic seizures, both sodium channel voltage-dependence of activation and steady-state inactivation shifted to more hyperpolarizing potentials, which resulted in the enlarged window current; the membrane density of sodium current decreased and the time constant of recovery from inactivation increased. CBZ displayed unchanged efficacy on sodium channels, with a similar binding rate to them, except that at higher concentrations, the voltage shift of inactivation was reduced. For the short-term kainate model rats, no differences were detected between the control and epilepsy groups., Conclusion: These results indicate that the properties of sodium channels in acutely dissociated hippocampal neurons could be changed following long-term epilepsy, but the alternation might not be enough to induce the channel resistance to CBZ.

Overexposure to corticosteroid hormones is harmful to hippocampal neuronal integrity, likely by perturbation of calcium homeostasis. To identify molecular mechanisms at the single-cell level, we characterized mRNA expression corresponding to voltage- and ligand-gated Ca channels in individual dissociated CA1 neurons in response to long-term corticosterone (CORT) exposure. Predominant mineralocorticoid receptor occupation (ADC-LO group) resulted in low levels of P/Q- and L-type Ca channel mRNAs, high levels of GluR-2 versus GluR-1, and a high ratio of NMDAR-2A to NMDAR-2B mRNA. Corresponding alterations in protein expression were consistent with the restriction of Ca influx. In contrast, additional glucocorticoid receptor occupation (ADC-HI group) altered the expression of these mRNAs in a manner consistent with enhanced Ca influx; interestingly, qualitatively similar alterations were seen in control ADX neurons. Electrophysiological data from the same neurons indicate that Ca current amplitudes also are modulated by CORT, although on a shorter time scale. Finally, principal components analysis (PCA) suggests that neuronal AMPA and NMDA receptor composition may be regulated by MR and GR activation in a complex manner. Therefore, our data implicate molecular events by which CORT may regulate Ca influx into CA1 hippocampal neurons.

Previously it was found that the amplitude of Ca currents in CA1 hippocampal neurons increases after adrenalectomy (ADX) of young adult rats. Preliminary data suggested that this effect of ADX is age-dependent. In the present study we therefore investigated the effect of ADX on Ca currents in three age groups: rats that were 1, 3, or 6 months old. It appeared that ADX of the youngest age group resulted in a selective enhancement of sustained, high threshold Ca currents. By contrast, ADX of the oldest rats enhanced only the low threshold, transient Ca current amplitude. The age dependency of the ADX-induced effects can be explained by developmental changes in Ca current properties in the adrenally intact rats with which ADX animals were compared. Data from acutely dissociated cells, which lack most of their dendrites, suggest that the ADX-induced changes of Ca current properties are at least partly targeted at currents generated in the distal dendrites. No significant changes were observed with age or after ADX for the mRNA expression of the alpha 1D Ca channel subunit, which forms the ionpore of the sustained high threshold L-type Ca channel. We can therefore presently not exclude that the altered amplitude of Ca current with age and ADX is due to changes in ion channel function rather than in the total number of these channels.

Consequences of corticosteroid receptor activation on voltage-dependent Na+ conductances were studied in acutely dissociated CA1 hippocampal neurons. This preparation was selected because of the compact electrotonic properties of dissociated neurons, allowing reliable voltage-clamp of the large and fast Na+ currents. The Na+ currents were studied in (i) neurons of adrenalectomized animals (no steroid receptors occupied), (ii) neurons from tissue of adrenalectomized rats treated in vitro with corticosterone and the glucocorticoid receptor antagonist RU38486 (selectively occupying the mineralocorticoid receptor), (iii) corticosterone-treated neurons of adrenalectomized animals (occupying both the mineralocorticoid and glucocorticoid receptors) and (iv) neurons of sham-operated animals. Activation and steady-state inactivation properties of the Na+ current recorded in neurons of adrenalectomized animals were slightly shifted (3-5 mV) to hyperpolarized potentials as compared to the Na+ currents from neurons of the other experimental groups. Furthermore, the removal from inactivation of the Na+ current in the group of neurons of adrenalectomized animals was relatively slow. Although small, these effects could influence neuronal properties like action potential generation and accommodation. Under the present experimental conditions, no apparent differences were seen between cells with predominant mineralocorticoid receptor activation and cells where both mineralocorticoid and glucocorticoid receptors were occupied. In contrast to Na+ currents, voltage-dependent Ca2+ currents displayed no steroid-dependent shifts in voltage-dependent properties. However, Ca2+ current amplitudes were increased by approximately 160% in CA1 neurons of adrenalectomized animals as compared to Ca2+ currents from neurons of the other experimental groups. We conclude that corticosteroid receptor activation affects various properties of voltage-dependent Na+ and Ca2+ conductances in CA1 neurons, indicating that the steroid receptors are involved in the modulation of neuronal excitability in these cells.

The effects of alaproclate on voltage-dependent K+ currents and N-methyl-D-aspartate (NMDA) and gamma-aminobutyric acidA (GABAA) receptor currents were investigated in cultured rat hippocampal neurons using whole-cell voltage clamp recording techniques. Alaproclate produced a concentration-dependent block of the sustained voltage-dependent K+ current activated by depolarization from -60 to +40 mV (IC50, 6.9 microM). At similar concentrations alaproclate also blocked the sustained voltage-dependent K+ current in fibroblast cells transformed to stably express Kv1.2 K+ channels. Analysis of tail currents and the voltage-dependence of the alaproclate block suggested an open-channel blocking mechanism. Alaproclate also produced a potent block of NMDA receptor currents in hippocampal neurons (IC50, 1.1 microM), but did not affect GABAA receptor currents (concentrations up to 100 microM). The alaproclate block of NMDA receptors occurred predominantly by an open-channel mechanism, although the drug was also able to block closed NMDA channels at a much slower rate. The interaction of alaproclate with NMDA receptors (activated by 10 microM NMDA) appeared to be governed by a first order binding reaction with forward and reverse rate constants of 6.7 x 10(3) M-1 s-1, and 0.025 sec-1, respectively (at -60 mV). At depolarized potentials the alaproclate-induced block of the NMDA receptor current was strongly reduced, a result opposite to that seen with the voltage-activated K+ currents, suggesting that the K+ channel block may occur at a superficial internal site, whereas the NMDA receptor block occurs at a deep external site. (+)-Alaproclate was a more potent blocker of K+ currents than (-)-alaproclate, whereas a reversed stereoselectivity was observed for NMDA receptor current, supporting the view that alaproclate block of the two channel types occurs at structurally distinct binding sites.