Cyclic nucleotides in rheumatoid arthritis

Cyclic 3,5-adenosine monophosphate (cAMP) and cyclic 3,5-guanosine monophosphate (cGMP) may influence important regulatory mechanisms in the rheumatoid inflammatory process. It has been claimed that fasting imporves the condition of the patient with rheumatoid arthritis (RA). The present study was designed to measure cAMP in plasma and urine and cGMP in urine in medically untreated RA patients. 12 female patients were investigated in a cross-over study during a control and a fasting period. They received no other drugs than analgesics during these periods.Levels of plasma and urinary cAMP found during the control period were somewhat lower than previously reported. However, the ratio cAMP/cGMP in urine was 10 to 1 which is reported to be normal. Clinical and laboratory variables of inflammatory activity were significantly improved during the 7-day fasting period. The ratio of cAMP/cGMP in urine was significantly increased on days 2–4 and coincided in time with the maximum of clinical improvement. Cyclic AMP concentrations were lowered both in plasma and urine during fasting. This is in contrast to fasting in normal and obese subjects reported in previous studies.     

Cyclic AMP decreases chemotaxis,invasiveness and lung colonization of H-ras transformed mouse fibroblasts

We transfected mouse 10T1/2 fibroblasts with the H-ras oncogene and isolated lines expressing H-ras. One of the lines exhibited a highly malignant phenotype with the ability to produce large tumors and to colonize the lung after tail vein injection. In addition, the cells of this line showed increased collagenase IV production, directed migration and invasiveness, properties associated with the ability of tumor cells to metastasize. Since cyclic adenosine 3,5-monophosphate (cAMP) is known to down-regulate ras expression, we exposed the malignant cells (Cl-1) to either N⁶, 2,0-dibutyryl cAMP (DB-cAMP) or 8-bromo cAMP (8-Br-cAMP), either with or without a phosphodiesterase inhibitor. We found that these treatments reduced the expression of ras, chemotaxis, invasiveness, and lung colonization of the ras-transformed cells. We therefore postulate that the malignancy of some cells may be regulated by alterations in the intracellular cAMP levels by suppressing ras expression and/or by reducing other activities required for the dissemination of tumor cells.     

Dissociation of acetylcholine- and cyclic GMP-induced currents inXenopu oocytes

InXenopus follicular oocytes, activation of muscarinic receptors evokes a slow potassium current (H-response); a similar current is evoked by intracellular injection of cyclic guanosine 3,5-monophosphate, cGMP (Dascal et al. 1984). We have tested the hypothesis that cGMP may be the second messenger that mediates the opening of K channel by acetylcholine (ACh). ACh elevated the intracellular level of cGMP with a time course similar to that of the development of the muscarinic H-response; maximal increase in cGMP concentration above the control was about 0.2 pmole/oocyte. The amount of injected cGMP that produced a detectable K current (threshold dose) varied between 0.5 and 3 pmole/oocyte. At low doses of cGMP, the slope of log dose-log response curve was about 2.5, suggesting involvement of a biochemical process with a positive cooperativity of at least 3. Higher doses of cGMP evoked, in addition to the outward current, an irregular, rapidly developing, long-lasting inward current, that never reached amplitudes comparable to those of ACh-evoked Cl currents. The K current elicited by cGMP was insensitive to elevation or depletion of external Ca. It was potentiated by isobutylmethylxanthine (IBMX). ACh strongly inhibited the cGMP-evoked K current when applied at the plateau of the latter. 4-Phorbol 12,13-dibutyrate (PDBu) (1 M) rapidly and completely inhibited the cGMP response. It is concluded, that most of the results presented in this report contradict the hyothesis that cGMP is the intracellular mediator of ACh-induced changes in membrane conductance in the oocytes.Abbreviations ACh acetylcholine - cAMP cyclic adenosine 3,5-monophosphate - cGMP cyclic guanosine 3,5-monophosphate - EGTA ethylenediaminetetraacetic acid - Hepes N-2-hydroxyethyl-piperazinc-N-2-hydroxypropanesulphonic acid - IBMX 3-isobutyl-l-methylxanthine - IP₃ inositol 1,4,5-trisphosphate - PDBu 4-phorbol 12,13-dibutyrate     

Vasoactive hormones and cAMP affect pericyte contraction and stress fibresin vitro

Summary Pericytes are contractile cells of the microvascular wall that may influence capillary haemodynamics and permeability. We examined the contractile responses of cultured pericytes to selected vasoactive agents and cAMP agonists. Morphological and biochemical changes associated with these responses were also studied. Pericytes seeded onto silicone rubber contracted when stimulated with histamine or serotonin, relaxed in response to the beta-adrenergic agonist isoproterenol and did not respond to epinephrine. Since hormonal-induced relaxation of vascular smooth muscle involves cAMP, we investigated the ability of cAMP, to modulate pericyte contraction. Dibutyryl cAMP and forskolin (an adenylate cyclase activator) both induced pericyte relaxation and elevated intracellular cAMP levels. Isoproterenol increased cAMP levels but epinephrine had no effect. However, when epiniphrine and isoproterenol were co-incubated with the phosphodiesterase inhibitor 3-isobutyl-l-methylxanthine (IBMX), cAMP was increased to levels above those elicited by these agonists alone. Serotonin and histamine in the presence of IBMX did not affect cAMP levels. These results suggest that certain vasoactive agents may relax pericytes by cAMP-dependent processes. We have shown previously that stress fibres are also involved in pericyte contraction. Hence, changes in the staining patterns of stress fibres in response to these selected agonists were studied. Histamine, serotonin and epinephrine had no apparent effect on stress fibre staining. Dibutyryl cAMP, forskolin, and isoproterenol, which relax pericytes and increase cAMP, disassembled fibres. In summary, the results demonstrate that the contractile activity of cultured pericytesin vitro can be regulated by vasoactive agonists and that changes in cAMP and stress fibres may mediate the regulation.Abbreviations cAMP cyclic adenosine 3,5-monophosphate - dBcAMP dibutyryl cyclic adenosine 35-monophosphate - DiL-Ac-LDL 1,1-dioctadecyl 1-3,3,3,3 tetramethylindo-carbocyanine perchlorate - DME Dulbecco's Modified Eagles Medium - FCS foetal calf serum - HBSS Hanks balanced salt solution - IBMX 3-isobutyl-1-methylxanthine - PBS phosphate buffered saline - R020-1724 (4-(3-butoxy-4-methoxybenzyl)-2-imidazolidinone - TCA trichloroacetic acid     

cAMP levels in monocytes of heroin addicts

Summary Intravenous heroin addicts have a high intracellular cAMP level in relation to control monocytes. Incubation in vitro with ascorbic acid normalizes the cellular content of cAMP. We discuss the role of cAMP in the pathogenesis of defective monocyte chemotaxis in intravenous heroin addicts.Abbreviations cAMP adenosine 3:5-cyclic monophosphate - IDAs intravenous drug abusers - PBS phosphate buffered saline - EDTA ethyldiaminotetraacetic - cGMP guanosine 3-5-cyclic monophosphate - CNS central Nervous System     

Effect of cyclic AMP on cells of experimental lymphatic leukemia L-5718

The effect of cyclic AMP (cAMP), dibutyryl-cAMP, and theophylline (an inhibitor of phosphodiesterase, the enzyme converting adenosine-3, 5-monophosphate into adenosine-5-monophosphate) on the intensity of proliferation (as reflected in the increase in the nucleic acid content in the culture), DNA synthesis (thymidine-H³ incorporation), and transplantation properties (ability to repopulatein vivo) of leukemic cells of strain L-5178 was studied. The experiments showed that cAMP in a concentration of 0.8 mM inhibits thymidine-H³ incorporation considerably, retards proliferation, and reduces the transplantability of the leukemic cells. Theophylline and dibutyryl-cAMP have comparatively weak ability to inhibit DNA synthesis and the proliferative activity and transplantation properties of the cells.Laboratory of Biochemistry and Laboratory of Experimental Therapy of Leukemias, Central Institute of Hematology and Blood Transfusion, Ministry of Health of the USSR. Laboratory for the Search for New Antibiotics, All-Union Research Institute for the Search for New Antibiotics, Academy of Medical Sciences of the USSR, Moscow. (Presented by Academician of the Academy of Medical Sciences of the USSR K. V. Bunin.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 81, No. 3, pp. 365–367, March, 1976.     

Concomitant increase in plasma atrial natriuretic peptide and cyclic GMP during volume loading

Summary To investigate the effects of fluid expansion on endogenous atrial natriuretic peptide (ANP) and cyclic 3,5-guanosine monophosphate (cGMP), four male volunteers were studied before, during and after intravasal volume loading. Volume expansion was performed by intravenous infusion of 2,000 ml isotonic saline solution within 30 min. Mean plasma ANP levels increased 2.5-fold from 31.2 pg/ml to 81.7 pg/ml 40 min after the start of infusion. Plasma cGMP levels paralleled the rise in ANP, shwoing a mean cGMP increment from 2.7 pmol/ml to a maximum of 8.2 pmol/ml. Both ANP and cGMP levels were back to basal levels 120 min after termination of the infusion. Stimulation of endogenous ANP release by volume loading suggests that ANP is involved in the regulation of fluid homeostasis in man. The parallel rise in plasma cGMP levels supports the idea that cGMP is a mediator for the effects of ANP.Abbreviations ANP atrial natriuretic peptid - cGMP cyclic 3,5-guanosine monophosphate - PRA plasma renin activity     

Effects of 5-hydroxytryptamine,dopamine, and acetylcholine on accumulation of cyclic AMP and cyclic GMP in the anterior byssus retractor muscle ofMytilus edulis L. (Mollusca)

We investigated in vitro accumulation of adenosine 3,5-monophosphate (induced by 5-hydroxytryptamine and dopamine) and of guanosine 3,5-monophosphate (induced by acetylcholine) in the anterior byssus retractor muscle ofMytilus.The response to 5-hydroxytryptamine exceeded that induced by equimolar concentrations of dopamine.1-methyl lysergic acid, a 5-hydroxytryptamine-blocking agent, diminished the 5-hydroxytryptamine-induced increase of cyclic AMP level. This parallels the effect of this amine on the contracted muscle.Acetylcholine, which causes a tonic contraction of the muscle, increased intracellular levels of cyclic GMP in a dose-dependent (max. 45-fold at 10^–4 M ACh) manner. The time course of the rise in cyclic GMP level was rapid and transient (peak concentration of cyclic GMP at 2 min).Mytolon was the most effective of all cholinergic blockers tested. It was concluded that cyclic nucleotides may play a role in the modulatory process of the transmitters. A direct relation to the relaxation-contraction process could not be established.Abbreviations ABRM anterior byssus retractor muscle - cyclic AMP adenosine 3,5-monophosphate - cyclic GMP guanosine 3,5-monophosphate - ACh acetylcholine - 5-HT 5-hydroxytryptamine, serotonin - DA dopamine - TCA trichloroacetic acid - UML 1-methyl lysergic acid hydrogen maleinate     

Regulation of the trehalose-6-phosphate synthase complex in Saccharomyces

Summary Trehalose-6-phosphate synthase is another example of an enzyme of carbohydrate metabolism, in Saccharomyces, which could be regulated by interconversion of forms. Deactivation was mediated both in vivo and in vitro by a cyclic AMP-dependent protein kinase. Reversibility of this process was obtained by a phosphatase treatment leading to an increase in activity. The phosphorylated, less active form of the enzyme proved to be more susceptible to activation by ATP.Mg. Mutants with well defined lesions in the cyclic AMP-dependent protein kinase system were used to corroborate our findings of a possible regulatory mechanism of trehalose-6-phosphate synthase activity by interconversion of forms.Abbreviations PMSF phenyl-methyl-sulfonyl fluoride - G-6-P glucose-6-phosphate - UDPG uridine-5-diphosphoglucose - PEP phosphoenol pyruvate - NAD⁺ -nicotinamine adenine dinueleotide - ATP adenonise 5-triphosphate - cAMP adenosine 2:3-cyclic monophosphate - MOPS 3 (N-morpholino) propanesulfonic acid     

Effects of activators and inhibitors of protein kinase A on increases in quantal size at the frog neuromuscular junction

Adrenaline, permeable cyclic adenosine monophosphate (cAMP) derivatives and insulin are known to elicit an increase in quantal size at the frog neuromuscular junction, primarily by increasing the amount of acetylcholine (ACh) per quantum. The quantal size increases produced by adrenaline or cAMP were antagonized by the protein kinase inhibitor H8 N-[2-(methyl-amino)ethyl]-5-isoquinolonesulfonamide. The increase in quantal size produced by insulin was not prevented by H8. Quantal size is also increased by pretreatment in hypertonic solution; this increase was also antagonized by H8. The H8 did not alter the increase in miniature endplate potential (MEPP) frequency produced by the hypertonic solution. A permeable cGMP derivative had no effect on quantal size. The diastereomer (Sp)-cAMPS (cyclic 3, 5-phosphothoate) activates protein kinase A(PKA). It elicited an increase in quantal size. The (Rp)-cAMPS isomer is known to inhibit PKA; it had no effect on quantal size. The increase in quantal size produced by hypertonic solution was antagonized by (Rp)-cAMPS but not by (Sp)-cAMPS. Brief exposure to a hypertonic solution containing a phosphodiesterase inhibitor followed by incubation in the inhibitor leads to an increase in quantal size. We conclude that one pathway for signaling for an increase in quantal size involves activation of PKA and that hypertonic pretreatment acts via this pathway.