Number Of Β-adrenergic Receptors Research Articles

Mechanism of endotoxin-induced reduction in the number of β-adrenergic receptors in dog livers: Role of phospholipase A

The role of phospholipase A on the endotoxin-induced reduction in the number of β-adrenergic receptors in dog liver plasma membranes was investigated. The results show that digestion of control liver plasma membranes with exogenous phospholipase A 2 ( 0.2 unit 200 μ g protein) decreased the specific binding of (−)-[ 3H]dihydroalprenolol by 37.3% ( P < 0.01) and reduced the number of receptor sites by 31.7% ( P < 0.05). These decreases in the specific binding and the number of β-adrenergic receptors were completely reversible by the addition of phosphatidylcholine (0.2 m m). Endotoxin administration (2 hr postendotoxin) decreased the specific binding by 36% ( P < 0.05) and reduced the number of β-adrenergic receptors by 33% ( P < 0.05), and these decreases were completely reversible by the addition of 0.2 m m phosphatidylcholine. Digestion of control liver membranes with exogenous phospholipase A 2 decreased phosphatidylcholine and phosphatidylethanolamine levels by 50.6 and 51.2%, respectively, but increased lysophosphatidylcholine and lysophosphatidylethanolamine levels by 12- and 8.4-fold, respectively. Endotoxin administration decreased phosphatidylcholine and phosphatidylethanolamine contents by 21.4 and 23.8%, respectively, but increased lysophosphatidylcholine and lysophosphatidylethanolamine contents by 2.1- and 1.4-fold, respectively. In addition, endotoxin administration increased endogenous phospholipase A activity by 73.5%. Based on these results, it is suggested that the decreases in the specific binding and the number of β-adrenergic receptors in dog livers during endotoxic shock are a result of phospholipase A activation.

The adrenergic beta-receptor adenylate cyclase system in heart and lymphocytes from streptozotocin-diabetic rats. In vivo and in vitro evidence for a desensitized myocardial beta-receptor.

The myocardial beta-receptor adenylate cyclase system was investigated in short-term streptozotocin-diabetic rats. Earlier reports of a decreased sensitivity of the myocardium to isoproterenol (ISO) in these animals were elucidated by measuring the in vivo production of cAMP after ISO. A substantial decrease was seen in diabetic animals compared with controls and starved animals, and thyroxine treatment, known to sensitize the myocardium to catecholamines, did not normalize the response. The desensitization was retained in a membrane fraction in such a way that ISO was unable to increase the cAMP production while stimulation via the nucleotide-binding protein (with NaF or GTP) leads to a normal cAMP response. As the beta-adrenergic receptor number and affinity turned out to be identical in control and diabetic animals, a functional uncoupling of the myocardial beta-receptor from productive adenylate cyclase activation seems thus to exist in experimental diabetes. It is unlikely that it has anything to do with the thyroid status of the animals, but the possibility of a catecholamine-induced densensitization cannot be excluded. The phenomenon is not universal as the beta-receptor-adenylate cyclase system is normal in isolated spleen lymphocytes. Whether the described phenomenon obtained in an animal study has any relevance for the increased incidence of heart failure in human diabetes mellitus is not known at present.

Roles of alpha 1- and beta-adrenergic receptors in adrenergic responsiveness of liver cells formed after partial hepatectomy

The adrenergic receptor involved in the action of epinephrine changed dramatically during the process of active proliferation which follows partial hepatectomy. In control or sham-operated animals, the stimulation of glycogenolysis, gluconeogenesis and ureogenesis by epinephrine was mediated through alpha 1-adrenergic receptors. In contrast, in hepatocytes obtained from animals partially hepatectomized 3 days before experimentation, the receptor involved in the stimulation of these metabolic pathways by epinephrine was of the beta-adrenergic type. Interestingly, the adrenergic receptor involved in the metabolic actions of epinephrine, in hepatocytes from rats partially hepatectomized 7 days before experimentation was again of the α 1-subtype. Thus, it appears that during the process of liver regeneration which follows partial hepatectomy there is a transition in the type of adrenergic receptor involved in the hepatic actions of catecholamines from β in the initial stages to later α 1. A similar transition seems to occur as the animal ages. Cyclic AMP accumulation in response to β-adrenergic stimulation was significantly enhanced in hepatocytes obtained from rats partially hepatectomized 3 days before the experiment, as compared to control hepatocytes or cells obtained from animals operated 7 days before experimentation. This enhanced β-adrenergic sensitivity is probably related to the increased number of β-adrenergic receptors observed at this stage. However, a clear dissociation between cyclic AMP levels and metabolic effects was evidenced when the different conditions were compared. The number and affinity (for epinephrine or prazosin) of α 1-adrenergic receptors did not change at any stage of the process, which indicates that the markedly diminished α 1-adrenergic sensitivity observed in hepatocytes obtained from rats partially hepatectomized 3 days before experimentation is probably due to defective generation or intracellular processing of the α 1-adrenergic signal, rather than to changes at the receptor level.

Effects of combined administration of imipramine and chlorpromazine on β- and α 2-adrenergic receptors in rat cerebral cortex

Administration of the combination of antidepressant and neuroleptic drugs has been reported to have a synergistic effect in the treatment of delusional depression. The effects of chronic coadministration of imipramine (IMIP) and chlorpromazine (CPZ) on β-adrenergic and α-adrenergic sites in rat cerebral cortex were studied. The combination caused the same reduction in the number of β-adrenergic receptors as IMIP alone. No changes in α-adrenergic receptors were observed with IMIP and/or CPZ.

Alloxan-induced diabetes reduces beta-adrenergic receptor number without affecting adenylate cyclase in rat ventricular membranes.

We have investigated alterations in beta-adrenergic receptors and adenylate cyclase activity in myocardial membranes from normal and alloxan-treated diabetic rats. Saturation curves of [3H]dihydroalprenolol binding yielded a Bmax of 96.3 +/- 3.9 fmol/mg protein in normal membranes and 47.6 +/- 3.9 fmol/mg protein in diabetic membranes. Decreased receptor number in membranes from diabetic animals was not accompanied by alteration in receptor affinity for either antagonists or agonists to the beta-receptor. We were unable to detect any alteration in adenylate cyclase activity in similar ventricular membranes. Adenylate cyclase activity in the basal state or in the presence of sodium fluoride, guanyl-5'-yl imidodiphosphate, or isoproterenol, with or without GTP, was not altered by the alloxan-induced diabetic state. Stimulation of adenylate cyclase activity by forskolin, the novel diterpene activator, also was not altered by diabetes. The results suggest that while diabetes reduced beta-receptor number, this is not reflected in any other component of the adenylate cyclase complex.

Effects of experimental insulin-dependent diabetes on the beta-adrenergic-receptor-coupled adenylate-cyclase system and lipolysis in fat cells of the rat.

According to the hypothesis put forward by Zumstein et al. [Eur. J. Biochem., (1980) 105, 187–194], we have investigated whether the increased sensitivity of adenylate cyclase to catecholamines they observed in streptozotocintreated rat adipocytes could be due to altered β-adrenergic receptor number, affinity or coupling to adenylate cyclase. The present report shows the following in streptozotocin-treated rat adipocytes. 1 A 60–70% decrease in the number of β-adrenergic receptors and in basal and isoproterenol maximally-stimulated adenylate cyclase activities, a 45% decrease in the 5′-nucleotidase activity and a 60% reduction in membrane protein content per cell. 2 An almost complete reversion of these alterations by insulin-treatment. 3 An unaltered β-adrenergic-receptor affinity towards β-agonists and antagonists, an unaltered sensitivity of adenylate cyclase to guanine nucleotides and to isoproterenol (in the presence of guanine nucleotides) and an unaltered adenylate cyclase responsiveness (expressed as the percentage of basal activity) to isoproterenol and guanine nucleotides. 4 A 60–70% reduction of the basal lipolytic activities per cell and those stimulated by isoproterenol, adrenocorticotropin and dibutyryladenosine 3′,5′-monophosphate, but no modification of either the sensitivity or the responsiveness (expressed as the percentage of basal activity) of lipolysis to these effectors. From this study, it can be concluded that in streptozotocin-treated rat adipocytes, the sensitivity of the β-receptor-coupled adenylate cyclase system to catecholamines is unimpaired, but the number of β-receptors and of adenylate cyclase catalytic-subunits per adipocyte are reduced, an effect which seems related to defective protein biosynthesis induced by insulin deficiency.

Hormonal regulation of β-adrenergic receptors in fetal rabbit lung in organ culture

Explants of fetal rabbit lung were established on the 25th day of gestation. These were maintained in serum-free medium for periods up to 10 days. During this time, the cultures exhibited morphological changes typical of terminal lung differentiation. Morphological evidence was also obtained for synthesis and secretion of pulmonary surfactant in these explants. β-Adrenergic receptors were identified in these lung explants. Exposure of the explants to 10 −7M dexamethasone on the third day of culture resulted in a significant increase in the number of β-adrenergic receptors in the tissue without a change in receptor affinity. The effect of dexamethasone in organ culture was dose-dependent, a maximum increase in receptor number being observed within 48 hours of incubation with a hormone concentration of 1 × 10 −7M. Exposure of the explant tissue to 1 × 10 −7M triiodothyronine resulted in no significant increase in the concentration of β-adrenergic receptors and no change in receptor affinity. These results suggest that glucocorticoids may potentiate the effects of β-adrenergic agents in the fetal lung by increasing the numbers of their receptors. The effects of triiodothyronine upon the fetal lung do not appear to be mediated by this mechanism.

Lymphocyte beta-adrenergic receptors are not altered in hyperthyroidism.

Binding of (-)3H]-dihydroalprenolol (3H-DHA) to lymphocytes from five thyrotoxic patients and five age- and sex-matched contents were examined to ascertain whether beta-adrenergic receptor number or binding affinity were altered in thyrotoxicosis. Whereas an increase in beta-adrenoceptor density has been reported in triiodothyronine-induced hyperthyroidism, we did not find changes in beta-adrenergic receptor density or binding affinity. In one patient studied sequentially, we did not find alteration in 3H-DHA bindings before or after the subject was rendered euthyroid. We conclude that lymphocyte beta-adrenergic receptor density and affinity are not altered in spontaneous hyperthyroidism.

Evidence for a defect in the number of β-adrenergic receptors and in the adenylate cyclase responsiveness to guanine nucleotides in fat cells after adrenalectomy

Receptor binding studies (−)-[ 3H]dihydroalprenolol as the ligand revealed, in adrenalectomized rat fat cells, a 50% decrease in the number of β-adrenergic receptors. er cell with no change in the receptor affinity for this ligand. Adrenalectomy caused no change in the binding affinity for isoproterenol of both high affinity and low affinity populations of the β-adrenergic receptors. Guanine nucleotide sensitivity of the agonist binding to β-receptors was also unaltered by adrenalectomy. Adrenalectomy caused a 30–40% decrease in the maximal response of adenylate cyclase to (−)-isoproterenol only when guanine nucleotides were present in the assay, without altering the (−)-isoproterenol concentration giving half-maximal adenylate cyclase stimulation ( K act values). The maximal response of adenylate cyclase to G pp(NH) p also was lower in adrenalectomized membranes, indicating a defect at the guanine nucleotide regulatory site. Removal of adenosine by addition of adenosine deaminase failed to reverse the decreased adenylate cyclase response to isoproterenol in adrenalectomized rats. However, in intact fat cells, in which cyclic AMP accumulation in response to isoproterenol was decreased by adrenalectomy, removal of adenosine almost completely corrected this defect. These results indicate that the observed changes in the number of β-adrenergic receptors and in the ability of guanine nucleotides to stimulate adenylate cyclase, though explaining the decreased adenylate cyclase responsiveness to catecholamines, do probably not contribute significantly to the mechanism by which adrenalectomy decreases the lipolytic responsiveness of adipocyte to catecholamines. In addition, this study also suggests that the increased sensitivity to adenosine of lipolysis reported in adipocytes from adrenalectomized rats may result from an action of adenosine at a post-adenylate cyclase step, possibly on the cyclic AMP phosphodiesterase.

Beta-Adrenergic receptors and catecholamine sensitive adenylate cyclase in developing rat ventricular myocardium: effect of thyroid status.

The present study was designed to describe the relationships among thyroid status, myocardial growth and myocardial beta-adrenergic receptors in the developing rat ventricle. In normal rat myocardium the beta-adrenergic binding capacity (Bmax) for (-)-[3H] DHA decreased with increasing age and heart size. In order to determine the effect of thyroid status on ventricular growth characteristics and beta-adrenergic receptors, animals were rendered: (1) hypothyroid with propylthiouracil (PTU), (2) euthyroid with PTU and daily thyroxine (T4) replacement, (3) hyperthyroid for several days with daily thyroxine injections or (4) normal controls with sham saline injections. Growth characteristics were similar in euthyroid and normal rat myocardium; ventricular weight, protein and DNA content were similar at postnatal days 5, 15 and 28. Growth in hypothyroid pups was normal until postnatal day 14 at which time the heart weight and protein content were significantly lower than in normal or euthyroid pups, whereas the number of beta-adrenergic receptors was decreased in hypothyroid myocardium at all ages studied. On postnatal day 5 the (-)-[3H]DHA binding (Bmax) was 37 +/- 9 in hypothyroid myocardium compared to 63 +/- 8 fmole per mg protein mean +/- S.D. in euthyroid myocardium. The function of the beta-adrenergic receptors was also decreased in hypothyroid as compared to euthyroid or normal myocardium as demonstrated by a decrease in maximal catecholamine sensitive adenylate cyclase activity in myocardial membranes at 28 days of age. Treatment of hypothyroid or normal pups with T4 resulted in an increase in heart size, protein content and beta-adrenergic receptors. Ventricular DNA content, which describes hyperplastic growth, was not decreased in hypothyroid rats demonstrating that postnatal hypertrophic but not hyperplastic ventricular growth is dependent on thyroid hormone.

Selective increases in the density of cerebellarβ 1-adrenergic receptors

Destruction of noradrenergic neurons by 6-hydroxydopamine or chronic blockade of β-adrenergic receptors with propranolol increased the density ofβ 1-adrenergic receptors two-fold in rat cerebellum but had no effect on the density ofβ 2-adrenergic receptors. The results suggest that even thoughβ 1 receptors comprise only 5–10% of the total number of β-adrenergic receptors in the cerebellum they are the receptors specifically innervated by noradrenergic neurons and they may thus be the physiologically important receptors.

Alterations induced by a prolonged fasting: opposite effects on the beta-adrenergic receptor-coupled adenylate-cyclase system and on lipolysis in fat cells from rat.

The maximal number of β‐adrenergic receptors in rat fat‐cell membranes from 72‐h‐fasted rats is about two‐times higher than in fed rats. However, the affinity of these receptors for β‐agonists and antagonists and the ability of guanosine 5′‐[β,γ‐imido]triphosphate (100 μM) to reduce the receptor binding affinity for β‐agonists are unaltered by fasting. Basal and fluoride‐stimulated activities of adenylate cyclase are similar in adipocyte membranes from 72‐h‐ fasted and fed animals. However, relative to the basal activity, maximal stimulation by (—)‐isoproterenol is 33 ±5 %higher in fasted than in fed adipocytes. The sensitivity of adenylate cyclase to (—)‐isoproterenol is also increased in fat‐cell membranes from fasted animals, as the concentration of (—)‐isoproterenol required for half‐ maximal activation is three‐times lower in fasted than in fed adipocytes. Basal adenylate cyclase activity is also about three‐times more sensitive to stimulation by guanosine 5′‐[β,γ‐imido]triphosphate in fasted than in fed adipocyte membranes. This increased sensitivity of adenylate cyclase to guanosine 5′‐[β,γ‐imido]triphosphate in fasting is much more pronounced when adenylate cyclase activity is stimulated by isoproterenol. In contrast, although basal lipolysis is unaltered by fasting, lipolysis stimulated in vitro by isoproterenol, adrenocorticotropin or dibutyryladenosine 3′,5′‐monophosphate is severely depressed in fat cells from 72‐h‐fasted rats. However, the concentration of (—)‐isoproterenol and adrenocorticotropin required for half‐maximal activation of lipolysis are similar in fasted and fed adipocytes. From this study, it is concluded that, contrary to the fasting‐induced decreased lipolytic responsiveness which is paradoxically observed in adipocytes in vitro, the fasting‐induced changes found in the adenylate cyclase‐ coupled‐β‐adrenergic receptor‐system of rat adipocyte are most probably physiologically relevant as these changes contribute to promote cyclic AMP synthesis and, consequently, to stimulate lipolysis, as is observed in vivo during fasting.

Differences in the β-adrenergic responsiveness between high and low passage rat glioma C6 cells

Responsiveness to catecholamines was studied in two different strains of rat glioma C6 cells. The C6 cells of low passage possessed a high capacity to accumulate cyclic AMP in response to (−)-isoproterenol. Cholera toxin was also able to stimulate cyclic AMP accumulation in these cells. High passage C6 cells were unresponsive to (−)-isoproterenol or to cholera toxin except in the presence of a high concentration of phosphodiesterase inhibitor. The affinity of β-adrenergic receptors on both strains for (−)[ 3H]dihydroalprenolol was similar; however, C6 low passage possessed several times the number of β-adrenergic receptors found in C6 high passage. This difference correlated with the difference found in (−)-isoproterenol-stimulated adenylate cyclase between C6 low passage and high passage. The sodium fluoride-stimualted adenylate cyclase was similar in both strains. Cyclic AMP phosphodiesterase activity was 2–3-times higher in homogenates of C6 high passage than in low passage. In intact cells, the rate of breakdown of cyclic AMP was 5-times faster in C6 high passage than in low passage. Thus, differences in β-adrenergic receptor number and phosphodiesterase activity explain in part the lack of responsiveness of C6 high passage. Our studies indicate that continuous subculturing of rat glioma C6 cells led to complex alterations in the β-adrenergic receptor-adenylate cyclase system.

Developmental aspects of beta-adrenergic receptors and catecholamine-sensitive adenylate cyclase in rat myocardium.

Summary: The ontogeny of β-adrenergic receptors and the characteristics of catecholamine sensitive adenylate cyclase activity were demonstrated in rat ventricular myocardium. β-Adrenergic receptors were identified by the binding of (—)-[3H]dihydroalprenolol, [(—)-[3H]DHA|, and (±)-[125I]iodohydroxy-benzylpindolol, (±)-[125[I]HYP. Heart weight, protein, and the number of β-adrenergic receptors per cell increased with advancing age; however, binding capacity (Bmax) per nig protein was lower in membrane from the adult rat (220 g body weight) than in membrane from the fetal or neonatal rat, 36.8 ±4.1 and 58.3 ± 6.3 fmoles·mg−1 protein, respectively, mean ± SE, P < 0.01. The age-dependent increase in the numbers of receptors per heart correlated well with the increases in ventricular weight. In contrast, protein content, Na+, K+-ATPase, 5′-nucleotidase, and adenylate cyclase content increased more rapidly than ventricular weight, and the developmental increase in [3H]DHA binding sites (per heart) did not keep up with increases in ventricular protein. Although specific activity of (—)-[3H]DHA binding was lower in the adult ventricle, the characteristics of the β-receptor did not vary with age; the subtype was β1 determined by classical agonist competition in fetal and adult ventricular membranes and the β1- and β2-adrenergic receptor subtypes were present in the ratio of 75% β1 and 25% β2 in both fetal and adult samples. Guanine nucleotides decreased agonist affinity for the inhibition of (±)-[125I]HYP binding equally in adult and fetal samples and enhanced the activation of adenylate cyclase by catecholamine. Basal adenylate cyclase activity was significantly higher in weanling, 13.4 ± 1.42, and adult, 14.2 ± 5.8, than in 21-day fetal or newborn ventricular membrane, 8.18 ± 1.5 pmoles cyclic adenosine 3′:5′-monophosphate produced mg−1· min−1 (mean ± S.D.). The percent activation by catecholamine (with and without GTP) was also slightly higher in samples from the older rats, P < 0.01. In the adult and weanling heart, both guanine triphosphate (GTP) and guanyl-5′-yl-imidodiphosphate [Gpp(NH)p] were effective in enhancing catecholamine sensitive activity. Maximal catalytic activation of adenylate cyclase was observed in the presence of (—)-epinephrine and Gpp(NH)p and was significantly higher (fold stimulation) in fetal or neonatal ventricular samples as compared to adult or weanling ventricular samples, P < 0.01. Speculation: The decreased specific activity (per mg protein) of β-adrenergic receptors in adult as compared to fetal and neonatal myocardium may therefore be related to the decreasing proportion of sarcolemmal protein versus total and particulate cell protein which occurs during hypertrophic cardiac growth, supporting the hypothesis that sarcolemmal surface area is a major determinant of β-receptor number. In contrast, sarcolemmal activities which mediate cellular functions by regulating the concentration of various substances within the cell, such as Na+, K+-ATPase, and adenylate cyclase, increased to a greater extent than (—)-[3H]DHA binding, and were more directly proportional to cell protein. The function and affinity of these β-adrenergic receptors in the rat myocardium are mediated by guanine nucleotides early in development, and age-dependent differences in the activation of catecholamine sensitive adenylate cyclase by GTP and Gpp(NH)p may relate to developmental differences in the properties of guanine nucleotide-dependent factors which mediate receptor occupancy and cyclic adenosine 3′:5′-monophosphate formation.

Increased sensitivity to epinephrine stimulated lipolysis during starvation: Tighter coupling of the adenylate cyclase complex

Adipocytes of rats starved for 72 hr showed three-fold increase in sensitivity of the lipolytic response to epinephrine. Adenylate cyclase sensitivity in adipocyte membranes from starved rats was increased almost seven-fold over control rats. Binding studies using [ 3H]-dihydroalprenolol showed that the number of β-adrenergic receptors per fat cell (1.1 × 10 5 and 1.1 × 10 5 receptors/cell, starved vs control) and the dissociation constant (5 vs 4 nM) of the receptor for the antagonist did not change following starvation. Increased sensitivity of adenylate cyclase activity in response to epinephrine without changes in the β-adrenergic binding parameters suggests that tighter coupling of adenylate cyclase with the receptor may mediate the more sensitive lipolytic response of starved rat adipocytes to epinephrine.

Increase in catecholamine-stimulated cyclic AMP and progesterone synthesis in rat granulosa cells during culture

Catecholamines stimulated cyclic AMP and progesterone synthesis in rat corpora lutea but not in the pre-ovulatory follicles. We collected granulosa cells from follicles of immature rats treated with pregnant mare's serum gonadotropin (PMSG) and cultured them either in monolayer or in suspension. Freshly collected granulosa cells responded to isoproterenol and epinephrine with 2-fold increases in cyclic AMP accumulation and progesterone synthesis. However, granulosa cells cultured in monolayer for 2 days responded to isoproterenol and epinepherine with a 90-fold and a 6-fold increase in cyclic AMP accumulation and progesterone synthesis, respectively. The accumulation of cyclic AMP in response to Catecholamines gradually increased in cells cultured in suspension, from 2-fold over control after 5 h, to 8-fold after 24 h. Granulosa cells isolated from hypophysectomized and diethylstilbestrol (Hx-DES) treated immature rats (containing pre-antral follicles) also showed an increase in cyclic AMP accumulation in response to Catecholamines during culture. Because these cells are devoid of an LH-responsive adenylate cyclase system, we conclude that luteinization of granulosa cells in culture is not necessarily the process responsible for the increased response to Catecholamines. During culture, the number of β-adrenergic receptors in granulosa cells rose from 6020 ± 400 per 10 6 cells shortly after isolation to 26400 ± 1800 after 2 days in culture. This increase in receptor density during culture may be responsible for the change in the responsiveness to Catecholamines, although other factors, such as changes in coupling efficiency between the hormone-receptor complex and the adenylate cyclase moiety and/or supersensitivity to Catecholamines, should also be considered.

Thyroid hormone regulation of beta-adrenergic receptor number in aging rats.

To evaluate the capacity of the myocardium in aged rats to respond to hyperthyroidism, we quantified myocardial beta-adrenergic receptors in female Fischer 344 rats of 3, 12, and 24 months of age. In T3-treated rats (500 microgram T3/kg . day for 3 days), myocardial beta-adrenergic receptors, as measured by [3H]dihydroalprenolol binding, were significantly increased (P less than 0.01) over controls in 3-, 12-, and 24-month-old animals. The data demonstrate that senescent rats retain the capacity to increase myocardial beta-adrenergic receptors in response to exogenous hyperthyroidism. In the myocardium, the mechanism of decreased catecholamine responsiveness in aging appears to be at other than the beta-adrenergic receptor site, since receptor density is unaltered with age, as is receptor modulation in response to hyperthyroidism.

Replacement of serum with a defined medium increases β-adrenergic receptor number in cultured glioma cells

β-Adrenergic receptors of C6 glioma cells were compared following growth in serum or in a serum-free defined medium. In the absence of serum there was an increase in β-receptor number by approx. 50% as measured by the binding of [ 125I]iodohydroxybenzylpindolol. Other receptor properties, such as the affinities for β-adrenergic agonists and antagonists, were not different in the two growth conditions. The alteration in receptor number took over 3 days to occur and was not readily reversible. The increase in β-adrenergic receptor number of cells grown in a serum free medium indicates that replacement of serum with a defined medium is able to alter the expression of at least one cell surface marker, the β-adrenergic receptor. These results imply that growth of cells in defined media yields cells with properties of the plasma membrane that differ from those of cells grown in serum-containing media.

Reversal of noradrenaline denervation-induced increase of β-adrenereceptor binding in rat neocortex by noradrenaline infusion

The effect of intraventricular infusion of (—)-noradrenaline (NA) on β-receptor binding in vitro to homogenates from 6-hydroxydopamine (6-OH-DA)-denervated and from normal rat cerebral cortex was studied. NA was infused with osmotic minipumps connected to cannulas placed in the right lateral ventricle, delivering 1 or 5 μg (—)-NA/h continously for 9 days. One day later the rats were sacrificed and cortical tissue taken for β-receptor (using [ 3H]dihydroalprenolol ([ 3H]DHA) as radioligand) and NA assay. The NA level in the cerebral cortex of 6-OH-DA treated rats was decreased to 70–80% of that of controls. No substantial change in the NA level was observed after infusion of 1 μg (—)-NA/h, whereas infusion of 5 μg/h led to a 40–60% increase compared to that of control rats infused with vehicle alone. Infusion of vehicle alone into control rats did not cause any change in [ 3H]DHA binding, whereas in denervated rats there was a 30–50% increase in [ 3H]DHA binding compared to that of controls. This increase was completely counteracted by infusion of 1 μg (—)-NA/h. Infusion of 1 μg (—)-NA/h to control rats did not cause any change, while infusion of 5 μg (—)-NA/h led to a significant decrease (—24%) in [ 3H]DHA binding. The present results further support the view that the availability of NA at the receptors controls the number of β-adrenergic receptors, thereby probably regulating the NA sensitivity of cells with β-receptors.

Ontogenetic development of isoproterenol subsensitivity of myocardial adenylate cyclase and β-adrenergic receptors in spontaneously hypertensive rats

[ 3H]Dihydroalprenolol binding and adenylate cyclase activity in the myocardial membranes of Kyoto Wistar normotensive rats and spontaneously hypertensive rats were compared at various stages of postnatal development ranging from 2 to 36 weeks. Basal as well as agonist-stimulated myocardial adenylate cyclase activity was consistently decreased in spontaneously hypertensive rats as compared to normotensive rats as early as 2 weeks of age with significant differences ( P < 0.05) observed after 6 weeks of age. When results were expressed as percent stimulation over the basal activity, only isoproterenol plus GTP-stimulated enzyme activity was reduced by 25–30% in spontaneously hypertensive rats, suggesting a specific loss of stimulation by isoproterenol in hypertensive animals. The number of [ 3H]dihydroalprenolol binding sites and K D for dihydroalprenolol binding were comparable between spontaneously hypertensive and normotensive rats at 3, 6 and 12 weeks of age. The competition of isoproterenol with [ 3H]dihydroalprenolol for the specific binding sites showed that the affinity of isoproterenol binding was decreased 3–4-fold in spontaneously hypertensive compared with normotensive rats. With postnatal development in age, basal as well as agonist-stimulated activities decreased progressively in both spontaneously hypertensive and normotensive rats. Similarly, the number of [ 3H]dihydroalprenolol binding sites decreased with the development in age, whereas affinity of dihydroalprenolol binding increased up to 12 weeks of age. These results therefore suggest that adenylate cyclase activity and the number of β-adrenergic receptors in rat heart, decrease with age and that in hypertension, specific decrease in isoproterenol stimulation of cyclase appears at all stages of development.