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LOCAL ACTIONS OF NGF ON NEURITE OUTGROWTH, EXAMINED BY DIFFERENTIATING CULTURE CONDITIONS BETWEEN SOMA AND AXONS (OR DENDRITES) BY USING A NEW TWO-CHAMBER CULTURE DISH. J N F K D AN K $< ic’ne Universitv of Tokvo. Bunkvo-ku. Tokvo 113. Jaoa

Local and trans-nuclei actions of NGF for neurite promotion were differentiated by separating culture conditions of somas and fibers of dorsal root ganglion (DRG) cells, by utilizing newly developed two-chamber culture dish. DRG were dissected from newborn rats (5-10 days) in attaching their peripheral and central nerves. The preparation was placed in the two-chamber dish: the somas were in one chamber (Iml) and their peripheral nerves in the other chamber (lml), being separated by Vaseline-gap of 300-500 pm: Culture conditions of peripheral nerves were completely differentiated from those of the somas (see Keino and Fukuda, 1991). An addition of NGF to the nerve-chamber caused failure of neurite outgrowth from the soma, and vice-versa. An addition of anti-NGF-antisera to soma-chamber caused no changes in neurite growth from peripheral nerves. K-252a which selectively inhibits NGF-dependent neurite outgrowth was ineffective across the Vaseline gap. Protein kinase inhibitors which suppress neurite growth were also ineffective across the gap. A breakdown of microtubules around the soma did not stop neurite outgrowth from the peripheral nerves. These lead to a conclusion that local actions of NGF (but not trans.nuclei actions) were crucial for neurite outgrowth from the primary sensory nerve cells.

EXPRESSION OF TRKB GENE IN THE RAT CENTRAL AND PERIPHERAL NERVOUS SYSTEM - IN SITU HYBRIDIZATION STUDY - YOSHIYUKI MASANA, AK10 WANAKA, AND MASAYA TOHYAMA, Department of Anatomv and Neuroscience, Osaka University Medical School, 2-2 Yamadaoka, Suita, Osaka, 565, Japan.

trkB is a member of the gene family that encodes putative receptors with tyro- sine kinase activity for NGF family. BDNF and NT-3 has been shown to bind to trkB gene product specifically, suggesting that trkB gene product may be the component of the high-affinity receptor for BDNF and NT-3. In the present study using oligonucleotide probe, we examined trkB gene expression in the nervous system of the newborn and adult rats by in situ hybridization histochemistry. The signals were detected mainly in the brain (cerebral cortex, ependymal lining of the third ventricle) and the dorsal root ganglion (DRG) both in the newborn and the adult. The intensity of the hybridization signals was stronger in the newborn than in the adult. These results suggest that trkB gene may be closely related to the differentiation of the neocortex and the devolopment and maintainance of the sensory nervous system.

MIGRATION OF NEUROPHiLIC CELLS IN VITRO. I. NEL’RONAL NATURE OF GUIDES FOR CEILULAR HELOCAT I ON. KATSUHIKO ONG AND AKIRA TOKIJNAGA. Third Department of Anatomy, Okayama University Medical School, Shikata-cho,Okayama 700, Japan.

During the formation of the central nervous system. neuronal and glial cells _. migrate from their sites of origin to their destinations. NeuronaI processes are known to play a guidance role in cell migration in the developing brainstem of vertebrates, and cells moving along neuronal guides are called neuroohilic cells. In the present study, explant culture was used to demonstrate neuroohil ic ccl 1 migration in vitro.

Small sections of tissue from the brainstem of fetal rats (13th or 14th day of gestation) were placed onto collagen-coated culture dishes and incubated in DMEM containing IO % fetal calf and l!) X norse sera for 3-15 days. Long processes extended from the explants, and numerous oval or fusiform cells with lo-20 pm long axis were observed to translocste along these processes at the rate of 2-30 pm/h. The processes he;_c positive for anti-neurofiiament antibodies. Antiserum to gila! fibril!ary acidic protein stained short and flattened processes to which rc!ocating cells were not ap=osed. bnder transmission electron microscope, the long processes appeared (1s fasciclts composed of many thin (1 pm or lesser in diameter) neurites wni.:n contained microtubules and sometimes intermediate filaments. ‘i’ t; e p r e s c n t study demonstrated that the long processes cxt,,nding from the explant were neuronal in nature, and acted as guidelines for cell migration.