DIRECT NITRIC OXIDE IMAGING IN CULTURED HIPPOCAMPAL NEURONS WITH DIAMINOANTHRAQUINONE AND CONFOCAL MICROSCOPY

✍ Scribed by Xiaochun Chen; Chao Sheng; Xiaoxiang Zheng

• Book ID: 102569377
• Publisher: Elsevier Science
• Year: 2001
• Tongue: English
• Weight: 265 KB
• Volume: 25
• Category: Article
• ISSN: 1065-6995
• DOI: 10.1006/cbir.2001.0734

✦ Synopsis

A method for the direct measurement of intracellular nitric oxide (NO) production stimulated by penicillin G (PG) in cultured hippocampal neurons with diaminoanthraquinone (DAA) using laser scanning confocal microscopy (LSCM) was developed. Intracellular DAA fluorescence could specifically represent NO production based on two facts: (1) 3-morpholinosydnonimine, a NO donor, could dose-dependently increase DAA fluorescence; and (2) haemoglobin, a NO scavenger, could inhibit the increase of DAA fluorescence. The PG dose-dependently increased the intercellular level of glutamate (Glu, 5 min after stimulation) and the intracellular NO production (30 min throughout stimulation). The increase of NO production could be reversed by N(w)-nitro- l -arginine (a NO synthase inhibitor), and also by d (-)2-amino-5-phosphonovaleric acid, a subtype of Glu receptor antagonist. These results revealed that DAA could be used to indicate real-time and kinetic intracellular NO production of hippocampal neurons with higher sensitivity, specificity and accuracy.