1992 Fiscal Year Final Research Report Summary
Analysis of specific neuronal circuit formation with in vitro preparations of mammalian prenatal nervous system.
| Project/Area Number |
03670053
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
Neurophysiology and muscle physiology
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| Research Institution | Tokyo Medical and Dental University |
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Principal Investigator |
SUZUE Toshihiko Tokyo Medical and Dental University, Department of Physiology, Instructor., 医学部, 助手 (40143565)
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| Project Period (FY) |
1991 – 1992
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| Keywords | Neuronal Circuit / Rat / Development / Mammals / Culture / Monoclonal Antibody / Fluorescent Dye |
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| Research Abstract |
The transplacental perfusion method, which was recently developed for in vitro maintenance of late-gestation rat fetuses, was further improved and was used for anterograde and retrograde labeling of fetal neurons with fluorescent dyes.
In order to overcome the difficulty of studying mammalian fetal development, a transplacental-perfusion method was devised for maintaining physiological activities of late gestation rat fetuses in vitro. Attempts were made to improve the condition of the preparation and it became possible to keep the viability of the in vitro fetuses for up to 36 hours. As an application of the method for the study of neuronal development, the method was used for fluorescent dye labeling of fetal neurons. In vitro rat fetus preparations were perfused, and small crystals of DiI were inserted into the thoracic spinal cord for anterograde labeling of peripheral spinal axons. After a 10-15-hour incubation, the thorax was isolated and observed with fluorescence microscopy. A bright staining of axons in intercostal nerves and/or the sympathetic chain were observed. For retrograde labeling, small crystals of DiI were inserted into peripheral tissues so that crystals would contact directly with peripheral nerve fibers. After in vitro perfusion, fetal nervous tissues were fixed and sections were made for microscopic observations. It was found that cell bodies of motoneurons and neurons in the dorsal root ganglia were brightly stained. The central and peripheral branches of the sensory nerves were also brightly stained. The results show that neurons can be stained retrogradely as well as anterogradely in the in vitro fetuses and suggest that the transplacental perfusion method will be useful in various ways for the observation of mammalian fetal neurons.
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