Ba++ stimulates accumulation of cyclic AMP in rat pancreatic islets.

In pancreatic islets prelabelled with (³H) adenine, Ba⁺⁺ augmented (³H) cyclic AMP in 1–10 min incubations. 3-isobutyl-l-methylxanthine markedly enhanced and prolonged the Ba⁺⁺-induced nucleotide as well as the insulin response. In the presence of the methyl xanthine 1.6 mM Ba⁺⁺ was a maximally and 0.4 mM a submaximally effective concentration both for the stimulation of (³H) cyclic AMP and insulin. A 5-fold excess of Ca⁺⁺ partly inhibited the Ba⁺⁺-induced nucleotide and — more profoundly — the insulin response. Increasing Mg⁺⁺ from 2 to 10 mM was also inhibitory. Stimulation by Ba⁺⁺ was observed in the absence as well as in the presence of D-glucose. It is concluded that the insulinotropic action of Ba⁺⁺ is at least partly mediated by cyclic AMP.     

Prostaglandin-E2 and cyclic adenosine 3'-5' monophosphate levels in the hypertrophied rat heart.

To assess whether prostaglandin-E2 (PGE2) and cyclic adenosine 3'-5'-monophosphate (cAMP) are involved in the cardiac response to chronic pressure overload, we measured by specific radioimmunoassay method the cardiac tissue and plasma concentrations of PGE2 and cAMP in an animal model of left ventricular hypertrophy. The cardiac hypertrophy was accompanied by a significant increase in PGE2 content, and a significant decrease in cAMP content, in the heart. In addition, we found elevated PGE2 and cAMP levels in arterial plasma samples from the rats with hypertrophied hearts compared to normal rats. These findings suggest a link between cardiac and vascular PGE2 and cAMP generation and the hemodynamic stresses of advanced cardiac overload.     

Studies of the cyclic adenosine monophosphate chemoreceptor ofParamecium

Summary A doublet of proteins (48,000M _r) from theParamecium cell body membrane fits several criteria for the external cAMP chemoreceptor. These criteria include: (i) selective elution from a cAMP affinity column, matching a specificity that could be predicted from the behavioral response and whole-cell binding; (ii) binding to wheat germ agglutinin indicating the presence of carbohydrate moieties indicating surface exposure; and (iii) selective inhibition of the intact cells' chemoresponse to cAMP by antibodies against the doublet. Additional evidence for the existence of a receptor, in general, comes from selective elimination of the cAMP chemoresponse by photoaffinity labeling of whole cells with 8-N₃-cAMP. The doublet proteins are not identical to the regulatory subunit of a cAMP-dependent protein kinase fromParamecium, theDictyostelium cAMP chemoreceptor, or the 42–45 kDa range proteins related to the large surface glycoprotein inParamecium. The doublet proteins are not readily separable and, as inDictyostelium, may represent two different covalent modification states of the same protein. Amino acid analysis indicates that the proteins are similar, but does not distinguish between the possibilities of proteolysis and covalent modification. Once cloned, this doublet may prove to be only the fifth external, eukaryotic chemoreceptor to be identified.     

Decreased K+ conductance produced by Ba++ in frog sartorius fibers

The action of Ba⁺⁺ on membrane potential (E_m) and resistance (R_m) of frog (R. pipiens) sartorius fibers was studied. In normal Cl^- Ringer''s, Ba⁺⁺ (<9 mM) did not depolarize or induce contractions, but increased R_m slightly above the control value of 3.8 ± 0.6 KΩ-cm². In Cl^--free Ringer''s (methane sulfonate) R_m was 28.8 ± 2.8 KΩ-cm², and low concentrations of Ba⁺⁺ (0.05–5.0 mM) depolarized and induced spontaneous contractions (fibrillation), even in tetrodotoxin. To stop disturbance of the microelectrodes, contractions were prevented by using two Cl^--free solutions: (a) twice hypertonic with sucrose (230 mM), or (b) high K⁺ (83 mM) partially replacing Na⁺. In the hypertonic solution, the fiber diameters decreased, E_m increased slightly, and R_m decreased to 9.0 ± 0.6 KΩ-cm² (perhaps due to swelling of sarcotubules). Ba⁺⁺ (0.5 mM) rapidly increased R_m to 31.3 ± 3.8, decreased E_m (e.g., to -30 mv), and induced spontaneous "action potentials;" Sr⁺⁺ had no effect. In the high K⁺ solution, the fibers were nearly completely depolarized, and R_m was decreased markedly to 1.5 ± 0.2 KΩ-cm²; Ba⁺⁺ increased R_m to 6.7 ± 0.5 KΩ-cm². The Ba⁺⁺ actions usually began within 0.5 min and reached a maximum within 5 min. Addition of SO₄ ⁼, to precipitate the Ba⁺⁺, rapidly reversed the increase in R_m. Ba⁺⁺ must act by decreasing K⁺ conductance (g_K). In Cl^- Ringer''s, the high g_Cl/g_K ratio masked the effect of Ba⁺⁺ on g_K. Thus, small concentrations of Ba⁺⁺ specifically and rapidly decrease g_K.     

Regulation of phospholipid synthesis inMycobacterium smegmatis by cyclic adenosine monophosphate

Forskolin, an adenylate cyclase activator and a cyclic AMP analogue, dibutyryl cyclic AMP have been used to examine the relationship between intracellular levels of cyclic AMP and lipid synthesis inMycobacterium smegmatis. Total phospholipid content was found to be increased in forskolin grown cells as a result of increased cyclic AMP levels caused by activation of adenylate cyclase. Increased phospholipid content was supported by increased [¹⁴C] acetate incorporation as well as increased activity of glycerol-3-phosphate acyltransferase. Pretreatment of cells with dibutyryl cyclic AMP had similar effects on lipid synthesis. Taking all these observations together it is suggested that lipid synthesis is being controlled by cyclic AMP in mycobacteria.     

A melanin-dispersing effect of cyclic adenosine monophosphate on Fundulus melanophores

Treatment of Fundulus melanophores with adenosine 3′,5′-monophosphate (cyclic AMP) is followed by reversible melanin dispersion in these cells. Adenosine 3′-monophosphate and adenosine 5′-monophosphate both have a similar, but weaker dispersing action. In addition, adenosine 5′-monophosphate also has a melanin aggregating effect. These results are interpreted to mean that nerve transmitters may act by controlling the level of cyclic AMP within the Fundulus melanophore.     

Modulation of postjunctional cholinergic sensitivity of rat diaphragm muscle by cyclic adenosine monophosphate.

Addition of 2.5 mM cyclic adenosine monophosphate (cAMP) to the solution bathing a rat diaphragm muscle alters the magnitude of depolarization responses to iontophoretic pulses of acetylcholine (ACh) at neuromuscular endplates. Alterations are repeatable with small variability on a given preparation for initial and repeat experiments on both hemidiaphragms, but are different on each preparation. Five min after addition of the nucleotide solution, increases (potentiations) of up to 30% above control levels and decreases (attenuations) to 50% below control levels are observed. The effects on sensitivity to ACh of dibutyryl cAMP (1.25 mM), monobutyryl cAMP (0.25 mM), and cAMP (2.5 mM) in Ca++ -free solution are a function of whether the experiment is an initial one on that preparation or a repeat experiment after 10 or more minutes of perfusion flow. In all three cases, initial exposure attenuates sensitivity (means at 5 min: --30, --10, and --20%, respectively) and repeat exposure potentiates sensitivity (mean: 20% at 5 min, 15% at 5 min, and 10% at 2 min respectively). A concentration of dibutyryl cAMP (0.25 mM) which is without effect on sensitivity alone, produces a large, transient potentiation (mean: 45% at 1 min) in conjunction with 0.5 mM theophylline. A decrease in the rate of desensitization is observed during exposure to 0.25 mM cAMP. The results are interpreted in terms of a physiological mechanism whereby receptor activity at the postjunctional membrane is modulated by cAMP formed from prejunctionally released ATP.     

Role of cyclic adenosine monophosphate in the induction of endothelial barrier properties

Cyclic adenosine monophosphate (AMP) has numerous important effects on cell structure and function, but its role in endothelial cells is unclear. Since cyclic AMP has been shown to affect transmembrane transport, cell growth and morphology, cellular adhesion, and cytoskeletal organization, it may be an important determinant of endothelial barrier properties. To test this we exposed bovine pulmonary artery endothelial cell monolayers to substances known to increase cyclic AMP and measured their effect on endothelial permeability to albumin and endothelial cell cyclic AMP concentrations. Cholera toxin (CT), a stimulant of the guanine nucleotide binding subunit of adenylate cyclase, led to a concentration-dependent 2-6-fold increase in cyclic AMP which was associated with a 3-10-fold reduction in albumin transfer across endothelial monolayers. The effect was not specific to albumin as similar barrier-enhancing effects were also noted with an unrelated macromolecule, fluorescein isothiocyanate (FITC)-dextran (MW 70,000). Barrier enhancement with cyclic AMP elevation was also observed with forskolin, a stimulant of the catalytic subunit of adenylate cyclase. The temporal pattern of barrier enhancement seen with these agents paralleled their effects on increasing cyclic AMP, and the barrier enhancement could be reproduced by incubation with either dibutyryl cyclic AMP or Sp-cAMPS, cyclic AMP-dependent protein kinase agonists. Furthermore, the forskolin effect on barrier enhancement was partially reversed with Rp-cAMPS, an antagonist of cyclic AMP-dependent protein kinase. Since endothelial actin polymerization may be an important determinant of endothelial barrier function, we sought to determine whether the cyclic AMP-induced effects were associated with increases in the polymerized actin pool (F-actin). Both cholera toxin and forskolin led to apparent endothelial cell spreading and quantitative increases in endothelial cell F-actin fluorescence. In conclusion, increased endothelial cell cyclic adenine nucleotide activity was an important determinant of endothelial barrier function in vitro. The barrier enhancement was associated with increased endothelial apposition and increases in F-actin, suggesting that influences on cytoskeletal assembly may be involved in this process.     

The effect of autonomic nerve stimulation cyclic adenosine monophosphate in skeletal muscle

The effect of electrical stimulation of autonomic nerves on c-AMP levels in the tibial muscle of rats was studied after pretreatment with a phosphodiesterase inhibitor and a muscle paralyzant. Cyclic-AMP levels in the skeletal muscle increase significantly. This increase is not the result of changes in muscle blood flow which might have resulted from autonomic nerve stimulation. These studies indicate that the adenyl cyclase-cyclic AMP system in skeletal muscle may be controlled by the autonomic nervous system.     

Restoration of voltage-dependent antibrain antibody-inhibited calcium current by cyclic adenosine monophosphate

A dual-microelectrode voltage clamp technique was used for recording voltage-dependent calcium current (I_ca) in unidentified neurons isolated fromHelix pomatia. Neither intracellular injection of cyclic adenosine monophosphate (cAMP; 10 nA, 5 min) nor intracellular application of dibutyril-cAMP (dcAMP; 1 mM, 10–20 min) induced a change in normal I_ca or produce a reversible 10–20% reduction in amplitude. Adding S-100 protein fraction antibodies to the external medium led to the onset of calcium-dependent inactivation of I_ca, bringing amplitude of I_ca down to 15±12% of its initial level. Either cAMP or dcAMP then restored inhibited I_ca to 50±11% of its original level. It was found that the effects of cAMP on I_ca of intact neurons depend on level of cytoplasmic Ca²⁺.Institute for Brain Research, All-Union Center for Mental Health Research, Academy of Medical Sciences of the USSR, Moscow. Translated from Neirofiziologiya, Vol. 21, No. 2, pp. 247–252, March–April, 1989.     

Estrogen synthetase in choriocarcinoma cell culture. stimulation by dibutyryl cyclic adenosine monophosphate and theophylline

The stimulation of estrogen biosynthesis by N⁶, O² -dibutyryl adenosine 3':5'-cyclic monophosphate and theophylline (dbT) in cultures of the JAr line of choriocarcinoma cells was investigated by measuring the specific activity and kinetic constants of estrogen synthetase (aromatase) in the various subcellular fractions after differential centrifugation of homogenized cells in isotonic sucrose. The low speed (900x$\text{g}$) pellet,from cells grown with or without dbT and homogenized in isotonic sucrose,contains the majority of the aromatase activity and the highest aromatase specific activity. The aromatase specific activity in the homogenate of cells grown with dbT and in the various subcellular fractions is 4- to 10-fold higher than in cells grown without dbT. The Vmax of androstenedione (4-androstene-3,17-dione) aromatization in homogenates from dbT-stimulated cells (6.9 pmol estrogen/min per mg protein) is significantly increased over that measured in the absence of dbT (1.5 pmol estrogen/min per mg protein); the Km values, however, are not significantly different (average of 43.8nM in dbT-stimulated fractions; 53.2nM in control fractions). These results suggest that the increased aromatase specific activity in dbT-stimulated cells results from an increase in amount of active enzyme, rather than from an increase in affinity of the enzyme for its substrate.