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Morphological and electrophysiological analysis of the development of cerebellar granule cells in the microexplant culture
s143
GROWTH CONES OF COMMISSURAL AXONS IN THE DEVELOPING RAT CEREBRUM: AN ANTEROGRADE TRACING STUDY WITH BIOCYTIN. WOSHI ISHII AND TOSHIO SHIRAI. Dept. of Anatomv, Yamanata Univ.Sch. of Medicine, Yamaxata OOO-23,Japan.
1235
We examined morphology of the growth cones of commissural axons(CAs) and their distribution in the cerebral medullary layer and the contralateral cerebral cortex to study pathway formation of CAs. We injected Biocytin into the right parietal cortices of rats anesthetized with sodium pentobarbital from postnatal day(PD)O to PD'?. At PDO growth cones of CAs were found in the ipsilateral and contralateral medullary layers through callosal body, and they showed various forms: complicated swelling tips with several filopodia,flattened, and spindle forming ones. At PDl,although many tips of CAs were still found in medullary layer and begun to make their branchings in it, a few tips of CAs appeared in the deep layer of the contralateral cerebral cortex. By PD5 they appeared in the superficial layer of the contralateral cerebral cortex in addition to in the deep layer of it and the medullary layer. Many of them formed oval or spindle tips. At PDT-9 many tips of CAs were found in the superficial layer and a few of them in the medullary layer. From these findings it suggests that the later commissural axons may still extend in the medullary layer about at PDT-O.
1236 Phvsiolonv,
TURNING OF NERVE GROWTH CONES INDUCED BY FOCAL ELECTRIC FIELDS IN Cal+ FREE MEDIUM S. TSUKADA. K. KEINO-MASU. T. IWAMOTO. AND J. FUKUDA Department of National Defense Medical School. Tokorozawa. Saitama. JAPAN.
Transmembrane Ca?-+ influx at nerve growth cones has been understood as essential for turning response of Here we report an opposite result that the turning response takes place in growing tips by electric currents. Dorsal root ganglia neurites growing in the absence of extracellar Ca2+; i.e., without Ca2+ influx in growth cones. dissected from newborn rats were placed in a two chamber dish so that their neurites grew in a Ca-free medium. DC current (50-250 nA) was focally applied to growth cones through a microelectrode placed approximately 100 pm distant from the growing tips. The turning responses that were induced in neurites bathed in normal Ca medium (2.5 mM) were still observed in neurites that were growing in the Ca-free medium; its Ca2+ concentration was not more than 0.01 nM. Since no Ca2+ influx is expected to occur in the growth cones in such a culture condition, we conclude that other mechanisms than Ca2+ influx at the growing ends are responsible for inducing the turning responses of rat DRG neurites.
MORPHOLOGICAL AND ELECTROPHYSIOLOGICAL ANALYSIS OF THE DEVELOPMENT OF CEREBELLAR GRANULE CELLS IN THE MICROEXPLANT CULTURE. YOSHIHIKO WAKAZONO’, SHIGEKI YUASA’, KENSUKE NAKAHIRA’, TAKASHI KURAHASHI’, AKIMICHI KANEKO’*’AND KAZUHIRO IKENAKA’, ‘Natl. Inst. for Physiol. Sci., 38 Nishigonaka, Myodaiji, Okazaki 444. 2Dept. of Physiology, School of Medicine, Keio University, Shinjuku-ku, Tokyo 160 Japan.
1237
Themode ofmigration andthedistribution of the granule cells were examined in the microexplant culture’) of mouse early postnatal cerebellum by immunocytochemical methods and retrovims mediated gene transfer. Granule cells were shown to migrate radially from the cerebellar explant by labeling the proliferating granule cells with bromodeoxyuridine. Migratory granule cells were apposed to the neurites which extended radially from the explants and formed small clusters as revealed by labeling with anti-Zic antibody), a specific marker for cerebellar granule cells. Furthermore, granule cells were identified morphologically by the retrovirus mediated transfer of fi -galactosidase gene. Patch-clamp method was applied to this microexplant culture to examine the generation of action potentials in neurons which were morphologically identified as granule cells. In view of these findings, the interelations between the generation of action potential and the morphological differentiation of the developing cerebellar granule cells will be discussed. 1) Nagata, I. and Nakatsuji, N. (1990) Dev. Brain Res. 52,63-73 2) Aruga, J. et al. (1994) J. Neurochem. 63, in press
GROWTH CONES OF COMMISSURAL AXONS IN THE DEVELOPING RAT CEREBRUM: AN ANTEROGRADE TRACING STUDY WITH BIOCYTIN. WOSHI ISHII AND TOSHIO SHIRAI. Dept. of Anatomv, Yamanata Univ.Sch. of Medicine, Yamaxata OOO-23,Japan.
1235
We examined morphology of the growth cones of commissural axons(CAs) and their distribution in the cerebral medullary layer and the contralateral cerebral cortex to study pathway formation of CAs. We injected Biocytin into the right parietal cortices of rats anesthetized with sodium pentobarbital from postnatal day(PD)O to PD'?. At PDO growth cones of CAs were found in the ipsilateral and contralateral medullary layers through callosal body, and they showed various forms: complicated swelling tips with several filopodia,flattened, and spindle forming ones. At PDl,although many tips of CAs were still found in medullary layer and begun to make their branchings in it, a few tips of CAs appeared in the deep layer of the contralateral cerebral cortex. By PD5 they appeared in the superficial layer of the contralateral cerebral cortex in addition to in the deep layer of it and the medullary layer. Many of them formed oval or spindle tips. At PDT-9 many tips of CAs were found in the superficial layer and a few of them in the medullary layer. From these findings it suggests that the later commissural axons may still extend in the medullary layer about at PDT-O.
1236 Phvsiolonv,
TURNING OF NERVE GROWTH CONES INDUCED BY FOCAL ELECTRIC FIELDS IN Cal+ FREE MEDIUM S. TSUKADA. K. KEINO-MASU. T. IWAMOTO. AND J. FUKUDA Department of National Defense Medical School. Tokorozawa. Saitama. JAPAN.
Transmembrane Ca?-+ influx at nerve growth cones has been understood as essential for turning response of Here we report an opposite result that the turning response takes place in growing tips by electric currents. Dorsal root ganglia neurites growing in the absence of extracellar Ca2+; i.e., without Ca2+ influx in growth cones. dissected from newborn rats were placed in a two chamber dish so that their neurites grew in a Ca-free medium. DC current (50-250 nA) was focally applied to growth cones through a microelectrode placed approximately 100 pm distant from the growing tips. The turning responses that were induced in neurites bathed in normal Ca medium (2.5 mM) were still observed in neurites that were growing in the Ca-free medium; its Ca2+ concentration was not more than 0.01 nM. Since no Ca2+ influx is expected to occur in the growth cones in such a culture condition, we conclude that other mechanisms than Ca2+ influx at the growing ends are responsible for inducing the turning responses of rat DRG neurites.
MORPHOLOGICAL AND ELECTROPHYSIOLOGICAL ANALYSIS OF THE DEVELOPMENT OF CEREBELLAR GRANULE CELLS IN THE MICROEXPLANT CULTURE. YOSHIHIKO WAKAZONO’, SHIGEKI YUASA’, KENSUKE NAKAHIRA’, TAKASHI KURAHASHI’, AKIMICHI KANEKO’*’AND KAZUHIRO IKENAKA’, ‘Natl. Inst. for Physiol. Sci., 38 Nishigonaka, Myodaiji, Okazaki 444. 2Dept. of Physiology, School of Medicine, Keio University, Shinjuku-ku, Tokyo 160 Japan.
1237
Themode ofmigration andthedistribution of the granule cells were examined in the microexplant culture’) of mouse early postnatal cerebellum by immunocytochemical methods and retrovims mediated gene transfer. Granule cells were shown to migrate radially from the cerebellar explant by labeling the proliferating granule cells with bromodeoxyuridine. Migratory granule cells were apposed to the neurites which extended radially from the explants and formed small clusters as revealed by labeling with anti-Zic antibody), a specific marker for cerebellar granule cells. Furthermore, granule cells were identified morphologically by the retrovirus mediated transfer of fi -galactosidase gene. Patch-clamp method was applied to this microexplant culture to examine the generation of action potentials in neurons which were morphologically identified as granule cells. In view of these findings, the interelations between the generation of action potential and the morphological differentiation of the developing cerebellar granule cells will be discussed. 1) Nagata, I. and Nakatsuji, N. (1990) Dev. Brain Res. 52,63-73 2) Aruga, J. et al. (1994) J. Neurochem. 63, in press