Identification and characterization of glucose transport proteins in plasma membrane- and Golgi vesicle-enriched fractions prepared from lactating rat mammary gland.

Previous studies have indicated that turkey erythrocyte and rat liver membranes contain endogenous alpha beta heterodimeric insulin receptors in addition to the disulphide-linked alpha 2 beta 2 heterotetrameric complexes characteristic of most cell types. We utilized 125I-insulin affinity cross-linking to examine the structural properties of insulin receptors from rat liver and turkey erythrocyte membranes prepared in the absence and presence of sulphydryl alkylating agents. Rat liver membranes prepared in the absence of sulphydryl alkylating agents displayed specific labelling of Mr 400,000 and 200,000 bands, corresponding to the alpha 2 beta 2 heterotetrameric and alpha beta heterodimeric insulin receptor complexes respectively. In contrast, affinity cross-linking of membranes prepared with iodoacetamide (IAN) or N-ethylmaleimide identified predominantly the alpha 2 beta 2 heterotetrameric insulin receptor complex. Similarly, affinity cross-linking and solubilization of intact turkey erythrocytes in the presence of IAN resulted in exclusive labelling of the alpha 2 beta 2 heterotetrameric insulin receptor complex, whereas in the absence of IAN both alpha 2 beta 2 and alpha beta species were observed. Turkey erythrocyte alpha 2 beta 2 heterotetrameric insulin receptors from IAN-protected membranes displayed a 3-4-fold stimulation of beta subunit autophosphorylation and substrate phosphorylation by insulin, equivalent to that observed in intact human placenta insulin receptors. Turkey erythrocyte alpha beta heterodimeric insulin receptors, prepared by defined pH/dithiothreitol treatment of IAN-protected membranes, were also fully competent in insulin-stimulated protein kinase activity compared with alpha beta heterodimeric human placenta receptors. In contrast, endogenous turkey erythrocyte alpha beta heterodimeric insulin receptors displayed basal protein kinase activity which was insulin-insensitive. These data indicate that native turkey erythrocyte and rat liver insulin receptors are structurally and functionally similar to alpha 2 beta 2 heterotetrameric human placenta insulin receptors. The alpha beta heterodimeric insulin receptors previously identified in these tissues most likely resulted from disulphide bond reduction and denaturation of the alpha 2 beta 2 holoreceptor complexes during membrane preparation.     

Evidence for imidazoline binding sites in basolateral membranes from rabbit kidney

[3H]-RX 781094 and [3H]-rauwolscine, two potent alpha 2-adrenergic antagonists, were used to characterize alpha 2 receptor in basolateral membranes from rabbit kidney. However, the following findings suggest that the imidazoline [3H]-RX 781094 binds to an heterogeneous population of binding sites: 1) dissociation plot was biphasic with a fast and slow component, 2) in saturation experiments, [3H]-RX 781094 labels 3.5 more binding sites than [3H]-rauwolscine (p less than 0.02), 3) competition studies showed that molecules with imidazoline structure completely inhibited the [3H] RX 781094 binding; in contrast, only 25% of binding was affected by non-imidazoline alpha 2 adrenergic compounds. These results suggest that in basolateral membranes from rabbit kidney, [3H] RX781094 labels alpha 2 adrenergic and non-adrenergic receptors which might be imidazoline-preferring binding sites.     

Identification of the Subunit Structure of Rat Pineal Adrenergic Receptors by Photoaffinity Labeling

The adrenergic receptors of rat pineal gland were investigated using radiolabeled ligand binding and photoaffinity labeling techniques. 125I-2-[beta-(4-hydroxyphenyl)ethylaminomethyl]tetralone (125I-HEAT) and 125I-cyanopindolol (125I-CYP) labeled specific sites on rat pineal gland membranes with equilibrium dissociation constants (KD) of 48 (+/- 5) pM and 30 (+/- 5) pM, respectively. Binding site maxima were 481 (+/- 63) and 1,020 (+/- 85) fmol/mg protein. The sites labeled by 125I-HEAT had the pharmacological characteristics of alpha 1-adrenergic receptors. 125I-CYP-labeled beta-adrenergic receptors were characterized as a homogeneous population of beta 1-adrenergic receptors. The alpha 1- and beta 1-adrenergic receptors were covalently labeled with the specific photoaffinity probes 4-amino-6,7-dimethoxy-2-(4-[5-(4-azido-3-[125I]iodophenyl) pentanoyl]-1-piperazinyl) quinazoline (125I-APDQ) and 125I-p-azidobenzylcarazolol (125I-pABC). 125I-APDQ labeled an alpha 1-adrenergic receptor peptide of Mr = 74,000 (+/- 4,000), which was similar to peptides labeled in rat cerebral cortex, liver, and spleen. 125I-pABC labeled a single beta 1-adrenergic receptor peptide with a Mr = 42,000 (+/- 1,500), which differed from the 60-65,000 peptide commonly seen in mammalian tissues. Possible reasons for these differences are discussed.     

Identification of alpha-2 adrenergic receptors in rabbit adipocytes

Adrenaline, an alpha and beta adrenergic agonist don't modify theophylline induced lipolysis in perirenal and epididymal adipose tissue of the rabbit. Clonidine an alpha 2 adrenergic agonist inhibits theophylline stimulated lipolysis in the two tissue indicating the existence of alpha 2 adrenergic responsiveness. Direct identification of these receptors by radioligand binding studies shows that (H3) yohimbine an alpha 2 adrenergic antagonist binds to rabbit's fat cell membranes with high affinity: KD = 1.7 +/- 0.1 nmoles. The maximal number of binding sites at saturation is low 16 +/- 29 fmoles/mg protein.     

Calcium dependent aggregation of marine sponge cells is provoked by leukotriene B4 and inhibited by inhibitors of arachidonic acid oxidation

The mechanism of agonist-induced desensitization of the beta adrenergic receptor coupled adenylate cyclase has been studied in a smooth muscle cell line, BC3H-1, which expresses both alpha and beta adrenergic receptors and nicotinic receptors. beta receptors have been investigated in intact cells using as radioligand 3HCGP-12177, an hydrophilic compound which labels only surface receptors. The treatment of BC3H-1 cells with the agonist Isoproterenol, at 37 degrees but not at 4 degrees, induced a dose dependent internalization of the beta adrenergic receptor. Agonist-induced internalization was very rapid, in the order of few minutes. beta adrenergic receptor internalization was very specific: the alpha adrenergic agonist Phenylefrine had almost no effect on beta receptor levels, while Isoproterenol treatment had no effect on the number of alpha adrenergic or nicotinic receptors expressed at the cell surface of these cells. beta adrenergic receptor internalization is probably the major mechanism responsible for catecholamine desensitization in smooth muscle cells.     

Effects of androgen on alpha- and beta-adrenergic receptors in membranes from the rat seminal vesicle.

The effects of castration and androgen-replacement on adrenergic receptors in membranes from the rat seminal vesicle were studied. Membranes from seminal vesicles showed saturable and high-affinity binding sites for the beta-adrenergic receptor antagonist, [3H]dihydroalprenolol ([3H]DHA), and the alpha 1-adrenergic receptor antagonist, [3H]prazosin. Castration markedly reduced beta-adrenergic receptors with decreasing the effect of GTP modulating the receptor-ligand affinity, suggesting defects in both the receptor per se and the guanine-nucleotides-regulating mechanism after castration. In contrast, castration increased alpha 1-adrenergic receptors and androgen-replacement reversed this change. The effects of GTP decreasing the alpha 1-receptor binding affinity to the radioligand were observed to a similar extent in the castrated and control membranes. These results demonstrate an inverse regulation by androgen on beta- and alpha 1-adrenergic receptors in membranes of the rat seminal vesicle.     

Glutaraldehyde pretreatment blocks phospholipase A2 modulation of adrenergic receptors

Treatment of rat cerebral cortical membranes with phospholipase A2 affects, in a parallel fashion, beta-, alpha 1- and alpha 2-adrenergic receptor binding, but not the affinity of these receptors for their respective ligands. Pretreatment of membranes with 0.1 percent glutaraldehyde blocks the effects of phospholipase A2 on adrenergic receptor binding. The results support the hypothesis that desensitization or "masking" of adrenergic receptors may involve changes in membrane lipid composition. Furthermore, glutaraldehyde may prove a useful tool in the investigation of the dynamic roles of lipids in receptor function and more specifically, their regulation and coupling to physiological events.     

Somatostatin: a metabolic regulator

Somatostatin, the hypothalamic release-inhibiting factor, has been found to stimulate gluconeogenesis in rat kidney cortical slices. Stimulation by somatostatin was linear and dose-dependent. Other bioactive peptides such as cholecystokinin, gastrointestinal peptide, secretin, neurotensin, vasoactive intestinal peptide, pancreatic polypeptide, beta endorphin and substance P did not affect the renal gluconeogenic activity. Somatostatin-induced gluconeogenesis was blocked by phentolamine (alpha adrenergic antagonist) and prazosin (alpha1 adrenergic antagonist) but not by propranolol (beta adrenergic antagonist) and yohimbine (alpha2 adrenergic antagonist) suggesting that the effect is via alpha1 adrenergic stimuli. Studies on the involvement of Ca2+ revealed that tissue depletion and omission of Ca2+ from the reaction mixture would abolish the stimulatory effect of somatostatin. Furthermore, somatostatin enhanced the uptake of 45calcium in renal cortical slices which could be blocked by lanthanum, an inhibitor of Ca2+ influx. It is proposed that the stimulatory effect of somatostatin on renal gluconeogenesis is mediated by alpha1 adrenergic receptors, or those which functionally resemble alpha1 receptors and that the increased influx of Ca2+ may be the causative factor for carrying out the stimulus.     

Epidermal growth factor binding, stimulation of phosphorylation, and inhibition of gluconeogenesis in rat proximal tubule

Epidermal growth factor and insulin share many biological activities, including stimulation of cell proliferation, ion flux, glycolysis, fatty acid and glycogen synthesis, and activation of receptor-linked tyrosine kinase activity. In the kidney, insulin has been shown to regulate transport processes and inhibit gluconeogenesis in the proximal tubule. Since the kidney represents a major source of EGF, the present studies investigated whether proximal tubule contained EGF receptors, whether EGF receptors were localized to apical or basolateral membranes, and whether EGF receptor activation participated in the regulation of an important proximal tubule function, gluconeogenesis. Specific EGF receptors were demonstrated in the basolateral membrane of proximal tubule. Following incubation with 125I EGF, basolateral membranes demonstrated equilibrium binding at 4 degrees C and 23 degrees C. There was 78 +/- 2% specific binding (n = 13). The dissociation constant (Kd) was 1.5 x 10(-9) M and maximal binding was 44 fmol/mg protein. There was ninefold more specific binding to proximal tubule basolateral membrane than to brush border membrane. In basolateral, but not brush border membranes, EGF induced phosphorylation of the tyrosine residues of intrinsic membrane proteins, including a 170 kDa protein, corresponding to the EGF receptor. In the presence of the gluconeogenic substrates, alanine, lactate, and succinate, proximal tubule suspensions synthesized glucose. EGF inhibited glucose production in a concentration-dependent manner over a concentration range of 3 x 10(-11) to 3 x 10(-9) M. In addition, EGF inhibited angiotensin II-stimulated glucose production in the proximal tubule suspensions. EGF did not significantly increase net glucose metabolism nor decrease cellular ATP concentrations. Therefore, these studies demonstrated that rat proximal tubule contained specific receptors for EGF that were localized to the basolateral membrane and linked to tyrosine kinase activity. EGF significantly inhibited proximal tubule glucose production without significantly increasing net glucose consumption.     

Insulin stimulates proteolysis of the alpha-subunit, but not the beta-subunit, of its receptor at the cell surface in rat liver.

Insulin receptors in rat liver plasma membranes contain two alpha- and two beta-subunits held together by interchain disulphide bonds ([alpha beta]2 receptors). Affinity-labelled receptors were digested with chymotrypsin or elastase and then exposed to dithiothreitol before solubilization from membranes and SDS/polyacrylamide-gel electrophoresis. This resulted in partial reduction and isolation of Mr-225,000 alpha beta, Mr-200,000 alpha 1 beta, Mr-165,000 alpha beta 1 and Mr-145,000 alpha 1 beta 1 receptor halves containing intact (alpha, beta) or degraded (alpha 1, beta 1) subunits. The ability to identify half-receptor complexes containing intact or degraded subunits made it possible to assay each subunit simultaneously for insulin-induced proteolysis in isolated plasma membranes or during perfusion of rat liver in situ with insulin. In liver membranes, insulin binding increased the fraction of receptors containing degraded alpha-subunits to about one-third of the total population during 2 h of incubation at 23 degrees C. beta-Subunit proteolysis increased only minimally during this time. Plasma membranes isolated from livers perfused with insulin at 37 degrees C contained degraded alpha-subunits but only intact beta-subunits, showing that insulin induced cell-surface proteolysis of the binding, but not the kinase, domain of its receptor. Since previous observations [Lipson, Kolhatkar & Donner (1988) J. Biol. Chem 263, 10495-10501] have shown that receptors containing degraded alpha-subunits are internalized but do not recycle, it is possible that cell-surface degradation may play a role in the regulation of insulin-receptor number in hepatic tissue. Proteolysis of the beta-subunit is not a likely mechanism by which receptor-kinase activity may be attenuated under physiological conditions.     

Regulation of alpha- and beta-adrenergic receptor subclasses by gonadal steroids in human myometrium

Both alpha- and beta-adrenergic receptors have been identified in the human myometrium by radioligand binding. Both adrenergic receptor subclasses have been shown to mediate the contractile response of the uterus upon catecholamine stimulation: alpha-adrenergic receptors cause uterine contraction while beta-adrenergic receptors induce relaxation. We have identified alpha 1- and alpha 2-adrenergic receptors in myometrial membranes using the newly developed radiolabelled specific antagonists [3H]-prazosin and [3H]-rauwolscine. This enabled us to characterize both receptor subclasses individually. Beta adrenergic receptors were identified using the radiolabelled antagonist (-)-[3H]-dihydroalprenolol. Binding of radioligands to the myometrial membrane receptors was rapid, readily reversible, of high affinity and stereoselective. The total number of alpha 1-, alpha 2- and beta-receptors was determined by Scatchard analysis of radioligand saturation binding and the beta/beta 2-receptor ratio was determined by computer analysis of the beta 2-selective antagonist ICI 118 551) (-)-[3H]-dihydroalprenolol competition binding curves. This enabled us to study the regulation of both alpha- and beta-receptor subclasses under various physiological and pharmacological conditions in the human, i.e., during different phases of the menstrual cycle, in postmenopausal women and during depo-progestin (Medroxyprogesterone acetate) therapy. Only the alpha 2- and beta 1-adrenergic receptor concentrations were found to be subjected to gonadal steroid regulation. The number of alpha 2- and beta 1-adrenergic receptors increased concomitantly with circulating plasma oestradiol levels. This effect was counteracted by progesterone. The number of alpha 1- and beta 2-adrenergic receptors was unaffected by the gonadal steroid environment. These results are an example of the heteroregulation of membrane receptors by oestrogens and progesterone and cast new light on the regulatory mechanisms involved in uterine contractility in the human.     

Comparison of rat and human left ventricle beta-adrenergic receptors: subtype heterogeneity delineated by direct radioligand binding

Beta adrenergic receptors were identified in rat myocardial left ventricle and human papillary muscle by using the antagonist radioligand 3H-dihydroalprenolol. The number (37.3 and 44.5 fmol/mg of protein, respectively in rat and man), and the KD (1.6 and 2.8 nM, respectively in rat and man) of beta receptors were not significantly different. Adrenergic receptors of both beta 1 and beta 2 subtypes were found to coexist in the left ventricle. The relative proportions of the two beta receptor subtypes were determined by the use of competition radioligand selective binding and computer modelling techniques employing the subtype selective antagonists ICI 118,551 (beta 2 selective) and atenolol (beta 1 selective) in rat or metoprolol (beta 1 selective) in man. The rat left ventricle contained about 74% beta 1 and 26% beta 2 adrenergic receptors, human left ventricle papillary muscles contained about 69% beta 1 and 31% beta 2. Human and rat left ventricles contain both beta 1 and beta 2 adrenergic receptors with similar affinities. Rat might be a model for the study of human myocardial beta adrenergic receptors.     

Genomic organization of adrenergic and serotonin receptors in the mouse: linkage mapping of sequence-related genes provides a method for examining mammalian chromosome evolution.

Five sequence-related genes encoding four adrenergic receptors and a serotonin receptor were localized to specific regions of four mouse chromosomes with respect to 11 other genetic markers. Linkage was established by the analysis of the haplotypes of 114 interspecific backcross mice. Adra2r (alpha 2-C10) and Adrb1r (beta 1) receptors mapped to the distal region of mouse chromosome 19. These genes were separated by 2.6 +/- 1.5 cM in a segment of mouse chromosome 19 that has a similar organization of these genes on the long arm of human chromosome 10. The Adra1r (alpha 1B), Adrb2r (beta 2), and Htra1 (5HT1A) genes mapped to proximal mouse chromosome 11, proximal mouse chromosome 18, and distal mouse chromosome 13, respectively. The organization of genes linked to these loci on regions of the three mouse chromosomes is consistent with the organization of homologous human genes on human chromosome 5. These findings further define the relationship of linkage groups conserved during the evolution of the mouse and human genomes. We have identified a region that may have been translocated during evolution and suggest that the human genomic organization of adrenergic receptors more closely resembles that of a putative primordial ancestor.     

Adrenergic receptor homologies in vertebrate and invertebrate species examined by DNA hybridization

The deduced protein sequences of the mammalian adrenergic receptors (ARs) suggest that these proteins have evolved by several ancient gene duplication events. To investigate in what species these events may have occurred DNA fragments encoding the family of adrenergic receptors from human (beta 1AR and alpha 2AR) and hamster (beta 2AR and alpha 1AR) were used to detect homologous sequences in other vertebrates, invertebrates and unicellular organisms by Southern blot hybridization analysis. Sequences homologous to hamster beta 2AR were detected in lower vertebrates, invertebrates and Dictyostelium, but not in yeast or bacteria. Within vertebrates, sequences strongly homologous to human beta 1AR and human platelet alpha 2AR were confined to the higher vertebrates only. In the invertebrates, only Drosophila contained sequences homologous to hamster alpha 1AR. Our results suggest that non-mammalian species may contain receptors homologous to the mammalian adrenergic receptors and that the sequences homologous to human beta 2AR have been the most strongly conserved.     

Cloning and expression of the mouse homolog of the human alpha 2-C2 adrenergic receptor.

Three subtypes of alpha 2 adrenergic receptors have been identified in the human and rat. The subtype located on human chromosome 2 (alpha 2-C2) is unique in that it is expressed mainly in the peripheral tissues and lacks sites for N-linked glycosylation. We isolated the gene encoding the mouse homolog of the human alpha 2-C2 adrenergic receptor (M alpha 2-2H). The deduced amino acid sequence of the M alpha 2-2H shows 82% and 96% identity to the human alpha 2-C2 and the rat RNG alpha 2 adrenergic receptors, respectively. Southern blot analysis demonstrated that the M alpha 2-2H was encoded by a single copy gene and was distinct from the mouse homologs of the alpha 2-C4 and alpha 2-C10 adrenergic receptors. When expressed in COS-7 cells, the M alpha 2-2H exhibited a pharmacological profile similar to the human alpha 2-C2 and rat RNG alpha 2 receptors.     

Hormone action at the membrane level. VIII. Adrenergic receptors in rat heart and adipocytes and their modulation by thyroxine.

The regulation of adrenergic receptors in rat heart was measured in rats made hyperthyroid by injection with thyroxine and made hypothyroid by addition of propylthiouracil to the drinking water. Hyperthyroid rats display cardiac hypertrophy and a decrease in epididymal fat pad weight. The maximal beta-receptor level of ventricular membranes, as determined by (-)-[3H]dihydroalprenolol binding, was increased 60% by thyroxine treatment and decreased about 30% by propylthiouracil treatment. The affinity of the beta receptor was unchanged after thyroxine or propylthiouracil treatment. The maximal activity of the isoproterenol-stimulated adenylate cyclase (EC 4.6.1.1) varied with thyroid state in a manner parallel to the increase in beta-adrenergic binding sites. Thyroxine treatment also increases by 2-fold the beta receptors in isolated rat fat cells. Propylthiouracil treatment lowered the level of alpha receptors in heart by 30% as measured by [3H]dihydroergocryptine binding, but increased the affinity about 2.5-fold. The highest level of alpha receptors was seen in control hearts. These studies indicate that thyroxine may control the turnover of beta-adrenergic receptors in heart and fat cells and regulate physiological responses in these tissues via a hormone-hormone interplay system. Thyroxine treatment reduced the activity of the membrane-bound Mg2+-ATPase (EC 3.6.1.3) and 5'-mononucleotidase (EC 3.1.3.5) but appears to increase the activity of the (Na+ + K+)ATPase (EC 3.6.1.4).     

Serotonergic and noradrenergic receptors in the rat brain: modulation by chronic exposure to ovarian hormones

Noradrenergic (alpha 1 and beta) and serotonergic (5HT1 and 5HT2) receptors were assayed in the brains of ovariectomized female rats treated for 2 weeks with estrogen, progesterone or a combination of both hormones. Estrogen treatment resulted in a decrease in the number of 5HT1 and beta adrenergic receptors, with a concomitant increase in 5HT2 receptors. Progesterone alone caused a smaller increase in 5HT2 receptors, a similar decrease in 5HT1 and had no significant effect on noradrenergic receptors. When given with estrogen, progesterone blocked the estrogen effect on 5HT2 receptors but did not inhibit the estrogen-mediated decrease in 5HT1 and beta adrenergic receptors. alpha 1 adrenergic receptors were not affected by any of the hormone treatment paradigms. beta adrenergic and 5HT2 receptors are often implicated in antidepressant action, and the modulation of these two receptor types by ovarian hormones might be relevant to hormone-linked affective changes such as premenstrual tension and post-partum depression.     

GTP-binding proteins in luminal and basolateral membranes from pars convoluta and pars recta of rabbit kidney proximal tubule.

The GTP-binding proteins on luminal and basolateral membrane vesicles from outer cortex (pars convoluta) and outer medulla (pars recta) of rabbit proximal tubule have been examined. The membrane vesicles were highly purified, as ascertained by electron microscopy, by measurements of marker enzymes, and by investigating segmental-specific transport systems. The [35S]GTP gamma S binding to vesicles, and to sodium cholate-extracted proteins from vesicles, indicated that the total content of GTP-binding proteins were equally distributed on pars convoluta, pars recta luminal and basolateral membranes. The membranes were ADP-ribosylated with [32P]NAD+ in the presence of pertussis toxin and cholera toxin. Gel electrophoresis revealed, for all preparations, the presence of cholera toxin [32P]ADP-ribosylated 42 and 45 kDa G alpha s proteins, and pertussis toxin [32P]ADP-ribosylated 41 kDa G alpha i1, 40 kDa G alpha i2 and 41 kDa G alpha i3 proteins. The 2D electrophoresis indicated that Go's were not present in luminal nor in basolateral membranes of pars convoluta or pars recta of rabbit proximal tubule.     

Adrenergic control of coronary arteries

This review of adrenergic control of coronary arteries is based on studies conducted on isolated vessels. Both alpha- and beta-adrenergic receptors are present in the coronary vasculature but with different distributions. The large coronary arteries have a larger percentage of alpha receptors, which mediate contraction, whereas the small coronary arteries are equipped almost exclusively with beta receptors, which mediate relaxation. The beta receptors are of the subclass beta 1. Both alpha 1 and alpha 2 receptors are present in the large coronaries. The alpha 2 receptors are prejunctional and inhibit neuronal release of norepinephrine (NE), and are also postjunctional and mediate contraction of the vascular smooth muscle. Evidence is reviewed suggesting that coronary alpha and beta receptors may be different conformational states of the same parent macromolecule in which the alpha state predominates at lower temperatures. Contrary to these results we have observed that whereas the response of the large coronary artery to NE at 39 C is predominantly constriction, it is predominantly dilation at 29 C. The characteristic of the adrenergic receptor is clearly temperature dependent.     

alpha 1- and beta 2-adrenergic receptor expression in the Madin-Darby canine kidney epithelial cell line

The Madin-Darby canine kidney (MDCK) cell line, derived from distal tubule/collecting duct, expresses differentiated properties of renal tubule epithelium in culture. We studied the expression of adrenergic receptors in MDCK to examine the role of catecholamines in the regulation of renal function. Radioligand-binding studies demonstrated, on the basis of receptor affinities of subtype-selective adrenergic agonists and antagonists, that MDCK cells have both alpha 1- and beta 2- adrenergic receptors. To determine whether these receptor types were expressed by the same cell, we developed a number of clonal MDCK cell lines. The clonal lines had stable but unique morphologies reflecting heterogeneity in the parent cell line. Some clones expressed only beta 2-adrenergic receptors and were nonmotile, whereas others expressed both alpha 1- and beta 2-receptors and demonstrated motility on the culture substrate at low cell densities. In one clone, alpha- and beta- receptor expression was stable for more than 50 passages. Catecholamine agonists increased phosphatidylinositol turnover by activating alpha- adrenergic receptors and cellular cyclic adenosine monophosphate accumulation by activating beta-adrenergic receptors. Guanine nucleotide decreased the affinity of isoproterenol for the beta 2- receptor but did not alter the affinity of epinephrine for the alpha 1- receptor. These results show that alpha 1- and beta 2-receptors can be expressed by a single renal tubular cell and that the two receptors behave as distinct entities in terms of cellular response and receptor regulation. Heterogeneity of adrenergic receptor expression in MDCK clones may reflect properties of different types of renal tubule cells.