RAT BRAIN TRYPTOPHAN HYDROXYLASE: PROPERTIES AND EFFECTS OF 5,7-DIHYDROXYTRYPTAMINE.
Item
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Title
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RAT BRAIN TRYPTOPHAN HYDROXYLASE: PROPERTIES AND EFFECTS OF 5,7-DIHYDROXYTRYPTAMINE.
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Identifier
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AAI8401922
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identifier
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8401922
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Creator
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CLEWANS, CAROL SYDNEY.
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Contributor
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Dr. E.C. Azmitia
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Date
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1983
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Language
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English
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Publisher
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City University of New York.
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Subject
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Biology, General
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Abstract
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It has previously been shown using anatomical methods that partial denervation of the rat hippocampus by removal of serotonergic (5-HT) fibers in the cingulum bundle induces sprouting of intact 5-HT fibers reaching the hippocampus in the fornix-fimbria (Azmitia et al., 1978; Zhou and Azmitia, 1981). The biochemical properties of sprouted fibers have remained largely uncharacterized. Thus, the rate-limiting enzyme in 5-HT synthesis, tryptophan hydroxylase (TPOH), was studied to determine whether new sprouts possess the ability to synthesize 5-HT.;Optimal assay conditions were established for measuring TPOH activity in the presence of tetrahydrobiopterin (BH(,4)) using the radiometric method of Ichiyama et al. (1970). The effects of reducing and phosphorylating conditions were studied. Enzyme activity was maximally stimulated by incubation in the presence of 2 mM DTT and 14 (mu)M Fe('++). Phosphorylating conditions produced a slight but variable increase in TPOH activity depending on the conditions of enzyme preincubation and on the cofactor used (BH(,4) or 6-methyltetrahydropterin). Neither a change in pH nor addition of an ATP-regenerating system or EDTA increased the amount of activation by phosphorylating conditions.;Unilateral stereotaxic injections of 5 (mu)g of the specific 5-HT neurotoxin 5,7-dihydroxytryptamine were made into the cingulum bundle of adult rats pretreated with desmethylimipramine in order to produce partial 5-HT deafferentation of the hippocampus. Three to 90 days after the injection, TPOH activity was measured in ipsilateral and contralateral hippocampi and in the midbrain, site of neuronal cell bodies of hippocampi and in the midbrain, site of neuronal cell bodies of hippocampal 5-HT projections. Enzyme activity was also measured in hypothalamus and septum/preoptic area 3 to 14 days post-lesion in order to estimate the extent of retrograde degeneration of 5-HT fibers.;In both hippocampi, enzyme activity declined gradually, reaching minimal levels 7 to 14 days post-lesion. This decrease in enzyme activity was paralleled by a decrease in the Vmax and an increase in the Km of TPOH for tryptophan. TPOH activity increased from 28 to 60 days and decreased again from 60 to 90 days post-lesion. In midbrain, enzyme activity gradually increased, reaching levels significantly greater than sham at 28 days post-lesion. Enzyme activity in the hypothalamus and septum/preoptic area was unaffected 3 to 14 days after the lesion.;The results indicate that 5-HT fibers remaining in the hippocampus following partial denervation are able to compensate biochemically for those removed by cingulum bundle lesions. Changes in hippocampal TPOH can be related to simultaneous changes in midbrain TPOH and to the appearance of sprouts from intact 5-HT fibers.
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Type
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dissertation
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Source
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PQT Legacy CUNY.xlsx
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degree
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Ph.D.
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Program
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Biomedical Sciences
