A protein fraction was isolated from rat mast cell granules. Disc gel electrophoresis and peptide mapping showed this fraction to be a mixture of 1 or 2 main components and 2–3 minor components. On ultracentrifugation the protein fraction behaved as an homogeneous material with a molecular weight of 5600. Amino acid analysis showed the presence of all the common amino acids (with the possible exception of tryptophan). High contents of lysine, aspartic acid, glutamic acid, proline, glycine and valine were noted. The content of cysteine was remarkably low. The isoelectric point of the main component(s) was found to be around 9.
Subcellular fractions containing adrenergic vesicles were obtained from bovine adrenal medulla and splenic nerve, rat and cat heart, and rat spleen. The fractions were washed free from lipids, digested with papain and the mucopolysaccharides (MPSs) then precipitated from the supernatant with ethanolic potassium acetate. The mpsswere identified by several different methods--microelectrophoresis on cellulose acetate in various media, cellulose column chromatography using elution solvents of increasing ionic strength, and treatment with specific enzymes followed by electrophoresis or column chromatography. The MPS precipitate from all the sources investigated contained dermatan sulphate or a dermatan sulphate-chondroitin sulphate hybrid. in addition, the precipitate from rat spleen was found to contain chondroitin sulphate. Heparan sulphate was found in the precipitates from rat heart and spleen and hyaluronic acid in that from bovine splenic nerve. The finding of sulphomucopolysaccharides in the adrenergic vesicles, probably in a complex with protein, raises the question of the functional significance of such complexes. They might, by analogy with the ion-exchange function of the heparin-protein complex in mast cell granules, play a role in the storage and release of the amines.
ANGGARD, E., U. BERGQVIST, B. HOGBERG, K. JOHANSSON, I. L. THON and B. UVNAS. Biologically active principles occurring on histamine release from cat paw, guinea pig lung and isolated rat mast cells. Acta physiol. scand. 1963. 59. 97-1 10. -Histamine release was induced from perfused cat paws and isolated rat mast cells with compound 48/80, and from sensitized guinea pig lungs with antigen. Concomitantly occurring biologically active substances were separated by various chromatographic procedures (silicic acid column, silicic acid impregnated paper, etc.). From all the species several spasmogenic principles were obtained. Chemical and biological properties indicated the presence of unsaturated fatty acids, phosphatidyl and phosphatidal choline and an "SRS" principle. The "SRS" principles from the three species seemed to be identical or very closely related. Phosphatidase A-like and chymotrypsin-like enzymes are present in mast cells. Such enzymes are the only enzymes shown to be able to degranulate mast cells. The possible connection between these enzymes and the formation of spasmogenic principles during histamine release is discussed. I n a series of papers we have presented evidence supporting the view that the histamine release that occurs i n the presence of compound 48/80, extracts from Cyanea a n d Ascaris (in the cat a n d rat), a n d antigens (in the guinea pig), is brought about by identical or very similar release processes in the mast cells of the three animal species UVNAS 1960, UVNAS 1961). T h e 7-633015. Acta physiol. scand. Vol. 59. 97
Rats treated at birth with 6-OH DA were sacrificed in adult age 24 h after an injection of 3H-NA. Synaptosomes were isolated from the cerebral cortex, the hypothalamus, the pons-medulla region and the cerebellum. The similar distribution pattern of 3H-NA and 35S on gradient centrifugation of the synaptosome preparations, the similar effect of 6-OHDA on the uptake of 3H-NA into slices of brain tissue in vitro and on their 35S content as well as the identification of chondroitin and heparan sulphate in the synaptosome fractions are observations which indicate a possible function of SMPSs in the storage of NA in adrenergic terminals.
The histamine in the granules of mast cells has been proposed to be stored as a heparin-zinc-histamine complex. Due to its chelating action zinc should increase (double) the histamine binding capacity of heparin. We have determined the zinc content of isolated rat peritoneal mast cells to 2.4-4.1 nmol/10-6 cells and of their granules to 13.3-21.4 nmol/mg dry weight. The corresponding amounts of histamine were 150 and 680 nmol respectively. The zinc content found is far (20 times) too low to allow for an adequate binding of histamine in a heparin-zinc-histamine complex. In vitro the granules take up zinc in the same manner as they take up other cations and zinc competes with histamine for granule storage sites. Consequently, Hi-uptake is reduced and not enhanced in the presence of zinc in the suspension medium. In summary no evidence was found for a storage function of zinc in a heparin-zinc-histamine complex.
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