摘要:

cDNA encoding the thyrotropin-releasing hormone receptor (TRH-R) was recently cloned in rat pituitary prolactin cells and in mouse thyrotropes. The molecular weights of the protein sequences obtained are 46.6 and 44.5 kD. However, TRH-R has not yet been purified to homogeneity and specific anti-TRH-R antibody could not yet be obtained by classical biochemical methods. We thus attempted to obtain antibodies specific for TRH-R using an anti-idiotypic approach. Rabbits of the same allotype were immunized using Igs (Ab1) extracted from rabbit polyclonal anti-TRH immune serum. Anti-idiotypic rabbit polyclonal anti-anti-TRH antibodies (Ab2) were obtained, as shown by their ability to inhibit the formation of TRH-anti-TRH complexes in a radioimmunoassay system. One of them, the poyclonal Ab2 R38/B12, was tested for its ability to recognize the TRH-R in rat pituitary, tumor-derived, GH3/B6 prolactin-secreting cells. Immunoreactive material was immunocytochemically detected in fixed and saponin-permeabilized GH3/B6 cells. The immunostaining was localized at the plasma membrane and on intracellular structures. It was not observed using non-anti-TRH Ab2 and was abolished in the presence of excess TRH. Furthermore, binding of [125I]R38/B12 on fixed and saponin-permeabilized GH3/B6 cells was partially inhibited by excess TRH. By immunoblot analyses of Triton X-l 14 cell extracts performed under reducing or nonreducing conditions, the polyclonal R38/B12 Igs revealed two main protein species of ≈98 and ≈76 kD as well as several proteins ≤46 kD. In the presence of excess TRH, the ≈98- and ≈42-kD bands were abolished, whereas the intensity of the other bands was faintly attenuated only. The ≈98-kD protein was also revealed in a two-dimensional PAGE analysis. Nevertheless, the effects of R38/B12 Igs on [3H]TRH binding by GH3/B6 cells and on basal or TRH-induced prolactin secretion were not markedly different from those elicited by control Ab2. These data suggest that we have characterized Ab2 antibodies which recognize a molecular entity that might be related to the TRH-R in

ISSN:0028-3835    

DOI:10.1159/000126696

出版商:S. Karger AG    

年代:1994

数据来源: Karger

摘要:

The mammalian anterior pituitary is known to be regulated by humoral factors only, no direct neural influence having ever been discovered. Our recent studies have shown in several species that there are substantial amounts of substance P and calcitonin-gene-related peptide-like immunoreactive (CGRP-LI) nerve fibers in the anterior pituitary and that they can make synaptic contacts with secretory cells, among which corticotropes. In the present study, we investigated changes in CGRP-LI nerve fibers of the anterior pituitary after bilateral adrenalectomy in the rat. The results show that the number of CGRP-LI nerve fibers increases significantly. In parallel, they cover a wider area of the gland after the operation, suggesting a direct neural influence.

ISSN:0028-3835    

DOI:10.1159/000126697

出版商:S. Karger AG    

年代:1994

数据来源: Karger

ISSN:0028-3835    

DOI:10.1159/000126698

出版商:S. Karger AG    

年代:1994

数据来源: Karger