dizocilpine-maleate and biocytin

Epilepsy, a common neurological disorder, is characterized by the occurrence of spontaneous recurrent epileptiform discharges (SREDs). Acquired epilepsy is associated with long-term neuronal plasticity changes in the hippocampus resulting in the expression of spontaneous recurrent seizures. The purpose of this study is to evaluate and characterize endogenous epileptiform activity in hippocampal-entorhinal cortical (HEC) slices from epileptic animals. This study employed HEC slices isolated from a large series of control and epileptic animals to evaluate and compare the presence, degree and localization of endogenous SREDs using extracellular and whole cell current clamp recordings. Animals were made epileptic using the pilocarpine model of epilepsy. Extracellular field potentials were recorded simultaneously from areas CA1, CA3, dentate gyrus, and entorhinal cortex and whole cell current clamp recordings were obtained from CA3 neurons. All regions from epileptic HEC slices (n=53) expressed SREDs, with an average frequency of 1.3Hz. In contrast, control slices (n=24) did not manifest any SREDs. Epileptic HEC slices demonstrated slow and fast firing patterns of SREDs. Whole cell current clamp recordings from epileptic HEC slices showed that CA3 neurons exhibited paroxysmal depolarizing shifts associated with these SREDs. To our knowledge this is the first significant demonstration of endogenous SREDs in a large series of HEC slices from epileptic animals in comparison to controls. Epileptiform discharges were found to propagate around hippocampal circuits. HEC slices from epileptic animals that manifest SREDs provide a novel model to study in vitro seizure activity in tissue prepared from epileptic animals.

Topics: Action Potentials; Animals; Cerebral Cortex; Disease Models, Animal; Dizocilpine Maleate; Electric Stimulation; Epilepsy; Excitatory Amino Acid Antagonists; Hippocampus; In Vitro Techniques; Lysine; Neural Pathways; Neurons; Patch-Clamp Techniques; Pilocarpine

Competitive synaptic remodeling is an important feature of developmental plasticity, but the molecular mechanisms remain largely unknown. Calcium/calmodulin-dependent protein kinase II (CaMKII) can induce postsynaptic changes in synaptic strength. We show that postsynaptic CaMKII also generates structural synaptic rearrangements between cultured cortical neurons. Postsynaptic expression of activated CaMKII (T286D) increased the strength of transmission between pairs of pyramidal neuron by a factor of 4, through a modest increase in quantal amplitude and a larger increase in the number of synaptic contacts. Concurrently, T286D reduced overall excitatory synaptic density and increased the proportion of unconnected pairs. This suggests that connectivity from some synaptic partners was increased while other partners were eliminated. The enhancement of connectivity required activity and NMDA receptor activation, while the elimination did not. These data suggest that postsynaptic activation of CaMKII induces a structural remodeling of presynaptic inputs that favors the retention of active presynaptic partners.

Topics: Alanine; Anesthetics, Local; Animals; Animals, Newborn; Aspartic Acid; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Calcium-Calmodulin-Dependent Protein Kinases; Cell Count; Cell Surface Extensions; Cerebral Cortex; Dizocilpine Maleate; Drug Interactions; Excitatory Amino Acid Antagonists; Excitatory Postsynaptic Potentials; Fluorescent Dyes; Gene Expression Regulation, Enzymologic; Green Fluorescent Proteins; Immunohistochemistry; Luminescent Proteins; Lysine; Mutation; Nerve Tissue Proteins; Neuronal Plasticity; Neurons; Organ Culture Techniques; Pyridinium Compounds; Quaternary Ammonium Compounds; Rats; Rats, Long-Evans; Synapses; Synapsins; Synaptic Transmission; Tetrodotoxin; Threonine; Transfection; Valine

NMDA receptor antagonists produce region-specific neurodegeneration by an undetermined mechanism, but one proposed mechanism involves disinhibition. In certain areas of the brain, NMDA receptors mediate excitatory drive onto inhibitory interneurons. Thus, NMDA receptor/channel antagonists may reduce inhibition (i.e., produce "disinhibition"). If a sufficient level of disinhibition is produced, enhanced vulnerability to excitotoxicity may result. Furthermore, if there are region-specific differences in NMDA antagonist-induced disinhibition, this could underlie region-specific NMDA antagonist-induced neurotoxicity. In the present study, we tested this hypothesis by exposing rat brain slices to the NMDA receptor antagonist dizocilpine maleate (MK-801) and measuring MK-801-induced disinhibition in areas of higher and lower vulnerability to neurodegeneration [posterior cingulate/retrosplenial cortices (PCC/RSC) and parietal cortex, respectively]. Using whole-cell patch-clamp techniques, bicuculline-sensitive GABA(A) receptor-mediated IPSCs were measured in biocytin-labeled pyramidal neurons in the PCC/RSC and parietal cortex. In the PCC/RSC, bath-applied MK-801 (10-40 microm) produced disinhibition, shown as a concentration-dependent decrease in spontaneous IPSC frequency and amplitude; MK-801 (40 microm) also reduced evoked IPSC amplitudes. In parietal cortex, MK-801 produced significantly less disinhibition. To determine whether disinhibition is caused by presynaptic or postsynaptic mechanisms, we tested the effects of MK-801 (40 microm) against miniature IPSC (mIPSC) frequency and amplitude in tetrodotoxin (TTX; 0.5 microm)-treated slices and found that MK-801 did not alter mIPSC frequency or amplitude. Taken together, these results suggest that NMDA receptors regulate activity of inhibitory interneurons and, consequently, GABA release in certain cortical areas. This region-specific reduction in inhibitory input to pyramidal cells could underlie the region-specific neurotoxicity of NMDA antagonists.

Topics: Animals; Axons; Dendrites; Dizocilpine Maleate; Electric Stimulation; Excitatory Amino Acid Antagonists; Excitatory Postsynaptic Potentials; GABA Antagonists; Gyrus Cinguli; In Vitro Techniques; Interneurons; Lysine; Male; Membrane Potentials; Neural Inhibition; Neurotoxicity Syndromes; Parietal Lobe; Patch-Clamp Techniques; Pyramidal Cells; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Tetrodotoxin

Activity-dependent synaptic plasticity triggered by N-methyl-d-aspartate (NMDA) receptor activation is a fundamental property of many glutamatergic synapses and may be critical for the shaping and refinement of the structural and functional properties of neuronal circuits during early postnatal development. Using a combined morphological and electrophysiological approach, we showed that chronic blockade of NMDA receptors in hippocampal slice cultures during the first two weeks of postnatal development leads to a substantial increase in synapse number and results in a more complex dendritic arborization of CA1 pyramidal cells. Thus, the development of excitatory circuitry in the hippocampus is determined by two opposing processes: NMDA receptor-independent synapse formation and NMDA receptor-dependent attenuation of synaptogenesis.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; Animals; Animals, Newborn; Cell Surface Extensions; Cells, Cultured; Dendrites; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Excitatory Postsynaptic Potentials; Hippocampus; Histocytochemistry; In Vitro Techniques; Ion Channels; Lysine; Microscopy, Confocal; Patch-Clamp Techniques; Piperazines; Pyramidal Cells; Rats; Rats, Wistar; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Synapses