6-cyano-7-nitroquinoxaline-2-3-dione and sodium-binding-benzofuran-isophthalate

Neuronal activity causes substantial Na+ transients in fine cellular processes such as dendrites and spines. The physiological consequences of such Na+ transients are still largely unknown. High-resolution Na+ imaging is pivotal to study these questions, and, up to now, two-photon imaging with the fluorescent Na+ indicator sodium-binding benzofuran isophthalate (SBFI) has been the primary method of choice. Recently, a new Na+ indicator dye, CoroNa Green (CoroNa), that has its absorbance maximum at 492 nm, has become available. In the present study, we have compared the properties of SBFI with those of CoroNa by performing Na+ measurements in neurons of hippocampal slices. We show that CoroNa is suitable for measurement of Na+ transients using non-confocal wide-field imaging with a CCD camera. However, substantial transmembrane dye leakage and lower Na+ sensitivity are clearly disadvantages when compared to SBFI. We also tested CoroNa for its suitability for high-resolution imaging of Na+ transients using a confocal laser scanning system. We demonstrate that CoroNa, in contrast to SBFI, can be employed for confocal imaging using a conventional argon laser and report the first Na+ measurements in dendrites using this dye. In conclusion, CoroNa may prove to be a valuable tool for confocal Na+ imaging in fine cellular processes.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Animals, Newborn; Benzofurans; Calibration; Ethers, Cyclic; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Fluorescent Dyes; Hippocampus; In Vitro Techniques; Indicators and Reagents; Membrane Potentials; Mice; Microscopy, Confocal; Neurons; Patch-Clamp Techniques; Potassium; Sodium

The dendrites of many types of neurons contain voltage-dependent Na+ and Ca2+ conductances that generate action potentials (see ref. 1 for review). The function of these spikes is not well understood, but the Ca2+ entry stimulated by spikes probably affects Ca(2+)-dependent processes in dendrites. These include synaptic plasticity, cytotoxicity and exocytosis. Several lines of evidence suggest that dendritic spikes occur within subregions of the dendrites. To study the mechanism that govern the spread of spikes in the dendrites of hippocampal pyramidal cells, we imaged Ca2+ entry with Fura-2 (ref. 9) and Na+ entry with a newly developed Na(+)-sensitive dye. Our results indicate that Ca2+ entry into dendrites is triggered by Na+ spikes that actively invade the dendrites. The restricted spatial distribution of Ca2+ entry seems to depend on the spread of Na+ spikes in the dendrites, rather than on a limited distribution of Ca2+ channels. In addition, we have observed an activity-dependent process that modulates the invasion of spikes into the dendrites and progressively restricts Ca2+ entry to more proximal dendritic regions.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; Animals; Benzofurans; Calcium; Dendrites; Electric Stimulation; Ethers, Cyclic; Fluorescent Dyes; Fura-2; Hippocampus; In Vitro Techniques; Male; Neurons; Pyramidal Tracts; Quinoxalines; Rats; Rats, Inbred Strains; Sodium; Spectrometry, Fluorescence; Tetraethylammonium; Tetraethylammonium Compounds; Tetrodotoxin