Planned collaborative project

Planned collaborative project themes are selected by NIPS, which are based on requests from researchers. Until 2007, there were two themes, “Physiological and neuroscientific studies into genetically modified model animals” and “Biomolecular sensors and physiological function”. Additional themes were added in 2008, with “Functional and morphological analyses of cells and tissues by multi-photon microscopy” and “Medical and biological applications of phase-contrast cryoelectron microscopy” (name changed to “Medical and biological applications of cutting edge electron microscopy” in 2011), and in 2009 with, “Behavioral analysis of mouse and rat”. Also, “Analysis of metabolic physiology for mouse and rat” began in 2011, while “Transfection study with primates”, “Analysis of fluctuations in function research in life science”, and “Multidisciplinary study of neural information” began in 2012. In addition, “Transfection study with viral vector neurological system” was started in 2013. All these themes cover the most talked about scientific topics today, and are areas where NIPS is considered to be a frontrunner in Japan. We expect to receive many new proposals. In regards to the proposal agenda, long discussions had been carried out at both faculty meetings and work meetings in 2012. The agreed requirements are as follows.

1) Proposals should clearly state the aim and experimental design of the research project, and should be completed within five years. However, depending on the state of the research, an extension period may be granted after the initial five years.
2) Proposals should specifically state the research area of interest. Broad themes will not be accepted.
3) There will be a limit to the number of proposals accepted. Each general collaborative research area category and research facility will accept five projects each at most, in principle. The details of the planned collaborative research are as follows.

The details of the planned collaborative research are as follows.

“Physiological and neuroscientific analysis of genetically modified model animals”

Genetically modified model animals help researchers studying physiology and brain science, where progress can only be made through studying individuals. The engineering required to create such model animals has taken huge leaps forward in recent years. Compared to other institutes, the Section of Mammalian Transgenesis at the Center for Genetic Analysis of Behavior in NIPS has made a large contribution to physiology and brain science, and reproductive biotechnology, by providing researchers all across the country with technology to produce genetically modified model animals. To support our cooperative studies, we provide the means to develop adoptive models such as transgenic or knock-out mice and rats. Genetically modified rats have been particularly difficult to produce in the past, but the recently accepted use of embryonic stem(ES)cells and induced pluripotent stem(iPS)cells have made it possible to create knock-out rats. Researchers at our lab have already been successful in establishing ES and iPS germ cell lines from rats, from which they and then created three strains of knock-out rats and one strain of knock-in rats. In a calendar year of 2013, we have created a total of 23 transgenic or knock-out lines in mice and rats under nine collaborative research projects. Successful application of artificial restriction enzymes to create knock-out/knock-in animals will facilitate our future contribution to requested task in NIPS.

“Behavioral analysis of mouse and rat”

Today it has become possible to associate genes to particular behaviors, thanks to genetically modified animals. However, such research requires a large number of different behavioral tests that are also reproducible. Having individual laboratories conduct these tests individually is both complicated and produces a lot of waste. The Section for Behavior Patterns was set up in the Center for Genetic Analysis of Behavior at NIPS to provide analytical information on animal behavior to all the researchers involved in our cooperative research studies. As an expert in mouse behavior, Adjunct professor Tsuyoshi Miyakawa was invited to the section,and in 2009 started the planned cooperative research on“Behavioral analysis of mouse and rat”. Currently, mouse analysis is being carried out, but the group plans to start rat analysis soon.
In 2013, NIPS carried out 11 planned collaborative projects with outside research institutes, and 1 projects within NIPS. A number of behavioral test analyses have been carried out on eight strains of genetically modified mice and mice administered with medication, and exhaustive behavior test batteries on five strains of mice. In this year, the protocol of the contextual and cued fear conditioning test was published as a video article (Shoji et al, JoVE, 2014). The ImageFZ, Program for the Contextual and Cued Fear Conditioning Test, the software used to analyze the behavior in the test has been publicly released. ImageTM, Program for the T-Maze Test (Shoji et al, JoVE 2012) was also released. Those softwares can be downloaded at the following website: Using the software should help researchers studying behavior to analyze pictures more efficiently.

“Analysis of metabolic physiology for mice and rats”

The Section of Metabolic Physiology was set up in 2010, and the planned collaborative research project,“Metabolic physiology analysis of mice and rats”, had started in 2011. Since then, researchers from within and outside NIPS have been looking at the following topics concerning genetically modified animals.
1) Measuring neural activity of individual neurons associated with motor function while awake.
2) Measuring the discharge of neurotransmitter substance in specific areas of the brain during freemovement.
3) Circuit behavior imaging of flavin and haemoglobin intrinsic signals in the brain using voltage sensitive dyes.
4) Measuring food intake and energy consumption during free movement.
5) Measuring body temperature, pulse rate, and blood pressure
Eight collaborative research projects with researchers outside NIPS were conducted in 2013.

”Ultrastructure analysis of biological specimens by cutting edge electron microscopy”

One phase-contrast electron microscope (PC-TEM) and two serial block-face scanning electron microscopes (SBF-SEMs) are mainly used for this joint research program. PC-TEM developed by NIPS shows the best performance when combined with a rapid-freezing sample preparation method. Under this condition, it is possible to study three-dimensional structures of unstained biological specimens, including isolated proteins, viruses, bacteria, cultured cells and tissues, to more or less their true state with 1 nm spatial resolution. On the other hand, SBF-SEMs are used for the studies of ultrastructural analysis of thick biological specimens, like a brain tissue. The specimens embedded in plastic resin are sliced by a diamond-knife and imaged by SEM continuously. Finally, the three-dimensional ultrastructure of the specimens is rebuild at dozens of nanometer resolution. The program support studies by using these state of the art electron microscopes. In 2013, four projects using PC-TEM and eight projects using SBF-SEMs were carried out.

“Functional and morphological analyses of cells and tissues by multi-photon excitation microscopy”

Two-photon excitation fluorescence microscope is a less invasive method for studying the microscopic structure and functions of cells in deep tissues of biological organisms. Currently, our institute has three upright two-photon excitation microscopes, and these allows us to observe the structure in depth of one millimeter with a spatial resolution of micrometer. Since the maintenance of two-photon microscpe is complicated, NIPS is the only institute which can provide the opportunity of collaborative research with the high quality experience. Furthermore, we recently build the two-photon fluorescence lifetime microscope system which enable us to observe the intermolecular interactions and the activity of signaling protein in living cell in deep tissue. We also working on single-molecule imaging using quantum dot in combination of fluorescence microscope. Using these "cutting-edge methods", we have conducted the collaborative researches. Recent successes are particularly in vivo Ca2+ imaging, and long-term imaging of neurons in living mice.
This year, three planned collaborative projects and seven preliminary experiments to find potential collaborative projects had been carried out. We also discussed about collaborative research with over ten groups, and introduced our multi-photon excitation microscopes to over 20 groups.

“Transfection study with primates”

Advances in technology to control molecular functions or change neural activity by inserting certain genes into primate brains using virus vectors can lead to major possibilities. Getting to do such research, however, requires a long list of equipment and facilities to enable researchers to develop do things such as develop vectors, or insert vectors. Planned collaborative research projects were launched in 2012 so that researchers could share their resources, and work together to unravel mysteries about higher brain functions and pathological conditions. In 2012, three projects were carried out and 5 projects were carried out in 2013. The first study looked at whether virus vectors could help find out how compensatory motor system circuits in macaque monkey brains causes a monkey with a damaged motor cortex to recover its function. The second study used virus vectors and immunotoxins to look at how the basal ganglia functioned and its pathological condition. The team was then able to selectively eliminate the hyperdirect pathway in the neural pathway of the basal ganglia. The last study used virus vectors in RNA interference to suppress gene expression in primates, all of which was observed using PET molecular imaging.

“Analysis of fluctuations in function research in life sciences”

“Decisions and fluctuations in functional life science” had been started in 2010, as part of the NIPS project to make the institute a hub for international research. Its goals are as follows. ‘ Maintaining stability and balance’ and‘ having the power to change occasionally’ are important to keep in mind when thinking about people’s decisions and evolution. ‘ Fluctuation’ can make both‘ stability’ and‘ occasional change’ possible. This project needs to look at the world from a‘ fluctuating and decisive’ perspective, and include everything from single molecules, polymolecular systems, to cells and whole biological bodies. It needs to understand that instability in life hierarchy is possible, and that jumps in functional life science decisions are a result of the valuable role that fluctuations play. From this, fluctuations in biologically functioning molecules and its interactions with others give birth to complicated life phenomena. Ultimately, we must strive to understand how to become man’s will. As part of this project, the planned collaborative study,
“Analysis of fluctuations in function research in life sciences”, was launched in 2012. One project was perofrmed in 2012, and 3 in 2013. Three projects are planned in 2014.

“Multidisciplinary study of neural information”

In 2010, this project was launched as one of two NIPS projects aimed at “forming a hub for international research in natural sciences”. Its goal is to study how the brain processes information by linking molecules, cells, circuits, and the brain hierarchy in humans and various model animals. We can do this by finding correlations between the brain’s structure and function, which would explain how the cranial nerve processes information. This would require running imaging experiments to connect the dots between hierarchy levels and animal types. Also, collaborating with researchers around the world would help establish an international hub dedicated to furthering our knowledge on neuroscience. In 2013, seven individual projects proposed by NIPS, two projects by NIBB, and one from IMS took part. The project also invited scientists from overseas, sent their own scientists overseas, and started holding symposiums with international guests.
Collaborative research proposals were also accepted from 2012 onwards.

“Gene transfer into nervous system using viral vectors”

Viral vector provides a very useful technology for gene transfer into nervous system. In Section of Viral Vector Development, we promote the collaboration with many laboratories by providing various serotypes of AAV vectors, conventional lentiviral vectors, and highly efficient retrograde gene transfer vectors. Moreover, we proceed with the collaboration to exploit the more advantageous viral vectors.
In 2013, we provided more than 100 viral vectors for other laboratories, and performed 2 planned collaborative research. At present, very intriguing research results are being obtained.
In future, we plan to promote the collaboration more actively by providing high-quality viral vectors for more and more laboratories.