Molecular cloning and expression of a prostaglandin E<sub>2</sub> receptor of the EP<sub>3β</sub> subtype from rat hepatocytes

Neuschäfer-Rube, Frank; DeVries, Christa; Hänecke, Kristina; Jungermann, Kurt; Püschel, Gerhard

doi:10.1016/0014-5793(94)00837-x

Cited by 29 publications

(19 citation statements)

References 23 publications

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“…Two of the rat EP3 isoforms decrease intracellular cAMP concentration and one isoform increases intracellular Ca2+ concentration [29,30]. Breyer et al demonstrated that rabbit EP3 was distributed in chromaffin cells of the adrenal medullae but not in the cortex [31].…”

Section: Discussionmentioning

confidence: 99%

Expression of Prostaglandin E Receptor EP4 Subtype in Rat Adrenal Zona Glomerulosa. Involvement in Aldosterone Release.

et al. 2000

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Abstract.We examined the role of prostaglandin (PG) E receptors in the secretion of aldosterone. PGE2 is known to exert its various biological functions by binding to PGE receptors.There are four subtypes of PGE receptors, EP1, EP2, EP3, and EP4. Among the PGE receptors EP2 and EP4 subtypes are coupled to Gs protein and stimulate adenylyl cyclase. In this study, PGE2 caused a dose-dependent increase in aldosterone production from the rat adrenal zona glomerulosa cells in vitro accompanied with an increase in intracellular CAMP concentration.A specific agonist for EP2, butaprost, did not increase the CAMP production or the aldosterone release, suggesting the possibility that EP4 mediates the secretion of aldosterone by PGE2. Northern blot hybridization analysis disclosed that EP4 gene was expressed in the rat adrenal gland but that EP2 gene was not. In situ hybridization revealed that EP4 mRNA is present abundantly in the zona glomerulosa of rat adrenal gland. These findings suggest that the PGE2-EP4 system is involved in the regulation of aldosterone secretion from the rat adrenal gland.

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Section: Discussionmentioning

confidence: 99%

Expression of Prostaglandin E Receptor EP4 Subtype in Rat Adrenal Zona Glomerulosa. Involvement in Aldosterone Release.

et al. 2000

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“…The number of EP3 splice isoforms is species specific, with 3 identified in mouse (EP3α, EP3β and EP3γ) Sugimoto et al, 1993), 4 in rat (Neuschaferrube et al, 1994;Oldfield et al, 2001;Takeuchi et al, 1993) and 9 isoforms (out of which 8 are functional) in humans (Kotani et al, 1997). The EP receptors bind PGE2 with varying affinities.…”

Section: Pge2 Receptorsmentioning

confidence: 99%

Talking to the Brain at the Blood-Brain Barrier through Inflammation-Induced Prostaglandin E2

Vasilache¹

2015

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The immune-to-brain signaling is a critical survival factor when the body is confronted by pathogens, and in particular by microorganisms. During infections, the ability of the immune system to engage the central nervous system (CNS) in the management of the inflammatory response is just as important as its ability to mount a specific immune response against the pathogen, since the CNS can provide a systemic negative feed-back to the immune activation by release of stress hormones and also can prioritize the usage of the energy resources by the vital organs. Prostaglandin E2 (PGE2) and pro-inflammatory cytokines were among the first mediators to be identified to participate in the immune-to-brain signaling, a process that is clinically recognized by the development of manifestations of common illness such as fever, anorexia, decreased social interactions, lethargy, sleepiness, and hyperalgesia.In this thesis the contribution of PGE2 to the immune-to-brain signaling was further characterized at the blood-brain-barrier (BBB) and in the anterior preoptic area (POA) of the hypothalamus (i.e. the thermoregulatory region or, in sickness, the fever generating region).BBB is the major interface region between peripheral circulating cytokines and the neuronal parenchyma and a critical source of PGE2. Using chimeric mice lacking the inducible enzyme for PGE2 synthesis, microsomal PGE synthase-1 (mPGES-1), in either hematopoietic or nonhematopoietic cells, we demonstrate in paper I that brain endothelial cells are the critical source of PGE2 in BBB during peripheral inflammation. For the demonstration of the mPGES-1 expression in the BBB cells we developed in paper I a method for enzymatic dissociation of these cells, followed by fluorescence activated cell sorting (FACS). Using the same method, we show in paper II that the BBB response to immune stimuli is towards an increased production of PGE2 in endothelial cells and an increased sensitivity of these cells for proinflammatory cytokines. These changes are supported by decreased PGE2 degradation and decreased synthesis of other prostanoids in perivascular macrophages, which hence respond in concordance with the endothelial cells in enhancing PGE2 signaling.Once released in the neuronal tissue, PGE2 has been shown to be critical for the fever response by acting on the type 3 PGE2 receptors (EP3) within POA. By laser capture microdissection (LCM) we extracted the EP3 receptor expressing region in POA, defined by in situ hybridization histochemistry, from mouse brain sections. We demonstrate in paper III that the predominant subtypes of the EP3 receptor in POA are EP3α and EP3γ. In paper IV we further analyze the effect of PGE2 on the LCM dissected EP-rich POA using gene expression microarrays. We demonstrate that PGE2 has a limited effect on the gene expression changes within POA, suggesting that the neuronal activity is modulated by PGE2 in a transcriptionindependent manner and that the profound gene expression changes that are seen in the CNS during inflammation ...

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“…Cloning of the rEP 3 pR cDNA was carried out as described [7]. For cloning of the I1EP4R total RNA was isolated from human T-lym-phocytes by CsCl gradient centrifugation [10].…”

Section: Cloning Of Rat Ep 3 P and Human Ep^ Receptor Cdnasmentioning

confidence: 99%

“…Stable expression of the rEP 3 pR in CHO cells was carried out as described previously [7]. The 1.66 kb Notl cDNA fragment for the rEP 3 hEP 4 R was subcloned into the eukaryotic expression vector pRc/ CMV (Invitrogen).…”

Section: Stable Expression Of Rat Ep 3 P Receptor In Cho and Rep^heptmentioning

confidence: 99%

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The C‐terminal domain of the G_s‐coupled EP₄ receptor confers agonist‐dependent coupling control to G_i but no coupling to G_s in a receptor hybrid with the G_i‐coupled EP₃ receptor

et al. 1997

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Prostaglandin E 2 receptors (EPR) belong to the family of G-protein-coupled receptors with 7 transmembrane domains. They form a family of four subtypes, which are linked to different G-proteins. EPiR are coupled to G q , EP 2 and EP 4 R to G s and EP 3 R to G;. Different C-terminal splice variants of the bovine EP 3 R are coupled to different G-proteins. A mouse EP 3 R whose C-terminal domain had been partially truncated no longer showed agonist-induced Gj -protein activation and was constitutively active. In order to test the hypothesis that the C-terminal domain confers coupling specificity of the receptors on the respective G-proteins, a cDNA for a hybrid rEP 3 hEP 4 R, containing the N-terminal main portion of the Gj-coupled rat EP 3 ßR including the 7th transmembrane domain and the intracellular C-terminal domain of the G s -coupled human EP 4 R, was generated by PCR. HEK293 cells transiently transfected with the chimeric rEP 3 hEP4R cDNA expressed a plasma membrane PGE 2 binding site with a slightly lower K d value for PGE 2 but an identical binding profile for receptorspecific ligands as cells transfected with the native rat EP 3 ßR. In HepG 2 cells stably transfected with the chimeric rEP 3 hEP 4 R cDNA PGE 2 did not increase cAMP formation characteristic of G s coupling but attenuated the forskolin-stimulated cAMP synthesis characteristic of Gj coupling. This effect was inhibited by pre-treatment of the cells with pertussis toxin. Thus, the hybrid receptor behaved both in binding and in functional coupling characteristics as the native rat EP 3ß R. Apparently, the intracellular C-terminal domain did not confer coupling specificity but coupling control, i.e. allowed a signalling state of the receptor only with agonist binding.

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Molecular cloning and expression of a prostaglandin E₂ receptor of the EP_3β subtype from rat hepatocytes

Cited by 29 publications

References 23 publications

Expression of Prostaglandin E Receptor EP4 Subtype in Rat Adrenal Zona Glomerulosa. Involvement in Aldosterone Release.

Expression of Prostaglandin E Receptor EP4 Subtype in Rat Adrenal Zona Glomerulosa. Involvement in Aldosterone Release.

Talking to the Brain at the Blood-Brain Barrier through Inflammation-Induced Prostaglandin E2

The C‐terminal domain of the G_s‐coupled EP₄ receptor confers agonist‐dependent coupling control to G_i but no coupling to G_s in a receptor hybrid with the G_i‐coupled EP₃ receptor

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Molecular cloning and expression of a prostaglandin E2 receptor of the EP3β subtype from rat hepatocytes

Cited by 29 publications

References 23 publications

Expression of Prostaglandin E Receptor EP4 Subtype in Rat Adrenal Zona Glomerulosa. Involvement in Aldosterone Release.

Expression of Prostaglandin E Receptor EP4 Subtype in Rat Adrenal Zona Glomerulosa. Involvement in Aldosterone Release.

Talking to the Brain at the Blood-Brain Barrier through Inflammation-Induced Prostaglandin E2

The C‐terminal domain of the Gs‐coupled EP4 receptor confers agonist‐dependent coupling control to Gi but no coupling to Gs in a receptor hybrid with the Gi‐coupled EP3 receptor

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Molecular cloning and expression of a prostaglandin E₂ receptor of the EP_3β subtype from rat hepatocytes

The C‐terminal domain of the G_s‐coupled EP₄ receptor confers agonist‐dependent coupling control to G_i but no coupling to G_s in a receptor hybrid with the G_i‐coupled EP₃ receptor