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Title: Characterization of beta1-adrenergic receptor expression in the rat anterior pituitary
Other Titles: Karakterisatie van de expressie van beta1-adrenerge receptoren in de hypofysesvoorkwab van de rat
Authors: Janssens, Kristel
Issue Date: 10-Nov-2007
Abstract: The two major effectors of the stress-response system are the hypothalamic-pituitary-adrenal axis and the autonomous nervous system that upon activation result in increased glucocorticoid and (nor)epinephrine production which are the driving forces for the so-called fight-or-flight reactions. In the present study we focussed on the action of these two signalling systems in the rat anterior pituitary and report the expression of the ß1-adrenergic receptor (ß1-AR) and the glucocorticoid-induced receptor (GIR), two G-protein-coupled receptors that are target genes of glucocorticoids. Messenger RNA of both receptors was maintained at low level in cell aggregate cultures and ß1-AR mRNA was strongly up-regulated in a dose- and time-dependent manner by dexamethasone (DEX). The response characteristics were different from that of the ß2-AR. Glucocorticoids also prevented down-regulation of ß1-AR mRNA levels. The ß1-AR protein, as detected by immunoblot, was up-regulated by DEX as well in cultured aggregates. DEX-dependent up-regulation of ß1-AR was found in a separated subpopulation consisting of small-sized cells enriched in lactotrophs and folliculo-stellate cells but not in a population containing large gonadotrophs and somatotrophs.
As examined by immunofluorescence confocal microscopy, ß1-AR-immunoreactivity (-ir) was detected in gonadotrophs but not in somatotrophs, lactotrophs, corticotrophs, thyrotrophs or folliculo-stellate cells. Remarkably, ß1-AR-ir cells were often surrounded by cup-shaped lactotrophs. The selective localization in gonadotrophs was confirmed by quantitative RT-PCR and immunoblot analysis on mRNA- and protein-extracts of anterior pituitary cell lines showing that ß1-AR is more abundantly expressed in αT3-1 (gonadotroph precursor) and LßT2 (LH gonadotroph) cells compared to GHFT (somatotroph precursor), GH3 (lactosomatotroph) and TtT/GF (folliculo-stellate) cells. All these cell lines, except GH3 cells, expressed ß2-AR mRNA as well, but without large differences in expression level.
The ß-AR agonist isoproterenol stimulated cAMP levels in pituitary aggregate cell cultures, GHFT, αT3-1, LßT2 and in TtT/GF cells but not in GH3 cells. As studied with the selective ß1-AR antagonists CGP 20712A and ICI 89,406 derivative 8a and the selective ß2-AR antagonist ICI 118,551, isoproterenol-induced cAMP levels were not mediated by ß1-ARs in GHFT and TtT/GF cells whereas in aggregates and in both gonadotroph cell lines, i.e. αT3-1 and LßT2, the ß1- and ß2-AR antagonists each blocked the response. Responses were glucocorticoid-independent in the hormonal cell lines but ß1-AR-mediated cAMP stimulation was DEX-dependent in TtT/GF cells. This response was only seen with very high doses of norepinephrine and not after ß1-AR blockade of isoproterenol-stimulated cAMP accumulation with CGP 20712A. Taken these data together we propose that the ß1-AR is expressed in a subpopulation of gonadotrophs with a special topographic relationship to lactotrophs in a DEX-independent way but that the DEX-dependent regulation of ß1-AR mRNA is likely not located in a specific cell type but is the consequence of paracrine interactions with non-hormonal cells (possibly folliculo-stellate cells).
In cultured anterior pituitary cell aggregates, but not in cell lines, CGP 20712A inhibits basal cAMP levels, an effect not seen with other ß-AR antagonists such as propranolol, betaxolol, ICI 89,406 derivative 5a or carvedilol which was described as a neutral ß1-AR antagonist. The effect was blocked by carvedilol and by pre-treatment with pertussis toxin (PTx), suggesting that ß1-ARs in the anterior pituitary are constitutively active and coupled to the adenylate cyclase system in a PTx-sensitive manner, on which CGP 20712A would act as an inverse agonist. Depletion of stored norepinephrine from secretory vesicles by pre-treatment of aggregates with reserpine blocked the inhibition of basal cAMP levels by CGP 20712A as well and caused an inhibition of basal cAMP accumulation by itself. Therefore, (part of) the effect of CGP 20712A might be based on interruption of an endogenous adrenergic agent tonically elevating basal cAMP accumulation in an autocrine/paracrine loop.
The present work has revealed the expression, cellular localization and activity of the ß1-AR in the rat pituitary and has shown a glucocorticoid-dependent drive that is lost in pituitary cell lines. This ß1-AR system may be involved in the complex paracrine network acting in this tissue.
Publication status: published
KU Leuven publication type: TH
Appears in Collections:Pharmacology Section (-)
Animal Physiology and Neurobiology Section - miscellaneous

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