Homologous β-adrenergic desensitization in isolated rat hepatocytes (original) (raw)
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European Journal of Pharmacology: Molecular Pharmacology, 1993
Preincubation of rat hepatocytes with isoproterenol induces homologous /3-adrenergic desensitization evidenced both in whole ceils (cyclic AMP accumulation) and membranes (adenylyl cyclase activity). This desensitization is associated with and quantitatively similar to a loss of/32-adrenoceptors from the plasma membrane. Desensitization did not alter the affinities of isoproterenol for the [azsI]iodocyanopindolol binding sites nor reduce the ability of guanine nucleotides to modulate agonist affinity, i.e., the receptors that remain in the surface of plasma membrane after desensitization (= 50%) retain their functional integrity. When membranes from isoproterenol-desensitized hepatocytes were treated with alkaline phosphatase, no attenuation of the desensitization was observed. Cholera toxin-catalyzed ADP-ribosylation was not decreased but rather slightly increased in membranes from desensitized cells as compared to the controls. Our data indicate that in hepatocytes, a loss of/32-adrenoceptors from the plasma membrane is closely associated to the homologous desensitization induced by isoproterenol.
Biochimica et Biophysica Acta (BBA) - Bioenergetics, 1988
Homologoos-heterologous desensitizations: Pertussis toxin in helmtocytes obtained from hypothyroid rats, phorbol myris~te acetate (PMA) and vasepress'm dln6nished the accumulation of cydic AMP and the stimulation of ureagenesis induced by isoprenal~ or Oucagen without altering significantly the accmnalation of cyclic AMP induced by fmskel~ Pretremment wlth PMA markecWy reduced the stimulation o! m~genesis and the accumulation of eyelk AMP induced by ~enaline or glucagon. In membranes from cells pretreated with PMA, the stimulatioa of adenylate cydase induced by isoprenaline + GTP, glucagoa + GTP or by GpplNH]p were dearly ~ as cmupared to the control, whereas forskulin-stimulated activity was not affected. The data indicate ~ desensitization of adenylate cydase, i! was also obser, ~ that the hon~ogoes (Garcia-SMuz J.A. and Michel, B. (1987) Biochem. J. 246, 331-3.36) and this heterologous ~-adrenergie desensitizations were ~e. Pertossis toxin treatment markedly reduced the heterulogous desensitization of adenylate eydase bat n~t the homologous ~-adkenersie desensitization. It is concluded that the homelolpms and heterulegmm tions involve different mechanisms. The homologous desensitization seems to occur at the receptor level, whereas the heterologous probably involves the guanine uudeotide.binding regulatory proteht, Ns.
The α-adrenergic mediated effect in rat liver
Biochemical Pharmacology, 1980
ABSTRACT Alpha-adrenergic receptors of purified rat liver plasma membranes were characterized by the direct binding of [3H]-dihydroergocryptine. Since glycogen phosphorylase (EC 2.4.1.1; 1,4-alpha-d-glucan: orthophosphate alpha-glucosyl transferase) activation in isolated rat hepatocytes is known to be mediated mainly through an alpha-adrenergic mechanism, we compared the abilities of various adrenergic agonists and antagonists in competing with [3H]-dihydroergocryptine for the binding to alphasites in purified membranes and in activating phosphorylase or in inhibiting its activation in isolated hepatocytes. The activation of phosphorylase by agonists displayed an order of potencies typical of alpha-adrenergic mediators: (−)norepinephrine (Ka = 0.31 μM) ⩾ (−)epinephrine (Ka = 0.76 μM) ⪢ (−)isoproterenol (Ka = 24 μM), and the stereospecificity expected: (−)epinephrine (Ka = 0.76 μM) > (+)epinephrine (Ka = 5.2 μM), (−)norepinephrine (Ka = 0.31 μM) > (+)norepinephrine (Ka = 3.3 μM). Alpha-adrenergic antagonists (phentolamine Kiapp =1.71 μM) were much more potent than betaadrenergic antagonists [(−)propranolol Kiapp = 113 μM] in preventing the activation of glycogen phosphorylase by epinephrine. In our system imidazoline derivatives such as clonidine appeared to be mainly antagonists for the alpha-adrenergic mediated events. Close correlations (r = 0.92, P < 0.01 and r = 0.91, P < 0.001) were observed, respectively, for adrenergic agonists and antagonists between either activation or prevention of the activation of the enzyme and competition with dihydroergocryptine for alpha-adrenoreceptors. These results validate the use of [3H]-dihydroergocryptine as a marker of the physiological alpha-adrenergic receptor in rat liver.
FEBS Letters, 1983
Adult male rat hepatocytes, which normally respond poorly to /3-adrenergic agents, acquire such responsiveness during primary monolayer culture. We here show that the rise in catecholamine-sensitive adenylate cyclase activity in hepatocytes in vitro is closely paralleled by an increase in the ability to bind the ,&adrenoceptor ligand [1251]cyanopindolol. The emergence of fl-adrenergic responsiveness did not require cell attachment or serum. Addition of dexamethasone, insulin, thyroxine or dihydrotestosterone to the cultures, singly or in combination, did not prevent the augmented&adrenergic responsiveness. The increase in catecholamine-sensitive adenylate cyclase activity and [r2'I]cyanopindolol binding could be blocked by cycloheximide or actinomycin D. Exposure of the cultures to isoproterenol at 3-hourly intervals led to a dose-dependent suppression of the rise in isoproterenol-responsive adenylate cyclase and prevented the increase in ,&adrenoceptor binding.
Biochimica et biophysica acta. Molecular cell research, 1985
We have previously suggested that the effects of al-adrenergic agents on hepatocyte metabolism involve two pathways: (a) a calcium-independent, insulin-sensitive process which is modulated by glucocorticoids; and (b) a calcium-dependent, insulin-insensitive process which is modulated by thyroid hormones. Cycloheximide stimulated ureogenesis through a prazosin-sensitive mechanism in liver cells (oq-adrenergic). The effect of cycloheximide was insulin-insensitive and calcium-dependent. Furthermore, a clear effect of cycloheximide was observed in hepatocytes obtained from adrenalectomized animals, whereas no effect was observed in cell from hypothyroid rats. The effects of epinephrine and cycloheximide were blocked by phorbol esters in all the conditions tested. Binding competition experiments indicated that cycloheximide interacts with only a fraction of the al-adrenergic sites labeled with [aHlprazosin. It is suggested that cycloheximide activates preferentially one of the pathways involved in the oq-adrenergic action in liver cells.
Journal of Autonomic Pharmacology, 1993
Using agonists (adrenaline, noradrenaline, isoprenaline, dopamine, epinine, procaterol) and antagonists (CGP 20712A and ICI 118,551) in radioligand binding and adenylate cyclase-stimulation studies we demonstrate that the human neuroblastoma cell line SK-N-MC contains a homogeneous population of P,-adrenoceptors coupling to stimulation of adenylate cyclase. 2 A 34 h treatment with isoprenaline reduced P,-adrenoceptor density, concentration-dependently, maximally by 69-+ 1 %. Functional PI-adrenoceptor desensitization was comparatively small. 3 Although 100 nM isoprenaline caused significantly greater receptor downregulation than 1 O,UM isoprenaline (47 * 2%), desensitization of CAMP accumulation in intact cells or of adenylate cyclase stimulation in cell membranes was attenuated similarly or even more with 1 0 ,~~ isoprenaline. 4 While both agonist concentrations did not significantly alter immunodetectable a-subunits for Gi or Go or G-protein P-subunits, 1 0 ,~~ (but not 1 0 0 n~) isoprenaline decreased immunodetectable Gsa. Accordingly, only treatment with 1 O,UM isoprenaline reduced receptor-independent adenylate cyclase stimulation by GTP, NaF or forskolin. Neither agonist concentration desensitized adenylate cyclase activation by MnCl,. 5 We conclude that isoprenaline produces a concentration-dependent regulation of P1-adrenoceptor number in SK-N-MC cells, which is weaker at higher than at intermediate agonist concentrations, and is accompanied by only minor desensitization of adenylate cyclase activity; whether this involves alterations of postreceptor events depends on the agonist concentration used.