Adrenomedullin expression and function in the rat carotid body

Abstract

Adrenomedullin (AM) immunoreactivity has been found in granules of the glomus (type I) cells of the carotid bodies in rats. The identity of these cells was ascertained by colocalization of immunoreactivities for AM and tyrosine hydroxylase in their cytoplasm. Exposure of freshly isolated carotid bodies to synthetic AM resulted in a concentration-and time-dependent degranulation of glomus cells as measured by dopamine (DA) release. DA release reached a zenith 30 min after exposure to AM (94·2% over untreated controls). At this time-point, the response to AM was similar to the one elicited by 5 min of exposure to 100 mM K + . Nevertheless, injection of 1 µl 60 nM AM/g body weight into the tail vein of the rats did not induce statistical differences in DA release from the carotid bodies. Exposure of the oxygen-sensitive cell line PC-12 to hypoxia elicited an increase in AM mRNA expression and peptide secretion into serum-free conditioned medium. Previous data have shown that elevation of AM expression under hypoxia is mediated through hypoxia-inducible factor-1, and that exposure of chromaffin cells to AM results in degranulation. All these data suggest that AM is an important autocrine regulator of carotid body function.

Sustained Exposure to Cytokines and Hypoxia Enhances Excitability of Oxygen-Sensitive Type I Cells in Rat Carotid Body: Correlation with the Expression of HIF-1α Protein and Adrenomedullin

“…Moreover, acute application of the former cytokine has been shown to inhibit K + -current and elevate [Ca 2 + ] I in type I cells (Shu et al, 2007). The present study extends these important findings by showing for the first time that sustained exposure to inflammatory cytokines increases levels of HIF-1a and the HIF-1-regulated peptide, ADM, suggesting that CH-induced inflammation in carotid body contributes to phenotypic adjustments in type I cells that are associated with chemoreceptor adaptation to hypoxia (Kline et al, 2002a;Martinez et al, 2003). Moreover, our data show that cytokine treatment elevates Ca 2 + -responses evoked by subsequent acute hypoxic challenges.…”

“…The increased presence of ADM following exposure to CH in vivo, as well as sustained hypoxia and cytokines in culture, provides further evidence that chemoreceptor adaptation involves highly specified HIF-1-directed phenotypic adjustments in type I cells, including the expression of known hypoxia-sensitive genes. Immunocytochemical experiments have previously localized ADM in these cells, and exposure of whole carotid bodies to the peptide elicits a dose-dependent release of dopamine (Martinez et al, 2003). CH is known to increase expression of TH, the rate-limiting enzyme for dopamine synthesis in carotid body ( Gonzalez et al, 1979;Fidone and Gonzalez, 1982;Fidone and Gonzalez, 1982).…”

Proadrenomedullin NH2-Terminal 20 Peptide Is a Potent Angiogenic Factor, and Its Inhibition Results in Reduction of Tumor Growth

“…Synthetic human adrenomedullin, PAMP, and PAMP (12)(13)(14)(15)(16)(17)(18)(19)(20) were purchased from Bachem (King of Prussia, PA). Recombinant human VEGF and basic fibroblast growth factor (bFGF) were obtained from R&D Systems (Minneapolis, MN).…”

“…Analysis and quantitation of angiogenesis was done using directed in vivo angiogenesis assay as described previously (10,16). In brief, 10-mm-long surgical-grade silicone tubes with only one end open (angioreactors) were filled with 20 L of Matrigel alone or mixed with adrenomedullin, bFGF, VEGF, PAMP, and/or PAMP (12)(13)(14)(15)(16)(17)(18)(19)(20) at the indicated concentrations. Human lung cancer cell lines (see below) were also premixed with Matrigel alone or in combination with PAMP (12)(13)(14)(15)(16)(17)(18)(19)(20) at 10,000 cells per angioreactor.…”

“…Total RNA was extracted using the RNeasy Mini kit from Qiagen (Valencia, CA) and reverse transcribed using the SuperScript First-Strand Synthesis system (Invitrogen). Quantification of gene expression was performed by real-time PCR as described previously (19). The PCR reaction was run in an Opticon cycler (MJ Research, Waltham, MA) using Sybr Green PCR master mix (Applied Biosystems, Foster City, CA).…”