NATIONAL INSITUTE FOR BASIC BIOLOGY

National Institute for Basic Biology

DIVISION OF CELL PROLIFERATION

(Adjunct)

Professor:
Motoya Katsuki (April 1, 1998 ~)
Research Associate:
Kei Ito (June 1, 1998 ~)
JST Technical Staff:
Kimiko Tanaka (October 1, 1998 ~)
Graduate Student:
Asami Kido (Ochanomizu University; July 1, 1998 ~)


The aim of this new adjunct division, started in June 1998, is to understand the basic rules by which elaborate neural circuits develop and function. To address these questions, an effective approach is to investigate the whole of an easily-accessible nervous system that shares certain of the architectural and functional features of the more complex vertebrate brains. With less than 105 neurones, and subject to powerful molecular and genetic techniques, the brain of the fruit fly Drosophila melanogaster is a good model system for such a study, particularly because it contains neural arrangements, such as those in the olfactory and visual pathways, that are extraordinarily similar to equivalent parts of vertebrate brains.


I. Developmental neuroanatomy

A comprehensive and detailed anatomical knowledge of the brain is a prerequisite for 1) analysing the phenotypes of nervous system-related mutants, 2) identifying the cells that express the cloned genes, 3) understanding the way information is processed in the brain, and 4) devising computer models that simulate brain functions. In spite of the hundred years of efforts using Golgi and other anatomical techniques, however, the circuit structure of higher order associative regions of the brain is still essentially unresolved.

The GAL4 enhancer-trap system, which is widely used for mutagenesis and gene cloning of Drosophila, is also a powerful tool for obtaining a vast array of molecular markers that label specific subsets of brain cells. Forming a consortium with eight other Drosophila laboratories in Japan, we have established more than 4000 GAL4 strains. Different aspects of the morphology of the GAL4-expressing cells can be visualised by crossing these strains with the strains that carry various UAS-linked reporter genes, such as nuclear-specific UAS-NLS-lacZ, axon-targeting UAS-tau, cytoplasmic UAS-lacZ and UAS-GFP, and presynaptic site-specific UAS-neuronal synaptobrevin-GFP.

The screening of the GAL4 strains is being performed by observing the expression pattern in the adult brain. Useful strains will be selected to analyse the information pathway of different sensory modalities in the higher-order brain regions, as well as to study how the labelled neurones form their elaborate fibre connections during neurogenesis.


II. Functional neuroanatomy

With the GAL4-UAS system, it is possible to alter the normal cell functions by ectopically expressing developmental switch genes specifically in the GAL4-expressing cells. If this results in a change of certain behaviour of the animal, it is likely that these altered cells may play major roles in controlling that behaviour.

We use UAS-linked transformer (tra) gene, which acts early in the sex determination cascade, to alter the sex of the GAL4-expressing cells. The courtship behaviour of Drosophila is very different between male and female. Male flies actively try to attract females by presenting stereotyped steps of courtship, whereas females stay passive. Does this male-specific behaviour disappear, if particular brain cells are feminised?

Previous studies by other investigators showed that feminisation of the mushroom body neurones may make the male flies bisexual. Although this is an interesting phenomenon, the male-specific behaviour did persist in these experiments. To find the GAL4 strains that can really suppress male behaviour, we performed a much larger behavioural screening. For this purpose, we abandoned any assumption to pre-select strains that drive tra gene in particular brain structures. Instead, we crossed all the available homozygous viable GAL4 strains with UAS-tra. Among the 446 strains tested so far, we found only two strains that show near-total suppression of male-specific courtship behaviour, regardless of the sex of the targets (Fig. 1).

Fig. 1 Reduced male-specific activity caused by the ectopic expression of the transformer gene
(A) Wild-type (CS) male flies actively court females when they are put together in small chambers. Within ten minutes, most (in this case four out of six) males reach the final step of the courtship: copulation. Other males are chasing and courting their partners. (B) Males of the GAL4 enhancer-trap strain Np218 crossed to UAS-tra, on the other hand, show no interests in females. (C) Quantitative comparison of the courtship behaviour. The duration of the time when a male fly engaged in any kind of the courtship-related behaviour (courtship index, CI), and the time when a male fly performed the courtship-specific wing vibration behaviour (sex appeal parameter, SAP), were recorded during the first ten minutes after the subject (male) and the target (either male or female) flies were put in the chamber. Wild type (CS) males spend long time courting females, but court males only occasionally. The males of the Np218 and Mz 490 strains crossed to UAS-tra spend only a very short period for courting both male and female.

The GAL4 expression patterns of these two strains, however, were not very specific. Rather, GAL4 is expressed in most of the brain cells. The results can be explained by assuming the following: 1: The courtship behaviour is controlled by various biochemical types of neurones, which may either be packed within a small brain area, or scattered in many regions; and 2: The male behaviour would manifest unless all the relevant neurones are female. Since these neurones may not share common enhancer activity, only GAL4 strains with near-ubiquitous expression pattern would be able to feminise all of these cells simultaneously.


III. Contribution to the science community

As the number of identified neurones grows, it becomes important to develop a system with which science community can easily access the record of complicated three-dimensional circuit structures. As a joint venture with German and US research groups, we maintain Flybrain, a web-based image database of the Drosophila nervous system (http://flybrain.nibb.ac.jp). Over 2000 images has already been stored and served worldwide.

Another database maintained here is Jfly, which is intended to help the exchange of information among Japanese-speaking Drosophila researchers (http://jfly.nibb.ac.jp). Archives of research-related discussions, images and experimental protocols, as well as meetings and job announcements, are provided.


Publication List:
Ito, K., Suzuki, K., Estes, P., Ramaswami, M., Yamamoto, D. and Strausfeld, N. J. (1998) The organization of extrinsic neurons and their implications regarding the functional roles of the mushroom bodies in Drosophila melanogaster Meigen. Learning and Memory 5, 52-77.
Strausfeld, N. J., Hansen, L., Li, Y., Gomez, R. S. and Ito, K. (1998) Evolution, discovery, and interpretations of Arthropod mushroom bodies. Learning and Memory 5, 11-37.

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Last Modified: 12:00, May 28, 1999