Acute administration of the small-molecule p75(NTR) ligand does not prevent hippocampal neuron loss or development of spontaneous seizures after pilocarpine-induced status epilepticus.

Neurotrophins, such as brain-derived neurotrophic factor (BDNF), are initially expressed in a precursor form (e.g., pro-BDNF) and cleaved to form mature BDNF (mBDNF). After pilocarpine-induced status epilepticus (SE), increases in neurotrophins regulate a wide variety of cell-signaling pathways, including prosurvival and cell-death machinery in a receptor-specific manner. Pro-BDNF preferentially binds to the p75 neurotrophin receptor (p75(NTR) ), whereas mBDNF is the major ligand of the tropomyosin-related kinase receptor. To elucidate a potential role for p75(NTR) in acute stages of epileptogenesis, rats were injected prior to and at onset of SE with LM11A-31, a small-molecule ligand that binds to p75(NTR) to promote survival signaling and inhibit neuronal cell death. Modulation of early p75(NTR) signaling and its effects on electrographic SE, SE-induced neurodegeneration, and subsequent spontaneous seizures were examined after LM11A-31 administration. Despite an established neuroprotective effect of LM11A-31 in several animal models of neurodegenerative disorders (e.g., Alzheimer's disease, traumatic brain injury, and spinal cord injury), high-dose LM11A-31 administration prior to and at onset of SE did not reduce the intensity of electrographic SE, prevent SE-induced neuronal cell injury, or inhibit the progression of epileptogenesis. Further studies are required to understand the role of p75(NTR) activation during epileptogenesis and in seizure-induced cell injury in the hippocampus, among other potential cellular pathologies contributing to the onset of spontaneous seizures. Additional studies utilizing more prolonged treatment with LM11A-31 are required to reach a definite conclusion on its potential neuroprotective role in epilepsy.

Egr3 stimulation of GABRA4 promoter activity as a mechanism for seizure-induced up-regulation of GABA(A) receptor alpha4 subunit expression.

GABA is the major inhibitory transmitter at CNS synapses. Changes in subunit composition of the pentameric GABA(A) receptor, including increased levels of alpha4 subunit in dentate granule cells and associated functional alterations such as increased zinc blockade of GABA currents, are hypothesized to be critical components of epileptogenesis. Here, we report that the minimal promoter of the human alpha4 subunit gene (GABRA4p), when used to drive reporter gene expression from adeno-associated viral vectors, controls condition-specific up-regulation in response to status epilepticus, defining a transcriptional mechanism for seizure-induced changes in levels of alpha4 subunit containing GABA(A) receptors. Transfection studies in primary hippocampal neurons show that inducible early growth response factor 3 (Egr3) up-regulates GABRA4p activity as well as the levels of endogenous alpha4 subunits. Given that Egr3 knockout mice display approximately 50% less GABRA4 mRNAs in the hippocampus and that increases in alpha4 and Egr3 mRNAs in response to pilocarpine-induced status epilepticus are accompanied by increased binding of Egr3 to GABRA4 in dentate granule cells, our findings support a role for Egr3 as a major regulator of GABRA4 in developing neurons and in epilepsy.

Long-term effects of diazepam and phenobarbital treatment during development on GABA receptors, transporters and glutamic acid decarboxylase.

Diazepam (DZ) and phenobarbital (PH) are commonly used to treat early-life seizures and act on GABAA receptors (GABAR). The developing GABAergic system is highly plastic, and the long-term effects of postnatal treatment with these drugs on the GABAergic system has not been extensively examined. In the present study, we investigated the effects of prolonged DZ and PH treatment during postnatal development and then discontinuation on expression of a variety of genes involved in GABAergic neurotransmission during adulthood. Rat pups were treated with DZ, PH or vehicle from postnatal day (P) 10-P40 and then the dose was tapered for 2 weeks and terminated at P55. Expression of GABAR subunits, GABAB receptor subunits, GABA transporters (GAT) and GABA synthesizing enzymes (glutamic acid decarboxylase: GAD) mRNAs in hippocampal dentate granule neurons (DGNs) were analyzed using antisense RNA amplification at P90. Protein levels for the alpha1 subunit of GABAR, GAD67, GAT1 and 3 were also assessed using Western blotting. At P90, mRNA expression for GAT-1, 3, 4, GABAR subunits alpha4, alpha6, beta3, delta and theta and GABAB receptor subunit R1 was increased and mRNA expression for GAD65, GAD67 and GABAR subunits alpha1 and alpha3 were decreased in DGNs of rats treated with DZ and PH. The current data suggest that prolonged DZ and PH treatment during postnatal development causes permanent alterations in the expression of hippocampal GABA receptor subunits, GATs and GAD long after therapy has ended.

Effects of status epilepticus on hippocampal GABAA receptors are age-dependent.

Long-term GABA(A) receptor alterations occur in hippocampal dentate granule neurons of rats that develop epilepsy after status epilepticus in adulthood. Hippocampal GABA(A) receptor expression undergoes marked reorganization during the postnatal period, however, and the effects of neonatal status epilepticus on subsequent GABA(A) receptor development are unknown. In the current study, we utilize single cell electrophysiology and antisense mRNA amplification to determine the effect of status-epilepticus induced by lithium-pilocarpine in postnatal day 10 rat pups on GABA(A) receptor subunit expression and function in hippocampal dentate granule neurons. We find that rats subjected to lithium-pilocarpine-induced status epilepticus at postnatal day 10 show long-term GABA(A) receptor changes including a two-fold increase in alpha1 subunit expression (compared with lithium-injected controls) and enhanced type I benzodiazepine augmentation that are opposite of those seen after status epilepticus in adulthood and may serve to enhance dentate gyrus inhibition. Further, unlike adult rats, postnatal day 10 rats subjected to status epilepticus do not become epileptic. These findings suggest age-dependent differences in the effects of status epilepticus on hippocampal GABA(A) receptors that could contribute to the selective resistance of the immature brain to epileptogenesis.

Role of excitatory amino acids in developmental epilepsies.

Altered excitatory amino acid (EAA) neurotransmission, mediated primarily by glutamate, is a major cause of the imbalance of excitation and inhibition which characterizes both early development and epileptogenesis. Glutamate's actions are mediated by three classes of receptors: NMDA, non-NMDA (AMPA and kainate), and metabotropic. Several features of normal EAA development contribute to hyperexcitability in the immature brain, making it more prone to development of seizures. These features include increased density of NMDA receptors, differences in NMDA receptor subunit composition and activation kinetics, which result in reduced voltage-dependent Mg(2+) blockade and longer receptor openings in early development. Also, the unique subunit composition of AMPA receptors present at synapses during early development results in increased Ca(2+) influx. These and other differences in EAA signaling, in combination with developmental alterations in inhibitory neurotransmission, contribute to the increased seizure susceptibility seen in young animals and children. In turn, seizures themselves may alter EAA neurotransmission in an age-dependent manner. Age related changes in excitatory neurotransmission may, therefore, lead to differences in basic mechanisms of epileptogenesis between the immature and mature brain, and may also alter the activity and efficacy of antiepileptic drugs in the pediatric age group.

Effects of vigabatrin on sleep-wakefulness cycle in amygdala-kindled rats.

PURPOSE: Our aim was to study the effect of prolonged administration of vigabatrin (VGB) on sleep-wakefulness cycle in kindled seizure-induced rats. METHODS: Adult male Wistar rats were implanted stereotaxically with electrodes for kindling and polysomnography. The rats were divided into two groups, kindled and VGB-treated kindled rats. VGB was administered intraperitonially every day for 21 days, and polysomnographic recordings were taken after doses 1, 7, 14, and 21. The drug effects were evaluated by comparing the records of kindled and drug-treated kindled rats. RESULTS: The VGB-administered kindled rats showed an increase in total sleep time (TST) due to an increase in total non-rapid eye movement (NREM) and light slow-wave sleep stage I (SI) with a decrease in wakefulness. The number of episodes and REM onset latencies were found to be decreased after drug treatment. CONCLUSIONS: It can therefore be concluded that VGB has a somnolence-inducing effect and that it might mediate its anticonvulsant effect by altering sleep architecture through sleep-regulating areas.

Sleep-wakefulness alterations in amygdala-kindled rats.

PURPOSE: Our aim was to study the relation between epilepsy and sleep-wakefulness cycles in the amygdala-kindling model of temporal lobe epilepsy. METHODS: Adult male Wistar rats were electrically kindled through bipolar electrodes implanted in the anterior amygdala. Polysomnographic recordings were taken before and after kindled seizures for 6 h. For the studies on the effects of a single, full-blown seizure, recordings were taken immediately after the seizure and daily thereafter until the recordings returned to baseline values. For studies on the effects of five full-blown seizures, recordings were taken immediately after the fifth seizure and then on day 1, 2, 3, 5, 7, 14, 21, and 28. RESULTS: Polysomnographic recordings taken immediately after the first full-blown seizure revealed an initial increase in the duration of deep slow-wave sleep (SII), a decrease in the light slow-wave sleep (SI) stage of non-rapid eye movement (NREM) sleep, and a decrease in the quiet wakefulness (W2) stage of wakefulness. All these parameters returned to baseline values after 24 h. The duration of rapid eye movement (REM) sleep increased and returned to the baseline value after 48 h. Five consecutive full-blown seizures caused an increase in the duration of SII from the day the seizures occured until day 28, whereas the duration of SI decreased for 72 h. The duration of REM sleep, decreased only on the day of the seizures and day 1, while decreases in the number of REM episodes were observed on the day of the seizure, day 2 and day 14. CONCLUSIONS: Our study indicates that even a single, full-blown seizure can cause alterations in the architecture of sleep-wakefulness cycles for a short duration, and that multiple seizures produce long-term effects.