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1308 Defective herpes simplex virus vectors for the study of l1 gene transfer into the nervous system
s143
DEFECTIVE HERPES SIMPLEX VIRUS VECTORS FOR THE STUDY OF Ll GENE TRANSFER INTO THE NERVOUS SYSTEM. TAKAHITO YAZAKI’, YOKO NAKAI’, KEIICHI UYEMURA’, ROBERT L. MARTUZA’, SAMUEL D. RABKIN*,‘Dept. of Physiol., Sch. of Med., Keio Univ., Tokyo 160, Japan. “Dept. of Neurosurg., Georgetown Univ. Med. Ctr. ) Washington, DC 20007, U.S.A.
1308
Multiple functions of the neural cell adhesion molecule Ll have been reported including cell-cell Ll plays axonal fasciculation, cell migration, and myelination. interactions, neurite elongation, important roles in neural development and neuronal regeneration in the peripheral nervous system (PNS). We constructed defective herpes simplex virus (HSV) vectors to express human or rat Ll in cultured primary rat cortical astrocytes. Rat cerebellar neurons co-cultured on a feeder layer of Llexpressing astrocytes demonstrated increased migration and neurite extension compared with neurons Because this vector system can be used to confer phenotypic co-cultured on control astrocytes. changes in primary neural cells, it will be useful for in vitro and in viva studies of neuronal regeneration and plasticity in the central nervous system (CNS).
1309 J NK A
LOCALIZATION OF E-CADHERIN IN PERIPHERAL GLL4 AFTER NERVE INJURY AND REPAIR. MITSUHJRO HASEGAWA. NAOKI MIJRAMATSU. TETSUMORI YAMASHIMA, H YAMA HITA wa. 920. JaDan.
Peripheral nerve injury results in histological and histochemical changes in neurons and glia We have recently found that Ca2+-dependent cell adhesion molecule E-cadherin plays an important role in the selective fasciculation of a particular subset of unmyelinated sensory fibers. In the present immunohistochemical and immunoblot analyses, the temporal profile of the subcellular expression of this molecule in spinal nerves was examined after crushing, transecting or ligaturing the sciatic nerve in mice with special attention paid to E-cadherin expression in glial cells. After axotomy of the sciatic nerve, distal axons of the proximal stump and the fibers of the distal stump degenerated, but E-cadherin was still detectable at the outer mesaxons of the myeliuated axons as long as they remained morphologically intact, Subsequently, Schwann cells proliferated and migrated to form Schwann cell columns (Btingner’s bands) as initial responses to denervation, and expressed E-cadherin at their site of contact with each other and later with sprouting axons. At the initial stage of myelin formation, slender processes of a single Schwaun cell interdigitated with and enveloped axons, and expressed E-cadherin at the contact site elaborated by a single Schwann cell. Immunoblot analysis on day 7 revealed that E-cadherin was detected in both the proximal nerve segments and the regenerative distal segments, but was negative in the degenerative distal segments. On the basis of present data, it is suggested that E-cadherin might be involved in the stabilization of peripheral glial network which provides the guidance of sprouting axons and myelination.
1310
DIFFERENTIAL 3-NITROPROPIONIC FUKUDA. A., DESHPANDE, .T.’ NISHIN H.l 467. Deot. ofstruct.
Hindu
Univ.5,
SUSCEPTIBILITY ACID (3-NPA) S.l*‘. FUJIMOTO,
CellBiol, Varanasi-221005,
NAIST'. India.
TO I.l,
INTRACELLULAR Ca” ( [Ca"] IN CULTURED NEURONS AND SHIMANO, Y.l, MURAMATSU,
Janan;
Deot.
ofphvsiol.,
.) INCREASES ASTROCYTES. K.l,‘, OKABE,
Inst.
ofMed.
PROMOTED A.l**,
Sci..
BY
IWASE,
Banaras
The effects of 3-NPA, a mitochondrial toxin, on [Ca2'li in cultured cortical and striatal astrocytes and neurons were examinedby [Ca2'li imaging technique using fura-2. 3-NPA (1.7 mM) increased [Ca"], in astrocytes. Pretreatment of cortical astrocytes with creatine (25 mM) significantly delayed the onset of 3-NPA-induced increase in [Ca2'].. The increase in [Ca2+li by 3-NPA was not observed in Ca-free or low-Na medium. Superfusion of 2 mM-Ca medium after exposing to 3-NPA in Ca-free medium increased the [Cai'll dramatically and reversibly. The lCa’+l iincreasewas attenuatedbyNi2' or amiloride. Themixture of several glutamate receptor antagonists hadno effectonthe [Ca2'liincreaseproducedbyreperfusion of Ca2'-containing medium. The results indicate that the 3-NPA-induced increase in [Ca"] in astrocytes was due to influx of Ca"perhaps by reverse operation of the Na-Ca exchanger system. This Ca*' influx may be responsible for the gliotoxic action of 3-NPA. On the other hand, the latency for the onset of [Ca2'11 rise in neurons was significantly longer than that of astrocytes.
DEFECTIVE HERPES SIMPLEX VIRUS VECTORS FOR THE STUDY OF Ll GENE TRANSFER INTO THE NERVOUS SYSTEM. TAKAHITO YAZAKI’, YOKO NAKAI’, KEIICHI UYEMURA’, ROBERT L. MARTUZA’, SAMUEL D. RABKIN*,‘Dept. of Physiol., Sch. of Med., Keio Univ., Tokyo 160, Japan. “Dept. of Neurosurg., Georgetown Univ. Med. Ctr. ) Washington, DC 20007, U.S.A.
1308
Multiple functions of the neural cell adhesion molecule Ll have been reported including cell-cell Ll plays axonal fasciculation, cell migration, and myelination. interactions, neurite elongation, important roles in neural development and neuronal regeneration in the peripheral nervous system (PNS). We constructed defective herpes simplex virus (HSV) vectors to express human or rat Ll in cultured primary rat cortical astrocytes. Rat cerebellar neurons co-cultured on a feeder layer of Llexpressing astrocytes demonstrated increased migration and neurite extension compared with neurons Because this vector system can be used to confer phenotypic co-cultured on control astrocytes. changes in primary neural cells, it will be useful for in vitro and in viva studies of neuronal regeneration and plasticity in the central nervous system (CNS).
1309 J NK A
LOCALIZATION OF E-CADHERIN IN PERIPHERAL GLL4 AFTER NERVE INJURY AND REPAIR. MITSUHJRO HASEGAWA. NAOKI MIJRAMATSU. TETSUMORI YAMASHIMA, H YAMA HITA wa. 920. JaDan.
Peripheral nerve injury results in histological and histochemical changes in neurons and glia We have recently found that Ca2+-dependent cell adhesion molecule E-cadherin plays an important role in the selective fasciculation of a particular subset of unmyelinated sensory fibers. In the present immunohistochemical and immunoblot analyses, the temporal profile of the subcellular expression of this molecule in spinal nerves was examined after crushing, transecting or ligaturing the sciatic nerve in mice with special attention paid to E-cadherin expression in glial cells. After axotomy of the sciatic nerve, distal axons of the proximal stump and the fibers of the distal stump degenerated, but E-cadherin was still detectable at the outer mesaxons of the myeliuated axons as long as they remained morphologically intact, Subsequently, Schwann cells proliferated and migrated to form Schwann cell columns (Btingner’s bands) as initial responses to denervation, and expressed E-cadherin at their site of contact with each other and later with sprouting axons. At the initial stage of myelin formation, slender processes of a single Schwaun cell interdigitated with and enveloped axons, and expressed E-cadherin at the contact site elaborated by a single Schwann cell. Immunoblot analysis on day 7 revealed that E-cadherin was detected in both the proximal nerve segments and the regenerative distal segments, but was negative in the degenerative distal segments. On the basis of present data, it is suggested that E-cadherin might be involved in the stabilization of peripheral glial network which provides the guidance of sprouting axons and myelination.
1310
DIFFERENTIAL 3-NITROPROPIONIC FUKUDA. A., DESHPANDE, .T.’ NISHIN H.l 467. Deot. ofstruct.
Hindu
Univ.5,
SUSCEPTIBILITY ACID (3-NPA) S.l*‘. FUJIMOTO,
CellBiol, Varanasi-221005,
NAIST'. India.
TO I.l,
INTRACELLULAR Ca” ( [Ca"] IN CULTURED NEURONS AND SHIMANO, Y.l, MURAMATSU,
Janan;
Deot.
ofphvsiol.,
.) INCREASES ASTROCYTES. K.l,‘, OKABE,
Inst.
ofMed.
PROMOTED A.l**,
Sci..
BY
IWASE,
Banaras
The effects of 3-NPA, a mitochondrial toxin, on [Ca2'li in cultured cortical and striatal astrocytes and neurons were examinedby [Ca2'li imaging technique using fura-2. 3-NPA (1.7 mM) increased [Ca"], in astrocytes. Pretreatment of cortical astrocytes with creatine (25 mM) significantly delayed the onset of 3-NPA-induced increase in [Ca2'].. The increase in [Ca2+li by 3-NPA was not observed in Ca-free or low-Na medium. Superfusion of 2 mM-Ca medium after exposing to 3-NPA in Ca-free medium increased the [Cai'll dramatically and reversibly. The lCa’+l iincreasewas attenuatedbyNi2' or amiloride. Themixture of several glutamate receptor antagonists hadno effectonthe [Ca2'liincreaseproducedbyreperfusion of Ca2'-containing medium. The results indicate that the 3-NPA-induced increase in [Ca"] in astrocytes was due to influx of Ca"perhaps by reverse operation of the Na-Ca exchanger system. This Ca*' influx may be responsible for the gliotoxic action of 3-NPA. On the other hand, the latency for the onset of [Ca2'11 rise in neurons was significantly longer than that of astrocytes.