Effects of diethylcarbamazine on the motility of Angiostrongylus cantonensis and Dirofilaria immitis

Effects of piperazine derivatives, especially of diethylcarbamazine (DEC) on adult Angiostrongylus cantonensis and Dirofilaria immitis were examined. Piperazine (3 X 10(-5)-10(-4) M) paralyzed A. cantonensis and the action was antagonized by picrotoxin. 1,1-dimethyl-4-phenylpiperazinium iodide (DMPP) (10(-5)-10(-4) M) caused contraction but little effect was produced by strychnine. An inhibitory effect on untreated preparations was caused by lower concentrations (3 X 10(-6)-10(-5) M) of diethylcarbamazine citrate (DEC) and also on the preparations contracted by eserine. A stimulatory effect was also seen when higher concentrations (10(-4)-3 X 10(-4) M) of this drug were applied to both preparations. The inhibitory action of DEC was antagonized by gabergic antagonists such as picrotoxin and bicuculline, but not by alpha-adrenergic antagonists like dibenamine and phentolamine. When the worm preparation was paralyzed by strychnine or hexylresorcinol (inhibitors of the release of acetylcholine in this worm), the stimulatory effect of DEC was blocked, but pyrantel (a nicotinic cholinergic agonist) contracted the paralyzed preparation. However, the effect of DEC on D. immitis (10(-7)-3 X 10(-4) M) was inhibitory, and this action was also antagonized by picrotoxin. These results suggest that the DEC inhibitory and stimulatory action is through the gabergic and cholinergic mechanisms in adult A. cantonensis and D. immitis.     

Assessment of the role of Ca2+ mobilization from intracellular pool(s), using dantrolene, in the glycogenolytic action of alpha-adrenergic stimulation in perfused rat liver

To identify the role of Ca2+ mobilization from intracellular pool(s) in the action of alpha-adrenergic agonist, the effects of dantrolene on phenylephrine-induced glycogenolysis were investigated in perfused rat liver. Dantrolene (5 X 10(-5) M) inhibited both glycogenolysis and 45Ca efflux induced by 5 X 10(-7) M phenylephrine. The inhibition by dantrolene was observed in the presence and absence of perfusate calcium. In contrast, dantrolene did not inhibit glycogenolysis induced by glucagon. To confirm the specificity of dantrolene action on calcium release in liver, experiments were also carried out using isolated hepatocytes. Dantrolene did not affect phenylephrine-induced production of inositol 1,4,5-trisphosphate. The compound did inhibit a rise in cytoplasmic Ca2+ concentration induced by phenylephrine both in the presence and absence of extracellular Ca2+. Thus, these results suggest that calcium release from an intracellular pool is essential for the initiation of alpha-adrenergic stimulation of glycogenolysis in the perfused rat liver.     

Cholinergic neuromodulation by prostaglandin D2 in canine airway smooth muscle

To determine whether prostaglandin D2 (PGD2) modulates cholinergic neurotransmission in airway smooth muscle and, if so, what the mechanism of action is, we studied bronchial segments from dogs under isometric conditions in vitro. PGD2 (10(-8)-10(-5) M) elicited dose-dependent muscle contraction, which was reduced after blockade of muscarinic receptors, so that 50% effective dose (ED50) increased from 1.3 +/- 0.3 X 10(-6) to 3.9 +/- 1.0 X 10(-6) M by atropine (10(-6) M) (mean +/- SE, P less than 0.05). Physostigmine, at a concentration insufficient to alter base-line tension (10(-8) M), enhanced the PGD2-induced contraction and decreased ED50 to 6.4 +/- 0.5 X 10(-7) M (P less than 0.05). When added at the highest doses that did not cause spontaneous contraction (1.9 +/- 0.5 X 10(-7) M), PGD2 increased the contractile response to electrical field stimulation (1-50 Hz) by 21.9 +/- 6.6% (P less than 0.001). In contrast to this effect, the response to administered acetylcholine was not affected by PGD2. On the other hand, PGD2-induced augmentation of the response to electrical field stimulation (5 Hz) was further increased from 23.6 +/- 3.0 to 70.4 +/- 8.8% in the presence of physostigmine (10(-8) M) and was abolished by atropine but not affected by the alpha-adrenergic antagonist phentolamine or the histamine H1-blocker pyrilamine. These results suggest that the contraction of airway smooth muscle induced by PGD2 is in in part mediated by a cholinergic action and that PGD2 prejunctionally augments the parasympathetic contractile response, likely involving the accelerated release of acetylcholine at the neuromuscular junction.     

Alpha-adrenergic inhibition of cyclic AMP accumulation in hamster adipocytes. Similarity of receptor with alpha-2 adrenergic receptors

The present communication shows the effects of several alpha-adrenergic agonists and antagonists on cyclic AMP levels in hamster epididymal adipocytes. In response to ACTH (30 mU/ml) in combination with 1-methyl-3-isobutylxanthine (0.10 mM) or adenosine deaminase (1.0 micrograms/ml), cyclic AMP levels increased to a maximum by 10 min and this level was maintained for another 20 min. Elevated cyclic AMP levels were partially suppressed by the alpha-adrenergic agents clonidine, methoxamine, methyl norepinephrine and phenylephrine. The lowest effective concentration of each of these agonists required to suppress cyclic AMP levels was 10 nM clonidine; 3 microM methoxamine; 10 microM methyl norepinephrine; 10 microM phenylephrine. Clonidine and methoxamine suppressed cyclic AMP levels by nearly 65% while phenylephrine and methyl norepinephrine caused only a 30% decline. Studies of the relative potencies of alpha-adrenergic blocking drugs on prevention of the inhibitor effect of clonidine on cyclic AMP levels disclosed that phentolamine and yohimbine were more potent blockers of clonidine action than phenoxybenzamine and prazosin. The rank order of potencies of agonists at causing suppression of cyclic AMP levels and the rank order of potencies of antagonists of clonidine action suggest similarity of the alpha-adrenergic receptors present on hamster adipocytes, which affect cyclic AMP accumulation to alpha-2 adrenergic receptors.     

Comparative effect of atropine on the adrenergic and muscarinic stimulation of phospholipid 32P labelling in isolated parotid cells: atropine, a possible blocker of alpha-adrenergic receptors

The inhibitory effect of atropine on phospholipid 32P labelling stimulated by muscarinic or alpha-adrenergic agonists was studied in isolated parotid cells. Atropine (10(-11) to 10(-4) M) had no effect on phospholipid 32P labelling in unstimulated cells. In contrast, 10(-8) to 10(-7) M atropine provoked a competitive inhibition of the cholinergic stimulation (i.e. this effect was completely wiped out at high agonist concentration). The atropine app. KD for the muscarinic receptor was 5 X 10(-9) M. Moreover, atropine inhibits the adrenergic stimulation of phospholipid 32P labelling by decreasing the efficacity and potency of the adrenergic agonists. The atropine app. KD for the alpha-adrenergic receptor can be estimated at 10(-5) M. This inhibition of alpha-adrenergic stimulation appears to be specific since atropine was without effect on the substance P or beta-adrenergic stimulation. At very low concentration (10(-10) - 10(-9) M) atropine seems to be a modulator (activator) of the muscarinic or adrenergic agonist-receptor complex. From the present data, it is suggested that atropine, besides its classical blocker effect at the muscarinic receptor, at high concentration is a specific alpha-adrenergic antagonist.     

Effect of phenylephrine on lipolysis in rat adipocytes: no evidence for an alpha-adrenergic mechanism

Phenylephrine, a strong alpha 1-adrenergic agonist, exerted a concentration dependent antilipolytic effect against isoproterenol-activated lipolysis in rat adipocytes with the effect decreasing as the isoproterenol concentration increased. The alpha-adrenergic antagonists phentolamine and phenoxybenzamine did not reverse phenylephrine's antilipolytic effect. Phenylephrine alone activated lipolysis at concentrations above 10(-5) M and at 5 X 10(-4) M the rate of lipolysis was increased 3.4-fold. Propranolol abolished this effect. In the presence of sub-maximum concentrations of dibutyryl cyclic-AMP (less than 10(-4) M), 10(-4) M phenylephrine increased the rate of lipolysis above that activated by dibutyryl cyclic-AMP alone. At maximum dibutyryl cyclic-AMP concentrations, or in the presence of propranolol, phenylephrine had no effect on dibutyryl cyclic-AMP-dependent lipolysis. There is no evidence to support an alpha 1-adrenergic mechanism for regulation of lipolysis in the rat adipocyte. All effects of the alpha-adrenergic agonist phenylephrine appear to be due to its weak beta-adrenergic activity.     

Role of alpha-adrenergic stimuli in the control of rat renal ammoniagenesis

The effects of phenylephrine on renal ammoniagenesis and the involvement of Ca2+ in phenylephrine action were investigated in isolated proximal fragments of rat-kidney tubules. Phenylephrine stimulated renal ammoniagenesis from 1 and 2 mM glutamine whereas no significant changes took place at a higher concentration of glutamine (20 mM). Stimulation of ammonia synthesis by phenylephrine was found to be linear with time and dose-dependent between 10(-9) and 10(-4) M. Phenylephrine-stimulated ammoniagenesis was blocked by phentolamine (10 microM) but not by propranolol (10 microM) confirming that the effect is mediated by alpha-adrenergic stimuli. No stimulatory effect of phenylephrine was observed in Ca2+ depleted proximal tubule fragments, suggesting that Ca2+ is required in this adrenergic response.     

Effects of epinephrine, clonidine, L-phenylephrine, and morphine on intestinal secretion mediated by Escherichia coli heat-stable enterotoxin in pig jejunum

Perfusion of pig jejunum with Escherichia coli heat-stable enterotoxin (strain 1261) reversed net absorption of water and electrolytes to net secretion. Addition of the alpha-adrenergic agonists clonidine (5 X 10(-7) M) or L-phenylephrine (5 X 10(-6) M), or the opiate agonist morphine (3.6 X 10(-6) M) to the perfusate reduced the secretory response to enterotoxin and stimulated absorption in normal jejunum. Epinephrine (5 X 10(-5) M) did not stimulate absorption in controls but reduced chloride loss in the presence of enterotoxin. Mucosal sodium--potassium adenosine triphosphatase was unchanged but disaccharidase activity was decreased in the presence of enterotoxin. The results suggest that alpha-adrenergic agonists and opiate agonists may exert an antidiarrheal action by increasing net transport across intestinal epithelium.     

Presynaptic alpha-adrenoceptor mediated inhibition of the neurogenic cholinergic contraction in the isolated intestinal bulb of the carp (Cyprinus carpio)

The effects of norepinephrine, epinephrine and clonidine on neurogenic cholinergic contraction were examined in the presence of a beta-adrenoceptor blocking agent, carteolol (5 X 10(-6) M), in the isolated intestinal bulb of the carp. Norepinephrine, epinephrine (10(-9)-10(-6) M) and clonidine (10(-8)-10(-5) M) inhibited the contraction induced by low frequency (2 or 5 Hz) transmural stimulation (TMS) without inhibiting the contraction induced by acetylcholine (ACh, 6 X 10(-8)-4 X 10(-7) M). Methoxamine (10(-4) M) and phenylephrine (10(-4) M) showed no such inhibitory effect on the TMS-induced contraction. The inhibitory effects of catecholamines and clonidine were decreased by phentolamine (5.4 X 10(-6) M) and yohimbine (10(-7)-10(-6) M) but not by prazosin (7 X 10(-7)-10(-6) M). Nicotine (10(-6)-10(-4) M) and serotonin (3 X 10(-8)-3 X 10(-6) M) caused contraction of the intestinal bulb indirectly by releasing endogenous ACh. This contraction was inhibited by norepinephrine, epinephrine and clonidine in a concentration-dependent manner. The present results suggest that catecholamines and clonidine inhibit cholinergic transmission via the activation of a presynaptic alpha-adrenoceptor (presumably of alpha-2 type) located on the cholinergic nerve terminals innervating the smooth muscle of the intestinal bulb of the carp.     

Inhibitory effect of [Asu1 . 7]-eel calcitonin on glucagon-induced glycogenolysis in perfused rat liver

In an attempt to elucidate the mechanism by which calcitonin modulates glucose metabolism, the effect of elcatonin ([Asu1 . 7]-eel calcitonin), a potent synthetic eel calcitonin analogue, on hepatic glycogenolysis was investigated by use of perfused liver from fed rats. Elcatonin, as infused into the portal vein at concentrations between 10 mU/ml and 200 mU/ml did not affect glucose output into the hepatic venous effluent. At concentrations higher than 100 mU/ml, it inhibited the glycogenolysis stimulated by submaximal concentrations of glucagon which was perfused concurrently. This aspect of elcatonin effect was reproduced by synthetic salmon calcitonin. Though elcatonin showed a marked inhibition of the glycogenolytic activity induced by glucagon at or less than 5.7 X 10(-11) M, the inhibitory effect became undetectable when higher concentrations of glucagon were employed. Elcatonin did not inhibit the glycogenolysis induced by dibutyryl cyclic AMP at near (0.5 microM) or less than half-maximally effective (0.2 microM) concentrations. In addition, it did not inhibit the glycogenolytic activity half-maximally stimulated by alpha-adrenergic agonist (phenylephrine, 0.4 microM) or vasopressin (0.2 mU/ml). Elcatonin inhibited the cyclic AMP production of the tissue induced by glucagon infusion. These data indicate that elcatonin modulates hepatic glycogenolysis by preventing the glucagon effect at a step before cyclic AMP production and with an apparently competitive kinetics. In view of the concept that Ca++ is involved in the glycogenolytic effect of alpha-adrenergic agonist and vasopressin, the fact that elcatonin did not influence the action of these agents suggests that Ca++ fluxes are not involved in the elcatonin effect on liver.     

Ivermectin, an unconventional agonist of the glycine receptor chloride channel

The effects of the antihelmintic, ivermectin, were investigated in recombinantly expressed human alpha(1) homomeric and alpha(1)beta heteromeric glycine receptors (GlyRs). At low (0.03 microm) concentrations ivermectin potentiated the response to sub-saturating glycine concentrations, and at higher (> or =0.03 microm) concentrations it irreversibly activated both alpha(1) homomeric and alpha(1)beta heteromeric GlyRs. Relative to glycine-gated currents, ivermectin-gated currents exhibited a dramatically reduced sensitivity to inhibition by strychnine, picrotoxin, and zinc. The insensitivity to strychnine could not be explained by ivermectin preventing the access of strychnine to its binding site. Furthermore, the elimination of a known glycine- and strychnine-binding site by site-directed mutagenesis had little effect on ivermectin sensitivity, demonstrating that the ivermectin- and glycine-binding sites were not identical. Ivermectin strongly and irreversibly activated a fast-desensitizing mutant GlyR after it had been completely desensitized by a saturating concentration of glycine. Finally, a mutation known to impair dramatically the glycine signal transduction mechanism had little effect on the apparent affinity or efficacy of ivermectin. Together, these findings indicate that ivermectin activates the GlyR by a novel mechanism.     

Unidirectional actions of insulin and Ca2+-dependent hormones on adipocyte pyruvate dehydrogenase

Norepinephrine and epinephrine, in the presence of the beta-adrenergic antagonist propranolol (10(-5) M), stimulated adipocyte pyruvate dehydrogenase at low concentrations but inhibited the enzyme at higher concentrations. The alpha-adrenergic agonist, phenylephrine, rapidly stimulated pyruvate dehydrogenase activity in a dose-dependent manner with maximal stimulation observed at 10(-6) M. The stimulation of pyruvate dehydrogenase by phenylephrine was mediated via alpha 1-receptors. Inhibition of pyruvate dehydrogenase by catecholamines was mediated via beta-adrenergic receptors, since the beta-agonist, isoproterenol, and dibutyryl cAMP produced similar effects. Like insulin, alpha-adrenergic agonists increased the active form of pyruvate dehydrogenase without changing the total enzyme activity and cellular ATP concentration. The effects induced by maximally effective concentrations of insulin and alpha-adrenergic agonists were nonadditive. The ability of phenylephrine and methoxamine to stimulate pyruvate dehydrogenase and phosphorylase and to inhibit glycogen synthase was not affected by the removal of extracellular Ca2+. Similarly, the stimulation of pyruvate dehydrogenase and glycogen synthase by insulin was also observed under the same conditions. However, when intracellular adipocyte Ca2+ was depleted by incubating cells in a Ca2+-free buffer containing 1 mM ethylene glycol bis(beta-amino-ethyl ether)-N,N,N' -tetraacetic acid, the actions of alpha-adrenergic agonists, but not insulin, on pyruvate dehydrogenase were completely abolished. Vasopressin and angiotensin II also stimulated pyruvate dehydrogenase in a dose-dependent manner with enhancement of glucose oxidation and lipogenesis. Our results demonstrate that the Ca2+ -dependent hormones stimulate pyruvate dehydrogenase and lipogenesis in isolated rat adipocytes, and the action is dependent upon intracellular, but not extracellular, Ca2+.     

Inhibition of lipolysis in hamster adipocytes with selective alpha-adrenergic stimuli. Functional characterization of the alpha-receptor

This communication shows the relative potencies of the alpha-agonists clonidine, methoxamine, methyl norepinephrine and phenylephrine in producing inhibition of lipolysis. At cell densities greater than 15 mg cell/ml lipolysis activated by either 1-methyl-3-isobutyl xanthine or adenosine deaminase was inhibited by alpha-adrenergic stimuli with a rank order of potency of clonidine greater than methoxamine greater than methyl norepinephrine; phenylephrine produced a further stimulation of lipolysis. At the same cell density isoproterenol-accelerated lipolysis was inhibited by alpha-adrenergic stimuli with a rank order of potency of phenylephrine greater than methoxamine greater than clonidine greater than methyl norepinephrine. When the density of fat cells was reduced to less than 5 mg/ml, clonidine was a more effective inhibitor of isoproterenol-activated lipolysis thatn phenylephrine. Lipolysis that was activated by dibutyryl cyclic AMP, ACTH or cholera enterotoxin was not reduced by any alpha-adrenergic agent. Under conditions when clonidine failed to inhibit catecholamine-activated lipolysis (i.e., at cell densities greater than 15 mg/ml), it failed to antagonize the antilipolytic activity of phenylephrine. The antilipolytic activities of clonidine and phenylephrine were most effectively antagonized by the blocking drugs phentolamine and yohimbine; in contrast, phenoxybenzamine and prazosin were less effective blockers. These data indicate that the alpha-adrenergic receptor on hamster fat cells is similar to presynaptic alpha-adrenergic receptors. The data further suggest the possibility that phenylephrine may exert its action through a separate alpha-adrenergic receptor mechanism.     

Relationships among airway reactivity, pupillary alpha-adrenergic and cholinergic responsiveness, and age

Healthy adult volunteers (n = 122), who denied personal history of lung disease or family history of cystic fibrosis or asthma, took no interfering medications, and had forced expiratory volume in 1 s greater than or equal to 80% predicted, underwent methacholine challenge and pupillary reactivity testing. Pupil diameter measured in dark and light test conditions declined with age (Pearson's r = -0.54 and -0.36). Pupillary alpha-adrenergic responsiveness (expressed as the concentration of phenylephrine required to dilate the pupil 1 mm) was significantly correlated with age. Older subjects required lower concentrations for dilation and therefore were more sensitive to phenylephrine. Pupillary cholinergic responsiveness (the concentration of carbachol required for 1-mm constriction) was not significantly correlated with age. Therefore the significantly smaller baseline pupil size in the elderly cannot be explained by failure of alpha-adrenergic receptor responses or by increased pupillary cholinergic responsiveness. We found no significant correlation of methacholine bronchial reactivity with age. In addition, there was no relation between airway reactivity and pupillary alpha-adrenergic or cholinergic responsiveness in this sample of healthy adults. These findings, taken with others in the literature, suggest that the contribution of alpha-adrenergic and cholinergic responsiveness to nonspecific airway reactivity in healthy persons is small, if it exists at all, and that there is no significant change in airway reactivity with age in healthy adults.     

Interaction of spin-labeled methacyne analog with butyrylcholinesterase

Interaction between spin-labeled methacyne (I) and butyrylcholinesterase (BChE) was studied by ESR and enzyme kinetic methods. The compound (I) was shown to be a competitive reversible inhibitor, the value of Ki appeared to be 1.3 X 10(-5) M. Insertion of nitroxyl fragment in the methacyne molecule results in a two-fold increase of its inhibitory activity. The ESR spectrum of the enzyme-inhibitor complex was registered. This complex dissociates under the action of eserine, tetramethylammonium and hexamethonium. Scatchard plot reveals two different types of binding sites with Kdiss values 1.5 X 10(-5) M and 2.6 X 10(-4) M. One type of binding sites is identified as the enzyme active centre. The restricted motion of (I) in complex with BChE proves the assumption that the enzyme active centre is located in the split of macromolecule surface.     

Effect of delta sleep-inducing peptide, anticonvulsant preparations and nicotinamide on generalized seizure activity

The influence of delta-sleep inducing peptide (DSIP) upon seizures induced by corazol, bicuculline, picrotoxin, strychnine, thiosemicarbazide were investigated in experiments on F1(CBA X C57 BL/6) mice. It was shown that DSIP increased the latency of first seizure manifestation which were induced by corazol, bicuculline and picrotoxin and also resulted in a suppression of seizure severity of corazol and bicuculline induced seizures. Anticonvulsant action of DSIP was evident under the condition of the mild severity seizures development. The effect of DSIP was mostly pronounced in range of its doses from 10 to 100 mcg/kg. DSIP when combined with phenobarbital, carbamazepine, diphenylhydantoin or nicotinamide enhanced the antiepileptic effects of these anticonvulsant drugs.     

Anticonvulsive effects of neurotropin

The antiepileptic action of neurotropin (Nippon Zoki Pharmaceutied Co. Ltd., Japan) was investigated in acute experiments on rats, mice and cats. The increasing of latent period of the first seizures which were picrotoxin-induced and enhancing of this type of seizures were observed correspondently in 1-24 hours and 8-9 days after intraventricular neurotropin injection. Intraperitoneal neurotropin injection (0.2-0.5 ml) to mice caused dosage-dependent antiseizure action on the model of generalized picrotoxin-induced seizures. +Anticonvulsant++ effect of neurotropin was observed during five days. The intraventricular neurotropin injection to cats led to strychnine (0.1%) induced cortical foci suppression.     

Studies on the alpha-andrenergic activation of hepatic glucose output. II. Investigation of the roles of adenosine 3':5'-monophosphate and adenosine 3':5'-monophosphate-dependent protein kinase in the actions of phenylephrine in isolated hepatocytes.

The effects of the alpha-adrenergic agonist phenylephrine on the levels of adenosine 3':5'-monophosphate (cAMP) and the activity of the cAMP-dependent protein kinase in isolated rat liver parenchymal cells were studied. Cyclic AMP was very slightly (5 to 13%) increased in cells incubated with phenylephrine at a concentration (10(-5) M) which was maximally effective on glycogenolysis and gluconeogenesis. However, the increase was significant only at 5 min. Cyclic AMP levels with 10(-5) M phenylephrine measured at this time were reduced by the beta-adrenergic antagonist propranolol, but were unaffected by the alpha-blocker phenoxybenzamine, indicating that the elevation was due to weak beta activity of the agonist. When doses of glucagon, epinephrine, and phenylephrine which produced the same stimulation of glycogenolysis or gluconeogenesis were added to the same batches of cells, there were marked rises in cAMP with glucagon, minimal increases with epinephrine, and little or no changes with phenylephrine, indicating that the two catecholamine stimulated these processes largely by mechanisms not involving cAMP accumulation. DEAE-cellulose chromatography of homogenates of liver cells revealed two major peaks of cAMP-dependent protein kinase activity. These eluted at similar salt concentrations as the type I and II isozymes from rat heart. Optimal conditions for preservation of hormone effects on the activity of the enzyme in the cells were determined. High concentrations of phenylephrine (10(-5) M and 10(-4) M) produced a small increase (10 tp 16%) in the activity ratio (-cAMP/+cAMP) of the enzyme. This was abolished by propranolol, but not by phenoxybenzamine, indicating that it was due to weak beta activity of the agonist. The increase in the activity ratio of the kinase with 10(-5) M phenylephrine was much smaller than that produced by a glycogenolytically equivalent dose of glucagon. The changes in protein kinase induced by phenylephrine and the blockers and by glucagon were thus consistent with those in cAMP. Theophylline and 1-methyl-3-isobutylxanthine, which inhibit cAMP phosphodiesterase, potentiated the effects of phenylephrine on glycogenolysis and gluconeogenesis. The potentiations were blocked by phenoxybenzamine, but not by propranolol. Methylisobutylxanthine increased the levels of cAMP and enhanced the activation of protein kinase in cells incubated with phenylephrine. These effects were diminished or abolished by propanolol, but were unaffected by phenoxybenzamine. It is concluded from these data that alpha-adrenergic activation of glycogenolysis and gluconeogenesis in isolated rat liver parenchymal cells occurs by mechanisms not involving an increase in total cellular cAMP or activation of the cAMP-dependent protein kinase. The results also show that phosphodiesterase inhibitors potentiate alpha-adrenergic actions in hepatocytes mainly by a mechanism(s) not involving a rise in cAMP.     

Studies on the alpha-adrenergic activation of hepatic glucose output. I. Studies on the alpha-adrenergic activation of phosphorylase and gluconeogenesis and inactivation of glycogen synthase in isolated rat liver parenchymal cells.

Epinephrine and the alpha-adrenergic agonist phenylephrine activated phosphorylase, glycogenolysis, and gluconeogenesis from lactate in a dose-dependent manner in isolated rat liver parenchymal cells. The half-maximally active dose of epinephrine was 10-7 M and of phenylephrine was 10(-6) M. These effects were blocked by alpha-adrenergic antagonists including phenoxybenzamine, but were largely unaffected by beta-adrenergic antagonists including propranolol. Epinephrine caused a transient 2-fold elevation of adenosine 3':5'-monophosphate (cAMP) which was abolished by propranolol and other beta blockers, but was unaffected by phenoxybenzamine and other alpha blockers. Phenoxybenzamine and propranolol were shown to be specific for their respective adrenergic receptors and to not affect the actions of glucagon or exogenous cAMP. Neither epinephrine (10-7 M), phenylephrine (10-5 M), nor glucagon (10-7 M) inactivated glycogen synthase in liver cells from fed rats. When the glycogen synthase activity ratio (-glucose 6-phosphate/+ glucose 6-phosphate) was increased from 0.09 to 0.66 by preincubation of such cells with 40 mM glucose, these agents substantially inactivated the enzyme. Incubation of hepatocytes from fed rats resulted in glycogen depletion which was correlated with an increase in the glycogen synthase activity ratio and a decrease in phosphorylase alpha activity. In hepatocytes from fasted animals, the glycogen synthase activity ratio was 0.32 +/- 0.03, and epinephrine, glucagon, and phenylephrine were able to lower this significantly. The effects of epinephrine and phenylephrine on the enzyme were blocked by phenoxybenzamine, but were largely unaffected by propranolol. Maximal phosphorylase activation in hepatocytes from fasted rats incubated with 10(-5) M phenylephrine preceded the maximal inactivation of glycogen synthase. Addition of glucose rapidly reduced, in a dose-dependent manner, both basal and phenylephrine-elevated phosphorylase alpha activity in hepatocytes prepared from fasted rats. Glucose also increased the glycogen synthase activity ratio, but this effect lagged behind the change in phosphorylase. Phenylephrine (10-5 M) and glucagon (5 x 10(-10) M) decreased by one-half the fall in phosphoryalse alpha activity seen with 10 mM glucose and markedly suppressed the elevation of glycogen synthase activity. The following conclusions are drawn from these findings. (a) The effects of epinephrine and phenylephrine on carbohydrate metabolism in rat liver parenchymal cells are mediated predominantly by alpha-adrenergic receptors. (b) Stimulation of these receptors by epinephrine or phenylephrine results in activation of phosphorylase and gluconeogenesis and inactivation of glycogen synthase by mechanisms not involving an increase in cellular cAMP. (c) Activation of beta-adrenergic receptors by epinephrine leads to the accumulation of cAMP, but this is associated with minimal activation of phosphorylase or inactivation of glycogen synthase...     

Fasciola hepatica: the effects of neuropharmacological agents upon in vitro motility

The effects of a wide range of neuropharmacological agents on the motility in vitro of Fasciola hepatica have been determined using an isometric transducer system. The neuromuscular blocking agents tubocurarine and decamethonium cause a long-term stimulation of the basal activity of the fluke. Acetylcholine causes an inhibition of activity. This effect is mimicked by the cholinergic agonists carbachol and nicotine, antagonised by the cholinergic blocking agents atropine and mecamylamine, and potentiated by eserine, a cholinesterase inhibitor. With nicotine and atropine the effects are accompanied by an increase in muscle tone at a concentration of 1 X 10(-2) M. Noradrenaline and adrenaline also cause some inhibition of activity, an effect antagonised by guanethidine, which blocks the release of noradrenaline. In contrast, dopamine stimulates fluke motility, whilst its antagonist dihydroergotamine causes an inhibition of activity. The monoamine oxidase inhibitors iproniazid and p-chloromercuribenzoic acid induce a stimulation of activity; with the latter there is an increase in muscle tone at a concentration of 1 X 10(-3) M. The amine depleting agents chloroamphetamine and reserpine, and the monoamine uptake inhibitors desipramine and nortriptyline produce an inhibition of fluke activity, as does the serotonin uptake inhibitor fluoxetine. High concentrations of chloroamphetamine (1 X 10(-2) M) and the uptake inhibitors (1 X 10(-3) M and above) also induce an increase in muscle tone. Serotonin causes a marked stimulation of motility. The pharmacological evidence is consistent with a neurotransmitter role of acetylcholine (inhibitory), dopamine (excitatory), and noradrenaline (inhibitory). The status of serotonin is discussed.