Effects of chronic morphine administration on catecholamines and beta-adrenergic receptors of the superior cervical ganglion and iris of the rat

The action of chronic morphine administration on catecholamine levels in the cervical (SCG) and iris of the rat pup was studied by radioenzymatic assay. Catecholamine levels in the SCG and iris were unaltered after chronic morphine administration and withdrawal. Norepinephrine (NE) uptake was unchanged throughout drug treatment. beta-adrenergic receptors were increased 30.8% in the iris of morphine-dependent animals. This increase in beta receptors closely parallels the increase in beta receptors found in the brainstem of adult morphine-dependent animals.     

Effects of chronic administration of doxorubicin on myocardial creatine phosphokinase and antioxidant defenses and levels of lipid peroxidation in tissues and plasma of rats

Tissue and plasma levels of thiobarbituric acid reactive substances (TBARS) were measured in rats treated chronically with doxorubicin. In addition, heart creatine phosphokinase and antioxidant defenses were examined. Male rats received doxorubicin (DXR) 2 mg/kg or vehicle weekly subcutaneously for 13 weeks and were sacrificed at 14 and 19 weeks, 1 and 6 weeks after the last dose, respectively. Histological evaluation in DXR-treated rats at 14 and 19 weeks found significant and progressive cardiac and renal lesions as compared to controls. Heart TBARS were unchanged from controls. Plasma and kidney levels of TBARS were elevated above controls at both 14 and 19 weeks. Lung levels of TBARS were significantly elevated above controls at 14 weeks. Liver levels of TBARS were elevated at 19 weeks. Heart creatine phosphokinase activity was significantly depressed from controls at both 14 and 19 weeks. Heart glutathione peroxidase and superoxide dismutase activities were unchanged from controls. Heart glutathione, glutathione reductase, glucose-6-phosphate dehydrogenase, and catalase were elevated above controls at both 14 and 19 weeks. The lack of change in heart TBARS suggests that changes in TBARS in other organs may be secondary processes. The depression of creatine phosphokinase suggests that levels of adenosine triphosphate may be insufficient to sustain the myocardial function and this may partly be responsible for DXR-induced cardiac myopathy.     

Effects of chronic administration of doxorubicin on myocardial alpha-adrenergic receptors, histamine, cyclic nucleotides, calcium, norepinephrine, calmodulin, and guanylate cyclase activity, and plasma catecholamines in rats

The present study examined changes in the levels of plasma catecholamines and myocardial histamine, guanylate cyclase activity, cyclic nucleotides, calcium, calmodulin, and norepinephrine following chronic administration of doxorubicin (DXR). In addition, changes in myocardial alpha 1-adrenergic receptor density and dissociation constant were measured. Rats received DXR (2 mg/kg) or vehicle weekly by the SC route for 2, 4, 8, and 13 weeks. Rats were sacrificed one week after their last dose. One group of rats treated for 13 weeks was sacrificed at 19 weeks, six weeks after the last dose. Heart histamine was unchanged at 3, 5, 9, and 19 weeks, yet at 14 weeks it was significantly elevated in DXR-treated rats over controls. Cardiac calcium, norepinephrine, and cyclic GMP levels were unchanged throughout the course of the study. Cardiac cAMP and calmodulin levels were unchanged at 3, 5, 9, and 14 weeks. At 19 weeks in DXR-treated rats, cAMP was depressed while calmodulin was elevated. Plasma catecholamines and myocardial guanylate cyclase activity examined at 14 weeks were unchanged. In contrast, alpha 1 receptor density examined at 14 weeks in DXR-treated rats was significantly depressed while the dissociation constant was unchanged. Changes in cAMP and calmodulin are suggestive of a redistribution of calcium, although total levels of calcium were unchanged. The depression of cAMP indicates damage to the membrane bound enzyme, adenylate cyclase, and that the membrane interaction of doxorubicin appears to be an integral part of the biochemical mechanism of its toxicity.     

Effects of chronic electroconvulsive shock on the expression of beta-adrenergic receptors in rat brain: immunological study

The purpose of the present study is to determine the effect of chronic electroconvulsive shock (ECS) on the expression of beta-adrenergic receptors in rat brain by Western blot using mAb beta CO2, a monoclonal antibody against beta-adrenergic receptors. Rats in ECS treated groups received maximal ECS (70 mA, 0.5 second, 60 Hz) through ear-clip electrodes for 12 consecutive days. The experiment was carried out in 14 discrete regions of brain. Chronic ECS reduced the expression of beta-adrenergic receptors in frontal cortex, temporal cortex, parietooccipital cortex, hippocampus and limbic forebrain, but not in other areas of brain. The regional specificity and the magnitude of the reduction of receptor expression are well correlated with those of the reduction of receptor ligand binding, which was determined using [3H]dihydroalprenolol. To the best of our knowledge, this is the first report to demonstrate that chronic ECS decreases the expression of receptor protein in specific regions of rat brain.     

Involvement of microtubules in the isoproterenol-induced 'down'-regulation of myocardial beta-adrenergic receptors

The number of cardiac beta-adrenergic receptors identified by [3H]dihydroalprenolol binding decreases in a concentration-dependent manner during prolonged administration of isoproterenol. Loss of membrane beta-receptors is paralleled by the appearance of [3H]dihydroalprenolol binding activity in the cytosol. This redistribution of receptors is prevented by colchicine and vinblastine but not lumicolchicine. Cardiac receptor desensitization is, therefore, dependent on microtubules and may be influenced by agents interfering with tubulin polymerization.     

Effects of chronic bifemelane hydrochloride administration on receptors for N-methyl-d-aspartate in the aged-rat brain

We assayed N-methyl-d-aspartate (NMDA) receptors [³H]3-(2-carboxypiperazin-4-yl)propyl-1-phosphonic acid ([³H]CPP) bindings) and evaluated their distribution in the brain by quantitative autoradiography in young adult and aged rats. In the young adult rats, NMDA receptors were present at relatively high concentrations in the cerebral cortex and hippocampus. In the aged rats, NMDA receptors were decreased in the nealy all areas of the brain, especially in the cerebral cortex and hippocampus. Chronic administration of bifemelane hydrochloride, a drug for sequela of cerebrovascular diseased, at a dose of 15 mg/kg/day for 14 days, markedly attenuated these decrease in NMDA receptors. Since NMDA receptors are considered to be involved in memory and learning processes, our results suggest that bifemelane hydrochloride may be applicable to the treatment of disturbed memory and learning.     

Increased number of beta-adrenergic receptors on cells of the rat adenohypophysis after thyroid hormone administration

The authors studied the effect of administration of thyroid hormones on the beta-adrenergic receptors of rat adenohypophyseal cells. The administration of triiodothyronine and thyroxine was followed by an increase in specific binding for 3H-dihydroalprenolol. No significant differences were found in cyclic adenosine monophosphate levels before and after isoprenaline stimulation. The significance of changes in these receptors for the hyperplastic reaction after oestrogens is discussed with reference to the inhibitory effect of the thyroid hormones on hyperplasia of the adenohypophyseal cells after the administration of oestradiol.     

Effects of chronic beta-adrenergic blockade on exercise training in dogs

To assess the role of beta-adrenergic stimulation in cardiovascular conditioning we examined the effects of a beta-adrenergic blocker, propranolol, in mongrel dogs during an 8-wk treadmill-training program. Seven dogs were trained without a drug (NP), six were trained on propranolol 10 mg.kg-1.day-1 (P), and five served as caged controls (C). Effective beta-adrenergic blockade was documented by a decrease in peak exercise heart rate of 54 +/- 11 (SE) beats/min (P less than 0.05) and a one-log magnitude of increase in the isoproterenol-heart rate dose-response curve. Testing was performed before drug treatment or training and again after training without the drug for 5 days. Submaximal exercise heart rate decreased similarly in both NP and P (-26 +/- 4 NP vs. -25 +/- 9 beats/min P, P less than 0.05 for both) but peak heart rate decreased only with NP (-33 +/- 9 beats/min, P less than 0.05). Treadmill exercise time increased similarly in both groups: 3.4 +/- 0.6 min in NP and 3.0 +/- 0.2 min in P (both P less than 0.05). Blood volume also increased after training in both groups: 605 +/- 250 ml (26%) in NP and 377 +/- 140 ml (17%) in P (both P less than 0.05). Submaximal exercise arterial lactates were reduced similarly in both groups but peak exercise lactate was reduced more in NP (-1.4 +/- 0.3 NP vs -0.3 +/- 0.12 mmol/l P, P less than 0.05). Lactate threshold increased in both groups but the increase was greater in NP (P less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of chronic administration of doxorubicin on cardiac adenylate cyclase activity in mice

Cardiac adenylate cyclase activity was examined in mice treated chronically with doxorubicin. Mice received a subcutaneous dose of either 2 or 4 mg/kg doxorubicin twice weekly for 5 weeks. Mice were sacrificed five weeks after the last injection. Basal cardiac adenylate cyclase activity was significantly elevated in both the 2 and 4 mg/kg DXR-treated groups over the control level. GTP, isoproterenol (plus GTP), NaF, and forskolin stimulated activities in both the 2 and 4 mg/kg DXR-treated groups were also significantly elevated over control levels.     

Effects of chronic and acute exercise on cardiovascular beta-adrenergic responses.

beta-Adrenergic receptor density and responsiveness may be increased in experimental animals by physical conditioning, and the opposite effects have been observed after a single bout of exercise. To determine whether the chronic and acute effects of exercise include similar alterations in cardiovascular function in humans, we characterized heart rate, blood pressure, and distal lower extremity blood flow responses to graded-dose isoproterenol infusion in 15 young healthy subjects before and after exercise training and with and without a single preceding bout of prolonged exercise of either low or high intensity (61 +/- 1 or 82 +/- 1% maximal heart rate). VO2max was increased 18% after exercise training (43.2 +/- 2.7 to 51.1 +/- 3.3 ml.kg-1.min-1; P less than 0.001). Despite a concomitant fall in resting heart rate (59 +/- 3 to 50 +/- 2 beats/min; P less than 0.001), chronotropic and lower extremity blood flow responses to isoproterenol remained unchanged. Similarly, 1 h of acute high-intensity treadmill exercise altered baseline heart rate (58 +/- 4 to 74 +/- 5 beats/min; P less than 0.02), but neither low- nor high-intensity acute exercise influenced heart rate or lower extremity blood flow responses to isoproterenol. In contrast, the systolic pressure response to isoproterenol was blunted after high- but not low-intensity prolonged exercise (P less than 0.02). These data indicate that cardiac chronotropic (primarily beta 1) and vascular (beta 2) adrenergic agonist responses are not altered in humans by training or acute exercise. The systolic blood pressure response to beta-adrenergic stimulation is decreased by a single bout of high-intensity prolonged exercise by mechanisms that remain to be defined.     

Functional integrity of desensitized beta-adrenergic receptors

The adenylate cyclase-coupled beta 2-adrenergic receptor of the frog erythrocyte has served as a useful model system for elucidating the mechanisms of catecholamine-induced densensitization. In this system, it has been previously demonstrated that agonist-induced refractoriness is associated with sequestration of the beta-adrenergic receptors in vesicles away from the cell surface and from their effector unit, the adenylate cyclase system (Stadel, J.M., Strulovici, B., Nambi, P., Lavin, T.N., Briggs, M.M., Caron, M.G., and Lefkowitz, R.J. (1983) J. Biol. Chem. 258, 3032-3038). These internalized beta-adrenergic receptors appear to be structurally intact as assessed by photoaffinity labeling, but their functional status has previously been unknown. In the present studies, we sought to assess the functionality of the sequestered vesicular receptors by fusing them to Xenopus laevis erythrocytes. This cell is suitable for such studies, since it has almost no detectable beta-adrenergic receptor or catecholamine-sensitive adenylate cyclase, but contains prostaglandin E1-stimulable adenylate cyclase. Fusion of beta-adrenergic receptor-containing vesicles from desensitized frog erythrocytes with X. laevis erythrocytes results in a 30-fold stimulation of the hybrid adenylate cyclase by the beta-adrenergic agonist isoproterenol. This effect was entirely blocked by the beta-antagonist propranolol. The catecholamine-sensitive adenylate cyclase activity established in the vesicle-Xenopus hybrids showed the characteristic agonist potency series of the donor frog erythrocyte beta 2-adrenergic receptor. Fusion of vesicles from desensitized frog erythrocytes in which the beta-adrenergic receptors had been inactivated with the group specific reagent dicyclohexylcarbodiimide, or of vesicles derived from control frog erythrocytes, which contain low amounts of beta-adrenergic receptor, did not establish catecholamine-sensitive adenylate cyclase activity in the hybrids. These data demonstrate that beta-adrenergic receptors internalized during desensitization retain their functionality when recoupled to an adenylate cyclase system from a different source. The functional uncoupling of these receptors during desensitization is thus more likely due to their sequestration away from the other components of the adenylate cyclase than to any alterations in the receptors themselves.     

Structural analysis of purified beta-adrenergic receptors

We have characterized the structure of purified beta-adrenergic receptors by a combination of photoaffinity labeling, high performance liquid chromatography (HPLC)-tryptic mapping, CNBr fragmentation, target size analysis, and electron microscopy of purified receptor molecules. Guinea pig lung beta-adrenergic receptors purified by affinity chromatography, ion exchange chromatography, and HPLC size exclusion chromatography or photoaffinity labeled with [125]-iodocyanopindolol diazirine displayed mobilities on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) that corresponded to Mr = 68,000. Purified, radioiodinated guinea pig lung beta-receptors were subjected to complete trypsin digestion and subsequent reverse-phase HPLC analysis, which revealed nine peptides. Active site labeling and tryptic digestion of partially purified hamster lung beta-receptors produced one peptide, whereas CNBr digestion of the same material produced two labeled fragments, yielding information about the location of the active site within the primary sequence. Purified guinea pig lung receptors were examined with transmission electron microscopy. Electron micrographs revealed slightly asymmetric, rod-shaped structures with an average length of 13 nm and width of 3.4 nm. Many receptors were arranged as apparent dimeric structures. These findings confirm data obtained from target size analysis of guinea pig lung beta-receptors in situ which suggest that receptors may exist as oligomeric arrays in the native membrane. Taken together, these data provide information about putative functional domains of the beta-adrenergic receptor and its quaternary structure.     

Effects of chronic glucagon administration on rat lipoprotein composition

Male adult rats of the Wistar strain received daily at 9 a.m. and 5 p.m. 10 micrograms of Zn-protamine glucagon (Novo) for 21 days by subcutaneous injections. Plasma levels of cholesterol, triacylglycerol and phospholipids were decreased by 47, 40 and 21%, respectively. Lipoproteins were separated by sequential ultracentrifugation. Concentrations of cholesterol, phospholipids and proteins were decreased in chylomicrons, VLDL, LDL2 (1.040-1.063 g/ml) and HDL, LDL2 being the most affected by glucagon treatment (-70%). Triacylglycerol levels were decreased only in chylomicrons and VLDL. The relative proportions of cholesterol, triacylglycerol, phospholipids and proteins in lipoproteins were virtually unchanged by glucagon, suggesting a reduced number of some lipoprotein particles in plasma. However, lipoproteins of glucagon-treated rats were depleted in cholesteryl esters, while the proportion of triacylglycerol increased in LDL and HDL. Apo E contents were decreased in plasma, LDL1 (1.006-1.040 g/ml), LDL2 and HDL, whereas apo B100 proportions increased in VLDL and LDL1 in glucagon-treated rats. Glucagon appeared to be a potent hypolipidemic agent affecting mainly the apo-E-rich lipoproteins.     

Agonist-induced changes in beta-adrenergic receptors on intact cells

Competition by beta-adrenergic agonists and antagonists for 125I-pindolol binding sites on intact cells (1321N1 human astrocytoma and C62B rat glioma) was measured using short time binding assays as previously described (Toews, M. L., Harden, T. K., and Perkins, J. P. (1983) Proc. Natl. Acad. Sci. U.S.A. 80, 3553-3557). Preincubation of cells with agonists converted about half of the cellular beta-adrenergic receptors from a form exhibiting high affinity for the agonists isoproterenol and epinephrine and the antagonist sotalol to a form exhibiting much lower apparent affinity for these ligands in short time assays. Exposure to agonists did not alter the affinity of receptors for the antagonist metoprolol. This change in the ligand binding properties of the receptor was rapid (t1/2 = 1-2 min following a lag of about 0.5 min), reversible (t1/2 = 6-8 min), and dependent on the agonist concentration present during the preincubation (K0.5 = 15 nM for isoproterenol). Both isoproterenol and sotalol attained equilibrium with the high affinity receptors very rapidly but equilibrated only slowly with those receptors exhibiting low apparent affinity in short time assays. These results are interpreted in terms of a model which postulates that both the low apparent affinity in short time assays and the subsequent slow equilibration of hydrophilic ligands with these receptors result from agonist-induced internalization of a fraction of cell surface beta-adrenergic receptors. The relationship of this change in receptor binding properties to other aspects of agonist-induced desensitization of the beta-adrenergic receptor-coupled adenylate cyclase system is discussed.     

Down-regulation of glucocorticoid and beta-adrenergic receptors on lectin-stimulated splenocytes

Activation of porcine splenocytes with the mitogen, concanavalin A, increases the number of glucocorticoid and beta-adrenergic receptors with no change in the apparent dissociation constant. Incubation of splenocytes with concanavalin A in the presence of hydrocortisone 21-sodium succinate prevented this mitogen-induced increase in glucocorticoid receptors. Isoproterenol also prevented the concanavalin A-induced increase in beta-adrenoceptors at 24 hr and reduced the binding affinity of these receptors at 48 hr. Neither agonist had any significant effect on the receptor number of binding affinity of nonstimulated cells. These data demonstrate that the increase in the number of glucocorticoid and beta-adrenergic receptors that occur on lymphoid cells after activation by a T-cell mitogen can be prevented by appropriate hormone agonists. Down-regulation of receptor number by appropriate agonists appears to be a common regulatory system that is shared by both the neuroendocrine and the immune systems.