Decline in beta adrenergic receptor-mediated vascular relaxation with aging in man

Beta adrenergic relaxation of vascular smooth muscle, mediated by cyclic AMP, is blunted with age in a variety of experimental animals. The applicability of these observations to man is uncertain. The dorsal hand vein technique provides an excellent method to examine the direct effects of aging on vascular responsiveness. Thirty-nine healthy male volunteers over the age range of 19 to 79 were studied. No differences in vascular responsiveness to phenylephrine, an alpha adrenergic agonist, were found for either the ED50 (dose producing 50% vasoconstriction) or Emax (maximum vasoconstriction attained). In marked contrast, vascular relaxation induced by isoproterenol, a beta adrenergic agonist, was significantly different in both the ED50 (dose producing 50% of maximum relaxation from a preconstricted state) and Emax (maximum relaxation attained). ED50 +/- S.E.M. for the youngest and oldest deciles were 8.9 +/- 2.3 and 60 +/- 17.0 ng/min, respectively (P less than .05); Emax +/- S.E.M. were 96.7 +/- 3.3 and 37.7 +/- 8.7%, respectively (P less than .001). Nitroglycerin, a smooth muscle relaxant whose effects are not mediated through the cyclic AMP system, was also used to examine the specificity of this blunted response to isoproterenol. Almost complete relaxation was achieved with the infusion of nitroglycerin in the older group. These results suggest that aging is associated with a specific decrease in beta adrenoreceptor-mediated vascular relaxation.     

Age-related decrease in beta adrenergic receptor-mediated vascular smooth muscle relaxation

Beta adrenergic receptor-mediated vascular smooth muscle relaxation decreases with increasing age. We have examined the mechanism responsible for this phenomenon using rat mesenteric arteries from young (5-6 weeks) and older (10-12 months) rats. The beta adrenergic agonist isoproterenol produced a dose-dependent relaxation of serotonin-constricted mesenteric artery rings from young rats, whereas the maximal ability of isoproterenol to relax arterial rings from the older rats was found to be reduced markedly (92.7 vs. 27.6%, P less than .0001). The relaxation responses caused by acetylcholine and nitroglycerin, which appear to act independently of cyclic AMP (cAMP), are similar in the two groups. The loss in responsiveness of the mesenteric artery to isoproterenol was not explained by a change in beta receptor number in the vessels (29 +/- 4 in young rats vs. 31 +/- 7 fmol/mg of protein in the older rats). The maximal stimulation of cAMP accumulation by isoproterenol was lower in the older vessels; forskolin activated cAMP accumulation equally in the two groups. However, the vessels from the older rats were less sensitive to forskolin-induced vascular relaxation. Also, the ability of dibutyryl cAMP to promote vascular relaxation was diminished in the older vessels. These data suggest that the diminished cAMP accumulation in older vessels in response to isoproterenol might not necessarily in itself explain completely the reduced physiological response and that an additional defect in the beta adrenergic-mediated relaxation in the vascular smooth muscle of older rats may lie at the level of cAMP-dependent protein kinase activation or more distally.     

The Adrenergic Control of Lower Esophageal Sphincter Function: AN EXPERIMENTAL MODEL OF DENERVATION SUPERSENSITIVITY

To evaluate the adrenergic regulation of lower esophageal sphincter (LES) function, LES pressure, LES relaxation during swallowing, and blood pressure were measured in the anesthetized opossum, Didelphis virginiana, during intravenous administration of alpha and beta adrenergic agonists and antagonists. Studies were done in controls and animals adrenergically denervated with 6-hydroxydopamine. Alpha adrenergic agonists (norepinephrine, phenylephrine) increased LES pressure and blood pressure, whereas a beta adrenergic agonist (isoproterenol) decreased both pressures. Alpha adrenergic antagonism (phentolamine) reduced basal LES pressure by 38.3+/-3.8% (mean +/-SEM) (P < 0.001). Beta adrenergic antagonism (propranolol) had no significant effect on either basal LES pressure or percent of LES relaxation with swallowing. After adrenergic denervation with 6-hydroxydopamine, basal LES pressure was reduced by 22.5+/-5.3% (P < 0.025) but LES relaxation during swallowing was unaltered. In denervated animals, both LES pressure and blood pressure dose response curves showed characteristics of denervation supersensitivity to alpha but not to beta adrenergic agonists. These studies suggest: (a) a significant portion of basal LES pressure is dependent upon alpha adrenergic stimulation; (b) LES relaxation during swallowing is not an adrenergically mediated response; (c) the LES pressure response to alpha adrenergic agonists after 6-hydroxydopamine may serve as a model of denervation supersensitivity in the gastrointestinal tract.     

Role of cyclic AMP-dependent protein kinase in the diminished beta adrenergic responsiveness of vascular smooth muscle with increasing age

Beta adrenergic receptor-mediated relaxation of blood vessels declines with age although the mechanism is unknown. We have utilized the mesenteric artery and aorta of young and older rats to investigate this problem. In vessels from 12-month-old rats there was a marked loss in relaxation mediated by beta adrenergic and adenosine receptors compared to younger rats whereas relaxation induced by muscarinic cholinergic receptors, [cyclic AMP (cAMP) independent], was not impaired. Maximal relaxation to forskolin and dibutyryl cAMP were intact in the vessels from older rats. Isoproterenol-stimulated cAMP accumulation and cAMP-dependent protein kinase activation were attenuated markedly in the vessels from the older rats. Maximal forskolin-stimulated cAMP accumulation and cAMP-dependent protein kinase activation were similar in older and young animals. There was an excellent correlation between cAMP-dependent protein kinase activity and relaxation and the relationship was similar in the two age groups. Continuous infusion of the beta adrenergic antagonist timolol for 1 week into older animals partially restored relaxation to beta adrenergic and adenosine receptor agonists in the aorta. These results suggest that the age-related loss of response to beta adrenergic receptor agonist-induced relaxation may be due in part to attenuated activation of cAMP dependent protein kinase and this change may be partially dependent on endogenous catecholamines.     

Desensitization of adenylate cyclase and down regulation of beta adrenergic receptors after in vivo administration of beta agonist

In vitro incubation of cells with catecholamines leads to both down regulation of beta adrenergic receptor number and desensitization of agonist-stimulated adenylate cyclase activity. These same parameters, down regulation of beta adrenergic receptor number and desensitization of adenylate cyclase activity were assessed in rat lung membranes after in vivo administration of metaproterenol, a beta-2 selective agonist. In vivo treatment with metaproterenol leads to: 1) reduced beta adrenergic receptor number; 2) reduced isoproterenol-stimulated adenylate cyclase activity; 3) unaffected NaF or 5'-guanylylimidodiphosphate-stimulated adenylate cyclase activity; and 4) reduced affinity of the receptor for isoproterenol similar to the affinity observed in the presence of 5'-guanylylimidodiphosphate. The date suggest that in vivo metaproterenol administration results in an uncoupled receptor-adenylate cyclase complex. The effects of in vivo administration of the glucocorticoid, methylprednisolone, to metaproterenol-pretreated animals were also assessed. Glucocorticoid treatment was associated with 1) increased beta adrenergic receptor number in rats in which the receptors have been down regulated, 2) increased isoproterenol responsiveness in agonist-desensitized rats and 3) no effect on agonist affinity in desensitized animals. These data suggest that the restoration of agonist responsiveness by glucocorticoids in the catecholamine refractive state is not simply a reversal of receptor down regulation or adenylate cyclase desensitization.     

Physiological approach of beta receptor coupling to adenylate cyclase in rat airways: ontogenical modification and functional antagonism

The relaxant effects of isoproterenol, forskolin and sodium nitroprusside were studied on tracheal pieces and lung parenchymal strips of Sprague-Dawley and Wistar rats according to age and functional antagonism with carbachol applied previously to induce the contraction. The beta receptor-related maximal relaxant effect of isoproterenol decreased from 4 to 11 weeks in Sprague-Dawley rat airways contracted with 10(-6) M carbachol. This maximal relaxant effect did not change with age in the Wistar strain. When lower carbachol concentrations were applied to Wistar trachea, the maximal relaxant effect of isoproterenol raised with a large decrease of the EC50 values. In the Sprague-Dawley strain, a similar diminution of carbachol concentration also allowed to increase the maximal amplitude of relaxation, but a smaller decrease of EC50 was observed as referred to the Wistar strain. These results suggest that the decrease with age of the maximal relaxation of Sprague-Dawley airways by isoproterenol might be linked to impaired functional antagonism between beta adrenergic and muscarinic stimulation in this rat strain. This hypothesis was strengthened by the observation of the effects of forskolin, an activator of adenylate cyclase, and sodium nitroprusside, a cyclic GMP-related relaxant drug, that did not show any modified effect in function of age in both rat strains. A modified regulation of adenylate cyclase complex with ontogenesis and with rat strain is suggested.     

Developmental changes in the rabbit sinus node action potential and its response to adrenergic agonists

We used standard microelectrode techniques to study developmental changes in the transmembrane potentials of sinus nodes from 1- to 10-day-old and adult rabbits superfused with Tyrode's solution at 37 degrees C. Maximum diastolic potential did not change with age, but action potential amplitude in day 1 to 3 hearts was lower than that in adults. Sinus cycle length (SCL) also was shorter in the young sinus nodes. In experiments on beta adrenergic stimulation the reduction of SCL in the young occurred at lower isoproterenol concentrations than in the adult, but the maximum reduction in SCL was the same at both ages (-40%). The alpha agonist, phenylephrine, had no age-dependent effect on SCL or action potential characteristics in either age group. In addition, the inhibitory effect of alpha stimulation demonstrated previously in automatic fibers of the ventricular specialized conducting system and atrium did not occur in sinus node. Studies of Purkinje fibers demonstrated this inhibitory effect to be voltage-dependent; it did not occur at membrane potentials in the range of those in the sinus node. Our results suggest that whereas developmental changes in the response to beta adrenergic stimulation are demonstrable in the sinus node, there are no such changes in the response to alpha adrenergic stimulation.     

Mechanism of isoproterenol-induced desensitization of tracheal smooth muscle.

Isolated rat tracheal smooth muscle became considerably less sensitive to the relaxing action of isoproterenol after being incubated with 5 x 10(-6) M isoproterenol for 30 minutes. Pretreatment of the tissue with propranolol, but not with methylprednisolone, clearly reduced the isoproterenol-induced desensitization. This suggested that propranolol by occupying the beta adrenergic receptor prevented isoproterenol from binding to this receptor, thereby preventing the isoproterenol-induced desensitization. Furthermore, an isoproterenol-desensitized tracheal preparation exhibited a diminished sensitivity to other beta agonists, but not to the spasmolytic actions of D600, hydralazine, sodium nitrite and aminophylline. These results suggested that the beta receptor is specifically involved in the desensitization induced by isoproterenol. A highly desensitized tissue could always be made to undergo complete relaxation by exposing it to sufficiently high concentrations of isoproterenol. Thus, there appeared to be no positive indication of a very large change in the apparent intrinsic activity of the isoproterenol in the desensitized tissue. However, the dissociation constant for the propranolol-beta receptor complex in the desensitized tissue was shown to be 180-fold larger than that in the normal tissue. These findings provide strong evidence that one demonstrable cellular change that occurs in the desensitized tissue is a pronounced reduction in the affinity of the beta receptors for isoproterenol.     

Thyroid status and adrenergic receptor subtypes in the rat: comparison of receptor density and responsiveness

The density and functional responsiveness of adrenergic receptor subtypes were determined in tissues from control, hyperthyroid and hypothyroid rats. There was a decrease in sensitivity to isoproterenol in spontaneously beating right atria, electrically driven left atria and field-stimulated vas deferens associated with hypothyroidism, with no change in maximum response. Hyperthyroidism increased the potency of isoproterenol in right atria, but not in left atria or vas deferens. The maximal response to isoproterenol was greatly reduced in hyperthyroid left atria. The potency of procaterol, a partial agonist at beta adrenergic receptors in right atria, was unaltered in hyper- or hypothyroidism, although the maximum stimulation by procaterol was increased in hyperthyroidism. Scatchard analysis of specific [125I]pindolol binding showed that beta adrenergic receptor density was greater in hyperthyroidism than in hypothyroidism in left atria, right atria, ventricles, vas deferens and cerebral cortex, although the proportions of beta-1 and beta-2 adrenergic receptor subtypes did not change. There was no change in the responsiveness of alpha-1 adrenergic receptors mediating contraction of caudal artery and vas deferens or mediating [3H]inositol phosphate accumulation in cerebral cortex in hyperthyroid or hypothyroid rats, although the maximal contraction of caudal artery was significantly reduced in hyperthyroidism. Scatchard analysis of specific [125I]BE 2254 binding showed that alpha-1 adrenergic receptor density was significantly decreased in the ventricles from hyperthyroid rats and increased in the ventricles of hypothyroid rats, but was unchanged in vas deferens, caudal artery and cerebral cortex. Alpha-2 adrenergic receptor density in cerebral cortex, determined by Scatchard analysis of specific [3H] rauwolscine binding, was not altered in hyperthyroid or hypothyroid rats.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of increasing age on the endothelium-mediated relaxation of rat blood vessels in vitro

The effect of age on cholinergic endothelium-mediated relaxation was examined in vitro using blood vessels from Fischer 344 rats ranging in age from 1 to 27 months. Although no differences were seen in contractile response to alpha adrenergic agonists, both aortic ring segments and perfused caudal arteries showed an increase in sensitivity of endothelium-mediated relaxation to the cholinergic agonist methacholine. This increase in sensitivity occurs between the ages of 6 and 12 months, with no further significant increase in sensitivity up to 27 months of age, suggesting it is a consequence of growth and development rather than old age. No difference with age in cholinergic relaxation was observed in the perfused mesenteric bed, indicating either no change of sensitivity in smaller resistance vessels or an effect that is hidden in this more complex perfused system. In contrast to findings with cholinergic stimulation, responses of the perfused caudal artery to the calcium ionophore A23187 were not altered with age. This suggests that the alteration with age in response to methacholine involves the muscarinic receptor or receptor-coupling mechanism rather than the generation of, or response to, endothelium-derived relaxing factor.     

Functional interactions in smooth muscle: kinetic characterization of the relaxation and desensitization responses to a beta adrenergic agonist in the rabbit aorta

Vascular smooth muscle tone is continuously modulated in vivo by the functional interaction of a variety of vasoconstrictor and vasodilator stimuli. Endogenous substances such as epinephrine simultaneously activate alpha adrenergic receptors that elicit muscle contraction and beta adrenergic receptors that relax the muscle. This study characterizes the beta adrenergic response in the isolated rabbit aorta precontracted with 1 microM phenylephrine. The beta adrenergic agonist isoproterenol (0.03-10 microM) produces a biphasic response that is composed of a rapid relaxation followed by a slower regaining of tension, which is identified as desensitization. An exploratory kinetic model that describes both the relaxation and the desensitization as first-order processes provides a good fit to the experimental data. The parameters used to describe the isoproterenol response are: 1) the observed rate constant for relaxation and its magnitude (krel and R, respectively), 2) the observed rate constant for desensitization and its magnitude (kdes and D, respectively) and 3) the observed delay in the onset of the desensitization response (td). Both the krel and the fractional relaxation were dependent on concentration of isoproterenol in a saturable manner (EC50 = 0.017 and 0.067 microM, respectively). No concentration dependence was observed for kdes, fractional desensitization and td (the average values +/- S.E.M. of these parameters are (4.7 +/- 0.2). 10(-3) sec-1, 0.83 +/- 0.02 and 191 +/- 6 sec, respectively). This work demonstrates that a kinetic approach is necessary to characterize the desensitization response and is also very useful in characterizing the kinetic and steady-state parameters of the relaxation response.(ABSTRACT TRUNCATED AT 250 WORDS)     

Alteration in sensitivity to isoproterenol and acetylcholine in the rat heart after repeated administration of isoproterenol

Alteration of sensitivity to the inotropic responses to isoproterenol and acetylcholine (ACh) and of saturation constants for the specific binding of [3H]dihydroalprenolol and [3H]quinuclidinyl benzilate to particulate fractions were investigated in rat hearts chronically treated with isoproterenol. The EC50 value for the inotropic response to isoproterenol in the atria of rats injected i.p. with isoproterenol (0.01 or 0.1 mg/kg) twice a day for 10 days, was 9.0-fold higher than in controls. Isoproterenol (0.17 mg/kg i.p.)-induced stimulation of cardiac ornithine decarboxylase activity was significantly (P less than .05) attenuated in atria from animals of the treated groups. Scatchard analysis of specific binding of [3H]dihydroalprenolol revealed that repeated isoproterenol injection produced a significant decrease in the maximum number of binding sites (Bmax), but not in the dissociation constant (KD) for this antagonist. In comparison to the isoproterenol-induced changes in beta adrenergic positive inotropic response, repeated treatment with isoproterenol decreased 2-fold the EC50 value for the negative inotropic response to ACh in isolated atria. Scatchard analysis of specific [3H]quinuclidinyl benzilate binding, however, indicated a significant (P less than .01) decrease in Bmax. Repeated isoproterenol injections also elicited a significant increase in cardiac weight. It is suggested that repeated administration of isoproterenol induces a decrease in density of beta adrenergic receptors which may in part underlie a cardiac hyposensitivity to isoproterenol and an increase in the number of muscarinic ACh receptors and inotropic responsiveness to ACh.     

Down regulation of beta adrenergic receptors in S49 lymphoma cells induced by atypical agonists

The ability of the atypical agonists celiprolol and pindolol to induce sequestration and down regulation of beta adrenergic receptors was investigated in S49 lymphoma cells. Sequestration was measured as the loss of binding sites for [3H]CGP-12177, a hydrophilic radioligand that binds only to surface beta adrenergic receptors at 6 degrees C. Down regulation was measured as the loss of binding sites for [125I]iodopindolol, a lipophilic radioligand which at 37 degrees C binds to both surface and sequestered receptors. Pindolol and celiprolol do not stimulate adenylate cyclase in membranes from wild-type (WT) S49 cells or do they induce the sequestration of beta adrenergic receptors on intact cells. Incubation of WT S49 lymphoma cells with isoproterenol for 24 hr resulted in the loss of 75% of total cellular beta adrenergic receptors (down regulation). Exposure of WT S49 cells to pindolol or celiprolol for 24 hr resulted in the loss of approximately half of the total cellular beta adrenergic receptors. In mutant S49 cells [cyc- (variant of S49 lymphoma cells which lacks Ns activity) and UNC (variant of S49 lymphoma cells in which Ns is present but cannot interact with beta adrenergic receptors)] in which interaction of beta adrenergic receptors with the stimulatory guanine nucleotide binding regulatory protein (Ns) does not occur, a 24 hr incubation with isoproterenol caused the loss of approximately half of the beta adrenergic receptors, whereas pindolol and celiprolol caused no change in the number of receptors. These results suggest that there are two mechanisms of down regulation of beta adrenergic receptors in S49 cells.(ABSTRACT TRUNCATED AT 250 WORDS)     

Modulation of beta adrenergic responsiveness by arachidonic acid metabolites in isolated bovine coronary arteries

The present study was designed to determine whether interference with endogenous arachidonic acid metabolism or exogenously administered cyclooxygenase and lipoxygenase products affects the relaxation of bovine coronary artery in response to isoproterenol. Rings of bovine coronary artery were suspended for isometric tension recordings in organ chambers filled with Krebs-Ringer bicarbonate solution (37 degrees C) gassed with 95% O2-5% CO2 (pH 7.4). In depolarized coronary artery rings (35 mM KCI) isoproterenol induced a dose-dependent relaxation, which was significantly augmented by the cyclooxygenase inhibitor indomethacin and depressed by arachidonic acid. The mixed lipoxygenase/cyclooxygenase inhibitor phenidone or the lipoxygenase products leukotriene D4 and C4 did not affect beta adrenergic responsiveness. Phenidone antagonized the facilitatory action of indomethacin. Exogenous arachidonic acid in the presence of indomethacin and phenidone depressed the relaxation induced by isoproterenol. Prostacyclin and prostaglandin E2 reduced beta adrenergic responsiveness, which was not affected by indomethacin. The data suggest that arachidonic acid depresses beta adrenergic responsiveness in the bovine coronary artery via cyclooxygenase and some noncyclooxygenase, nonlipoxygenase metabolites. Lipoxygenase products, other than leukotrienes D and C, may have a facilitatory action.     

Influence of age on relaxation of rat blood vessels to atriopeptin II

The ability of atriopeptin II to relax isolated ring segments of the thoracic aorta and renal, mesenteric and carotid arteries was studied in Fischer 344 rats aged 6, 12, 20 and 24 to 27 months. Vessels were precontracted with norepinephrine (aorta, mesenteric and carotid arteries) or serotonin (renal artery) in concentrations that produced approximately 50% of the maximal contractile response. Agonist concentrations used and magnitude of induced tone did not change with age, except in the carotid artery, where tone induced in vessels from the oldest group of animals was significantly greater. Thus it was possible to obtain contractile responses that were relatively stable with age in order to test the effect of age on relaxation responses effectively. There were significant differences in sensitivity to atriopeptin II and maximum relaxation among the vessels studied when comparisons were made within a single age group. Thus the thoracic aorta and carotid artery were more sensitive to atriopeptin II and had greater maximum relaxation responses than the renal and mesenteric arteries. In terms of the influence of age, for the most part neither sensitivity to atriopeptin II nor maximum relaxation response changed with age. However, there were some exceptions. Particularly in the thoracic aorta at several concentrations of atriopeptin II, relaxation in vessels from animals 12 months of age was significantly greater than in the other age groups. This difference is most likely due to an effect of maturation, as there was no further alteration in atriopeptin sensitivity into senescence.(ABSTRACT TRUNCATED AT 250 WORDS)     

Age-related changes in relaxant response of vascular smooth muscles to atrial natriuretic peptide

The effect of aging on responses of vascular smooth muscles to atrial natriuretic peptide (ANP) and other vasodilator substances was investigated in isolated rat aortae, rat renal arteries and monkey renal arteries which were precontracted with norepinephrine. There was no significant difference in the ANP-induced maximum relaxation between young and old rat aortae. However, the concentration of agonists causing a 50% relaxation (ED50) value for the old rats was 7.3 times greater than that for the young ones. In rat and monkey renal arteries, the ED50 ratios were 6.2 and 3.8, respectively. The relaxant responses of the rat aortae to isoproterenol and acetylcholine also decreased with increasing age. The ED50 ratios for isoproterenol and acetylcholine were more than 40 and 17, respectively. The maximum relaxation induced by 10(-5) M isoproterenol also decreased significantly in the aortae from the older rats. On the other hand, the ED50 for nitroprusside, nifedipine- and potassium-induced relaxation was not affected by increasing age. These results suggest that ANP-induced relaxation of vascular smooth muscles is reduced with increasing age in rat aortae, rat renal arteries and monkey renal arteries. The mechanisms by which the ANP-induced relaxation decreased in association with the aging process may be quite different from those in acetylcholine-induced and beta adrenoceptor-induced relaxation.     

Effects of protein kinase inhibitor, 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine, on beta-2 adrenergic receptor activation and desensitization in intact human lymphocytes

To investigate the role of protein kinases in agonist-mediated beta-2 adrenergic receptor regulation, the effects of the protein kinase A and C inhibitor, H-7 [1-(5-isoquinolinylsulfonyl)-2-methylpiperazine], on isoproterenol-induced beta adrenoceptor activation and desensitization have been studied in intact human lymphocytes. In the presence of the phosphodiesterase inhibitor, 3-isobutyl-1-methylxanthine, H-7 potentiated 10(-8) to 10(-4) M isoproterenol or prostaglandin E1-induced cyclic AMP (cAMP) accumulation in a dose-dependent manner. We failed to observe any effect of H-7 on forskolin-induced cAMP accumulation. These effects of H-7 are probably not due to its inhibition of phosphodiesterase. In addition, whereas up to 10(-3) M H-7 had no beta adrenergic receptor blocking effect, preincubation of intact cells with 10(-3.5) M H-7 partially prevented 50 nM isoproterenol-induced beta-2 adrenergic receptor desensitization in terms of decreases in beta adrenoceptor density (maximum binding), isoproterenol-mediated cAMP responsiveness and high affinity receptor binding for agonist. Interestingly, 10(-3.5) M H-7 alone treated cells also showed an up-regulation of cell surface beta receptor density (maximum binding) and increased cAMP responsiveness to isoproterenol stimulation. The mechanisms are unclear. If these effects occur as a result of inhibition by H-7 of protein kinase A and/or C, it may suggest an important role of protein kinase A and/or C in agonist-induced beta-2 adrenergic receptor regulation.     

Adrenergic regulation of glycogenolysis in rat liver after cholestasis. Modulation of the balance between alpha 1 and beta 2 receptors.

The effects of extrahepatic cholestasis upon adrenergic regulation of glycogenolysis and upon the numbers of adrenoceptors in rat liver were studied using isolated hepatocytes and plasma membranes, respectively. A 60% decrease in the number of alpha 1 adrenoceptors (285 vs. 680 fmol/mg protein) and a simultaneous 2.7-fold increase in the number of beta adrenergic sites (67 vs. 25 fmol/mg protein) were observed beginning 36 h after bile flow obstruction and persisted for at least 68 h. The reciprocal modification of the numbers of alpha 1 and beta adrenoceptors was accompanied by a change in the manner of stimulation of glycogen phosphorylase by catecholamines in hepatocytes; originally alpha 1 adrenergic in normal rats (phenylephrine Ka = 0.9 microM, isoproterenol Ka = 7.1 microM), the stimulation became predominantly beta adrenergic in cholestatic animals (phenylephrine Ka = 3.7 microM, isoproterenol Ka = 0.06 microM). In normal rats, activation of the enzyme by epinephrine was inhibited by the alpha blocker phentolamine, without inhibition by the beta blocker propranolol. In contrast, propranolol was more effective than phentolamine in cholestatic rat hepatocytes. Modification of the regulation of glycogenolysis after cholestasis did not seem to be secondary to an alteration in the metabolism of thyroid hormones or in the action of glucocorticoids. However, cholestasis provoked a 10-fold increase in the number of hepatic mitoses and in the incorporation of thymidine into liver DNA of cholestatic animals. Similar changes were observed in regenerating livers, following two-thirds hepatectomy. We propose that the changes following extrahepatic cholestasis might, as well, be explained by a regenerative process.     

Rat jugular vein relaxes to norepinephrine, phenylephrine and histamine.

Circular muscle of the rat external jugular vein contracted to serotonin, angiotensin and potassium chloride but not to norepinephrine, phenylephrine, histamine or carbamylcholine. In contrast, rabbit and guinea-pig jugular veins contracted to norepinephrine, phenylephrine and histamine, although contractions to norepinephrine were small in guinea-pig jugular veins. Norepinephrine, phenylephrine and histamine produced a concentration-dependent sustained relaxation of serotonin-induced contractions in the rat jugular vein, as did isoproterenol, nitroglycerin and papaverine. Propranolol blocked relaxation to norepinephrine, phenylephrine and isoproterenol whereas metiamide, a H2 receptor antagonist blocked relaxation to histamine. alpha adrenergic receptor blockade with phentolamine or prazosin resulted in greater relaxation to norepinephrine whereas cocaine did not enhance norepinephrine-induced vasodilation. This study supports the premise that norepinephrine may exert prominent beta adrenergic receptor stimulation in some blood vessels and that this effect may be more apparent in veins than arteries.     

Hypothyroidism modulates beta adrenergic receptor adenylate cyclase interactions in rat reticulocytes.

We have investigated alterations in beta adrenergic receptor binding sites of rat reticulocytes occurring in animals rendered hypothyroid by thyroidectomy. Beta adrenergic receptor interactions were assessed by measuring the number of (-)[3H]-dihydroalprenolol binding sites and the ability of an agonist to compete for occupancy of the receptors. The number of receptors was significantly reduced in cells from the hypothyroid animals. In addition, there were significant agonist-specific alterations in binding. Using computer assisted curve fitting techniques, it was found that the ability of (-)isoproterenol to stabilize a high affinity guanine nucleotide sensitive "coupled" form of the receptor was impaired. Reticulocytes from hypothyroid animals have, in addition, a reduction in the concentration of the nucleotide regulatory protein as assessed by the number of 42,000 Mr substrates for cholera toxin catalyzed ADP ribosylation. These alterations are associated with reductions in catecholamine and NaF stimulated adenylate cyclase activity. Diminished coupling of beta adrenergic receptors with other regulatory components of the adenylate cyclase system represents a mechanism by which altered thyroid states modulate beta adrenergic receptor function and beta adrenergic responsiveness of tissues.