Alveolar Epithelial Type I Cells Express ß2-Adrenergic Receptors and G-protein Receptor Kinase 2
Will Rogers Institute Pulmonary Research Center, Division of Pulmonary and Critical Care Medicine, University of Southern California, Los Angeles, California
Correspondence to: Janice M. Liebler, MD, Div. of Pulmonary and Critical Care Medicine, U. of Southern California, IRD 620, 2020 Zonal Avenue, Los Angeles, CA 90033. E-mail: liebler{at}usc.edu
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Summary |
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Key Words: alveolar epithelium ß2-adrenergic receptor G protein-coupled receptor kinase 2
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Introduction |
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Catecholamines and agents that are selective for the ß2-adrenergic receptor (ß2AR) stimulate transepithelial Na+ transport in the lung under normal and pathological conditions (Charron et al. 1999; Frank et al. 2000
; Saldias et al. 2000
). Increases in alveolar fluid clearance are associated with high endogenous plasma catecholamine levels in such diverse conditions as septic shock in rats (Pittet et al. 1994
) and neurogenic pulmonary edema in dogs (Lane et al. 1998
). Exogenously-administered ß adrenergic agonists have similarly led to accelerated clearance of edema fluid in rats with increased left atrial pressure (Azzam et al. 2001
; Frank et al. 2000
), after hyperoxic exposure (Saldias et al. 1999
), and after ventilator-associated lung injury (Saldias et al. 2000
). Adenovirus-mediated overexpression of ß2AR in normal rats increased clearance of alveolar fluid when measured in an isolated lung model (Dumasius et al. 2001
). Recently, investigators (Sartori et al. 2002a
) demonstrated that inhalation of a ß2-adrenergic agonist, salmeterol, attenuated development of high-altitude pulmonary edema in human subjects predisposed to that condition. Therefore, there is considerable experimental evidence to suggest that ß2-adrenergic agonists may be useful in accelerating clearance of pulmonary edema in human patients.
Despite these promising results, a potential concern is that continued stimulation of ß2ARs by endogenous catecholamines or by drugs that stimulate ß2ARs might lead to decreased responsiveness to the beneficial effects of these agents over time (Liggett 1997). ß2AR signaling occurs via agonist binding to the receptor, which then interacts with a membrane-bound G-protein. Desensitization, defined as a decline in cell response despite the continued presence of a stimulus of constant intensity, is a feature of G-protein-coupled receptors (GPCRs), such as ß2AR. Decreased responsiveness of GPCRs can occur by several mechanisms, including receptor phosphorylation (mediated by cAMP-dependent protein kinase A or by non-cAMP-dependent G-protein-coupled receptor kinases; GRKs), sequestration or internalization of receptor, or decreasing receptor number (Liggett 1999
; Ruiz-Gomez and Mayor 1997
). GRKs are of particular interest because their expression is altered in certain disease states, such as congestive heart failure (Lefkowitz et al. 2000
), hypothyroidism (Penela et al. 2001
), and cystic fibrosis (Mak et al. 2002
). An important GRK for desensitization of ß2ARs appears to be G-protein receptor kinase 2 (GRK2), also called ß-adrenergic receptor kinase 1 (ßARK1) (Liggett 1997
; Ruiz-Gomez and Mayor 1997
). It is possible that GRK2 plays an active role in regulation of responsiveness of alveolar epithelium to ß2-adrenergic agents over time.
AT2 cells have been reported to express ß2AR (Fabisiak et al. 1987). Given the large surface area covered by AT1 cells and their potential contribution to alveolar Na+ and water removal, we were interested in learning if AT1 cells also express ß2ARs. Here we used immunofluorescence (IF) techniques in isolated AT1 and AT2 cells, in alveolar epithelial cells (AECs) in primary culture, and in whole lung to localize ß2AR and GRK2 within the alveolar epithelium of normal rat lung. We further evaluated expression of ß2ARs and GRK2 in freshly isolated AT1 and AT2 cells, compared with AT2 cells in primary culture during transdifferentiation towards the AT1 cell phenotype.
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Materials and Methods |
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To obtain freshly isolated preparations partially enriched for AT1 cells, lungs from adult male SpragueDawley rats (300 g) were digested with 8 U/ml elastase and 10 mg/ml collagenase type 1 (Worthington), chopped, and filtered through 100 µM strainers. Approximately 20% of the cells obtained in this fashion are AT1 cells, as determined by immunoreactivity with the mouse monoclonal Ab (MAb) VIIIB2, a marker for rat AT1 cells (Danto et al. 1992) (see below). Cytocentrifuged preparations were processed for IF after fixation in 100% cold methanol for 10 min.
Processing of Lung Tissue and Cultured Cells for IF
Normal rat lung tissue was inflated and fixed in 4% paraformaldehyde. After embedding in paraffin, samples were cut into 4-µm sections and placed on glass slides. AT2 cells were grown on polycarbonate filters. At day 5, cells on filters were fixed in 4% paraformaldehyde, washed in Tris-buffered saline (TBS, pH 7.5), and prepared for IF studies as described below.
Antibodies
The anti-ß2-adrenergic receptor (ß2AR) Ab is an affinity purified rabbit polyclonal Ab developed against a peptide from the carboxy terminus of mouse ß2AR (Santa Cruz Biotechnology; Santa Cruz, CA). The manufacturer reports that this Ab reacts with ß2ARs and ß3ARs of mouse and rat origin but does not react with ß1ARs. ß3AR has not been previously localized to lung tissue (Evans et al. 1996; Mak et al. 1996
; Sartori et al. 2002b
). VIIIB2 is a mouse MAb that recognizes an epitope in the apical membrane of AT1 cells (Danto et al. 1992
). Anti-GRK2 (= ßARK1) Ab is a rabbit polyclonal Ab raised against a peptide from the carboxy terminus of human GRK2 that crossreacts with rat GRK2 (Santa Cruz). Normal rabbit IgG (Vector Laboratories; Burlingame, CA) was used a control for primary rabbit Abs at the same concentrations as the primary Abs. For Western blotting analysis, a mouse MAb to GRK2 was used (Santa Cruz). This MAb was raised against a recombinant peptide of human GRK2 origin and specifically reacts with rat GRK2. Polyclonal goat anti-surfactant protein C (SP-C) Ab (Research Diagnostics; Flanders, NJ) was used as an AT2 cell marker.
Immunofluorescence
Whole Lung
After deparaffinization and rehydration through graded alcohols, lung tissue sections on slides underwent microwave antigen retrieval (Antigen Unmasking Solution; Vector). Slides were then sequentially double labeled with goat anti-SP-C (1:75) and either rabbit anti-ß2AR (1:300) or anti-GRK2 (1:300) Abs, or with normal rabbit IgG as a negative control. Anti-SP-C Ab was amplified using an avidinbiotin system with fluorescein isothiocyanate (FITC; Vector). After an avidinbiotin blocking step, anti-ß2AR Ab or anti-GRK2 Ab was amplified using an avidinbiotin system with Texas Red (Vector). Tissue sections were treated with Vectashield Mounting Medium (Vector) containing 4',6 diamidino-2-phenylindole (DAPI), which counterstains nuclei blue.
AECs in Primary Culture
Paraformaldehyde-fixed AECs grown on polycarbonate filters were washed in TBS with 0.05% Tween-20 (TBS-T), treated with CAS block (Zymed Laboratories, South San Francisco, CA), incubated with rabbit anti-ß2AR Ab, anti-GRK2 Ab, or normal rabbit IgG overnight at 4C, and washed in TBS-T. Signal was amplified using biotinylated anti-rabbit secondary Ab (Vector), washed in TBS-T, and labeled with avidin-linked FITC (Vector). Cells on filters were treated with Vectashield containing propidium iodide, which counterstains nuclei red, and mounted on glass slides.
Freshly Isolated AECs
Methanol-fixed cytocentrifuged cells on slides were blocked in PBS, pH 7.2, with 3% BSA overnight, incubated with primary Abs for 1 hr at room temperature, rinsed in PBS, incubated with secondary Ab linked to either rhodamine (red) or FITC (green), and rinsed in PBS. Cells were identified by double labeling with AT1 or AT2 cell markers and Abs to ß2AR or GRK2. To minimize crossreactivity of primary Abs, Ab combinations were chosen so that both Abs were taken from different animal species. Slides were treated with Vectashield containing DAPI.
Slides were viewed using an Olympus BX60 microscope equipped with epifluorescence optics. Images were captured separately using a cooled CCD camera (Magnafire; Olympus, Lake Success, NY) with filters for DAPI, FITC, or rhodamine/Texas Red. The images were merged and imported into Adobe Photoshop (Adobe Systems; Mountain View, CA) as TIFF files. Where indicated, slides were also viewed with a confocal scanning laser microscope (Nikon Eclipse, TE 300; Melville, NY) equipped with an argon blue laser (excites at 488 nm) and an HeNe green laser (excites at 543 nm). Confocal images were processed with a Nikon PCM2000 laser scanner.
RNA Isolation and Northern Analysis
Total RNA was isolated from cultured AECs by the acid phenolguanidiniumchloroform method (Chomczynski and Sacchi 1987). Equal amounts of RNA (5 or 10 µg) were denatured with formaldehyde, size-fractionated by agarose gel electrophoresis under denaturing conditions, and transferred to nylon membranes. RNA was immobilized by UV crosslinking. Hybridization was performed in hybridization buffer for 16 hr at 65C. Blots were probed with a 1.8-kb cDNA probe for rat ß2AR (from American Type Culture Collection; Manassas, VA) labeled with [
32P]-dCTP (Roche Molecular Biochemicals; Indianapolis, IN) or biotin (Kirkegaard & Perry Laboratories; Gaithersburg, MD) using the random primer method. Blots were washed at high stringency (0.5x SSC-0.1% SDS at 55C). Differences in RNA loading were normalized using a 24-mer oligonucleotide probe for 18S rRNA end-labeled with [32P]-ATP. Signal intensity was quantified using a Storm Phosphorimager (Bio-Rad; Hercules, CA).
Western Analysis
Freshly isolated AT2 cells and AECs in culture on days 68 were solubilized in 2% SDS sample buffer. GRK2 expression was similar in AT1-like cells at days 6 or greater in culture, so results were pooled for analysis. Protein concentrations were determined with the Bio-Rad DC Protein Assay kit. Commercially available control sample for GRK2 (Ramos cell lysate; Santa Cruz) was loaded with the AEC samples. Equal amounts of protein in sample buffer were resolved by SDS-PAGE and transferred to Immobilon-P membranes (Millipore; Marlborough, MA). Membranes were blocked in 5% non-fat dry milk in TBS-T overnight before incubation with the mouse anti-GRK2 Ab. After washing, membranes were incubated with anti-mouse IgG Ab conjugated to horseradish peroxidase (Promega; Madison, WI). Complexes were visualized using enhanced chemiluminescence (ECL; Amersham, Arlington Heights, IL). Signal intensity was quantified using the Fluorchem Imaging System (Alpha Innotech; San Leandro, CA).
Statistical Analysis
Results are expressed as mean ± SEM. Significance of differences (p<0.05) was determined by Students's t-test or, where multiple time points were compared, by one-way ANOVA with the TukeyKramer adjustment for multiple comparisons.
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Results |
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To further characterize ß2AR expression in AECs, peripheral rat lung cells were enzymatically dispersed and isolated cells cytocentrifuged onto glass slides. Cytocentrifuged cell preparations were concurrently labeled with anti-ß2AR Ab and a primary Ab specific for AT1 cells, VIIIB2. AT1 cells, identified by reactivity to VIIIB2 (Figure 1IIIA, red), were immunoreactive to anti-ß2AR Ab (Figure 1IIIB, green). Other cells in the crude lung cell mixture also reacted with the ß2AR antibody, including small cells that appear morphologically to be AT2 cells. No reactivity was seen when control rabbit IgG was substituted for rabbit anti-ß2AR Ab in double-labeling experiments (Figure 1IIIC and IIID). Cell nuclei were identified using DAPI (blue).
ß2AR Expression in AECs in Primary Culture
As shown in this representative Northern blot, freshly isolated AT2 cells and AEC grown on polycarbonate filters for one and eight days express ß2AR mRNA (Figure 2A)
. Densitometric analysis of three separate preparations (Figure 2B) demonstrated a 40% increase in ß2AR mRNA from day 0 to day 8 (p<0.05) when corrected for levels of 18S rRNA. These findings indicate increased expression of ß2AR mRNA as cells transdifferentiate from the AT2 to the AT1 cell phenotype.
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Discussion |
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Our laboratory and others have shown that active Na+ transport across cultured AECs is enhanced by exposure to catecholamines and agents that are selective for ß2ARs (Kim et al. 1991; Minakata et al. 1998
; Bertorello et al. 1999
). In experimental lung models, increased Na+ transport after stimulation of ß2ARs has resulted in enhanced clearance of pulmonary edema fluid (Saldias et al. 1999
,2000
; Frank et al. 2000
; Azzam et al. 2001
). Several mechanisms have been proposed to account for these effects of ß2-adrenergic agonists, including an increase in phosphorylation of the Na+,K+-ATPase
-subunits and quantity of the subunit at the basolateral membrane, enhanced Na+ channel open probability, and migration of epithelial Na+ channel (ENaC) components to the apical membrane (Crandall and Matthay 2001
). Subacutely, increases in Na+,K+-ATPase
-subunit mRNA and protein and
ENaC mRNA expression have been described (Minakata et al. 1998
). Indirect enhancement of Na+ absorption after stimulation of ß2ARs by promoting apical chloride conductance has also been suggested (Jiang et al. 2001
). AT1 cells, by virtue of the fact that their long cytoplasmic processes cover most of the gas exchange surface of the lung, are likely candidates to respond to ß2-adrenergic agonists by stimulating Na+ and water clearance. This study demonstrates that AT1 cells possess the receptor for ß2-adrenergic agonists.
Previous attempts to localize ß2AR in lung used radioligand receptor labeling or in situ hybridization (Carstairs et al. 1985; Hamid et al. 1991
; Mak et al. 1996
). Those studies localized ß2AR activity and mRNA to alveolar walls but were unable to precisely localize it to AT1 vs AT2 cells. Only recently have satisfactory Abs for immunolocalization of ß2AR become readily available (Boivin et al. 2001
) that have been used to localize ß2AR in other types of cells. In rat kidney, ß2AR was localized to apical and subapical areas of proximal and distal renal tubule epithelial cells, with only faint immunoreactivity seen on basolateral membranes (Boivin et al. 2001
). Furthermore, localization of ß2AR was found to correspond to radioligand binding, indicating that the receptor had intact ß2AR activity. In human lung fibroblasts and vascular smooth muscle cells, strong ß2AR reactivity was found primarily at the plasma membrane but also occurred in the cytoplasm (Ruiz-Gomez and Mayor 1997
). It was speculated that the non-membrane-associated ß2AR might reflect cytoplasmic recirculation of the receptor (Ruiz-Gomez and Mayor 1997
). Consistent with these previous studies, we found that ß2AR immunoreactivity was most intense on the AEC membranes but was also scattered throughout the cytoplasm of the cells.
ß2AR signaling occurs via agonist binding to the receptor, which then interacts with the membrane-bound G-protein (Liggett 1997). One feature of G-protein-coupled receptors is the ability to modulate function in response to differing conditions (Liggett 1997
). Phosphorylation of G-protein-coupled receptors in general and of ß2ARs in particular constitutes a major mechanism for desensitization of these receptors. GRKs are a family of at least six members that differ in tissue distribution, substrate interactions, and regulation (Liggett 1997
). The most important GRK that mediates desensitization of ß2AR is believed to be GRK2 (McGraw and Liggett 1997
). McGraw and Liggett (1997)
found significant lung cell-type variation in expression of GRK2 in cultured mast cells, bronchial epithelial cells, and airway smooth muscle cells. They further found a direct relationship between GRK2 expression and the degree of short-term desensitization of these cells to continued agonist exposure. Therefore, even within a single organ such as the lung, cells may experience different degrees of desensitization to ß2-adrenergic agents. In this study we demonstrate similar expression of GRK2 in freshly isolated AT2 cells and in AT1-like cells in culture, suggesting that ß2AR in both cell types may be influenced by GRK2 in a similar fashion.
Previous investigators have looked for evidence of desensitization of the response to ß2-adrenergic agents in vivo and in vitro (Nishikawa et al. 1994; Charron et al. 1999
; Kelsen et al. 2000
; Morgan et al. 2002
). Studies of prolonged exposure to ß-adrenergic agonists in rats showed that enhanced clearance of alveolar fluid persisted over 4 hr (Charron et al. 1999
), while 48 hr of exposure resulted in a decline in the ability of a ß2-adrenergic agonist to stimulate alveolar fluid clearance (Morgan et al. 2002
). After 7 days of an inhaled ß2-adrenergic agonist, airway epithelial cells harvested from human subjects showed desensitization to this agonist, with an increase in immunoreactivity to GRK2 (Kelsen et al. 2000
). These studies suggest that the relative responsiveness of ß2ARs of AT1 or AT2 cell origin may diminish over time and may be mediated, at least in part, by GRK2.
In summary, ß2AR was immunolocalized to AT1 and AT2 cells in normal rat lung. We further found mRNA for ß2AR in AECs in primary culture that had undergone transdifferentiation towards an AT1 cell-like phenotype. These findings suggest a role for AT1 cells in the increase in alveolar Na+ and fluid clearance observed after exposure to catecholamines or more selective ß2- adrenergic agents. Both AT1 and AT2 cells in whole lung express immunoreactive GRK2. The effect of GRK2 on relative responsiveness of different cells in alveolar epithelium to continued ß2-adrenergic agonists awaits further study. We speculate that the net effect of ß2-adrenergic agonists on clearance of pulmonary edema fluid will depend on the balance among the numbers of ß2ARs available on each cell type, the numbers of each cell type present, and the relative impact of ß2AR regulators, such as GRK2, on each cell type. Unknown at this point is the degree to which this dynamic interplay of cellsreceptorsdesensitizers occurs in the setting of lung injury, when the usual relationship between AT1 and AT2 cells is disrupted and hyperplastic AT2 cells may be the predominant type of epithelial cell present. Characterization of these features in AT1 and AT2 cells under normal conditions and in lung injury will lead to a better understanding of the potential role of ß2-adrenergic agonists in the management of pulmonary edema.
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Acknowledgments |
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We note with appreciation the excellent technical support of Zerlinde Balverde and Susie Parra. The Microscopy Sub core at the USC Center for Liver Diseases (NIH 1 P30 DK48522) provided the confocal microscope used for these studies. Dr Crandall is Hastings Professor and Norris Chair of Medicine.
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Footnotes |
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Literature Cited |
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Azzam ZS, Saldias FJ, Comellas A, Ridge KM, Rutschman DH, Sznajder JI (2001) Catecholamines increase lung edema clearance in rats with increased left atrial pressure. J Appl Physiol 90:10881094
Bertorello AM, Ridge KM, Chibalin AV, Katz AI, Sznajder JI (1999) Isoproterenol increases Na+-K+-ATPase activity by membrane insertion of -subunits in lung alveolar cells. Am J Physiol 276:L2027[Medline]
Boivin V, Jahns R, Gambaryan S, Ness W, Boege F, Lohse MJ (2001) Immunofluorescent imaging of ß1- and ß2-adrenergic receptors in rat kidney. Kidney Int 59:515531[CrossRef][Medline]
Borok Z, Danto SI, Zabski SM, Crandall ED (1994) Defined medium for primary culture de novo of adult rat alveolar epithelial cells. In Vitro Cell Dev Biol Anim 30A:99104[Medline]
Borok Z, Liebler JM, Lubman RL, Foster MJ, Zhou B, Li X, Zabski SM, et al. (2002) Na transport proteins are expressed by rat alveolar epithelial type I cells. Am J Physiol 282:L599608
Carstairs JR, Nimmo AJ, Barnes PJ (1985) Autoradiographic visualization of beta-adrenoceptor subtypes in human lung. Am Rev Respir Dis 132:541547[Medline]
Charron PD, Fawley JP, Maron MB (1999) Effect of epinephrine on alveolar liquid clearance in the rat. J Appl Physiol 87:611618
Cheek JM, Evans MJ, Crandall ED (1989) Type I cell-like morphology in tight alveolar epithelial monolayers. Exp Cell Res 184:375387[Medline]
Chomczynski P, Sacchi N (1987) Single step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem 162:156159[CrossRef][Medline]
Crandall ED, Matthay MA (2001) Alveolar epithelial transport. Am J Respir Crit Care Med 162:10211029
Danto SI, Zabski SM, Crandall ED (1992) Reactivity of alveolar epithelial cells in primary culture with type I cell monoclonal antibodies. Am J Respir Cell Mol Biol 6:296306[Medline]
Dobbs LG (1990) Isolation and culture of alveolar type II cells. Am J Physiol 258:L134147[Medline]
Dumasius V, Sznajder JI, Azzam ZS, Boja J, Mutlu GM, Maron MB, Factor P (2001) ß2-Adrenergic receptor overexpression increases alveolar fluid clearance and responsiveness to endogenous catecholamines in rats. Circ Res 89:907914
Evans BA, Papaioannou M, Bonazzi VR, Summers RJ (1996) Expression of beta 3-adrenoreceptor mRNA in rat tissues. Br J Pharmacol 117:210216[Abstract]
Fabisiak JP, Vesell ES, Rannels DE (1987) Interactions of beta adrenergic antagonists with isolated rat alveolar type II pneumoncytes. Analysis, characterization and regulation of specific beta adrenergic receptors. J Pharmacol Exp Ther 241:722727[Abstract]
Frank JA, Wang Y, Osorio O, Matthay MA (2000) ß-Adrenergic agonist therapy accelerates the resolution of hydrostatic pulmonary edema in sheep and rats. J Appl Physiol 89:12551265
Hamid QA, Mak JCW, Sheppard MN, Corrin B, Venter JC, Barnes PJ (1991) Localization of ß2-adrenoreceptor messenger RNA in human and rat lung tissue using in situ hybridization: Correlation with receptor autoradiography. Eur J Pharmacol 206:133138[CrossRef][Medline]
Jiang X, Ingbar DH, O'Grady SM (2001) Adrenergic regulation of ion transport across adult alveolar epithelial cells: effects on Cl channel activation and transport function in cultures with an apical air interface. J Membr Biol 181:195204[Medline]
Johnson MD, Widdicombe JH, Allen L, Barbry P, Dobbs LG (2002) Alveolar epithelial type I cells contain transport proteins and transport sodium, supporting an active role for type I cells in regulation of lung liquid homeostasis. Proc Natl Acad Sci USA 99:19661971
Kelsen SG, Aksoy MO, Brennan K, Ciccolella D, Borbely B (2000) Chronic effects of inhaled albuterol on ß-adrenergic system function in human respiratory cells. J Asthma 37:361370[Medline]
Kim KJ, Cheek JM, Crandall ED (1991) Contribution of active Na+ and Cl fluxes to net ion transport by alveolar epithelium. Respir Physiol 85:245256[CrossRef][Medline]
Lane SM, Maender KC, Awender NE, Maron MB (1998) Adrenal epinephrine increases alveolar liquid clearance in a canine model of neurogenic pulmonary edema. Am J Respir Crit Care Med 158:760768
Lefkowitz RJ, Rockman HA, Koch WJ (2000) Catecholamines, cardiac ß-adrenergic receptors, and heart failure. Circulation 101:16341637
Liggett SB (1997) Molecular basis of G protein-coupled receptor signaling. In Crystal RG, West JB, Weibel ER, Barnes PJ, eds. The Lung: Scientific Foundations 2nd ed. Philadelphia, LippincottRaven, 1936
Liggett SB (1999) Molecular and genetic basis of ß2-adrenergic receptor function. J Allergy Clin Immunol 104:S4246[Medline]
Mak JCW, Chuang T-T, Harris CA, Barnes PJ (2002) Increased expression of G protein-coupled receptor kinases in cystic fibrosis lungs. Eur J Pharmacol 436:165172[CrossRef][Medline]
Mak JCW, Nishikawa M, Haddad E-B, Kwon OJ, Hirst SJ, Twort CHC, Barnes PJ (1996) Localization and expression of ß-adrenoceptor subtype mRNAs in human lung. Eur J Pharmacol 302:215221[CrossRef][Medline]
McGraw DW, Liggett SB (1997) Heterogeneity in ß-adrenergic receptor kinase expression in the lung accounts for cell-specific desensitization of the ß2-adrenergic receptor. J Biol Chem 272:73387344
Minakata Y, Suzuki S, Grygorczyk C, Berthiaume Y (1998) Impact of ß-adrenergic agonist on Na+ channel and Na+-K+-ATPase expression in alveolar type II Cells. Am J Physiol 275:L414422[Medline]
Morgan EE, Hodnichak CM, Stader SM, Maender KC, Boja JW, Folkesson HG, Maron MB (2002) Prolonged isoproterenol infusion impairs the ability of ß2-agonists to increase alveolar liquid clearance. Am J Physiol 282:L666674
Nishikawa M, Mak JCW, Shirasaki H, Harding SE, Barnes PJ (1994) Long-term exposure to norepinephrine results in down-regulation and reduced mRNA expression of pulmonary ß-adrenergic receptors in guinea pigs. Am J Respir Cell Mol Biol 10:9199[Abstract]
Penela P, Barradas M, Alvarez-Dolado M, Muñoz A, Mayor F Jr (2001) Effect of hypothyroidism on G protein-coupled receptor kinase 2 expression levels in rat liver, lung, and heart. Endocrinology 142:987991
Pittet JF, Weiner-Kronish JP, McElroy MC, Folkesson HG, Matthay MA (1994) Stimulation of lung epithelial liquid clearance by endogenous release of catecholamines in septic shock in anesthetized rats. J Clin Invest 94:663671[Medline]
Ruiz-Gomez A, Mayor F Jr (1997) ß-Adrenergic receptor kinase (GRK2) colocalizes with ß-adrenergic receptors during agonist-induced receptor internalization. J Biol Chem 272:96019604
Saldias FJ, Comellas A, Ridge KM, Lecuona E, Sznajder JI (1999) Isoproterenol improves ability of lung to clear edema in rats exposed to hyperoxia. J Appl Physiol 86:3035
Saldias FJ, Lecuona E, Comellas AP, Ridge KM, Rutschman DH, Sznajder JI (2000) ß-Adrenergic stimulation restores rat lung ability to clear edema in ventilator-associated lung injury. Am J Respir Crit Care Med 162:282287
Sartori C, Allemann Y, Duplain H, Lepori M, Egli M, Lipp E, Hutter D, et al. (2002a) Salmeterol for the prevention of high-altitude pulmonary edema. N Engl J Med 346:16311636
Sartori C, Fang X, McGraw DW, Koch P, Snider ME, Folkesson HG, Matthay MA (2002b) Selected contribution: long-term effects of ß2-adrenergic receptor stimulation on alveolar fluid clearance in mice. J Appl Physiol 93:18751880