Spatiotemporal laser inactivation of inositol 1,4,5-trisphosphate receptors using synthetic small-molecule probes.
A malachite green-conjugated inositol 1,4,5-trisphosphate (MGIP(3)) induces specific inactivation of IP(3) receptor (IP(3)R) in tissue samples upon laser irradiation. To verify potential usefulness of the method for studies of cellular Ca(2+) signaling, we conducted laser inactivation at the single-cell level and show that IP(3)R was inactivated with extremely high spatiotemporal resolution. In the presence of MGIP(3), the Ca(2+) release function of IP(3)R in single B lymphoma cells decayed exponentially with increasing duration of laser irradiation with a time constant of 3.4 s. Moreover, by confining laser irradiation to a spatially distinct region of differentiated PC12 cells, subcellular inactivation of IP(3)R was attained, as revealed by a loss of local Ca(2+) signal. Such real-time inactivation of IP(3)R only within a subcellular region may provide a powerful method for investigating spatiotemporal dynamics of Ca(2+) signaling.
Inoue
Takanari
T
Graduate School of Pharmaceutical Sciences, Graduate School of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, 113-0033, Tokyo, Japan.
Kikuchi
Kazuya
K
Hirose
Kenzo
K
Iino
Masamitsu
M
Nagano
Tetsuo
T
D000818:Animals; D015220:Calcium Channels / Q000187:drug effects / Q000378:metabolism* / Q000528:radiation effects; D020013:Calcium Signaling / Q000187:drug effects / Q000528:radiation effects; D002645:Chickens; D015544:Inositol 1,4,5-Trisphosphate / Q000737:chemistry / Q000494:pharmacology; D053496:Inositol 1,4,5-Trisphosphate Receptors; D015640:Ion Channel Gating; D007834:Lasers; D016393:Lymphoma, B-Cell; D008722:Methods; D015335:Molecular Probes; D016716:PC12 Cells; D051381:Rats; D018160:Receptors, Cytoplasmic and Nuclear / Q000187:drug effects / Q000378:metabolism* / Q000528:radiation effects; D012394:Rosaniline Dyes / Q000737:chemistry; D013997:Time Factors
D015220:Calcium Channels; D053496:Inositol 1,4,5-Trisphosphate Receptors; D015335:Molecular Probes; D018160:Receptors, Cytoplasmic and Nuclear; D012394:Rosaniline Dyes; C005095:malachite green; D015544:Inositol 1,4,5-Trisphosphate