Binding and internalization of a iodinated substance P analog by cultured anterior pituitary cells

Larsen, Philip J.; Mikkelsen, Jens D.; Mau, Søren Mads; Særmark, T.

doi:10.1016/0303-7207(89)90169-x

Cited by 27 publications

(19 citation statements)

References 46 publications

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“…Finally, it is possible to arrest enzymatic breakdown and internalization of the ligand, which otherwise could cause serious interpretational problems. As evaluated from the present study and earlier HPLC analysis of the incubation medium after 30 min of incubation to O' C, the [ lZ51]-BH-SP ligand remains intact and is not susceptible to internalization (Larsen et al, 1989b). When using in vitro techniques involving tissue sections, to obtain acceptable cell morphology one must use lightly fixed sections.…”

Section: Discussionmentioning

confidence: 71%

“…The combination with immunocytochemistry has made it possible to confine the binding of a radioligand to biochemically defined cells. The number of SP binding sites within the anterior pituitary gland is fairly low, and the receptor ligand complex is internalized and recycled (Larsen et al, 1989b). Therefore, it is likely that the synthesis rate of SP receptors in this organ is very low, and this could explain why a recent attempt to demonstrate mRNA coding for the SP receptor in anterior pituitary tissue extracts found specific mRNA content to be below the detection limit (Hershey et al, 1991).…”

Section: Discussionmentioning

confidence: 99%

“…6 " . ' 5 ' I ' not at 0°C (Larsen et al, 1989b). After incubation of primary anterior pituitary cultures at 0°C with [12SI]-BH-SP, many cells showed binding of this radioligand when subjected to autoradiography.…”

Section: Methodsmentioning

confidence: 99%

“…Adult male Wistar rats were decapitated, the pituitaries removed, and the neurointermediate lobe separated from the anterior lobe. The anterior lobes were cut into about 10 smaller fragments and isolated pituitary cells were prepared by trypsinizing the organs according to the method of Gillies and Lowry (1978), as modified by Larsen et al (1989b). The dispersed cell sample was re-suspended in Dulbecco's modified Eagle's medium (DMEM) containing 10% fetal calf serum, 15 pg benzylpenicillinlml, and 25 pg streptomycinlml to a final concentration of 1 x lo6 cellslml.…”

Section: Methodsmentioning

confidence: 99%

“…The binding experiments were performed essentially as described earlier (Larsen et al, 1989b), with minor variations. To abolish internalization of the ligand and minimize the action of enzymatic degradation, pre-incubation and incubation with the radioactive ligand were performed at O' C. Primary anterior pituitary cell cultures were washed twice with Earl's balanced salt solution (EBSS) containing 0.2 % bovine serum albumin (BSA), followed by pre-incubation for 20 min in 0.5 ml EBSS containing 0.2% BSA and 3 m M MnC12 per well (Buffer A).…”

Section: Methodsmentioning

confidence: 99%

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Binding of an iodinated substance P analogue to cultured anterior pituitary prolactin- and luteinizing hormone-containing cells.

Larsen

Særmark²,

Mau³

1992

J Histochem Cytochem.

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Several lines of anatomic, biochemical, and pharmacological evidence suggest that the neuropeptide substance P has a direct action on cells of the anterior pituitary lobe via a specific neurokinin-1 receptor. In the present study we confiied this association by combining Bolton-Hunter iodinated substance Preceptor autoradiography with immunocytochemistry on cultured anterior pituitary cells. Radiolabeled substance P was bound to living cell cultures at O"C, and after a brief wash the cultures were fued and processed immunocytochemically for prolactin and luteinizing hormone. A large proportion of cultured anterior pituitary cells possessed substance P binding Sites. When receptor autoradiography was combined with immunocytochemistry, it

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Section: Discussionmentioning

confidence: 71%

Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

See 3 more Smart Citations

Binding of an iodinated substance P analogue to cultured anterior pituitary prolactin- and luteinizing hormone-containing cells.

Larsen

Særmark²,

Mau³

1992

J Histochem Cytochem.

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Distribution of substance P‐immunoreactive elements in the preoptic area and the hypothalamus of the rat

Larsen

1992

J of Comparative Neurology

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The localization and morphology of neurons, processes, and neuronal groups in the rat preoptic area and hypothalamus containing substance P-like immunoreactivity were studied with a highly selective antiserum raised against synthetic substance P. The antiserum was thoroughly characterized by immunoblotting; only substance P was recognized by the antiserum. Absorption of the antiserum with synthetic substance P abolished immunostaining while addition of other hypothalamic neuropeptides had no effect on the immunostaining. The specificity of the observed immunohistochemical staining pattern was further confirmed with a monoclonal substance P antiserum. The distribution of substance P immunoreactive perikarya was investigated in colchicine-treated animals, whereas the distribution of immunoreactive nerve fibers and terminals was described in brains from untreated animals. In colchicine-treated rats, immunoreactive cells were reliably detected throughout the preoptic area and the hypothalamus. In the preoptic region, labeled cells were found in the anteroventral periventricular and the anteroventral preoptic nuclei and the medial and lateral preoptic areas. Within the hypothalamus, immunoreactive cells were found in the suprachiasmatic, paraventricular, supraoptic, ventromedial, dorsomedial, supramammillary, and premammillary nuclei, the retrochiasmatic, medial hypothalamic, and lateral hypothalamic areas, and the tuber cinereum. The immunoreactive cell groups were usually continuous with adjacent cell groups. Because of the highly variable effect of the colchicine treatment, it was not possible to determine the actual number of immunoreactive cells. Mean soma size varied considerably from one cell group to another. Cells in the magnocellular subnuclei of the paraventricular and supraoptic nuclei were among the largest, with a diameter of about 25 microns, while cells in the supramammillary and suprachiasmatic nuclei were among the smallest, with a diameter of about 12 microns. Immunoreactive nerve fibers were found in all areas of the preoptic area and the hypothalamus. The morphology, size, density, and number of terminals varied considerably from region to region. Thus, some areas contained single immunoreactive fibers, while others were innervated with such a density that individual nerve fibers were hardly discernible. During the last decade, knowledge about neural organization of rodent hypothalamic areas and mammalian tachykinin biochemistry has increased substantially. In the light of these new insights, the present study gives comprehensive morphological evidence that substance P may be centrally involved in a wide variety of hypothalamic functions. Among these could be sexual behavior, pituitary hormone release, and water homeostasis.

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Internalization of [3H]Substance P Analogues in NK-1 Receptor Transfected CHO Cells

Sagan¹,

Lavielle²

2001

Biochemical and Biophysical Research Communications

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Binding and internalization of a iodinated substance P analog by cultured anterior pituitary cells

Cited by 27 publications

References 46 publications

Binding of an iodinated substance P analogue to cultured anterior pituitary prolactin- and luteinizing hormone-containing cells.

Binding of an iodinated substance P analogue to cultured anterior pituitary prolactin- and luteinizing hormone-containing cells.

Distribution of substance P‐immunoreactive elements in the preoptic area and the hypothalamus of the rat

Internalization of [3H]Substance P Analogues in NK-1 Receptor Transfected CHO Cells

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