Number Of Beta-adrenergic Receptors Research Articles

Catecholamine-induced adipocyte lipolysis in human hyperthyroidism.

Increased lipid mobilization in thyrotoxicosis is attributed to amplification of catecholamine action in fat cells by thyroid hormones. We investigated the adrenergic regulation of lipolysis in isolated sc abdominal fat cells obtained from 14 patients with thyrotoxicosis and 18 control subjects. Ten of the hyperthyroid subjects were also reinvestigated after antithyroid treatment. The thyrotoxic state was associated with a 3-fold increase in maximum norepinephrine-induced lipolysis (P < 0.005), unaltered sensitivity to dobutamine (selective beta 1-adrenoceptor agonist) and clonidine (selective alpha 2-adrenoceptor agonist), but 15 times enhanced sensitivity to terbutaline (selective beta 2-adrenoceptor agonist; P < 0.01). Moreover, thyrotoxicosis was accompanied by a 3-fold increase in beta 2-adrenoceptor number (P < 0.005), but unchanged beta 1-adrenoceptor levels. Further, the lipolytic effects of dibutyryl cAMP (activating protein kinase A and thereby hormone-sensitive lipase) and forskolin (activating adenylate cyclase) were about 60% enhanced (P < 0.005). No change in the maximum activity of the hormone-sensitive lipase could be demonstrated in the hyperthyroid state compared to that in the euthyroid state. The observed abnormalities in lipolysis and beta 2-adrenoceptor number were normalized after antithyroid treatment. It is concluded that in human hyperthyroidism, the interactions between thyroid hormone and catecholamines in adipocytes involve abnormalities at both receptor and postreceptor levels. The former mechanism seems to be a selective increase in the expression of the beta 2-adrenoceptors. The latter mechanism involves increased ability of cAMP to activate hormone-sensitive lipase, but not a change in maximum enzyme capacity.

Influence of nutritional weaning on porcine adipocyte beta-adrenergic and adenosine A1 receptors.

Obese and crossbred (X-Bred) pigs were removed from sows at 14 d of age and given ad libitum access to a high-fat, milk-based diet. A subset of pigs fed this diet were killed at 28 and 49 d of age. At 28 d, another subset of pigs was given ad libitum access to a low-fat, grain-based diet and were killed at 31, 35, and 49 d of age (nutritionally weaned for 3, 7, and 21 d, respectively). Dorsal subcutaneous adipose tissue was obtained at death and adipocytes were prepared by incubation with collagenase. A portion of the cells was fixed with osmium to determine size and number; remaining cells were lysed in hypotonic media and centrifuged to yield a crude membrane fraction. The beta-adrenergic receptor (beta-AR) and A1 adenosine receptor (A1R) affinity and number were measured in the membranes by equilibrium saturation ligand binding. Obese pigs had a lower body weight than X-Bred pigs at all ages (P < .05). Obese pigs tended (P for volume > .1 but < .2) to have larger adipocytes than X-Bred pigs. The beta-AR affinity did not differ between obese and X-Bred pigs. There were fewer beta-AR per milligram of membrane protein in obese than in X-Bred pigs at 28 and 49 d of age when fed the high-fat, milk-based diet (P < .01). However, beta-AR number expressed per cell or unit cell surface area did not differ between genetic groups. As pigs of either genetic group continued to be fed the low-fat, grain-based diet, the beta-AR decreased when expressed per milligram of protein or unit cell surface area (P < .05) and tended to decrease when expressed per cell. Obese and X-Bred pigs fed the high-fat, milk-based diet had more beta-AR than respective pigs fed the low-fat, grain-based diet when data were expressed per milligram of protein (P < .01) but not when expressed per cell or unit surface area. The A1R were only detectable in 2 of 16 X-Bred litters but were more developed in adipocytes of obese pigs, being measurable in 8 of 14 litters. The A1R number, expressed per milligram of protein, was lower in obese pigs fed the milk-based diet than in those fed the grain-based diet (P < .05). These findings suggest the decreased beta-AR number after nutritional weaning, or the transition from a high-fat to low-fat diet, may contribute to fat accretion in pigs. Furthermore, the lower number of beta-AR in obese than in X-Bred pigs may contribute to the obesity.

Dexamethasone increases beta 2-adrenoceptor-regulated phosphatidylcholine secretion in rat alveolar type II cells.

Insufficient production of pulmonary surfactant in alveolar type II cells is relevant to many lung diseases. To cure its deficiency, glucocorticoid is commonly used in clinical areas. In the present study, we investigated the effect of dexamethasone on the secretion of phosphatidylcholine, a major phospholipid of pulmonary surfactant, in a primary culture of rat alveolar type II cells. Dexamethasone had no effect on the basal secretion rate of phosphatidylcholine. Dexamethasone augmented both the phosphatidylcholine secretion and the cyclic AMP formation increased by terbutaline. Furthermore, dexamethasone increased the number of beta-adrenoceptors and mRNA expression of beta 2-adrenoceptors in type II cells. These findings indicate that dexamethasone increases pulmonary surfactant secretion through an enhancement of beta 2-adrenoceptor gene expression.

Thyroid-thymus interaction on beta-adrenoceptor regulation during development and ageing.

Up-regulation of brain cortex beta-adrenoceptors (beta ARs) can be induced by very acute stimulation with a single injection of T4 or T3 in young Balb/c-nu mice. We have also shown that this very rapid receptor increase can also be demonstrated in ageing animals when stimulated with T3 but not T4 injection. The aim of the present paper was to verify the capability of the thymus to reverse these impairments which we often observed in other experiments on old mice and young athymic nudes. The up-regulation induced by T4 and T3 was studied in normal and athymic nude young, normal old, and normal old and athymic nude young mice grafted with neonatal thymus 1 month earlier. In addition, since brain cortex bears both beta AR subpopulations, the eventual differential behavior of beta 1AR and beta 2AR subtypes was also investigated. Animals were injected intraperitoneally with saline, or saline containing 0.5 microgram T3 or 6.4 micrograms T4 per g body weight and killed 15 or 60 min after injection. Results show that thymus can recover the modifications of basal levels as well as T3-induced up-regulation of beta ARs in nude and old mice. On the contrary, impaired response to T4 stimulation is corrected in nude but not in old mice. The peripheral conversion of T4 into T3 can explain their differential influence since a correct conversion only occurs in presence of an efficient beta-adrenergic function. Thus, a vicious circle may occur with a decreasing number of beta-adrenoceptors causing in old age altered T4 to T3 conversion, in turn responsible for altered beta-adrenergic responsiveness to T4.

In vivo quantification of human pulmonary beta-adrenoceptors: effect of beta-agonist therapy.

In human subjects, chronic beta2-agonist dosing reduces mononuclear leukocyte (MNL) beta-adrenoceptor numbers. The aim of this study was to investigate whether this downregulation also occurs in the lung. Seven healthy male subjects were treated for 2 wk with oral (up to 16 mg/d) and inhaled (up to 1.6 mg/d) albuterol (salbutamol in Europe). Pulmonary maximal beta-adrenoceptor binding capacity (Bmax) was determined in vivo using positron emission tomography (PET) and the beta-receptor antagonist ligand, 11C-labeled CGP-12177, before and after the 2-wk chronic dosing. MNL Bmax was also measured, using a radioligand binding assay and 3H-labeled CGP-12177. Bronchodilator responses to the beta2-agonist were determined after each PET scan by measuring the change in specific airway conductance (SGaw) after increasing doses of inhaled albuterol. Pulmonary and MNL Bmax fell by 22% +/- 14% (p < 0.05) and 42% +/- 19% (p < 0.05) respectively. The changes in pulmonary and MNL Bmax were correlated (r = 0.9, p < 0.05). There was also a reduction in the bronchodilator response to inhaled albuterol. In a further six subjects, pulmonary and MNL Bmax did not change during an acute infusion of albuterol (2 to 4 microg/kg/h). The reduction in pulmonary beta-adrenoceptor numbers after chronic albuterol dosing may be predictable from the changes observed in circulating MNL cells.

Transgenic mice carrying the human beta 2-adrenergic receptor gene with its own promoter overexpress beta 2-adrenergic receptors in liver.

Up to now, transgenic mice models created to study the physiological impact of alterations in the human beta-adrenoceptor system have only focused on cardiac tissues and carried hybrid transgenes with strong cardiac promoters. We have developed a transgenic mouse strain (F28) carrying the human beta 2-adrenoceptor gene with its natural promoter region with the aim of producing a model that more closely reproduces the natural human beta 2-adrenoceptor tissue expression pattern. By means of northern blot analyses, using the appropriate probes, we have obtained evidence that (a) the human beta 2-adrenoceptor's structural gene is transcribed in several tissues of F28 mice; (b) the tissue distribution pattern of human beta 2-adrenoceptor mRNA in F28 mice completely differs from that of mouse beta 2-adrenoceptor mRNA; and (c) the tissue distribution pattern of mouse beta 2-adrenoceptor mRNA in F28 mice is very similar to that observed in their non-transgenic littermates. Like humans, F28 mice express human beta 2-adrenoceptor mRNA in liver, lung, brain, heart, and muscle. However, unlike humans, F28 mice do not accumulate human beta 2-adrenoceptor mRNA in kidney and spleen. By using [125I]iodocyanopindolol to label all beta-adrenoceptors and ICI 118,551 to discriminate between the binding to beta 2- and beta 1-adrenoceptors we have demonstrated that the beta 2-adrenoceptor binding activity increases over control values in F28 mouse tissues that accumulate transgenic mRNA. Accordingly, the number of beta 2-adrenoceptors increased slightly over the control values in muscle, heart, brain, and lung of F28 mice, while in liver these receptors were strongly overexpressed. We further showed that transgene beta 2-adrenoceptors couple to GTP-binding proteins, mediate beta-adrenoceptor agonist-stimulated adenylyl cyclase activation, and cause a strong enhancement of this response in liver membranes of F28 versus control mice. Finally, F28 mice show a phenotype of depressed ponderal development and perturbed hindquarter movements. This unique model should be useful to further investigate beta 2-adrenoceptor causal relationships with human pathologies.

Growth hormone treatment of hypophysectomized rats increases catecholamine-induced lipolysis and the number of beta-adrenergic receptors in adipocytes: no differences in the effects of growth hormone on different fat depots.

Growth hormone (GH) has a lipolytic effect in adipose tissue but this effect may differ in adipose tissue from various fat depots. This latter possibility was investigated in the present study, in which the effects of GH in vivo on catecholamine-induced lipolysis and the number of beta-adrenergic receptors in isolated adipocytes from different fat depots of hypophysectomized rats were investigated. Female and male Sprague-Dawley rats were hypophysectomized or sham-operated at 45 days of age. One week after the operation, hormonal replacement therapy with L-thyroxine and hydrocortisone acetate was given. In addition, groups of rats were treated with GH (1.33 mg/kg per day, given as two daily subcutaneous injections). After 1 week of hormonal treatment, adipocytes were isolated from the parametrial, epididymal and inguinal fat pads, and glycerol release after catecholamine-stimulation and 125I-cyanopindolol binding were measured. Hypophysectomy resulted in a marked decrease in the lipolytic response to catecholamines. GH treatment significantly increased catecholamine-induced lipolysis with similar effects in adipocytes from parametrial or epididymal and inguinal fat depots in both female and male rats. There were no differences between norepinephrine compared with isoproterenol-induced responses. 125I-cyanopindolol binding was reduced after hypophysectomy and normalized by GH treatment, without differences between parametrial and inguinal adipose tissue regions. We conclude that the lipolytic effects of GH in the rat may partly be mediated by a stimulatory effect on beta-adrenergic receptors in adipocytes. In addition, GH exerted similar effect on catecholamine induced lipolysis and beta-adrenergic receptors in adipocytes from parametrial, epididymal and inguinal fat depots.

Lack of effect of chronic antidepressant treatment on Gs and Gi α-subunit protein and mRNA levels in the rat cerebral cortex

Experimental evidence indicates that chronic antidepressant treatment in rats modifies the central nervous system beta-adrenoceptor signaling pathway at multiple sites including receptor, G-protein, adenylyl cyclase, and protein kinase A. In the present study, we examined the postreceptor effect of antidepressant treatment on the protein and mRNA levels of stimulatory and inhibitory C protein alpha-subunits (G alpha s and G alpha i) and beta-subunits in rats infused continuously with various antidepressants for 21 days. Chronic treatment with tricylic (desipramine and amitriptyline) and monoamine oxidase inhibiting (tranylcypromine) antidepressants did not significantly affect the immunoreactivity levels of G alpha s (both 45- and 52-kDa species), G alpha i1, G alpha i2, G beta 36, and beta 35 in rat cerebral cortex. Similarly, the levels of mRNA encoding these G protein subunits remained unchanged subsequent to these drug treatments. In contrast, cortical beta-adrenoceptor number was significantly decreased by these treatments. These results suggest that the adaptive changes of rat cerebral cortical beta-adrenoceptor-adenylyl cyclase system often seen after chronic antidepressant treatment are not accompanied by changes in the abundance and gene expression of G alpha s, G alpha i, or G beta proteins.

Impaired activation of adenylyl cyclase in lung of the Basenji-greyhound model of airway hyperresponsiveness: decreased numbers of high affinity beta-adrenoceptors.

1. To evaluate mechanisms involved in the impaired beta-adrenoceptor stimulation of adenylyl cyclase in tissues from the Basenji-greyhound (BG) dog model of airway hyperresponsiveness, we compared agonist and antagonist binding affinity of beta-adrenoceptors, beta-adrenoceptor subtypes, percentage of beta-adrenoceptors sequestered, and coupling of the beta-adrenoceptor to Gs alpha in lung membranes from BG and control mongrel dogs. We found that lung membranes from the BG dog had higher total numbers of beta-adrenoceptors with a greater percentage of receptors of the beta 2 subtype as compared to mongrel lung membranes. 2. Agonist and antagonist binding affinity and the percentage of beta-adrenoceptors sequestered were not different in BG and mongrel dog lung membranes. However, the percentage of beta-adrenoceptors in the high affinity state for agonist was decreased in BG lung membranes suggesting an uncoupling of the receptor from Gs alpha. 3. Impaired coupling between the beta-adrenoceptor and G protein documented by the decreased numbers of beta-adrenoceptors in the high affinity state in BG lung membranes, is a plausible explanation for the reduced stimulation of adenylyl cyclase and the resultant reduction in airway smooth muscle relaxation in this model.

Circulating noradrenaline and beta-adrenergic receptors in children with congestive heart failure.

This study was designed to investigate changes in plasma catecholamine concentrations and the number of beta-adrenoceptors (beta-AR) of circulating lymphocyte in 94 noncyanotic congenital heart patients. In 43 patients with congestive heart failure, beta-AR density was significantly lower (p < 0.001) and plasma noradrenergic levels were significantly higher (p < 0.001) compared with corresponding values in 51 patients without heart failure. A significant negative correlation between lymphocyte beta-AR density and plasma noradrenergic levels was observed (r = -0.61, p < 0.001). The degree of left-to-right shunt and pulmonary pressure was correlated directly with noradrenaline level and inversely with lymphocyte beta-AR density. Both plasma noradrenaline level and lymphocyte beta-AR density return to normal in children with heart failure after surgical repair. Our results support the idea that changes in noradrenaline level and lymphocyte beta-adrenoceptor density occur concurrently with the presence and severity of heart failure in children.

Adipocyte lipolysis in normal weight subjects with obesity among first-degree relatives.

In this study we investigated whether fat cell lipolysis could be involved in the aetiology of obesity by comparing non-obese subjects with (Hob) or without (Hnorm) a family trait for overweight. A family history of obesity was present when at least one of the first-degree relatives had body mass index of 27 kg/m2 or more. Twenty-seven healthy, drug-free non-obese adult subjects were investigated; 13 were Hob and the remaining 14 were Hnorm. Eleven Hob had at least one obese parent. Isolated fat cells from abdominal subcutaneous adipose tissue were incubated in vitro. Glycerol release (lipolysis index), mRNA levels and enzymatic activity of hormone-sensitive lipase and radioligand binding to beta 1- and beta 2-adrenoceptors were determined. The lipolytic effects of noradrenaline (major endogenous lipolytic agent), isoprenaline (a non-selective beta-adrenoceptor agonist), forskolin (a direct activator of adenylyl cyclase) and dibutyryl cyclic AMP (activating protein kinase and thereby hormone-sensitive lipase) were reduced by about 50% (p from 0.001 to 0.01). The maximum activity of hormone-sensitive lipase was reduced 50% in Hob (p < 0.05) and correlated with the lipolytic responsiveness of fat cells in the whole population (r = 0.71). However, there was no difference between the groups in steady-state mRNA levels for the enzyme. Beta 1-->, beta 2- and alpha 2-adrenoceptor sensitivity as well as beta 1- and beta 2-adrenoceptor numbers were normal in Hob. Fasting plasma insulin was 49.1 and 32.6 pmol/l, respectively in Hob and Hnorm (p = 0.01). There was, however, no significant correlation between lipolysis in vitro and plasma insulin. Thus, lipolytic catecholamine resistance in fat cells, at least partly due to impaired function of hormone-sensitive lipase, is an adipocyte abnormality associated with a family tendency to obesity.

Desipramine administration in the olfactory bulbectomized rat: changes in brain β‐adrenoceptor and 5‐HT2A binding sites and their relationship to behaviour

1. The effects of repeated administration of the tricyclic antidepressant drug, desipramine (DMI), on behaviour (locomotor activity and rearing) and the number and affinity of brain beta-adrenoceptor and 5-HT2A receptor binding sites were examined in olfactory bulbectomized (OB) and sham-operated control rats. 2. Locomotor activity and rearing were increased in OB rats compared to sham-operated controls. The effect of various doses of DMI (administered orally twice daily for 21 days) on these behavioural measures was examined. A dose of 7.5 mg kg-1 provided optimal reversal of hyperlocomotion and increased rearing in OB rats, without changing these measures in sham-operated controls. 3. The time course of DMI (7.5 mg kg-1) on behavioural and neurochemical measures was examined. locomotion and rearing in OB rats were not significantly altered after 7 days, were significantly attenuated after 14 days and were normalized after 21 days. 4. After 7 days of DMI administration the number of beta-adrenoceptors was lower in frontal and occipital cortex and hippocampus. This reduction was largely restricted to the beta 1-adrenoceptor subtype. Administration of DMI for 14 or 21 days did not further reduce the number of beta-adrenoceptors. The DMI induced reduction in beta-adrenoceptors did not differ in OB and sham-operated control rats. 5. DMI administration for up to 21 days produced a progressive reduction in the number of 5-HT2A receptors in frontal cortex, without significant alterations in occipital cortex. 6. The time course of the reduction in the number of 5-HT2A receptors was similar to that of the DMI-induced behavioural changes whereas that for the reduction in beta-adrenoceptors was clearly different. 7. The present results suggest that the action of DMI in this animal model is unlikely to be directly related to a reduction in beta-adrenoceptors but may be related to a reduction in frontal cortical 5-HT2A receptors.

Developmental changes in beta-adrenergic and muscarinic modulations of Ca2+ currents in fetal and neonatal ventricular cardiomyocytes of the rat.

Developmental changes of the beta-adrenergic and muscarinic modulations acting on the slow Ca2+ channels in rat ventricular cardiomyocytes were investigated using Ba2+ as the charge carrier (IBa(L)). Single cardiomyocytes from fetal and neonatal rats were freshly isolated, and currents were recorded by whole-cell voltage clamp in Na(+)-free, K(+)-free external solution. Basal current densities showed a large increase between fetal days 15 and 18, and a small decrease after fetal day 20. The ability of isoproterenol (ISO, 1 microM) to enhance IBa(L) increased dramatically between fetal days 18 and 20, and remained almost constant thereafter during the neonatal period. The ability of forskolin (FOR, 10 microM) to stimulate IBa(L) also increased markedly between fetal day 18 and neonatal day 1. Carbachol (CCh), in a dose-dependent manner, reversed the stimulation of the IBa(L) produced by ISO in neonatal cardiomyocytes. The IC50 values for CCh inhibition of IBa(L) was almost the same on all neonatal days tested (neonatal days 1, 5 and 10). The results indicate that there are developmental changes in beta-adrenergic modulation of IBa(L) in the late fetal period, including a large jump in sensitivity to ISO on about fetal day 19. These findings are consistent with the steep increase in number of beta-adrenergic receptors that occurs between fetal days 18 and 20 (Kojima, Sperelakis & Sada, 1990a: Kojima, Ishima, Taniguchi, Kimura, Sada & Sperelakis, 1990C).

Digoxin reduces beta-adrenergic contractile response in rabbit hearts. Ca(2+)-dependent inhibition of adenylyl cyclase activity via Na+/Ca2+ exchange.

Whereas mobilization of intracellular Ca2+ stimulates neuronal adenylyl cyclase via Ca2+/calmodulin, mobilized Ca2+ directly inhibits adenylyl cyclase in other tissues. To determine the physiologic role of the Ca(2+)-dependent interaction between Na+/Ca2+ exchange and beta-adrenergic signal transduction in the intact heart, digoxin (0.3 mg/kg) was administered intravenously in rabbits. 30 min after the administration, digoxin impaired the peak left ventricular dP/dt response to dobutamine infusions by up to 38% as compared with control rabbits. This impairment was not caused by changes in either beta-adrenergic receptor number or in the functional activity of stimulatory guanine nucleotide-binding protein. It was associated with 33-36% reductions in basal and stimulated adenylyl cyclase activities. Animals treated with calcium gluconate (20 mg/kg/min for 30 min) also demonstrated similar reductions in adenylyl cyclase activities. In addition, increasing the free Ca2+ concentration progressively inhibited adenylyl cyclase activity in the control, digoxin-treated, and calcium gluconate-treated sarcolemma preparations in vitro. Moreover, digoxin and calcium gluconate pretreatment blunted the increase in cAMP in myocardial tissue after dobutamine infusion in vivo. Thus, digoxin rapidly reduces beta-adrenergic contractile response in rabbit hearts. This reduction may reflect an inhibition of adenylyl cyclase by Ca2+ mobilized via Na+/Ca2+ exchange.

Myocardial adenylyl cyclase type V and VI mRNA: differential regulation with age.

Myocardial beta-adrenergic signal transduction diminishes with age. This decrease is not due to a decrease in the number of beta-adrenoceptors, but may be a result of an impaired capacity to activate adenylyl cyclase (AC). Forskolin-stimulated AC activity is diminished, and the number of forskolin binding sites is decreased, suggesting that the decrease in signal transduction with age is a result of fewer AC catalytic units. To investigate whether the decrease in AC with age is associated with diminished AC mRNA, we assessed AC type V and type VI mRNA in ventricles from 6-, 11-, and 24-month-old F-344 rats. The predominant mRNA species, type V, increased by 45% between 6 and 11 months of age but decreased to just below the 6-month level by age 24 months. In contrast, type VI mRNA decreased by 44% between 6 and 11 months of age and then increased to the 6-month level at age 24 months. The changes in type V and type VI mRNA did not parallel the decreases in the AC activity or forskolin binding sites with senescence. These data indicate that the steady-state levels of type V and VI AC mRNA are not reliable predictors of the efficacy of forskolin stimulation of AC activity at different ages.

Beta2-Adrenoceptor Density of Human Lymphocytes After Nitroprusside-Induced Hypotension

The present study was undertaken to assess the influence of nitroprusside-induced hypotension on beta2-adrenoceptor density. Twenty-four patients undergoing noseseptum corrections under general anesthesia were allocated randomly to a nitroprusside or control group. beta2-Receptor density on lymphocytes was measured by binding studies using (-)125 I-iodocyanopindolol. Lymphocyte subpopulations B, T, T (helper), Tsuppressor, and natural killer cells were determined simultaneously by flow cytometry. Five of 12 nitroprusside-treated patients developed significant intraoperative increases of epinephrine levels (+ 69% versus preoperatively) which were not seen in the remaining seven patients. In these five patients, beta2-receptor density of unfractionated lymphocytes was 26% lower (P < 0.05) on the first day after surgery compared with preoperative values. Since no changes in proportions of lymphocyte subpopulations were observed, these results are not caused by redistribution phenomena inducing a decrease of subsets with a high number of beta receptors. These findings suggest that beta2-adrenergic responsiveness might be diminished after nitroprusside treatment in some patients. (Anesth Analg 1995;81:1250-4)

Alterations in G-protein-regulated transmembrane signalling induced in murine myocardium by coxsackievirus B3 infection

Cardiomyopathy is usually associated with marked alterations in myocardial transmembrane signalling. Although acute viral myocarditis may result in chronic cardiomyopathy in some cases, the possible consequences of viral infection on function of the myocardial signal-transducing complex have not been explored. Therefore, the present study was designed to investigate the G-protein-regulated adenylyl cyclase signalling system in murine myocardium during myocarditis induced by coxsackievirus B3 (CVB3) infection. We examined the functional characteristics of adenylyl cyclase as well as the function and distribution of beta-adrenoceptors, m-cholinoceptors and G-proteins in myocardial plasma membranes isolated from the hearts of mice with acute (7 days pi) or late phase (21 days pi) myocarditis and the obtained results were compared with the corresponding data determined in age-matched controls. While the basal adenylyl cyclase activity was not significantly altered, the ability of forskolin, sodium fluoride and GTP gamma S to activate adenylyl cyclase was lowered by about 20% in samples from virus-infected animals. The level of Gs alpha in myocardial plasma membranes as well as the functional activity of Gs alpha was not affected by viral infection, but the Gi alpha content was increased by about 20%. The total number of beta-adrenoceptors in myocardial plasma membranes increased by about 12-15% due to higher content of the beta 2-adrenoceptor subtype. Although the agonist-binding parameters of beta-adrenoceptors were not significantly altered, the ability of these receptors to mediate stimulation of adenylate cyclase was markedly diminished (by 56-80%). The total number of m-cholinoceptors in samples derived from virus-infected mice increased considerably (by 29-59%) and a significant proportion of the receptors shifted to a higher affinity status, but their ability to transduce agonist signals was impaired. These data are the first to demonstrate that several different sites of the myocardial G-protein-regulated adenylyl cyclase signalling complex are significantly altered in acute as well as in late phase of CVB3-induced myocarditis.

Evidence for mast cell, leukotriene and nitric oxide involvement in the regulation of the adrenoceptor number of inflamed small intestine in guinea pigs

Changes in the populations of neurotransmitter receptors involved in the control of intestinal smooth muscle function have been associated with the altered motility of the inflamed gut. Thus, trinitrobenzenesulphonic acid (TNBS)-induced gut inflammation is accompanied by an increase in alpha- and a decrease in beta-adrenoceptor numbers in guinea pig small intestine. In the present study, we investigated the effects of anti-inflammatory compounds (cyclooxygenase inhibitor indomethacin, lipooxygenase inhibitor MK-886, nitric oxide synthase inhibitor NG-nitro-L-arginine methylester (L-NAME), mast cell stabilizer doxantrazole) on TNBS-induced adrenoceptor changes. Smooth muscle adrenoceptor populations, labelled by subtype-specific radioligands 6 days after TNBS, were significantly different from those of sham-treated controls: alpha 1- and alpha 2-adrenoceptor numbers increased by more than 50%, while beta-adrenoceptor numbers decreased by more than 50%. These changes, associated with severe inflammation as assessed histologically and by myeloperoxidase assay, were prevented by doxantrazole or L-NAME, and only partly by MK-886. In contrast, indomethacin did not prevent these changes. It appears then that: (a) mast cell mediators, nitric oxide and leukotrienes are likely to contribute to TNBS-induced changes in adrenoceptor populations in the guinea pig inflamed intestine; (b) there is no evidence for prostanoid involvement in this process. It was suggested that changes in smooth muscle adrenoceptor populations may be an important mechanism by which gut inflammation alters intestinal motility.

Vasopressin resistance in chronic renal failure. Evidence for the role of decreased V2 receptor mRNA.

Studies were performed to determine the mechanism underlying deficient arginine vasopressin (AVP)-stimulated adenylyl cyclase activity in chronic renal failure (CRF). As compared to control, principal cells cultured from the inner medullary collecting tubule of rats previously made uremic by 5/6 nephrectomy fail to accumulate cAMP when stimulated with AVP. CRF cells do respond normally to forskolin or cholera toxin and the defect in AVP responsiveness is not prevented by treatment with pertussis toxin, by cyclooxygenase inhibition, or by inhibition or down-regulation of protein kinase C. In contrast to their lack of responsiveness to AVP, CRF cells respond normally to other agonists of adenylyl cyclase such as isoproterenol or prostaglandin E2. Plasma membranes prepared from the inner medullae of CRF rats exhibit a marked decrease in apparent AVP receptor number but no change in the apparent number of beta adrenergic receptors. Reverse transcriptase PCR of total RNA from the inner medullae of CRF animals reveals virtual absence of V2 receptor mRNA; mRNA for alpha s is present in normal abundance. These studies indicate that AVP resistance in CRF is due, at least in part, to selective down-regulation of the V2 receptor as a consequence of decreased V2 receptor mRNA.

Effects of dietary restriction on motor learning and cerebellar noradrenergic dysfunction in aged F344 rats.

Fisher 344 rats were fed either ad libitum or with a diet containing a 40% reduction of calories beginning at 4 months of age. At 14 months and 22 months male rats were tested for their ability to learn a complex motor skill. At both ages the diet restricted rats reached criterion of performing 10 successful crosses in 10 min at an earlier time than ad libitum fed controls. At 22 months of age the diet restricted rats showed improved acquisition of running times for the task. Male rats at 14 and 22 months and female rats at 24 months were examined electrophysiologically for the ability of isoproterenol to augment the action of GABA in the cerebellum when both substances were applied iontophoretically from an extracellular multibarreled glass electrode. In all 3 age and sex groups there was an improvement in the beta-adrenergic receptor modulation of GABA responses in the dietary restricted vs. ad libitum rats. However, no difference was observed between dietary restricted and ad libitum rats when the number and affinity of cerebellar beta-adrenergic receptors was assessed with 125I-iodopindolol binding. Overall, there was a significant improvement in cerebellar noradrenergic function in the dietary restricted rats and this was accompanied by an improvement in motor learning.