title: Boosting bioluminescence neuroimaging: an optimized protocol for brain studies.
abstract: Bioluminescence imaging is widely used for optical cell tracking approaches. However, reliable and quantitative bioluminescence of transplanted cells in the brain is highly challenging. In this study we established a new bioluminescence imaging protocol dedicated for neuroimaging, which increases sensitivity especially for noninvasive tracking of brain cell grafts. Different D-Luciferin concentrations (15, 150, 300 and 750 mg/kg), injection routes (i.v., i.p., s.c.), types of anesthesia (Isoflurane, Ketamine/Xylazine, Pentobarbital) and timing of injection were compared using DCX-Luc transgenic mice for brain specific bioluminescence. Luciferase kinetics was quantitatively evaluated for maximal photon emission, total photon emission and time-to-peak. Photon emission followed a D-Luciferin dose-dependent relation without saturation, but with delay in time-to-peak increasing for increasing concentrations. The comparison of intravenous, subcutaneous and intraperitoneal substrate injection reflects expected pharmacokinetics with fastest and highest photon emission for intravenous administration. Ketamine/Xylazine and Pentobarbital anesthesia showed no significant beneficial effect on maximal photon emission. However, a strong difference in outcome was observed by injecting the substrate pre Isoflurane anesthesia. This protocol optimization for brain specific bioluminescence imaging comprises injection of 300 mg/kg D-Luciferin pre Isoflurane anesthesia as an efficient and stable method with a signal gain of approx. 200% (compared to 150 mg/kg post Isoflurane). Gain in sensitivity by the novel imaging protocol was quantitatively assessed by signal-to-noise calculations of luciferase-expressing neural stem cells grafted into mouse brains (transplantation of 3,000-300,000 cells). The optimized imaging protocol lowered the detection limit from 6,000 to 3,000 cells by a gain in signal-to-noise ratio.
authors: In-Vivo-NMR Laboratory, Max-Planck-Institute for Neurological Research, Cologne, Germany.
authors: Markus
authors: M
authors: Aswendt
authors: Joanna
authors: J
authors: Adamczak
authors: Sebastien
authors: S
authors: Couillard-Despres
authors: Mathias
authors: M
authors: Hoehn
mesh_terms: D000818:Animals; D052160:Benzothiazoles / Q000378:metabolism*; D001921:Brain / Q000166:cytology* / Q000378:metabolism; D002478:Cells, Cultured; D000089662:Doublecortin Protein; D008156:Luciferases / Q000378:metabolism*; D008163:Luminescent Measurements; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008819:Mice, Nude; D008822:Mice, Transgenic; D058953:Neural Stem Cells / Q000637:transplantation*; D059906:Neuroimaging; D017785:Photons; D059629:Signal-To-Noise Ratio
chemical_list: D052160:Benzothiazoles; C532924:D-luciferin; C000714847:Dcx protein, mouse; D000089662:Doublecortin Protein; D008156:Luciferases