Adrenomedullin (1-50), rat

Adrenomedullin receptor is found exclusively in noradrenaline-secreting cells of the rat adrenal medulla

Adrenomedullin, originally identified in the adrenal medulla, has binding sites in the adrenal gland; however, its role in the adrenal medulla is unclear. This study was designed to characterise adrenomedullin binding sites in the rat adrenal medulla, using ligand binding studies, immunocytochemistry, and mRNA analysis. A single population of specific adrenomedullin receptors was identified in adrenal medullary homogenates. 125I-Adrenomedullin was displaced only by adrenomedullin1-50 and not by calcitonin gene-related peptide or amylin at concentrations up to 100 nmol/L. The receptor K(D) was 3.64 nmol/L with a receptor density of 570 fmol/mg of protein. Analysis of mRNA revealed that the genes encoding both the putative adrenomedullin receptors, termed calcitonin receptor-like receptor (CRLR) and L1, were expressed in the rat adrenal medulla. Dual-colour indirect-labelled immunofluorescence was used to localise phenylethanolamine N-methyltransferase (PNMT) and the adrenomedullin receptor in the same section. PNMT is the enzyme that converts noradrenaline to adrenaline and is not expressed in noradrenaline-secreting cells. These studies revealed that both CRLR and L1 were expressed only in cells that did not express PNMT, suggesting that adrenomedullin receptors are only found in noradrenaline-secreting cells. Further evidence to support this conclusion was provided by the demonstration of colocalisation of adrenomedullin receptors with dopamine beta-hydroxylase, confirming the presence of the receptors in medullary chromaffin cells. Taken together, these data suggest that adrenomedullin acts through a specific adrenomedullin receptor in the rat adrenal medulla. RT-PCR and northern blot analysis revealed greater abundance of mRNA for L1 than for CRLR, possibly suggesting that L1 may be the major adrenomedullin receptor expressed in this tissue. As it has been reported that adrenomedullin is synthesised predominantly by adrenaline-secreting cells, it appears likely that adrenomedullin is a paracrine regulator in the adrenal medulla.

[Effect of adrenomedullin 1-50 on chronic hypoxic pulmonary hypertension in rats]

Objective: To explore the effect of adrenomedullin(1-50) (ADM(1-50)) on hypoxic pulmonary hypertension and pulmonary vascular structural remodeling and the plasma concentration of nitric oxide (NO) and hydrogen sulfide (H(2)S) in rats.
Methods: Twenty male Wistar rats were randomly divided into control group (n=7), hypoxic group (n=6) and hypoxic with ADM(1-50) group (n=7). ADM(1-50) was subcutaneously administered into rats of hypoxic with ADM(1-50) group by mini-osmotic pump (300 ng/h). After two weeks' hypoxic challenge, mean pulmonary arterial pressure (mPAP) was evaluated by using a right cardiac catheterization procedure. The ratio of right ventricular mass to left ventricular plus septal mass [RV/(LV+S)] was detected. Pulmonary vascular microstructure was measured and the ultrastructural changes in intra-acinar pulmonary arteries were observed. Meanwhile, plasma concentrations of NO and H(2)S were measured.
Results: mPAP was significantly increased in hypoxic rats than that in controls [(24.9+/-6.8) mmHg vs (14.3+/-2.4) mmHg, P<0.01,1 mmHg=0.133 kPa]; RV/(LV+S) was also significantly increased in hypoxic rats than that in controls [(0.318+/-0.054) vs (0.182+/-0.007), P<0.01]. Microstructure and ultrastructure of pulmonary arteries changed obviously in hypoxic rats with the development of hypoxic pulmonary vascular structural remodeling. Meanwhile, plasma NO and H(2)S concentrations in hypoxic rats were markedly decreased compared with controls. However, mPAP was significantly decreased in hypoxic rats treated with ADM(1-50) than that in hypoxic rats [(14.9+/-3.0) mmHg vs (24.9+/-6.8) mmHg, P<0.01]; RV/(LV+S) was also significantly decreased than that in hypoxic rats [(0.185+/-0.011) vs (0.318+/-0.054), P<0.01]. ADM(1-50) ameliorated pulmonary vascular structural remodeling of hypoxic rats in association with an increase in plasma NO and H(2)S concentrations.
Conclusion: ADM(1-50) plays an important role in regulation of the development of hypoxic pulmonary hypertension and hypoxic pulmonary vascular structural remodeling, through promoting NO and H(2)S production in hypoxic rats.

Intrathecal administration of adrenomedullin induces mechanical allodynia and neurochemical changes in spinal cord and DRG

This study investigated the effect of adrenomedullin (AM) on mechanical pain sensitivity and its possible mechanisms. Intrathecal injection of AM receptor agonist AM_1-50 (20 μg) once per day briefly reduced mechanical pain threshold on days 1 and 2 but induced prolonged mechanical allodynia on day 3. However, AM_1-50 did not change mechanical pain sensation when the AM receptor antagonist AM_22-52 (20 μg) was intrathecally co-administered. Daily administration of AM_1-50 (20 μg) for 3 days increased expression of phosphorylated extracellular signal-regulated protein kinase (pERK) and neuronal nitric oxide synthase (nNOS) in the spinal dorsal horn. The AM-induced increase in pERK and nNOS was inhibited by the co-administration of AM_22-52. The chronic administration of AM_1-50 also increased expression of microglial maker Iba1 and astrocytic marker GFAP (glial fibrillary acidic protein) in the spinal dorsal horn in an AM_22-52-sensitive manner. Furthermore, the application of AM_1-50 (10 nM, 3 h) to dorsal root ganglion (DRG) explant cultures induced an increase in the expression of transient receptor potential vanilloid 1 (TRPV1). The treatment with AM_1-50 did not change TRPV1 expression in DRG in the presence of AM_22-52 (2 μM). These results suggest that the increased AM bioactivity induced mechanical allodynia and may contribute to the mechanical pain hypersensitivity under pathological conditions. The mechanisms may involve the activation of ERK signaling pathway and spinal glia as well as the recruitment of nNOS and TRPV1 in the spinal dorsal horn or DRG. The present study indicates that inhibition of the activation AM receptor might provide a fruitful strategy to relieving chronic pain.

Adrenomedullin mediates tumor necrosis factor-α-induced responses in dorsal root ganglia in rats

Adrenomedullin (AM), a member of the calcitonin gene-related peptide (CGRP) family, has been demonstrated to be a pain peptide. This study investigated the possible involvement of AM in tumor necrosis factor-alpha (TNF-α)-induced responses contributing to neuronal plasticity in the dorsal root ganglia (DRG). Exposure of the DRG explant cultures to TNF-α (5nM) for 48h upregulated the expression of AM mRNA. The treatment with TNF-α also increased the level of CGRP, CCL-2 and MMP-9 mRNA in the cultured DRG. This increase was attenuated by the co-treatment with the selective AM receptor antagonist AM22-52 (2μM). The blockade of AM receptors inhibited TNF-α-induced increase of the glial fibrillary acidic protein (GFAP), interleukin-1β (IL-1β), phosphorylated cAMP response element binding protein (pCREB) and nuclear factor kappa B (pNF-κB) proteins. On the other hand, the treatment with the AM receptor agonist AM1-50 (10nM) for 96h induced an increase in the level of GFAP, IL-1β, pCREB and pNF-κB proteins. The inhibition of AM activity did not change TNF-α-induced phosphorylation of extracellular signal-related kinase (pERK) while the treatment with AM1-50 still increased the level of pERK in the cultured DRG. Immunofluorescence assay showed the colocalization of AM-like immunoreactivity (IR) with TNF-α-IR in DRG neurons. The present study suggests that the increased AM receptor signaling mediated the many, but not all, TNF-α-induced activities, contributing to peripheral sensitization in neuropathic pain.

Vasodilation and glomerular binding of adrenomedullin in rabbit kidney are not CGRP receptor mediated

The polypeptide adrenomedullin (ADM) was infused systemically to conscious rabbits to elucidate its actions on overall circulation and especially the renovascular bed and the formation and/or release of hormones important for body fluid homeostasis, including adrenocortical steroids. ADM lowered mean arterial pressure from 71.5 +/- 3.2 to 64.7 +/- 3.2 mmHg only at the highest dose of 25 pmol.min-1.kg-1 infused intravenously for 20 min and concomitantly induced tachycardia, possibly due to both baroreflex activation and direct cardiostimulatory effects. Renal blood flow (RBF) determined in rabbits chronically equipped with a perivascular ultrasonic flow probe increased from 55.4 +/- 2.1 to 67.4 +/- 2.7 and from 58.2 +/- 3.5 to 75.2 +/- 6.0 ml/min at ADM infusions of 5 and 25 pmol.min-1.kg-1, respectively. The elevation in RBF persisted even in the presence of the calcitonin gene-related peptide (CGRP1 receptor antagonist CGRP-(8-37). Of all osmoregulatory hormones tested, only corticosterone (Cort) plasma concentration increased in response to the highest ADM dose from 17.6 +/- 3.1 to 38.9 +/- 6.2 ng/ml, probably due to haroreflex activation. Subdepressor doses of ADM, however, caused a mild reduction in circulating Cort. Expression of functional high-affinity binding sites specific for ADM in vitro could be demonstrated for the renal artery and outer cortical glomeruli using 125I-labeled rat ADM as radioligand and determination of cellular adenosine 3',5'-cyclic monophosphate (cAMP) formation within the glomeruli. The ineffectiveness of CGRP-(8-37) to displace radiolabeled ADM from its binding sites, to inhibit ADM-induced glomerular cAMP formation, and to prevent ADM-induced renal vasodilation supports the hypothesis of ADM altering renal hemodynamics by interacting with ADM- and not CGRP-specific membrane receptors.