TAILIEUCHUNG - Báo cáo khoa học: Receptor- and calcium-dependent induced inositol 1,4,5-trisphosphate increases in PC12h cells as shown by fluorescence resonance energy transfer imaging

The production and further metabolism of inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] require several calcium-dependent enzymes, but little is known about subsequent calcium-dependent changes in cellular Ins(1,4,5)P3. | ỊFEBS Journal Receptor- and calcium-dependent induced inositol 1 4 5-trisphosphate increases in PC12h cells as shown by fluorescence resonance energy transfer imaging Mitsuhiro Morita1 2 Fumito Yoshiki1 Akira Nakane1 Yoshiumi Okubo1 and Yoshihisa Kudo1 1 Laboratory of Cellular Neurobiology Schoolof Life Science Tokyo University of Pharmacy and Life Science Japan 2 Department of Neurosurgery University of New Mexico Albuquerque NM USA Keywords calcium fluorescent resonance energy transfer inositol 1 4 5-trisphosphate muscarinic acetylcholine receptor phospholipase C Correspondence M. Morita Laboratory of Cellular Neurobiology School of Life Science Tokyo University of Pharmacy and Life Science 1432-1 Horinouchi Hachioji 192-0392 Tokyo Japan Fax 81 426 76 8841 Tel 81 426 76 8963 E-mail moritam@ Received 15 May 2007 revised 4 August 2007 accepted 8 August 2007 doi The production and further metabolism of inositol 1 4 5-trisphosphate Ins 1 4 5 P3 require several calcium-dependent enzymes but little is known about subsequent calcium-dependent changes in cellular Ins 1 4 5 P3. To study the calcium dependence of muscarinic acetylcholine receptor-induced Ins 1 4 5 P3 increases in PC12h cells we utilized an Ins 1 4 5 P3 imaging system based on fluorescence resonance energy transfer and using green fluorescent protein variants fused with the pleckstrin homology domain of phospholipase C-S1. The intracellular calcium concentration monitored by calcium imaging was adjusted by thapsigargin pretreatment or alterations in extracellular calcium concentration enabling rapid receptor-independent changes in calcium concentration via store-operated calcium influx. We found that Ins 1 4 5 P3 production was increased by a combination of receptor- and calcium-dependent components rather than by calcium alone. The level of Ins 1 4 5 P3 induced by the receptor was found to be half that induced by the combined receptor and calcium components.

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