"Pertussis toxin induces tachycardia and impairs the increase in blood pressure produced by alpha 2-adrenergic agonists"

Administration of purified pertussis toxin to rats induced persistent tachycardia, (observed in conscious rats but not after pithing); as little as 0.05 microgram/100 g produced a significant effect. Pertussis toxin-treatment did not affected the pressor response produced in the pithed rats by the alpha 2-adrenergic agonist methoxamine but markedly diminished the pressor effect of the alpha 2-adrenergic agonists clonidine and azepexole. A role of adenylate cyclase inhibition in the action of postsynaptic vascular alpha 2-adrenergic receptors is suggested.     

Functional alpha 2-adrenoceptors in rat adipocytes: inhibition of cyclic AMP accumulation

Alpha 2-adrenoceptor activation inhibits cyclic AMP accumulation in fat cells from many species. However, the presence of alpha 2-adrenoceptors in rat adipocytes has been difficult to demonstrate. We observed that alpha 2-adrenergic activation inhibits forskolin-stimulated cyclic AMP accumulation both in rat and hamster adipocytes; UK 14304, p-amino clonidine and clonidine were the agents with higher efficacy. The effect of UK 14304 was blocked by yohimbine but not by prazosin demonstrating the involvement of alpha 2-adrenoceptors. Pertussis toxin blocked the alpha 2-adrenergic effect. Our results demonstrate the presence in rat fat cells of alpha 2-adrenoceptors coupled to adenylate cyclase via "Gi".     

Cycloheximide: An adrenergic agent

Cycloheximide, a widely used inhibitor of protein synthesis, stimulates glycogenolysis, gluconeogenesis and ureogenesis in isolated rat hepatocytes. The effects of cycloheximide were compared to those of norepinephrine. Both agents, cycloheximide and norepinephrine, produced slight increases in the levels of cyclic AMP (30% increases) which were blocked by propranolol. Interestingly, it was found that the metabolic actions of norepinephrine and cycloheximide (stimulation of glycogenolysis, gluconeogenesis and ureogenesis) were only slightly diminished by the β adrenergic antagonist propranolol but abolished by the selective α₁ adrenergic antagonist prazosin. The ability of cycloheximide to inhibit protein synthesis was not affected by either prazosin or propranolol. It is concluded that the stimulation of glycogenolysis, gluconeogenesis and ureogenesis by cycloheximide in rat hepatocytes, is an effect of the antibiotic independent of its ability to inhibit protein synthesis and that is mediated through activation of α₁ adrenoceptors. The adrenergic activity of cycloheximide should be considered when this drug is used as an inhibitor of protein synthesis.     

Role of alpha 1 adrenoceptors in the turnover of phosphatidylinositol and of alpha 2 adrenoceptors in the regulation of cyclic AMP accumulation in hamster adipocytes

The incorporation of radioactive phosphate into phosphatidylinositol was stimulated by epinephrine in hamster fat cells. This action was inhibited by alpha-adrenergic antagonists in the potency order: Prazosin?phentolamine>yohimbine. Methoxamine, but not clonidine, was able to mimic the effect of epinephrine. These data indicate that the phosphatidylinositol effect in fat cells is due to activation of alpha₁ adrenoceptors. On the other hand, the accumulation of cyclic AMP due to epinephrine was potentiated by alpha-adrenergic antagonists in the potency order phentolamine>yohimbine ?prazosin, in hamster fat cells. Clonidine significantly decreased the accumulation of cyclic AMP due to isoproterenol or ACTH in hamster fat cells, suggesting that the alpha-adrenergic modulation of cyclic AMP levels in hamster fat cells is mediated by alpha₂ adrenoceptors. Radioligand binding studies with plasma membranes from hamster adipocytes demonstrated the presence of both alpha₁ and alpha₂ adrenoceptors but about 90% of the binding sites were alpha₂. These data support the hypothesis that alpha₂ effects of catecholamines are due to inhibition of adenylate cyclase while the increases in phosphatidylinositol turnover that seem to be involved in the mobilization of calcium are linked exclusively to alpha₁ adrenoceptor activation.     

Differential effect of pertussis toxin on the affinity state for agonists of renal alpha 1- and alpha 2- adrenoceptors

The effect of pertussis toxin treatment on the guanine nucleotide-induced modulation of the affinity of renal alpha 1- and alpha 2-adrenergic receptors was investigated. Pretreatment of rats with pertussis toxin did not induce any change in the number of or affinity for antagonists of alpha 1- or alpha 2-receptors studied using [3H]prazosin and [3H]yohimbine, respectively. Guanyl-5'-yl imidodiphosphate induced an "up-shift" in the number of alpha 2-adrenergic receptors; this up-shift was not observed for alpha 1-adrenergic receptors. Pertussis toxin treatment decreased the affinity of epinephrine for the [3H]yohimbine-binding sites and reduced the ability of guanine nucleotides to modulate alpha 2-adrenoceptor agonist affinity. The regulation by guanine nucleotides of alpha 1-adrenoceptor affinity for agonists was not altered. These results suggest that the modulation of alpha 1- and alpha 2-adrenoceptors by guanine nucleotides is probably exerted through different molecular entities.     

Effect of pertussis toxin on the heart muscarinic-cholinergic receptors and their function

Administration of pertussis toxin to rats induced a significant increase in heart rate that was evident as soon as 24 hours after the administration of the toxin and that persisted for at least 15 days. Electrical stimulation of the vagus decreased dramatically the heart rate of control animals but was unable to do it so in rats treated with pertussis toxin. In cardiac membranes muscarinic agonists decreased adenylate cyclase activity (approximately equal to 20-25%); no effect was observed in membranes obtained from toxin-treated animals. Agonist displacement of antagonist binding [( 3H] Quinuclidinyl benzilate) indicated that treatment with pertussis toxin decreased the proportion of receptors in the high affinity state for agonists. All these data suggest that blockade of the parasympathetic tone plays a key role in the induction of tachycardia by pertussis toxin.     

α1B-Adrenergic Receptors Differentially Associate with Rab Proteins during Homologous and Heterologous Desensitization

Internalization of G protein-coupled receptors can be triggered by agonists or by other stimuli. The process begins within seconds of cell activation and contributes to receptor desensitization. The Rab GTPase family controls endocytosis, vesicular trafficking, and endosomal fusion. Among their remarkable properties is the differential distribution of its members on the surface of various organelles. In the endocytic pathway, Rab 5 controls traffic from the plasma membrane to early endosomes, whereas Rab 4 and Rab 11 regulate rapid and slow recycling from early endosomes to the plasma membrane, respectively. Moreover, Rab 7 and Rab 9 regulate the traffic from late endosomes to lysosomes and recycling to the trans-Golgi. We explore the possibility that α_1B-adrenergic receptor internalization induced by agonists (homologous) and by unrelated stimuli (heterologous) could involve different Rab proteins. This possibility was explored by Fluorescence Resonance Energy Transfer (FRET) using cells coexpressing α_1B-adrenergic receptors tagged with the red fluorescent protein, DsRed, and different Rab proteins tagged with the green fluorescent protein. It was observed that when α_1B-adrenergic receptors were stimulated with noradrenaline, the receptors interacted with proteins present in early endosomes, such as the early endosomes antigen 1, Rab 5, Rab 4, and Rab 11 but not with late endosome markers, such as Rab 9 and Rab 7. In contrast, sphingosine 1-phosphate stimulation induced rapid and transient α_1B-adrenergic receptor interaction of relatively small magnitude with Rab 5 and a more pronounced and sustained one with Rab 9; interaction was also observed with Rab 7. Moreover, the GTPase activity of the Rab proteins appears to be required because no FRET was observed when dominant-negative Rab mutants were employed. These data indicate that α_1B-adrenergic receptors are directed to different endocytic vesicles depending on the desensitization type (homologous vs. heterologous).     

Photosynthetic performance in cyanobacteria with increased sulphide tolerance: an analysis comparing wild-type and experimentally derived strains

Sulphide is proposed to have influenced the evolution of primary stages of oxygenic photosynthesis in cyanobacteria. However, sulphide is toxic to most of the species of this phylum, except for some sulphide-tolerant species showing various sulphide-resistance mechanisms. In a previous study, we found that this tolerance can be induced by environmental sulphidic conditions, in which two experimentally derived strains with an enhanced tolerance to sulphide were obtained from Microcystis aeruginosa, a sensitive species, and Oscillatoria, a sulphide-tolerant genus. We have now analysed the photosynthetic performance of the wild-type and derived strains in the presence of sulphide to shed light on the characteristics underlying the increased tolerance. We checked whether the sulphide tolerance was a result of higher PSII sulphide resistance and/or the induction of sulphide-dependent anoxygenic photosynthesis. We observed that growth, maximum quantum yield, maximum electron transport rate and photosynthetic efficiency in the presence of sulphide were less affected in the derived strains compared to their wild-type counterparts. Nevertheless, in ¹⁴C photoincoporation assays, neither Oscillatoria nor M. aeruginosa exhibited anoxygenic photosynthesis using sulphide as an electron donor. On the other hand, the content of photosynthetic pigments in the derived strains was different to that observed in the wild-type strains. Thus, an enhanced PSII sulphide resistance appears to be behind the increased sulphide tolerance displayed by the experimentally derived strains, as observed in most natural sulphide-tolerant cyanobacterial strains. However, other changes in the photosynthetic machinery cannot be excluded.