An activated renin–angiotensin system maintains normal blood pressure in aryl hydrocarbon receptor heterozygous mice but not in null mice
Graphical abstract
Introduction
The aryl hydrocarbon receptor (AHR) is a ligand-activated, basic helix-loop-helix/Per-ARNT-Sim transcription factor best known for mediating the toxicity of environmental pollutants, typified by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). More recently, studies of the ahr null mouse (ahr−/−) have established that the AHR has physiological functions in the absence of exogenous ligands and many of these functions relate to the vasculature. ahr−/− mice exhibit a significant reduction in liver weight, which results from the persistence of the fetal ductus venosus after birth [1]. Conditional deletion of ahr in a cell type-dependent manner demonstrates that AHR expression in endothelial/hematopoietic cells, but not hepatocytes, is necessary for normal postnatal closure of this fetal vascular structure [2]. The loss of ahr also has been shown to enhance new blood vessel development following skeletal muscle ischemia [3].
Given the effects of AHR deficiency on angiogenesis and vascular remodeling, others and we have studied blood pressure regulation in adult ahr−/− mice. ahr−/− mice exhibit increased plasma levels of two potent vasoconstrictors, angiotensin II (Ang II) and endothelin-1 (ET-1), and are hypertensive at modest altitude (1600 m), but hypotensive at sea level [4], [5], [6]. The hypertension at modest altitude is associated with hypoxemia and is normalized by exposure to sea level oxygen conditions [5]. Interestingly, after maintaining a colony of ahr−/− mice for multiple generations at modest altitude, our current studies reveal that the ahr−/− mice have adapted and now exhibit hypotension similar to that reported for ahr−/− mice at sea level. Nonetheless, since ahr−/− mice exhibit vascular defects during fetal development [1], it has been suggested that these developmental vascular changes might contribute to the disruption of cardiovascular homeostasis in adulthood [7].
Numerous studies have suggested that the intrauterine fetal environment can contribute to programming of blood pressure in adulthood, although most studies demonstrate hypertension in the adult offspring. Nonetheless, fetal programming of hypotension in adult offspring has been reported, including following prenatal dexamethasone exposure, placental insufficiency, and a shift from a protein to carbohydrate diet [8], [9]. To further evaluate the functional role of AHR in blood pressure regulation in adulthood and the potential contribution of abnormal fetal vascular development, we have conducted studies with mice lacking a single ahr allele, i.e. ahr heterozygous mice (ahr+/−). In contrast to ahr−/− mice, the liver size of ahr+/− mice is normal and the ductus venosus closes properly [10], [11], [12], [13]. In fact, no developmental phenotypic changes have been ascribed to the loss of a single ahr allele, suggesting that one copy of ahr is sufficient for normal development, including the vasculature.
In these studies we test the hypothesis that blood pressure regulation would be normal in ahr+/− mice in which fetal vascular abnormalities have not been reported, compared to ahr−/− mice in which numerous fetal vascular abnormalities have been described.
Section snippets
ET-1 receptor A (ETA) antagonist, PD155080
The orally active ETA antagonist, PD155080, was synthesized as described previously [14]. The sodium salt of PD155080 was dissolved in water, added to transgenic dough (Bio-Serv, Frenchtown, NJ) along with 0.1% bromophenol blue, and the mixture was folded together until the dye was evenly distributed throughout the dough. Pregelatinized corn starch was added to absorb excess water and reduce stickiness. The dough was then formed into 100 mg tablets using a pill mold (Gallipot, St. Paul, MN)
Functional AHR levels as measured by TCDD-inducible CYP1A1 mRNA expression
Since inducible expression of CYP1A1 mRNA is controlled by AHR, we used this as a sensitive, quantitative assessment of functional AHR levels. Neither constitutive nor TCDD-inducible cardiac CYP1A1 mRNA expression was detectable in ahr−/− mice (Fig. 1). While the constitutive expression of cardiac CYP1A1 mRNA was very low and not significantly different between ahr+/− and ahr+/+ mice, the TCDD-inducible cardiac CYP1A1 mRNA expression was significantly reduced in ahr+/− mice (3× fold lower),
Discussion
Although ahr+/− mice do not exhibit overt vascular abnormalities during development [12], [13] and are normotensive in adulthood, our studies show that blood pressure regulation in ahr+/− mice is different from both ahr−/− and ahr+/+ mice. Notably, ahr+/− mice exhibit an activated RAS and increased ETA signaling, which are responsible for maintaining their blood pressure at normotensive levels. In contrast, ahr−/− mice do not have an activated RAS and are significantly less responsive to ACE
Conflict of interest
None.
Acknowledgements
This study was supported by a grant from the National Institutes of Health [R01 HL078914 to M.K.W.] and a Pfizer summer internship program. The authors wish to thank Drs. Changjian Feng and Laura Gonzalez Bosc for their technical assistance, and critical analysis and interpretation of the data.
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Present address: Department of Internal Medicine, University of Michigan, Ann Arbor, MI, USA.