The adrenal medulla: A model for studies of hormonal and neuronal storage and release mechanisms

Helle, Karen B.; Serck‐Hanssen, Guldborg

doi:10.1007/bf01732006

Cited by 57 publications

(19 citation statements)

References 87 publications

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“…All of the sucrose solutions used in the fractionation procedures were buffered with 5 mM potassium phosphate (pH 7.3). Adrenal medulla was fractionated according to the following modification of the procedure of Helle and Serck-Hanssen (16). The medullae from 10 bovine adrenals or 20 rat adrenals were dissected in the cold and homogenized with an Ultra-Turrax for 90 sec in 5 vol of ice-cold 0.25 M sucrose.…”

Section: Methodsmentioning

confidence: 99%

Cellular and subcellular localization of protein I in the peripheral nervous system.

Fried¹,

Nestler²,

Camilli³

et al. 1982

Proc. Natl. Acad. Sci. U.S.A.

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The cellular and subcellular distribution of protein I, a major brain phosphoprotein, has been studied in the peripheral nervous system. The levels of protein I in various peripheral nerves and innervated peripheral tissues were determined by radioimmunoassay and radioimmunolabeling of polyacrylamide gels. The results indicated that protein I is present throughout the peripheral nervous system. Denervation studies ofadrenal medulla and iris suggested that the protein I contained in peripheral tissues is localized to the neuronal elements innervating those tissues. Protein I was found to be enriched in neurotransmitter vesicle fractions of peripheral nervous tissue. Moreover, protein I appeared to be transported from cell bodies to axon terminals at least partly in association with neurotransmitter vesicles.Protein I is a neuron-specific protein (1) that is a prominent endogenous substrate in brain for both cyclic AMP-dependent (1) and calcium/calmodulin-dependent (2-4) protein kinases. The state of phosphorylation of protein I has been shown to be regulated by impulse conduction (5), neurotransmitters (6, 7), and depolarizing agents (6-8) in intact preparations of neuronal tissue. The distribution of protein I in the central nervous system has been studied by light and electron immunocytochemistry (9, 10), subcellular fractionation (11, t), and radioimmunoassay (12). Protein I is present throughout the central nervous system. It is concentrated in presynaptic nerve terminals, where it is associated at least partially with neurotransmitter vesicles. Protein I has also been detected immunocytochemically, apparently in axon terminals, throughout the peripheral nervous system (13).In the present study we have determined the cellular and subcellular distribution of protein I in various peripheral nervous tissues which have the advantage over brain of being better defined, less complex, and more easily manipulated experimentally. MATERIALS AND METHODSMaterials. Pure protein I was kindly supplied by Louis J.DeGennaro of our laboratory. "2I-Labeled protein A (specific activity, 30 mCi/mg; 1 Ci = 3.7 X 1010 becquerels) was purchased from Amersham and protein A-bearing Staphylococcus aureus cells (SAC) (Pansorbin) were from Calbiochem-Behring.Denervation. Rats with denervated adrenal glands (achieved by transsection of the splanchnic nerve), as well as sham-operated control rats, were obtained from Zivic-Miller Laboratories (Allison Park, PA). Individual adrenal glands were analyzed 2-4 weeks after surgery, at which time denervation is complete (14). Irides were sympathetically denervated by superior cervical ganglionectomy of Sprague-Dawley rats from Anticimex (Stockholm, Sweden). This procedure causes complete degeneration of iris sympathetic nerves within 48 hr (15). Sixteen normal irides and 30 denervated irides, taken 5 days after surgery, were pooled for protein I analysis. Subcellular Fractionation. All of the sucrose solutions used in the fractionation procedures were buffered with 5 mM potassium phosphat...

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Section: Methodsmentioning

confidence: 99%

Cellular and subcellular localization of protein I in the peripheral nervous system.

Fried¹,

Nestler²,

Camilli³

et al. 1982

Proc. Natl. Acad. Sci. U.S.A.

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“…This selection of vesicles was from a mixed population consisting primarily of LDV 700-800 A diameter in mature cattle and fewer small dense core vesicles (SDV) ~ 45o A diameter, both with contents ranging from translucent to opaque i971 ;Tranzer,I973). Interestingly, LDV were remarkably resistant to osmotic shock and freeze-thaw procedures (Euler,I958;Stj~rne,I964;I974;I975), which are highly effective techniques to release the content of adrenomedullary vesicles (Hillarp and Nilsson,I954;Hillarp,I958;Stjfirne,I964;Poisner et al,I967;Winkler et al, I97O;Taugner, I97I;Helle, I973;Taugner and W~hler, I974;Helle and Serck-Hanssen, 1975).…”

Section: Introductionmentioning

confidence: 96%

Morphological effects of osmolarity on purified noradrenergic vesicles

1975

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Large dense core vesicles (LDV) were purified from bovine splenic nerve homogenates by the sucrose-D2O density gradient method. Vesicles were subjected to a 50% increase and decrease in osmolarity from the control 330 mosmol 1(-1) by adjusting sucrose or potassium phosphate buffer during pre- and/or postfixation. Control vesicles with a mean diameter of 717 A readily swelled to approximately 1050 A and shrunk to approximately 600 A in the hypotonic and hypertonic media, respectively, with either sucrose or phosphate buffer. The dense core responded similarly but to a lesser degree. Prefixation in glutaraldehyde had little effect on vesicle sensitivity to subsequent tonicity change, not did the fixative per se exert an obvious osmotic effect. Thus, final vesicle size was largely determined by the OsO4 postfixation medium and principally by the vehicle rather than the fixative. In controls there was a mixture of spherical to oblate vesicles mostly filled with an electron-dense matrix. Upon swelling, more vesicles became spherical and nearly all had a prominent translucent halo between core and membrane. Upon shrinking, more vesicles became oblate, the halo was obliterated and the electron-density of the matrix increased. Frequency distributions of vesicle diameters at different tonicity clearly indicated that the diameter of LDV could overlap the 400-500 A range characteristic of small dense core vesicles; however, there was no suggestion of a population of the latter in the purified LDV fraction. Implications are discussed concerning the biochemical and morphological identification of 'light' and 'heavy' density peaks of noradrenaline and dopamine beta-hydroxylase from mixed vesicle populations and the possible relevance of changes in vesicle shape to a functional state in situ.

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“…4A was not consistently seen and was therefore considered to be an artifact. Chromogranin A has been reported to be identical with the monomer form of DBH (HELLE & SERCK-HANSSEN, 1975;AUNIS et al, 197%) and on this basis might be expected to exhibit lectinbinding properties similar to DBH. However, other work has cast doubt upon the identity of these two proteins (GEISSLER et al, 1977;WINKLER, 1976;AUNIS;personal communication), and the results presented here clearly show that at least in respect to the carbohydrate compostion and the lectin-binding properties these two proteins are not identical.…”

mentioning

confidence: 95%

Soluble and Membrane Lectin‐binding Glycoproteins of the Chromaffin Granule

Cahill

Morris

1979

Journal of Neurochemistry

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Abstract— Fluorescein isothiocyanate‐labelled lectins were used to identify lectin‐binding glycoproteins of the chromaffin granule after electrophoresis of the membrane and soluble granule proteins on sodium dodecyl sulphate polyacrylamide slab gels. The glycoprotein nature of all lectin‐binding bands was confirmed by staining the gels for carbohydrates, and the specificity of the lectin‐binding was demonstrated by hapten sugar inhibition of binding. In samples of granule membrane proteins reduced with dithiothreitol 10 concanavalin A (Con A), 5 wheat germ agglutinin, 8 Ricinus communis agglutinin‐60, and 7 Ricinus communis agglutinin‐120 (RCA‐120) binding glycoproteins were identified. Molecular weights of these glycoproteins varied from 20,000 to 200,000 daltons. All but two of the Con A‐binding bands and one of the RCA‐120 binding bands appeared to react with more than one lectin, suggesting possible carbohydrate heterogeneity in these membrane glycoproteins. The band identified as dopamine β‐hydroxylase reacted most intensely with all four lectin tested, and in the soluble core material this enzyme was the sole significant lectin binding glycoprotein.

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The adrenal medulla: A model for studies of hormonal and neuronal storage and release mechanisms

Cited by 57 publications

References 87 publications

Cellular and subcellular localization of protein I in the peripheral nervous system.

Cellular and subcellular localization of protein I in the peripheral nervous system.

Morphological effects of osmolarity on purified noradrenergic vesicles

Soluble and Membrane Lectin‐binding Glycoproteins of the Chromaffin Granule

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