Akisa Nemoto,1 Reona Kobayashi,1,2 Sho Yoshimatsu,1,2,3 Yuta Sato,2,4 Takahiro Kondo,1,2 Andrew S. Yoo,5 Seiji Shiozawa,1,6 and Hideyuki Okano1,2
1 Department of Physiology, School of Medicine, Keio University, Tokyo (Japan); 2 Laboratory for Marmoset Neural Architecture, RIKEN Center for Brain Science, Saitama (Japan); 3 Laboratory for Proteolytic Neuroscience, RIKEN Center for Brain Science, Saitama (Japan); 4 Graduate School of Science and Technology, Keio University, Kanagawa (Japan); 5 Department of Developmental Biology, Washington University School of Medicine, St. Louis (USA); 6 Institute of Animal Experimentation, School of Medicine, Kurume University, Fukuoka (Japan).
Calcium Imaging, Medical Research, Neuroscience
The common marmoset (Callithrix jacchus) has attracted considerable attention, especially in the biomedical science and neuroscience research fields, because of its potential to recapitulate the complex and multidimensional phenotypes of human diseases, and several neurodegenerative transgenic models have been reported. However, there remain several issues as (i) it takes years to generate late-onset disease models, and (ii) the onset age and severity of phenotypes can vary among individuals due to differences in genetic background. In the present study, we established an efficient and rapid direct neuronal induction method (induced neurons; iNs) from embryonic and adult marmoset fibroblasts to investigate cellular-level phenotypes in the marmoset brain in vitro. We overexpressed reprogramming effectors, i.e., microRNA-9/9*, microRNA-124, and Achaete-Scute family bHLH transcription factor 1, in fibroblasts with a small molecule cocktail that facilitates neuronal induction. The resultant iNs from embryonic and adult marmoset fibroblasts showed neuronal characteristics within two weeks, including neuron-specific gene expression and spontaneous neuronal activity. As directly reprogrammed neurons have been shown to model neurodegenerative disorders, the neuronal reprogramming of marmoset fibroblasts may offer new tools for investigating neurological phenotypes associated with disease progression in non-human primate neurological disease models.
Movies were captured at 20 frames/s using an IX83 inverted microscope (Olympus, Tokyo, Japan) equipped with an electron multiplying CCD camera (Hamamatsu Photonics, Hamamatsu, Shizuoka, Japan) and pE-4000 LED illumination system (CoolLED, Andover, UK).
Product Associated Features
The pE-4000 Universal Illumination System offers 16 selectable wavelengths from 365 - 770 nm, which covers a wide variety of fluorophores such as Fluo-8 for calcium imaging.
Year of Publication
Country of Publication