6-cyano-7-nitroquinoxaline-2-3-dione and zinc-chloride

Protracted elevation in intracellular calcium caused by the activation of the N-methyl-d-aspartate receptor is the main cause of glutamate excitotoxic injury in stroke. However, upon excitotoxic injury, despite the presence of calcium entry antagonists, calcium unexpectedly continues to enter the neuron, causing extended neuronal depolarization and culminating in neuronal death. This phenomenon is known as the calcium paradox of neuronal death in stroke, and it represents a major problem in developing effective therapies for the treatment of stroke. To investigate this calcium paradox and to determine the source of this unexpected calcium entry after neuronal injury, we evaluated whether glutamate excitotoxicity activates an injury-induced calcium-permeable channel responsible for conducting a calcium current that underlies neuronal death. We used a combination of whole-cell and single-channel patch-clamp recordings, fluorescent calcium imaging, and neuronal cell death assays in a well characterized primary hippocampal neuronal culture model of glutamate excitotoxicity/stroke. Here, we report activation of a novel calcium-permeable channel upon excitotoxic glutamate injury that carries calcium current even in the presence of calcium entry inhibitors. Blocking this injury-induced calcium-permeable channel for a significant time period after the initial injury is still effective in preventing calcium entry, extended neuronal depolarization, and delayed neuronal death, thereby accounting for the calcium paradox. This injury-induced calcium-permeable channel represents a major source for the initial calcium entry following stroke, and it offers a new target for extending the therapeutic window for preventing neuronal death after the initial excitotoxic (stroke) injury.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; Animals; Animals, Newborn; Apoptosis; Calcium; Calcium Channel Blockers; Calcium Channels; Cells, Cultured; Chlorides; Cobalt; Dizocilpine Maleate; Dose-Response Relationship, Drug; Electric Impedance; Ethosuximide; Gadolinium; Glutamic Acid; Membrane Potentials; Neurons; Nifedipine; omega-Conotoxins; Patch-Clamp Techniques; Rats; Rats, Sprague-Dawley; Sodium; Stroke; Zinc Compounds

In the kindling model of temporal lobe epilepsy, several physiological indicators of inhibition by gamma-aminobutyric acid (GABA) in the hippocampal dentate gyrus are consistent with an augmented, rather than a diminished, inhibition. In brain slices obtained from epileptic (kindled) rats, the excitatory drive onto inhibitory interneurons was increased and was paralleled by a reduction in the presynaptic autoinhibition of GABA release. This augmented inhibition was sensitive to zinc most likely after a molecular reorganization of GABAA receptor subunits. Consequently, during seizures, inhibition by GABA may be diminished by the zinc released from aberrantly sprouted mossy fiber terminals of granule cells, which are found in many experimental models of epilepsy and in human temporal lobe epilepsy.

Topics: 2-Amino-5-phosphonovalerate; 6-Cyano-7-nitroquinoxaline-2,3-dione; Animals; Chlorides; Dentate Gyrus; Epilepsy, Temporal Lobe; Excitatory Amino Acid Antagonists; GABA-A Receptor Antagonists; gamma-Aminobutyric Acid; Humans; In Vitro Techniques; Interneurons; Kindling, Neurologic; Male; Neural Inhibition; Pyridines; Rats; Rats, Wistar; Receptors, GABA-A; Receptors, GABA-B; Synaptic Transmission; Zinc; Zinc Compounds; Zolpidem

Zn2+ is believed to be an endogenous modulator of glutamatergic excitation. It has been shown to attenuate NMDA receptor-mediated excitation and to increase AMPA-induced excitatory transmission. The dual activity of Zn2+ on ionotropic excitatory neurotransmission suggests that Zn2+ plays a role in the modulation of excitatory neurodegenerative events. Stimulation of rat primary cortical cultures with the combination of 50 microM AMPA and 300 microM Zn2+ for 30 min induced approximately 50% cell death compared with only approximately 20% cell death induced by AMPA alone. The degree of neurotoxicity 48 h after the incubation was reproducible and was attenuated by CNQX, EDTA, EGTA, diltiazem and DHP-type Ca2+ channel blockers but not by MK-801. These findings suggest that an initial depolarization induced by AMPA and a subsequent influx of Ca2+ and Zn2+ ions through voltage-operated L-type Ca2+ channels are crucial events which finally lead to neuronal death. Racemic nimodipine and its (+)- and (-)-enantiomers had remarkable in vitro neuroprotective efficacies, the IC50 values being 4 nM for the racemate, 11 nM for the (+)- and 1 nM for the (-)-enantiomer. This suggests a possible therapeutic role for Ca2+ channel blockers in neurodegenerative diseases which are characterized by a disturbance of cellular Ca2+ homeostasis.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Calcium Channels; Cell Death; Cells, Cultured; Cerebral Cortex; Chlorides; Diltiazem; Edetic Acid; Egtazic Acid; Female; Fetus; Kinetics; Membrane Potentials; Neurons; Neurotoxins; Pregnancy; Rats; Rats, Wistar; Synaptic Transmission; Time Factors; Zinc Compounds