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Dopaminergic involvement in the pharmacological actions of Buspirone
66
PosterSession IE. Neurotoxicology
in cultured neurons. The objective of this study was to determine the mode of neuronal cell death (apoptosis or necrosis) in areas of the brain sensitive to cyanide. KCN (6 mglkg, i.p.) administered to mice for 1-12 days was associated with apoptotic cell death in the cortex as determined by in situ detection of DNA strand breaks (TUNEL assay). Maximal apoptosis was observed by day 3. Necroticcell death was evident by day 3 in the substantia nigra (SN) and was progressive to day 12. Glial activation (as determined by GFAP proliferation) was most pronounced on day 12. Pretreatment of mice with MK801, nitric oxide inhibitors and cyclooxygenase-2 inhibitors markedly attenuated the cortical apoptosis, whereas SN necrosis was blocked only by nitric oxide inhibitors. These results indicate that activation of the NMDA receptor initiates intracellular cascades leading to oxidative stress and eventually apoptosis in the cortex. However, in the SN the necrotic response to cyanide appears to involvea mechanism independentof the NMDA receptor. (Supported by NIH grant ES(4140).
astrocytesand it serves as a marker for glial differentiation. Our data suggest that tiazofurin has both anti-proliferative and differentiating effects on C6 rat glioma cells.
IP1 E120 I EXPRESSION COORDINATED REGULATION OF GENE FROM THE ACETYLCHOLINESTERASE LOCUS M. Shapira *1, M. Korner" , L. Bosgraaf", B. Hinzmanrr", A. Rosenthal'", H. Soreq1 • J Department ofBiological Chemistry, The HebrewUniversity ofJerusalem, 91904. Israel; 2Department of GenomeAnalysis. lnstitutfur Molekulaire Biotechnologie, lena. Germany
IP1E1191 DIFFERENTIATION EFFECT OF TIAZOFURIN ON PROLIFERATION AND OF C6 RATGLIOMA CELLS
The human (h) ACHEgene encodes for acetylcholinesterase (AChE), a key enzyme in drug responses, poison scavengingand psychological stress processes. To define genomic elements regulating AChE expression, we sequenced a 35 Kb cosmid including the hACHE gene, 20 Kb upstream and ca. 8 Kb downstream sequences. Within 2 Kb downstream to the ACHE gene and in an inverse orientation, another cluster of exons exhibited 81% homology to the ARS2 Chinese hamster gene presumed to confer resistance to the toxic element arsenite, which is also a specific cholinesterase inhibitor. A "virtual" hARS cDNA sequence assembled by combining database-deposited ESTs with genomic DNA sequences extends beyond this cosmid and encodes a protein with a hydrophobic N-terminus, a charged a-helix and an intriguing amphipathic a-helix. hARS displays no resemblanceto any knownprotein or peptide motif, and is expressed ubiquitously in a pattern that overlaps with that of ACHE. This and the genomic co-localization of the hACHE and hARS genes, suggested co-regulation. Indeed, hARS mRNA levels were lower in brains of C571b6J mice, known for their paucity of cholinergic neurons and lower total AChE levels, compared with control FVBIN mice. Also, the upstream cosmid sequence includes multiple STRE elements associated with cellular stress responses; and semi-quantitative RT-PCR experiments demonstrated elevated levels of both brain ARS mRNA and ACHE mRNA in psychologically stressed mice compared with controls. While the function of hARS yet needs to be explored, our findings suggest that the reported increases in ARS mRNA levels under arsenite exposure are a secondary outcome of ACHEupregulationin responseto this poisoning, reflecting coordinated expressionof at least two stress responsivegenes.
V. Piperski*, M. Vracar, K. Drabek, M. Papovic, S. Jovanovic, M. Jokanovic, Lj . Rakic. ICN Yugoslavia Institute, Centerfor biomedical research, Pasterova 2, 11()()() Belgrade, Yugoslavia
IP1 E121 I CORPUS STRIATUM LONG-LAST1NG EFFECTS IN ADULT MICEPRENATALLY TREATED WITH
IP1 E1181 DOPAMINERGIC INVOLVEMENT IN THE
PHARMACOLOGICAL ACT10NS OF BUSPIRONE
M.M. Badawi *, N.T. El-Mossalamy, A.M. Shehata. Faculty of science, Cairouniv ami NODCAR. Cairo, Egypt Buspirone has demonstrable anticonflict actions in several animal models predictivefor its anxiolytic activity in man. The mechanism by which Buspironeexerts its pharmacological effects is not clear. In an attempt to explore the neurochemicalbasisof its action, Buspirone was administered I.P (10 mglkg) in adult male albino rats and its effect on the dopaminergic neurotransmissionwas investigated. Animals were subjectedto neuro-behavioral test to analyse the behavior, congitive functions and emotions. The animals were sacrificed after I, 2, 3 and 4 weeks of continous drug administration. Dopamine was measured in plasma and in five brain areas. It was found that Buspirone minimizethe stimulatory effects of the dopaminergic agonist, Bromocriptine mesylate, on the dopaminergic neurotransmission in rats. The study may suggest that Buspirone exert its effects, at least, through its antidoparninergie activity.
Drugs that promote malignant cell differentiation are in the focus of interest for cancer treatment. Tiazofurin (2-p-D-ribofuranosylthiasole-4-carboxamide) inhibits activity of inosine-monophosphate-dehydrogenase, the rate-limiting enzyme in GTP biosynthesis, decreases GTP concentrationin malignant cells and exibits anti-proliferative and differentiating effect on several malignant cell lines. The purpose of this study was to investigate the effect of tiazofurin on proliferation and phenotypic characteristics of C6 rat glioma cells, using cell counting and immunocytochemistry with a panel of antibodies against cytosceletal and other proteins: glial fibrillary acidic protein (GFAP), S-IOO protein, cytokeratines (CK) 8, 18 and 19, and vimentin. Tuazofurin treatment induced marked decrease in cell number and this effect was dose and time dependent. More than 95% of cells were viable, as evidencedby trypan blue exclusion. Untreated C6 cells were GFAP-negative, S-IOO-positive, CK8-negative, CK-18 and CKl9-positive and vimentin-positive. Incubation with tiazofurin (I5--{j0 p.M) for 24 h changed morphology of C6 cells and immunostainingpattern. The most important finding was induction of GFAP immunoreactivity which became evident after 24 h incubation and persisted after 48 h and 72 h. GFAP is the major constituent of intermediate filaments in normal, reactive and neoplastic
DIAZEPAM
A. Marquez-Orozco -. M.e. Marquez-Orozco, M.V. Gazca-Ramfrez, G. de la Fuente-Juarez. EmbryolDept.Faculty of Medicine UNAM. POB 70-553Mexico 04510 D.F.. Mexico Diazepam (DZ) produces in fetal mice corpus striatum a delay in the neuroblastic and nerve fibers differentiation. We investigate if the histologicalalterations of the fetal corpus striatum occur in adult mice. Single daily doses (2.7 mglkg/sc) of DZ were administrated to CD-l strain female mice, from gestation day 6 to 17. A second group (S) received saline solution and a third group was non-treated (NT). The offspring's were wet-nursed by NT mice, weaned and kept for 240 days. Mice were deeply anesthetized, perfused with 10% formalin, and the brains were removed, fixed and stained for myelin, nerve cells, and MOns identification; a selected blocks were impregnated with the fast-Golgi technique and observed under light microscope. In the DZ was evident an increased nuclear density of neurons per area (p < 0.05), less number of nerve fibers, and they were coarse and irregular. Such alterations could revealdisruption in cell multiplication and also it could be attributed to DZ inhibition protein synthesis. The results suggest that DZ produce permanent
PosterSession IE. Neurotoxicology
in cultured neurons. The objective of this study was to determine the mode of neuronal cell death (apoptosis or necrosis) in areas of the brain sensitive to cyanide. KCN (6 mglkg, i.p.) administered to mice for 1-12 days was associated with apoptotic cell death in the cortex as determined by in situ detection of DNA strand breaks (TUNEL assay). Maximal apoptosis was observed by day 3. Necroticcell death was evident by day 3 in the substantia nigra (SN) and was progressive to day 12. Glial activation (as determined by GFAP proliferation) was most pronounced on day 12. Pretreatment of mice with MK801, nitric oxide inhibitors and cyclooxygenase-2 inhibitors markedly attenuated the cortical apoptosis, whereas SN necrosis was blocked only by nitric oxide inhibitors. These results indicate that activation of the NMDA receptor initiates intracellular cascades leading to oxidative stress and eventually apoptosis in the cortex. However, in the SN the necrotic response to cyanide appears to involvea mechanism independentof the NMDA receptor. (Supported by NIH grant ES(4140).
astrocytesand it serves as a marker for glial differentiation. Our data suggest that tiazofurin has both anti-proliferative and differentiating effects on C6 rat glioma cells.
IP1 E120 I EXPRESSION COORDINATED REGULATION OF GENE FROM THE ACETYLCHOLINESTERASE LOCUS M. Shapira *1, M. Korner" , L. Bosgraaf", B. Hinzmanrr", A. Rosenthal'", H. Soreq1 • J Department ofBiological Chemistry, The HebrewUniversity ofJerusalem, 91904. Israel; 2Department of GenomeAnalysis. lnstitutfur Molekulaire Biotechnologie, lena. Germany
IP1E1191 DIFFERENTIATION EFFECT OF TIAZOFURIN ON PROLIFERATION AND OF C6 RATGLIOMA CELLS
The human (h) ACHEgene encodes for acetylcholinesterase (AChE), a key enzyme in drug responses, poison scavengingand psychological stress processes. To define genomic elements regulating AChE expression, we sequenced a 35 Kb cosmid including the hACHE gene, 20 Kb upstream and ca. 8 Kb downstream sequences. Within 2 Kb downstream to the ACHE gene and in an inverse orientation, another cluster of exons exhibited 81% homology to the ARS2 Chinese hamster gene presumed to confer resistance to the toxic element arsenite, which is also a specific cholinesterase inhibitor. A "virtual" hARS cDNA sequence assembled by combining database-deposited ESTs with genomic DNA sequences extends beyond this cosmid and encodes a protein with a hydrophobic N-terminus, a charged a-helix and an intriguing amphipathic a-helix. hARS displays no resemblanceto any knownprotein or peptide motif, and is expressed ubiquitously in a pattern that overlaps with that of ACHE. This and the genomic co-localization of the hACHE and hARS genes, suggested co-regulation. Indeed, hARS mRNA levels were lower in brains of C571b6J mice, known for their paucity of cholinergic neurons and lower total AChE levels, compared with control FVBIN mice. Also, the upstream cosmid sequence includes multiple STRE elements associated with cellular stress responses; and semi-quantitative RT-PCR experiments demonstrated elevated levels of both brain ARS mRNA and ACHE mRNA in psychologically stressed mice compared with controls. While the function of hARS yet needs to be explored, our findings suggest that the reported increases in ARS mRNA levels under arsenite exposure are a secondary outcome of ACHEupregulationin responseto this poisoning, reflecting coordinated expressionof at least two stress responsivegenes.
V. Piperski*, M. Vracar, K. Drabek, M. Papovic, S. Jovanovic, M. Jokanovic, Lj . Rakic. ICN Yugoslavia Institute, Centerfor biomedical research, Pasterova 2, 11()()() Belgrade, Yugoslavia
IP1 E121 I CORPUS STRIATUM LONG-LAST1NG EFFECTS IN ADULT MICEPRENATALLY TREATED WITH
IP1 E1181 DOPAMINERGIC INVOLVEMENT IN THE
PHARMACOLOGICAL ACT10NS OF BUSPIRONE
M.M. Badawi *, N.T. El-Mossalamy, A.M. Shehata. Faculty of science, Cairouniv ami NODCAR. Cairo, Egypt Buspirone has demonstrable anticonflict actions in several animal models predictivefor its anxiolytic activity in man. The mechanism by which Buspironeexerts its pharmacological effects is not clear. In an attempt to explore the neurochemicalbasisof its action, Buspirone was administered I.P (10 mglkg) in adult male albino rats and its effect on the dopaminergic neurotransmissionwas investigated. Animals were subjectedto neuro-behavioral test to analyse the behavior, congitive functions and emotions. The animals were sacrificed after I, 2, 3 and 4 weeks of continous drug administration. Dopamine was measured in plasma and in five brain areas. It was found that Buspirone minimizethe stimulatory effects of the dopaminergic agonist, Bromocriptine mesylate, on the dopaminergic neurotransmission in rats. The study may suggest that Buspirone exert its effects, at least, through its antidoparninergie activity.
Drugs that promote malignant cell differentiation are in the focus of interest for cancer treatment. Tiazofurin (2-p-D-ribofuranosylthiasole-4-carboxamide) inhibits activity of inosine-monophosphate-dehydrogenase, the rate-limiting enzyme in GTP biosynthesis, decreases GTP concentrationin malignant cells and exibits anti-proliferative and differentiating effect on several malignant cell lines. The purpose of this study was to investigate the effect of tiazofurin on proliferation and phenotypic characteristics of C6 rat glioma cells, using cell counting and immunocytochemistry with a panel of antibodies against cytosceletal and other proteins: glial fibrillary acidic protein (GFAP), S-IOO protein, cytokeratines (CK) 8, 18 and 19, and vimentin. Tuazofurin treatment induced marked decrease in cell number and this effect was dose and time dependent. More than 95% of cells were viable, as evidencedby trypan blue exclusion. Untreated C6 cells were GFAP-negative, S-IOO-positive, CK8-negative, CK-18 and CKl9-positive and vimentin-positive. Incubation with tiazofurin (I5--{j0 p.M) for 24 h changed morphology of C6 cells and immunostainingpattern. The most important finding was induction of GFAP immunoreactivity which became evident after 24 h incubation and persisted after 48 h and 72 h. GFAP is the major constituent of intermediate filaments in normal, reactive and neoplastic
DIAZEPAM
A. Marquez-Orozco -. M.e. Marquez-Orozco, M.V. Gazca-Ramfrez, G. de la Fuente-Juarez. EmbryolDept.Faculty of Medicine UNAM. POB 70-553Mexico 04510 D.F.. Mexico Diazepam (DZ) produces in fetal mice corpus striatum a delay in the neuroblastic and nerve fibers differentiation. We investigate if the histologicalalterations of the fetal corpus striatum occur in adult mice. Single daily doses (2.7 mglkg/sc) of DZ were administrated to CD-l strain female mice, from gestation day 6 to 17. A second group (S) received saline solution and a third group was non-treated (NT). The offspring's were wet-nursed by NT mice, weaned and kept for 240 days. Mice were deeply anesthetized, perfused with 10% formalin, and the brains were removed, fixed and stained for myelin, nerve cells, and MOns identification; a selected blocks were impregnated with the fast-Golgi technique and observed under light microscope. In the DZ was evident an increased nuclear density of neurons per area (p < 0.05), less number of nerve fibers, and they were coarse and irregular. Such alterations could revealdisruption in cell multiplication and also it could be attributed to DZ inhibition protein synthesis. The results suggest that DZ produce permanent