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THE SUBCELLULAR DISTRIBUTION OF ENDOGENOUS AND EXOGENOUS SEROTONIN IN BRAIN TISSUE: COMPARISON OF SYNAPTOSOMES STORING SEROTONIN, NOREPINEPHRINE, AND γ‐AMINOBUTYRIC ACID 1
Author(s) -
Kuhar M. J.,
Shaskan E. G.,
Snyder S. H.
Publication year - 1971
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1111/j.1471-4159.1971.tb11962.x
Subject(s) - differential centrifugation , centrifugation , endogeny , sucrose , serotonin , chemistry , synaptosome , monoamine oxidase , isopycnic , chromatography , pargyline , biophysics , distribution (mathematics) , cell fractionation , cytoplasm , biochemistry , biology , enzyme , receptor , membrane , mathematical analysis , mathematics
— We have studied the subcellular distribution of exogenous and endogenous serotinin in slices from the hypothalamus and midbrain of several species. In a procedure which appears to label the endogenous pools, tissue slices were incubated with low concentrations of [ 3 H]5‐HT (5 × 10 ‐8 M), for 45 min, when there was apparent equilibrium between [ 3 H]5‐HT of tissue and medium. After the tissue slices were homogenized in 0‐32 M‐sucrose and subjected to differential centrifugation, the distribution of exogenous and endogenous 5‐HT in pellets and supernatant fluid was similar. In some experiments, the crude mitochondrial pellets were resuspended in 0‐32 M‐sucrose, layered on linear, continuous density gradients of sucrose (1 ‐5‐0‐32 M), and centrifuged for short times (incomplete equilibrium centrifugation). The subcellular distribution of particulate tritium, total tritium, and particulate endogenous 5‐HT was the same in portions of the gradients containing synaptosomes. The peak distribution of [ 3 H]5‐HT in sucrose gradients was separable from the peak for [ 14 C]GABA by four to five fractions; potassium (a marker for cytoplasm occluded within synaptosomes) occurred in the regions of the gradients containing most of the labelled compounds. The distribution of monoamine oxidase activity (a mitochondrial marker) overlapped the distribution of [ 3 H]5‐HT after a 15 min centrifugation but appeared in denser regions of the gradient after centrifuging for 2 h. Particles containing [3H]5‐HT and [I4C]NE were slightly but consistently separable in synaptosomal fractions isolated from the hypothalamus or midbrain of rat, guinea pig and hamster.