IN VITRO DESENSITIZATION OF β-ADRENOCEPTORS IN GUINEA PIG TRACHEA: INTERACTIONS BETWEEN β-ADRENOCEPTOR AGONISTS AND INFLUENCE OF ADENOSINE AND OTHER DRUGS

The aim of this study was to investigate quantitatively the action of and the interaction between beta-adrenergic receptor agonists in desensitizing guinea pig isolated trachea. It was also to evaluate the influence of substances whose effects on desensitization are either disputed (theophylline, indomethacin, ketotifen, hydrocortisone) or unknown (nicardipine, Bay K 8644, fenspiride, adenosine). Tracheal strips were contracted with histamine (5 x 10(-5) M) or acetylcholine (5.10(-5) M) and concentration-response (C/R) curves for various beta-adrenoceptor agonists were determined before and after incubation (20 min to 4 h) with the same beta-adrenoceptor agonist (autodesensitization), with other beta-adrenoceptor agonists (cross-desensitization), or with a beta-adrenoceptor agonist and another substance. Our results show that the autodesensitization induced by isoprenaline is concentration dependent and that concentration dependence is more pronounced with salbutamol and fenoterol than with isoprenaline and adrenaline with respect to autodesensitization: shifts (log unit) of the C/R curves were 0.59 +/- 0.06 (N = 5) for salbutamol (10(-5) M), 0.78 +/- 0.09 (N = 5) for fenoterol (10(-6) M), 0.30 +/- 0.04 (N = 9) for isoprenaline (10(-5) M), and 0.33 +/- 0.05 (N = 5) for adrenaline (10(-5) M). Our studies of cross-desensitization (desensitization to isoprenaline, adrenaline, salbutamol, and fenoterol induced by incubation with isoprenaline 10(-5) M) showed a significantly greater shift in the C/R curves for fenoterol (0.56 +/- 0.08, N = 5) and salbutamol (0.62 +/- 0.05, N = 5) than for adrenaline (0.35 +/- 0.07, N = 5) and isoprenaline itself (0.30 +/- 0.05, N = 9). Of the substances we studied, none modified the desensitization induced by isoprenaline except hydrocortisone and adenosine. Hydrocortisone (10(-8) M) reduced it significantly, although to a negligible extent. Adenosine (3 x 10(-4) M) did not shift the C/R curve to isoprenaline by itself, but incubation of tracheal strips with adenosine and isoprenaline caused a significantly greater shift of C/R curves to isoprenaline (0.30 +/- 0.04) than incubation with isoprenaline alone (0.20 +/- 0.04) (P less than 0.05, N = 5). These experiments suggest that adenosine may have increased the uncoupling and/or down-regulation phenomena induced by isoprenaline, or modified adenylate cyclase-cAMP activity.     

Glucocorticoid modulation of muscarinic and β-adrenergic receptors in guinea pig lung

Summary— We investigated the effect of the in vivo treatment of guinea pigs with methylprednisolone, 10 mg/kg daily, on lung muscarinic and β-adrenergic receptors. Receptor densities were assessed by saturation experiments of tritiated N-methylscopolamine and dihydroalprenolol binding to lung membranes. After 3 h of treatment, methylprednisolone induced a decrease of 19.2% ( P < 0.05) of muscarinic receptors but was without effect on β-adrenergic receptor density. After 24 h, an increase of 39.7% ( P < 0.01) and 16.9% ( P < 0.05) was observed for muscarinic and β-adrenergic receptors, respectively. For muscarinic receptors, this increase reached 53.4% ( P < 0.01) within 48 h and stayed at this level until 96 h. The increase of β-adrenergic receptors was maximal (24.9%) after 72 h and returned to the control value after 96 h. The dissociation constant (K_d) values of both ligands were not affected by the glucocorticoid treatment. Functional studies showed that the 96 h treatment did not affect the contractile response of guinea pig lung parenchymal strips to carbachol since the 50% concentration value (EC₅₀) and the maximal contraction value (E_max) were not significatively different from control values. These data show that glucocorticoids control the expression of both muscarinic and β-adrenergic receptors in guinea pig lung but with different time courses and to a larger extent for muscarinic receptors. The glucocorticoid treament did not modify the contractile response of lung strips to carbachol, confirming the absence of effect on the affinity of muscarinic receptors and suggesting that the receptor reserve exceed the increase of their density by the steroid.     

Differences in β-adrenergic receptor density and adenylate cyclase activity between normal and leukaemic leukocytes

Abstract. An identical class of high-affinity binding sites for the ¹²⁵I-labelled β-adrenergic antagonist hydroxybenzylpindolol, was identified on intact human normal and leukaemic peripheral blood leukocytes. On normal unfractionated lymphocytes, polymorphonuclear leukocytes, and monocytes, receptor density did not differ significantly (1200–1400 receptors per cell; P > 0.3), but it was higher on B- than on T-lymphocytes ( P < 005). In leukaemia, monocytic blast cells expressed highest receptor numbers, whereas very low receptor density was seen on the pathologic B-cells from chronic lymphocytic leukaemia. Among normal leukocytes, adenylate cyclase activation by hormones (isoproterenol, prostaglandin E₁, histamine) and sodium fluoride was strongest in plasma membranes from monocytes, but very weak in polymorphonuclear leukocytes either due to uncoupling of hormone receptors from adenylate cyclase or to low catalytic activity. In T-cells, enzyme activity was significantly lower than in B-cells. Loss of adenylate cyclase sensitivity to hormones and fluoride occurred in leukaemic cells from chronic and acute lymphocytic leukaemia.     

Identification and characterization of α1- and β2-adrenergic receptors in human liver

Adrenergic receptors were identified in healthy human hepatic tissue from thirty-nine subjects undergoing elective abdominal surgery by using the specific alpha 1-antagonist [3H]-prazosin and the beta adrenergic antagonist [3H]-dihydroalprenolol ([3H]-DHA). [3H]-prazosin binding to plasma membranes was rapid, of high affinity, saturable and stereospecific with a maximal binding capacity (Bmax) of 74.1 +/- 5.5 fmol mg-1 of protein. The displacement curve for (-)-norepinephrine was better explained by a one-site binding and after addition of GTP 0.1 mM the curve was not right-shifted, suggesting the majority of alpha receptors in healthy human liver are of the alpha 1 subtype and not linked to a GTP-binding protein. [3H]-DHA binding to liver plasma membranes was also rapid, of high affinity, saturable and stereospecific with a Bmax 96.5 +/- 10.3 fmol mg-1 of protein of receptors. Computer aided analysis of the displacement curve of ICI 118,551, a subtype selective beta 2-antagonist (IC50 = 62 +/- 2 nM), indicated a one-site binding, thus, showing that beta adrenergic receptors are of the beta 2 subtype. The displacement curve of [3H]-DHA for (-)-isoproterenol was right shifted by GTP indicating that beta 2 adrenergic receptors are linked to a GTP-binding protein in human liver. These results indicate that alpha 1- and beta 2-receptors co-exist in human liver but only beta 2-receptors are linked to a GTP-binding protein.     

β-adrenergic priming of rats in vivo modulates the effect of β-agonist in vitro on surfactant phospholipid metabolism of isolated lungs

Abstract. To evaluate the effects of multiple β -adrenergic stimulations on pulmonary surfactant phospholipids, perfused lungs from β -adrenergic primed and non-primed rats were challenged with the β -agonist terbutaline in vitro . Cell-free lung lavage, lavagable alveolar cells and lung tissue were analysed for phospholipid content and incorporation of precursors. In lung lavage, terbutaline in vitro doubled the incorporation of ¹⁴C-choline and ³H-palmitate into total phosphatidylcholine (PC) and of ³H-palmitate into phosphatidylglycerol (PG). β -adrenergic priming in vivo prior to terbutaline in vitro lowered the increase of precursor incorporation. For lavagable cells, terbutaline in vitro increased the incorporation of ³H-palmitate into PC. Priming in vivo reduced this effect and diminished the specific ³H-choline incorporation into lavagable cell PC below control level. For lung tissue, priming increased the amounts of PC and disaturated PC (DSPC) whereas terbutaline in vitro decreased DSPC in both primed and non-primed lungs. Terbutaline in vitro slightly increased the incorporation of ¹⁴C-choline and ³H-palmitate into PC and DSPC in non-primed but not in primed lungs. β -adrenergic blockade by ICI 118.551 prevented all effects but generally increased ³H-palmitate incorporation into the phospholipids and, in lavagable cells, the amount of PC. We conclude that long-term β -adrenergic treatment may alter the metabolism of pulmonary surfactant phospholipids by increasing tissue PC and DSPC and by decreasing the secretion of newly-synthesized PC.     

β3-adrenoceptors and intestinal motility

Summary— Early substantial evidence of the low susceptibility to β-adrenoceptor antagonists of non α-adrenergic responses reducing gut motility and tone was reluctantly accepted as indicating a third β-receptor subtype different from the β₁ and β₂. This applied likewise to lipolysis until new selective “lipolytic” β-agonists poorly effective at established β-receptors were introduced. Shortly afterwards these “lipolytic” as well as certain newer and even more selective β-adrenoceptor agonists were shown to be potent inhibitors of intestinal motility. The latter are the “gut-specific” phenylethanolaminotetralins whose availability as pure isomers attested to the stringent stereochemical requirements for selectivity at non-β₁, non-β₂ β-adrenoceptors. Acceptance of the functionally based concept of a β₃-adrenoceptor was boosted on structural grounds by molecular biology studies. Sequence analysis indicated the existence in humans and rodents of genes coding for a third subtype of β-receptor that, when expressed in transfected heterologous cells, had a pharmacological profile distinct from the previously established subtypes. Finally, aryloxypropanolaminotetralins have been prepared as the first selective antagonists of β₃-adrenoceptors, thus providing unambiguous conclusive evidence of the distinctive functional features of those abundant in the rat colon. The therapeutic potential in gastroenterology of the newer compounds targetable on the β₃-adrenoceptor is suggested by their potent intestinal action in vivo in animal models without any of the cardiovascular or other unwanted effects of conventional β₃-adrenoceptor agonists and antagonists, and by the clinically confirmed importance of β-adrenergic control of motor function throughout the alimentary canal. However, open questions include the incidence of species-related differences in β₃-adrenoceptors, and as yet there are no data on gastrointestinal functions in humans under the influence of drugs designed to act selectively at these receptors.     

Cardiac β-adrenoceptor sensitivity and Parkinson''s disease

Certain clinical manifestations of Parkinson's disease (PD) (speech or/and balance disturbances) are not linked to brain dopamine deficiency. The purpose of the present study was to search for a possible relationship between those so-called "non-dopamine-dependent" extrapyramidal manifestations and the sensitivity of cardiac beta-adrenoceptors. Fourteen patients aged 51 to 69 were included in the study after having given their informed consent. Any factor or pathology susceptible to modify receptor sensitivity entailed exclusion. In the absence of a reference model for measuring the reactivity of central beta-adrenoceptors, a computation of the isoprenalin dose necessary to increase the resting heart rate by 20 bpm was used as an index for beta-adrenergic system reactivity. In addition to that test, other parameters were recorded: disease duration, motor status scale (Columbia), some cognitive functions (MMS and image differed recall). The cardiac beta-receptor decrease in reactivity to isoproterenol is correlated to PD duration (r = 0.8, P less than 0.001). Conversely, the sensitivity of these receptors appeared to be unrelated to the extrapyramidal severity of the disease, hence to the degree of the so-called "non dopamine-dependent" disturbances. Furthermore, such results raise the meaning of the impairment of peripheral aminergic receptors in the cognitive disturbances linked to ageing and/or PD.     

Myocardial β-adrenergic reactivity in volume overload-induced cardiac hypertrophy in the rat

Summary— The purpose of this study was to evaluate the changes in myocardial β-adrenergic reactivity in animals undergoing a 4 week cardiac volume overload. Aortocaval shunt (ACS) or sham operation (sham) were performed in male Wistar rats, and 4 weeks later, isoproterenol dose-effects (chronotropic, inotropic and lusitropic properties) were studied after pithing. Noradrenaline (NA) and adrenaline (A) concentrations and NA turn-over index were evaluated in plasma and heart ventricles, while β-adrenoceptor characteristics in ventricle homogenates and slices with [¹²⁵I]iodocyanopindolol, and the β(1)/β(2)-adrenoceptor ratio were estimated. Four weeks of cardiac volume overload resulted in a 55% increase in ventricle weight/body weight ratio (from 2.5 ± 0.1 to 3.9 ± 0.1 mg/g in sham and ACS rats, respectively) and a 20% increase in protein contents (from 11.3 ± 0.7 to 13.8 ± 1.1 mg/100 mg ventricles in sham and ACS rats, respectively). Furthermore, NA and A concentrations and NA turn-over index were increased in ACS rats (14, 40 and 80% versus sham, respectively). A shift to the right of the responses in heart rate, left ventricular systolic pressure, +d P /d t _max and -d/ P /d t _max responses following increasing doses of isoproterenol was observed, without change in the dose inducing maximum effect. Total β-adrenoceptor characteristics and β(1)/β(2) ratio were unchanged. However, β(1)-adrenoceptor density increased in epicardium while decreasing in endocardium of left ventricle from ACS rats. Rightward shift at lower doses of isoproterenol-induced cardiac responses in volume-overloaded rats are not likely due to overall β-adrenoceptor density changes.     

Myocardial β-adrenergic reactivity in pressure overload-induced cardiac hypertrophy in the rat

Summary— The purpose of this study was to evaluate the changes in myocardial β-adrenergic reactivity in animals undergoing a 4 week cardiac pressure-overload. Abdominal aortic constriction (AAC) or sham operation (sham) were performed in male Wistar rats, and 4 weeks later, isoprenaline dose-effects (chronotropic, inotropic and lusitropic properties) were studied after pithing. Noradrenaline (NA) and adrenaline (A) concentrations and NA turn-over index (DHPG /NE ratio) were evaluated in heart ventricles, while β-adrenoceptor characteristics in ventricle homogenates and slices with [¹²⁵I]iodocyanopindolol and the β(1)/β(2)-adrenoceptor ratio were estimated. Four weeks of cardiac pressure overload resulted in a 70% increase in ventricle weight/body weight ratio (from 2.5 ± 0.1 to 4.2 ± 0.3 mg/g in sham and AAC rats, respectively) and a 24% increase in protein contents (from 11.3 ± 0.7 to 14.0 ± 1.1 mg/100 mg ventricle in sham and AAC rats respectively). The ventricle NA content was similar in AAC and sham, while the ventricle A content and NA ***turn-over index were significantly increased in AAC rats (35 and 80% vs sham, respectively). Dose response of isoprenaline was significantly shifted to the right for all studied effects in AAC rats. However, maximal response (in relative values) was similar in AAC and sham rats only for heart rate but not for parameters depending on left ventricle contractile response. The β-adrenoceptor density was significantly decreased in AAC by 30% without apparent affinity change and due to decreases in β(1)-sites in septum and to β(1)- and β(2)-adrenoceptors in left ventricle endocardium. Decreases in isoprenaline-induced cardiac responses in AAC rats are associated with β(1)-adrenoceptor density reduction and modification of β(1)- and β(2)-adrenoceptor ratio. These modifications are not the only reason for such dose response changes, at least for contractile response.     

Differential activation of β1-, β2- and β3-adrenoceptors by catecholamines in white and brown adipocytes

Summary— This review summarizes the experiments performed by various groups to determine how the activation of the different β-adrenoceptors (β-ARs) is ordinated when they are present in the same fat cell and involved in the same biological event. When expressed after the transfection of their genes in Chinese hamster ovary cell (CHO cells), β₁- and β₂-ARs present a higher affinity for catecholamines than β₃-ARs. In vitro, the lipolytic effect induced by low concentrations of catecholamines in dog and rat white fat cells is due to the selective activation of β₁- and/or β₂-ARs. Higher concentrations only are able to activate β₃-ARs. Similar results have been obtained in rat brown adipocytes. On the other hand, the lipolytic effect of catecholamines in human and primate adipocytes does not involve a β₃-AR component whatever the concentration used. In vivo experiments in the dog have also shown that lipomobilization induced by low doses of isoprenaline only involved β₁- and β₂-AR activation, this effect being blocked by β₁-/ β₂-antagonist pretreatment. However, in the same blockade conditions, perfusion of a 10-fold higher dose of isoprenaline revealed a β₃-AR contribution in the lipomobilizing effect. These data showed that brown and white adipocyte β₃-ARs possess a lower affinity for catecholamines than β₁- and β₂-ARs and are only recruited by high concentrations of the amines. Thus, even if the β₃-AR plays an indisputable role in the white and brown adipose tissue of some species, its physiological status and its expression are still subject to debate in human white fat cells.     

Effects of AT1- and β-adrenergic receptor antagonists on TGF-β1-induced fibrosis in transgenic mice

Background Transforming growth factor‐β1 (TGF‐β1) is involved in interstitial remodelling promoting collagen synthesis and suppressing collagen degradation by inhibition of collagenases. TGF‐β1 mediates angiotensin II‐dependent effects and modulates β1‐adrenergic signalling. To study the effect of neuroendocrine antagonism on TGF‐β‐induced hypertrophic and fibrotic phenotype, we treated TGF‐β1 (Cys223,225Ser) transgenic mice (TGF‐β1‐TG) with either the β1‐receptor blocker metoprolol (MET), the angiotensin II type I (AT1)‐receptor antagonist telmisartan (TEL) or an antibody blocking TGF‐β1 signalling (TGFβ1‐sR‐Ab). Material and Methods Transforming growth factor‐β1‐TG mice (8 weeks) overexpressing TGF‐β1 were treated with either TEL (10 mg kg^?1), MET (350 mg kg^?1) or a soluble TGF‐β1 receptor antibody (1 mg kg^?1) for 6 weeks. Morphological analyses of interstitium and cardiomyocytes were related to expression of matrix metalloproteinases (MMPs) and their inhibitors (TIMPs) by immunoblotting and zymography. Results In TGF‐β1‐TG mice, myocardial interstitial total collagen content was fourfold elevated compared to that of controls (P < 0·05) and was lowered under the treatment with TEL (P < 0·05). Protein expression of TIMP‐1 and ‐4 was increased in TGF‐β1‐TG but inhibited by TEL (TIMP‐1 and TIMP‐4) and MET (TIMP‐1), while collagenase activity was decreased in TGF‐β1‐TG and normalized by treatment with TEL (MMP‐1 and MMP‐13) and MET (MMP‐1) (P < 0·05). Morphometric measurements of cardiomyocyte diameter and area demonstrated similar antihypertrophic effects for all treatment groups. Conclusion The AT1‐antagonist TEL reduced myocardial hypertrophy and interstitial fibrosis in TGF‐β1‐TG mice by normalizing MMP/TIMP ratio. β1‐Adrenergic inhibition by MET as well as TGF‐β1 antagonism induced antihypertrophic rather than antifibrotic effects. Inhibition of both renin‐angiotensin system and β1‐adrenergic system may exert different but synergistic effects to reduce myocardial remodelling.     

Flerobuterol: a potential antidepressant drug related to β-adrenergic agonists. Experimental profile in mice

The potential antidepressant effect of flerobuterol (dl-(fluoro-2 phenyl)-1 t-butylamino-2 ethanol), a new drug related to beta-adrenoceptor agonists, was evaluated and compared with imipramine and salbutamol using classical psychopharmacological tests in mice. Like imipramine and salbutamol, flerobuterol (0.5-32 mg kg-1, ip) fully prevented apomorphine (16 mg kg-1, sc)- and partly reversed reserpine- and oxotremorine-induced hypothermia. At higher doses (16-32 mg kg-1), flerobuterol enhanced the toxic effects of yohimbine. Unlike imipramine, flerobuterol and salbutamol did not reduce immobility duration in the behavioural despair test. Salbutamol and flerobuterol decreased locomotor activity. Flerobuterol did not induce mydriasis, did not prevent oxotremorine-induced tremors or salivary and lacrimal gland secretion and did not reduce reserpine-induced palpebral ptosis. Propranolol (8 mg kg-1, ip) but not alpha-methyl-paratyrosine (75 mg kg-1, ip) prevented the flerobuterol-induced antagonism of apomorphine-induced hypothermia. Our results suggest that flerobuterol demonstrates potential antidepressant activity, which could be related to beta-adrenoceptor activation in mice.     

Increased α2- but similar β-adrenergic receptor activities in subcutaneous gluteal adipocytes from females compared with males

alpha 2 and beta-adrenergic binding and action were studied in subcutaneous adipocytes from the gluteal region in females and males. The beta-selective antagonist [3H]dihydroalprenolol and the alpha 2-selective antagonist [3H]yohimbine were used to identify the beta- and the alpha 2-receptor, respectively. The biological effects mediated by these receptors were determined by measuring the glycerol release induced by isoproterenol (beta-receptor agonist) and by clonidine (alpha 2-receptor agonist). The study consisted of health volunteers (eighteen females and fifteen males). Compared to men the alpha 2-receptor binding was increased by 73% (P less than 0.01) in adipocytes from females. From Scatchard analysis it was determined that the increased binding was due to an increased receptor number (438 v. 262 fmol mg-1 protein, P less than 0.001) with unaltered Kd (1.18 v. 1.35 nmol l-1, P greater than 0.05). This increased alpha 2-receptor binding in female and adipocytes was associated with a significantly increased sensitivity (P less than 0.05) and maximal antilipolytic effect of clonidine (P less than 0.05). The beta-receptor binding was similar in adipocytes from females and males. However, isoproterenol was significantly more lipolytic in female adipocytes (P less than 0.01). Since the combination of theophylline and adenosine deaminase also was significantly more lipolytic in female adipocytes the enhanced effect of isoproterenol then seemed to be due to mechanisms not directly related to the hormone-beta-receptor binding. Finally, the mixed beta- and alpha 2-receptor agonist, adrenaline showed biphasic effects on lipolysis, with stimulatory effect at low concentrations (less than 500 nmol l-1) and pronounced inhibitory effect at higher concentrations (greater than 1 mumol l-1).(ABSTRACT TRUNCATED AT 250 WORDS)     

Four week administration of an ACE inhibitor and a cardioselective β-blocker in healthy volunteers: no influence on insulin sensitivity

Abstract. In most, but not all, studies antihypertensive treatment with angiotensin converting enzyme inhibitors (ACE inhibitors) improves insulin sensitivity, whereas β-blockers decrease insulin sensitivity. However, there was a significant increase in body weight with β-blockers and changes in the body potassium homeostasis with ACE inhibitors. In order to compare the drug specific metabolic effects of an ACE inhibitor and a cardioselective β-blocker controlling these factors, we measured insulin sensitivity in a randomized, double-blind cross-over study in 22 healthy volunteers (age 27 ± 3 years; BMI 22.0± 1.5kg m^-2 (mean ± SD)) during euglycaemic glucose clamps before and after 4 weeks' administration of 5mg Lisinopril or 5mg Bisoprolol. Both drug phases were separated by 4 weeks of no drug administration. During the insulin sensitivity measurements potassium concentrations were clamped at basal levels by means of a variable i.v. potassium infusion. Body weight was monitored at weekly intervals and kept constant within ± 1 kg of the subjects' baseline weight throughout the entire study period. Insulin sensitivity did not change significantly during either drug administration period. The insulin sensitivity index of the 22 volunteers after administration of the ACE inhibitor was 7.9±2.4 mL min^-1 m²μU^-1 mL^-1 (basal index 8.3 ± l.9mL min^-1 m²μ^-1 mL^-1, and 7.5 ± 2.1 mL min^-1 m²μU^-1 mL^-1 after administration of the β-blocker (basal index 8.2 ± l.9mL min^-1 m²μ^=-1 mL^-1; NS). In order to maintain blood potassium concentrations during the glucose c lamps at basal levels of 3.9 ± 0.3 mmol l^-1 a mean of 7.9±4.2 mval of potassium had to be infused during the low insulin step and a mean of 24.1 ±9.7mval during the high insulin step. Systolic and diastolic blood pressure were lowered significantly by Lisinopril (5±8 mmHg and 7 ± 9 mmHg) and Bisoprolol (7 ± 7 mmHg and 4 ± 8 mmHg; NS between both drugs), respectively. In healthy normo-tensive subjects administration of Lisinopril or Bisoprolol for 4 weeks does not influence insulin sensitivity, when appropriate experimental conditions are maintained.     

Distribution of β-endorphin in normal and schizophrenic human brains

beta-Endorphin was measured by radioimmunoassay in post-mortem human brains. Samples of brain were taken from five discrete areas, both from control brains and brains of schizophrenic patients. No difference in beta-endorphin levels was found in these two groups of brains. beta-Endorphin was confirmed to be widely distributed in the brain, but there were considerable differences in the concentrations in different areas.     

Platelet aggregation induced by the C-terminal peptide of thrombospondin-1 requires the docking protein LAT but is largely independent of αIIb/β3

Summary.  Thrombospondin-1 (TSP1) is abundantly secreted during platelet activation and plays a role in irreversible platelet aggregation. A peptide derived from the C-terminal domain of TSP1, RFYVVMWK (RFY) can activate human platelets at least in part via its binding to integrin-associated protein. Although integrin-associated protein is known to physically interact with αIIb/β3, we found that this major platelet integrin had only a partial implication in RFY-mediated platelet aggregation. Accordingly, RFY induced a significant Glanzmann type I thrombasthenic platelet aggregation. The αIIb/β3-dependent part of platelet aggregation induced by RFY was mainly due to secreted ADP and thromboxane A2. In the absence of αIIb/β3 and fibrinogen, RFY stimulated a rapid tyrosine phosphorylation of a set of proteins, including Syk, linker for activation of T cells (LAT) and phospholipase Cγ2. This signaling pathway was critical for RFY-mediated platelet activation as revealed by the use of pharmacological inhibitors as well as LAT-deficient mouse platelets. Phosphoinositide 3-kinase activation was also required for RFY-mediated platelet aggregation. Our results unravel a new αIIb/β3 and fibrinogen-independent mechanism for platelet aggregation in response to the active peptide from the C-terminal domain of TSP1.     

β-Adrenoceptor density in patients with left-sided valvular regurgitation

Summary— Aortic regurgitation differs from mitral regurgitation in that it is a result of combined volume and pressure overload, while the latter represents an almost pure volume overload. In this study, we tested the possibility that these two forms of left ventricular volume overload exert different effects on β-adrenoceptor density. Lymphocyte (n = 33) and myocardial (n = 22) β-adrenoceptor densities were evaluated by [¹²⁵I]-iodocyanopindolol binding in volume-overloaded patients with left heart valvular disease, compared with 31 healthy donor blood and 15 donor heart controls, made available as a result of failing to get matching recipient. The total lymphocyte (LC) β-adrenoceptor density decreased from 43.4 ± 5.5 fmol mg^?1 protein in controls to 9.2 ± 2.7 fmol (P < 0.001) in heart valvular patients. In the myocardial controls, the left ventricular (LV)-receptor density was 126.7 ± 19.5 fmol; right ventricular (RV), 123.1 ± 14.6 fmol; left atrial (LA), 81.6 ± 10.5 fmol; and right atrial (RA), 108.1 ± 14.5 fmol mg^?1 protein. Compared to this group, the total LV-receptor density of the patients decreased by 63%, RV by 54%, LA by 31% and RA by 34%. The decrease in receptor density exhibited a positive correlation with increasing ejection fractions in both the left (r = 0.38) and right (r = 0.44) ventricles, indicating that the former was dependent on the extent of the disease. These changes were accompanied by a 44% increase in plasma epinephrine, 13% in norepinephrine and a 27% decrease in dopamine levels. Based on the predominant left ventricular volume overload classified as aortic regurgitation (AVR), mitral regurgitation (MVR) and mixed aortic and mitral regurgitation (MOL), the attenuation in myocardial-receptor densities showed the following trend: MOL > MVR > AVR. The results show a global reduction in myocardial and LC β-adrenoceptor density, which depends on the origin and the gravity of the LV volume overload.     

Neurohormonal Profile Before and After β-Blockade in Patients with Neurocardiogenic Syncope

Our objective was to evaluate the effects of beta-blockers on the neurohormonal profile, particularly vasopressin (VP) release, in vasovagal syncope and to gain further insight on the pathophysiology of this syndrome. Patients (< or =75 years) with no cardiovascular, neurological disorders, or contraindications to the use of isoproterenol or beta-blockers and being explored for unexplained syncope were included. An 80 degrees HUT was performed under identical conditions. After a 25-min period of passive tilt, isoproterenol was infused at a rate of 1-5 microg/mn if required. Two groups matched for age and sex were considered: a HUT-positive and a HUT-negative group. The HUT-positive group was then given beta-blockers, subsequently reassessed, and divided into two subgroups: alpha beta-blocker nonresponder group and a beta-blocker responder group. Blood samples for assays of norepinephrine (NE), epinephrine (E), and VP were taken at baseline and the end of the procedure. In all, 44 subjects entered the study, 22 in each group. The HUT-positive group exhibited an obvious lesser increase in plasma NE and a clear-cut rise in plasma E and VP compared to the HUT-negative group (P < 0.05). Even though no patient in the HUT-positive group reported recurrent symptoms under treatment, the second HUT could distinguish two subgroups: a beta-blocker nonresponder group (n = 12) whose HUT remained positive and a beta-blocker responder group (n = 10) whose HUT was normalized. The time course of plasma E and VP during the second HUT was similar to that for the HUT-positive and HUT-negative groups. In conclusion, the efficacy of beta-blockers is associated not only with a reduction of the sympathoadrenal stimulation seen in vasovagal syncope but also with a lower release of VP suggesting that low-pressure baroreceptors might be involved in VP release.     

Evaluation of β-Endorphin Secretion in Patients Suffering From Episodic Cluster Headache

In order to obtain data regarding peripheral levels of β-endorphin in head pain syndromes, we evaluated the plasma β-endorphin secretory pattern in 12 adult male patients suffering from cluster headache. Blood samples were drawn every 2 hours for a 24-hour period, and in addition at 30-minute intervals for 120 minutes during cluster attacks. The same sampling was repeated during an asymptomatic period. Cluster headache patients showed no significant β-endorphin circadian rhythm and a delayed acrophase during cluster periods compared with that recorded in the remission period and in normal subjects. Eighteen cluster headache attacks were recorded during the study day, 13 (72%) of which were followed by a significant increase in β-endorphin levels. No correlation was found between β-endorphin maximum net increase and intensity and/or duration of pain. These data suggest the hypothesis of a temporary alteration of β-endorphin circadian secretion, probably related to involvement of neural structures controlling biorhythm pacemakers.