guanosine-triphosphate and Pheochromocytoma

Elongation factor 2 (eEF-2) is a 100-kD protein that catalyzes the ribosomal translocation reaction, resulting in the movement of ribosomes along mRNA. eEF-2 is the target for a very specific Ca2+/calmodulin-dependent eEF-2 kinase. Phosphorylation of eEF-2 makes it inactive in translation, which suggests that protein synthesis can be regulated by Ca2+ through eEF-2 phosphorylation. Recent data demonstrate that eEF-2 phosphorylation can be involved in cell-cycle regulation and other processes where changes of intracellular Ca2+ concentration induce a new physiological state of a cell. The main role of eEF-2 phosphorylation in these processes is temporary inhibition of overall translation in response to transient elevation of the Ca2+ concentrations in the cytoplasm. Temporary inhibition of translation may trigger the transition of a cell from one physiologic state into another because of the disappearance of short-lived repressors and thus the activation of expression of new genes.

Topics: Adrenal Gland Neoplasms; Amino Acid Sequence; Animals; Calcium; Calcium-Calmodulin-Dependent Protein Kinases; Calmodulin; Elongation Factor 2 Kinase; Globins; Guanosine Triphosphate; Interphase; Mitosis; Molecular Sequence Data; Nerve Growth Factors; Oogenesis; Peptide Chain Elongation, Translational; Peptide Elongation Factor 2; Peptide Elongation Factors; Pheochromocytoma; Phosphoprotein Phosphatases; Phosphorylation; Protein Kinases; Protein Processing, Post-Translational; Rabbits; Sequence Homology, Nucleic Acid; Signal Transduction; Transcription, Genetic; Tumor Cells, Cultured; Vertebrates; Xenopus laevis

The guanine base is prone to oxidation by free radicals regardless of the cellular moiety it is bound to. However, under conditions of oxidative stress, 8-oxoguanosine triphosphate (oxo(8)GTP) formation has been shown to occur without oxidation of the guanine base in DNA. In vitro studies have suggested that oxo(8)GTP could impact G-protein signaling and RNA synthesis. Whether increased levels of oxo(8)GTP translate into cellular malfunction is unknown. Data presented herein show that oxo(8)GTP is formed in cell-free preparations as well as in PC12 cells after exposure to physiologically relevant oxidative conditions generated with 10 microM copper sulfate and 1 mM L-ascorbic acid (Cu/Asc). We also determined that oxo(8)GTP has biological activity as a potent inhibitor of nitric oxide-stimulated soluble guanylyl cyclase (sGC). The increase in oxo(8)GTP formation in purified GTP and PC12 cells exposed to Cu/Asc caused a significant reduction in the product of sGC activity, cGMP. This oxidation of GTP was attenuated by the addition of reduced glutathione under these same Cu/Asc conditions, thus preventing the decrease in sGC activity. This suggests that oxo(8)GTP is produced by free radicals in vivo and could have significant impact on cell functions regulated by sGC activity such as synaptic plasticity in the central nervous system.

Topics: Animals; Ascorbic Acid; Cell Extracts; Cell Line; Central Nervous System; Chromatography, High Pressure Liquid; Copper Sulfate; Cyclic GMP; Enzyme Activation; Free Radicals; Guanosine Triphosphate; Guanylate Cyclase; In Vitro Techniques; Nitric Oxide; Oxidation-Reduction; Oxidative Stress; Pheochromocytoma; Rats

To clarify gene alterations in functional human adrenal tumors, we performed molecular analysis for p53 abnormalities in 23 cases with adrenal neoplasms. The immunohistochemical study with anti-p53 monoclonal antibody pAb1801 demonstrated that 10 of 23 (43.5%) cases overexpressed p53 protein in the tumor cells. Using a polymerase chain reaction-single strand conformation polymorphism study, 5 of 6 (83.3%) pheochromocytoma tissues (1 malignant and 5 benign) and 11 of 15 (73.3%) adrenocortical adenomas (2 with Cushing's syndrome and 13 with primary aldosteronism, all benign) showed an apparent electrophoretic mobility shift between the tumor and its paired adjacent normal adrenal tissue. Such differences were detected in exon 4 (12 cases), exon 5 (2 cases), and exon 7 (3 cases). The types of these mutations in exon 4 were a substitution from threonine (ACC) to isoleucine (ATC) at codon 102 in 5 cases, from glutamine (CAG) to histidine (CAC) at codon 104 in 1 case, from glycine (GGG) to alanine (CGG) at codon 117 in 1 case, from glutamate (GAG) to glutamine (CAG) at codon 68 in 1 case, and single base changes resulting in a premature stop codon at codon 100 in 2 cases. A 2-basepair deletion at codon 175 in exon 5 resulting in a frame shift was identified in 1 case. A single point mutation was identified, resulting in the substitution of glutamine (CAG) for arginine (CGG) at codon 248 of exon 7 in 1 case. A single basepair deletion at codon 249 resulted in a frame shift in 2 cases. There was 1 case with malignant pheochromocytoma that combined a single point mutation in exon 4 and a single base deletion in exon 7. Only 2 of 23 cases showed a loss of a normal allele encoding in the p53 gene. Northern blot analysis with 1.8-kilobase p53 cDNA revealed that p53 mRNA was overexpressed in 6 cases. Our results indicate that high frequencies of p53 gene mutation, especially in exon 4, exist in functional adrenal tumors. As p53 protein is a regulator of guanine nucleotide synthesis, the loss of normal inhibitory regulation by the p53 mutation would serve to increase the availability of GTP for the transduction of signals essential for increased cell growth and hormone expression in the adrenal tumors. These findings suggest that the p53 gene mutation may play a role in the tumorigenesis of benign and functional human adrenal tumors.

Topics: Adenoma; Adrenal Gland Neoplasms; Adult; Base Sequence; Blotting, Northern; Blotting, Southern; Cloning, Molecular; DNA, Neoplasm; Exons; Female; Gene Amplification; Genes, p53; Guanosine Triphosphate; Heterozygote; Humans; Immunohistochemistry; Male; Middle Aged; Molecular Sequence Data; Mutation; Pheochromocytoma; Polymerase Chain Reaction; RNA, Messenger; Sequence Analysis, DNA; Signal Transduction; Tumor Suppressor Protein p53

To determine the amino acid residues required for the signal-transducing activity of the human c-Ha-Ras protein, we introduced point mutations at residues 45-54 near the 'effector region' (residues 32-40). We transfected PC12 cells with these mutant genes and also micro-injected the mutant proteins, bound with an unhydrolyzable GTP analog, into PC12 cells. Both procedures showed that Val45----Glu and Gly48----Cys mutations impaired the ability of the Ras protein to induce morphological change of PC12 cells. These mutations did not affect the guanine nucleotide-binding activity or GTPase activity in the absence or presence of bovine GTPase-activating protein (GAP). Therefore, the Val45 and Gly48 residues should be included by definition in the effector region responsible for the signal transduction, while only a subset of the effector-region residues is required for enhancement of the GTPase activity by GAP.

Topics: Adrenal Gland Neoplasms; Amino Acids; Genes, ras; GTP Phosphohydrolases; GTPase-Activating Proteins; Guanosine Diphosphate; Guanosine Triphosphate; Mutagenesis, Site-Directed; Pheochromocytoma; Proteins; Proto-Oncogene Proteins p21(ras); ras GTPase-Activating Proteins; Signal Transduction; Structure-Activity Relationship; Transfection; Tumor Cells, Cultured

The effect of the hydrolysis-resistant GTP analogs, guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S) and guanylyl imidodiphosphate (GMPPNP), on norepinephrine (NE) secretion from digitonin-permeabilized rat pheochromocytoma cells, PC12, was examined. Although secretion in the presence of saturating Ca2+ (10 microM) was not affected by GTP gamma S or GMPPNP, secretion in the absence of Ca2+ was stimulated by these GTP analogs. Secretion induced by saturating concentrations of GTP gamma S or GMPPNP was approximately 80% of that induced by 10 microM Ca2+. Half-maximum stimulation was induced by 30 microM GTP gamma S or GMPPNP. Both Ca2(+)-stimulated and GTP gamma S-stimulated secretion were ATP dependent and inhibited by N-ethylmaleimide. The GTP gamma S-stimulated secretion of NE from permeabilized PC12 cells does not appear to result from either the release of Ca2+ or the activation of protein kinase C. Activation of protein kinase C by pretreatment of intact cells with 12-O-tetradecanoylphorbol 13-acetate caused a 50% increase in both Ca2(+)-stimulated and GTP gamma S-stimulated secretion. Cholera and pertussis toxins did not affect Ca2(+)-stimulated or GTP gamma S-stimulated NE secretion. Guanosine 5'-O-(2-thiodiphosphate) (GDP beta S) and GTP inhibited GTP gamma S-stimulated secretion but not Ca2(+)-stimulated secretion. The inability of GDP beta S to inhibit Ca2(+)-stimulated secretion indicates that the process affected by GTP gamma S is not an essential step in the Ca2(+)-stimulated pathway.

Topics: Adenosine Triphosphate; Adrenal Gland Neoplasms; Animals; Calcium; Cell Membrane Permeability; Digitonin; Ethylmaleimide; GTP-Binding Proteins; Guanosine 5'-O-(3-Thiotriphosphate); Guanosine Triphosphate; Guanylyl Imidodiphosphate; Hydrolysis; Kinetics; Norepinephrine; Pheochromocytoma; Rats; Signal Transduction; Tetradecanoylphorbol Acetate; Thionucleotides; Tumor Cells, Cultured

Angiotensin II receptor binding sites in rat liver and PC12 cells differ in their affinities for a nonpeptidic antagonist, DuP 753, and p-aminophenylalanine6 angiotensin II. In liver, which primarily contains the sulfhydryl reducing agent-inhibited type of angiotensin II receptor, which we refer to as the AII alpha subtype, DuP 753 displays an IC50 of 55 nM, while p-aminophenylalanine6 angiotensin II displays an IC50 of 8-9 microM. In PC12 cells, which primarily contain the angiotensin II receptor type whose binding affinity is enhanced by sulfhydryl reducing agents (AII beta), DuP 753 displays an IC50 in excess of 100 microM, while p-aminophenylalanine6 angiotensin II displays an IC50 of 12 nM. p-Aminophenylalanine6 angiotensin II binding affinity in liver is decreased in the presence of guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S) suggesting that this analogue is an agonist.

Topics: Adrenal Gland Neoplasms; Angiotensin II; Animals; Binding, Competitive; Cell Membrane; Guanosine 5'-O-(3-Thiotriphosphate); Guanosine Triphosphate; Imidazoles; In Vitro Techniques; Liver; Losartan; Mercaptoethanol; Pheochromocytoma; Rats; Receptors, Angiotensin; Tetrazoles; Thionucleotides

The major component of whole-cell Ca2+ current in differentiated, neuron-like rat pheochromocytoma (PC12) cells and sympathetic neurons is carried by dihydropyridine-insensitive, high-threshold-activated N-type Ca2+ channels. We show that these channels have unitary properties distinct from those of previously described Ca2+ channels and contribute both slowly inactivating and large sustained components of whole-cell current. The N-type Ca2+ currents are modulated by GTP binding proteins. The snail toxin omega-conotoxin reveals two pharmacological components of N-type currents, one blocked irreversibly and one inhibited reversibly. Contrary to previous reports, neuronal L-type channels are insensitive to omega-conotoxin. N-type Ca2+ channels appear to be specific for neuronal cells, since their functional expression is greatly enhanced by nerve growth factor.

Topics: Acetylcholine; Animals; Calcium; Calcium Channels; Dihydropyridines; GTP-Binding Proteins; Guanosine 5'-O-(3-Thiotriphosphate); Guanosine Triphosphate; Ion Channel Gating; Mollusk Venoms; Neurons; omega-Conotoxins; Pheochromocytoma; Rats; Thionucleotides; Tumor Cells, Cultured

Nerve growth factor (NGF) rapidly increases the cyclic GMP (cGMP) level about 2-3-fold and enhances the cGMP phosphodiesterase (PDE) activity about 2-fold in rat pheochromocytoma PC12 cells. No changes in the level of cyclic AMP (cAMP) and in the activity of cAMP PDE were found. GTP and a nonhydrolysable analog of GTP, GMP-PCP, at 100 microM, were able to mimic the effect of NGF on the cGMP PDE activity. These results suggest that the cGMP system may be one of the second messengers of NGF action in PC12 cells.

Topics: 1-Methyl-3-isobutylxanthine; 3',5'-Cyclic-AMP Phosphodiesterases; 3',5'-Cyclic-GMP Phosphodiesterases; Adrenal Gland Neoplasms; Animals; Cell Line; Cyclic AMP; Cyclic GMP; Enzyme Activation; Guanosine Triphosphate; Guanylyl Imidodiphosphate; Kinetics; Nerve Growth Factors; Pheochromocytoma

The mechanism of action of lithium regarding its therapeutic effects has not yet been established, despite many years of clinical use and scientific investigations. We recently reported that lithium stimulates the phospholipase C of NGF differentiated PC12 cells membranes. In view of the coupling between growth factor receptors, G proteins and phospholipase C, we investigated the effects of lithium on the binding of GTP to the membranes of PC12 cells cultured with NGF. Lithium (1.1 mM) increased 4-5-fold the Bmax of the binding of [3H]GTP to the PC12 membranes. NaF did not induce a similar stimulation.

Topics: Animals; Chlorides; GTP-Binding Proteins; Guanosine Triphosphate; Lithium; Lithium Chloride; Nerve Growth Factors; Pheochromocytoma; Rats; Sodium Fluoride; Subcellular Fractions; Tumor Cells, Cultured

The direct effects of chronic ethanol administration on adenylate cyclase, Na,K-ATPase, and Mg-ATPase activities in a cell containing neuronal characteristics were investigated using PC12 pheochromocytoma cells. Exposure of PC12 cells to 0, 75, and 150 mM ethanol for 4 days caused a dose-dependent increase in the stimulation of adenylate cyclase by in vitro ethanol without altering activation of the enzyme by GTP, NaF, MnCl2, or 2-chloroadenosine. Conversely, a 4-day treatment with 150 mM ethanol increased Na,K-ATPase and Mg-ATPase activities without altering the inhibitory effects of in vitro ethanol. The increase in Na,K-ATPase activity was associated with an increase in Vmax without any change in the Km for KCl. Chronic ethanol exposure also increased the amount of [3H]ouabain specifically bound to PC12 cell membranes. Except for the increase in Mg-ATPase activity, the above results were also observed when chronic ethanol treatment was carried out in the presence of pyrazole. Although ethanol slowed PC12 cell growth, observed changes were not due to an ethanol-induced reduction in cellular density. A 4-day exposure of a nonneuronal cell line (Madin Darby canine kidney cell) to 150 mM ethanol did not alter adenylate cyclase or ATPase activities. The present study indicates that the direct effects of chronic ethanol exposure of a neuronal-like cell involve an increase in the density of sodium pumps per cell and an enhanced sensitivity of adenylate cyclase to activation by ethanol.

Topics: 2-Chloroadenosine; Adenosine; Adenylyl Cyclases; Adrenal Gland Neoplasms; Ca(2+) Mg(2+)-ATPase; Chlorides; Enzyme Activation; Ethanol; Guanosine Triphosphate; Guanylyl Imidodiphosphate; Manganese; Manganese Compounds; Neurons; Ouabain; Pheochromocytoma; Sodium Fluoride; Sodium-Potassium-Exchanging ATPase; Tumor Cells, Cultured

LiCl stimulated the formation of inositol monophosphate in PC12 cells that had been exposed to nerve growth factor (NGF) for 4-5 days. Half-maximal accumulation was observed at approximately 8 mM LiCl. Stimulation of formation of inositol bisphosphate plus inositol trisphosphate was half-maximal at approximately 1 mM LiCl. With membranes isolated from PC12 cells differentiated with NGF, the hydrolysis of added phosphatidylinositol 4,5-bisphosphate (PIP2) was stimulated by LiCl in a biphasic manner, with the first stimulation half-maximal at approximately 0.7 mM and the second half-maximal at approximately 15 mM LiCl. The apparent Km for PIP2 was lowered in the presence of 1.1 mM LiCl from approximately 200 to approximately 70 microM. Membranes from cells grown in the absence of NGF did not respond to LiCl. Although observations with intact cells are difficult to interpret without ambiguity, the results obtained with isolated membranes support our interpretation of the stimulatory action of lithium in the intact PC12 cells.

Topics: Adrenal Gland Neoplasms; Animals; Cell Membrane; Chlorides; Guanosine 5'-O-(3-Thiotriphosphate); Guanosine Triphosphate; Inositol 1,4,5-Trisphosphate; Inositol Phosphates; Lithium; Lithium Chloride; Nerve Growth Factors; Pheochromocytoma; Phosphatidylinositol 4,5-Diphosphate; Phosphatidylinositols; Rats; Sodium Fluoride; Thionucleotides; Tumor Cells, Cultured; Type C Phospholipases

The ras gene product (p21) specifically binds GDP or GTP. In analogy with the reaction mechanism of other GTP-binding proteins, only the GTP-bound conformation is believed to be the biologically active one. Previously, we reported that not only oncogenic p21(Val-12) but also proto-oncogenic p21(Gly-12) could induce morphological differentiation in rat pheochromocytoma PC12 cells when microinjected in the complexed form with GTP gamma S [(1987) Mol. Cell. Biol. 7, 4553-4556]. In the present report we transformed PC12 cells with the oncogenic ras gene placed under the metallothionein I promoter. It was found that the transformed cells, when induced with Cd2+, differentiated in the absence of NGF. Then we analyzed the guanine nucleotide bound to p21 in the intact PC12 cells. It was found that conditionally induced p21(Val-12) was mostly present in the GTP-bound form, whereas the endogenous p21(Gly-12) was in the GDP-bound form. These results indicate again that p21.GTP induces the morphological differentiation of PC12 cells.

Topics: Animals; Cell Line, Transformed; Guanine Nucleotides; Guanosine Diphosphate; Guanosine Triphosphate; Immunoassay; Pheochromocytoma; Plasmids; Proto-Oncogene Proteins; Proto-Oncogene Proteins p21(ras); Rats; Transfection; Tumor Cells, Cultured

Type C1 and D toxins produced by Clostridium botulinum caused ADP-ribosylation of a protein of 24 kDa in membrane preparations of rat clonal pheochromocytoma cells (PC12) and of proteins of 25 and 26 kDa in neuron-rich culture of fetal rat brain cells. The ADP-ribosylation reaction was dependent on the presence of MgCl2, GTP and GTP gamma S. The results obtained suggested that the ADP-ribosylation reaction is responsible for the development of the biological activity of the botulinum neurotoxins and that the target of this reaction may be novel GTP-binding proteins localized on cell membranes.

Topics: Adenosine Diphosphate Ribose; Botulinum Toxins; Brain; Cells, Cultured; Clostridium botulinum; GTP-Binding Proteins; Guanosine 5'-O-(3-Thiotriphosphate); Guanosine Triphosphate; Magnesium; Magnesium Chloride; Membrane Proteins; Molecular Weight; Nerve Tissue Proteins; Neurotoxins; Pheochromocytoma; Thionucleotides

Two possible cellular pathways of catecholamines from the chromaffin vesicles of PC12 cells to the surrounding medium are explored in this study. The direct one circumventing the cytoplasm can be activated in alpha-toxin-permeabilized cells with micromolar levels of free Ca2+. Catecholamine metabolites formed in the cytoplasm (i.e., 3,4-dihydroxyphenylacetic acid and 3,4-dihydroxyphenylethanol) are neither formed nor released from the cells under these conditions. However, when vesicular catecholamines were discharged into the cytoplasm by addition of the ionophore nigericin, such metabolites are formed and released into the medium independent of Ca2+. Both types of experiments provide direct evidence for the operation of Ca2+-induced exocytosis of dopamine and noradrenaline in permeabilized PC12 cells. The Ca2+ dependence of dopamine or noradrenaline release, as measured by the determination of the endogenous catecholamines using the high-performance liquid chromatography technique, exhibits two different phases. One is already activated below 1 microM free Ca2+ and plateaus at 1-5 microM free Ca2+, while a second occurs in the presence of larger amounts of free Ca2+ (10-100 microM). Ca2+-induced catecholamine release from the permeabilized cells can be modulated in different ways: It is enhanced by the phorbol ester 12-O-tetradecanoylphorbol 13-acetate and the diacylglycerol 1-oleyl-2-acetylglycerol provided Mg2+/ATP is present, and it is inhibited by guanosine 5'-O-(3-thiotriphosphate). The latter effect is abolished by pretreatment of the cells with pertussis toxin but not by cholera toxin. Thus, it appears that Ca2+-induced exocytosis can be modulated via the protein kinase C system, as well as via GTP binding proteins.

Topics: Adrenal Gland Neoplasms; Animals; Bacterial Toxins; Calcium; Cell Line; Cell Membrane Permeability; Dopamine; Guanosine 5'-O-(3-Thiotriphosphate); Guanosine Triphosphate; Hemolysin Proteins; Kinetics; Neurotoxins; Nigericin; Norepinephrine; Pheochromocytoma; Tetradecanoylphorbol Acetate; Thionucleotides

Rat pheochromocytoma (PC12) cells differentiate to neuronal cells in response to nerve growth factor. It has been shown that microinjection of oncogenic but not proto-oncogenic p21 protein induces morphological differentiation in PC12 cells (D. Bar-Sagi and J. R. Feramisco, Cell 42:841-848, 1985). In this paper we describe a recombinant human proto-oncogenic Ha-ras protein which can effectively induce neurite extension of PC12 cells when microinjected as a complex with guanosine-5'-O-(3-thiotriphosphate). The protein was found to be less effective when complexed with GTP. On the other hand, an oncogenic ras protein coinjected with guanosine-5'-O-(2-thiodiphosphate) was entirely inactive. These results indicate that the binary p21-GTP complex, but not the p21-GDP complex, is effective in inducing differentiation in PC12 cells, irrespective of the oncogenic or the proto-oncogenic protein.

Topics: Adrenal Gland Neoplasms; Animals; Axons; Cell Differentiation; Guanosine 5'-O-(3-Thiotriphosphate); Guanosine Diphosphate; Guanosine Triphosphate; Microinjections; Pheochromocytoma; Proto-Oncogene Proteins; Proto-Oncogene Proteins p21(ras); Rats; Recombinant Proteins; Thionucleotides; Tumor Cells, Cultured

In previous studies from this laboratory (Yu, M.W., Tolson, N. W., and Guroff, G. (1980) J. Biol. Chem. 255, 10481-10492) nerve growth factor treatment of PC12 cells was shown to increase the phosphorylation of a specific nonhistone nuclear protein. In the present work these whole-cell observations have been pursued and a cell-free system developed, based on the detergent treatment devised by Lenk et al. (Lenk, R., Ransom, L., Kaufmann, Y., and Penman, S. (1977) Cell 10, 67-78), in order to explore the nerve growth factor-sensitive phosphorylation system in biochemical detail. Using this preparation it has been shown that treatment of the whole cells with nerve growth factor for 30 min or more leads to a marked increase in the subsequent cell-free phosphorylation of the same nonhistone nuclear protein. A characterization of this phosphorylation indicates that it is quite labile to heat and to structural disruption, that it prefers ATP as phosphate donor, and that it requires Mg2+, but is inhibited by high Mg2+ levels as well as by certain other divalent cations. The site of phosphorylation appears to be on serine residues of the protein, as was the phosphorylation observed previously in whole cells. The use of various inhibitors and stimulators suggests that the kinase catalyzing this phosphorylation is not cAMP-dependent, nor is it similar to protein kinase C or casein kinase. The increased phosphorylation produced by nerve growth factor is not transient, the stimulation being constant for at least 3 days in the continuous presence of nerve growth factor. Increases in the phosphorylation of the same nuclear protein can be seen upon treatment of the cells with other effectors such as epidermal growth factor and dibutyryl cyclic AMP, the latter in spite of the fact that cAMP-dependence could not be established in the cell-free system. Finally, a similar system, with a similar stimulation of phosphorylation due to nerve growth factor treatment, can be prepared from sympathetic ganglia from neonatal animals.

Topics: Adenosine Triphosphate; Adrenal Gland Neoplasms; Amino Acids; Animals; Cell Line; Cell-Free System; Ganglia, Sympathetic; Guanosine Triphosphate; Magnesium; Nerve Growth Factors; Nucleoproteins; Pheochromocytoma; Phosphorylation; Rats

Most beta-adrenergic effects are mediated by activation of the enzyme adenylate cyclase. Hormone binds to the receptor leading to an accelarated binding of GTP to the coupling protein, the N-protein, which is activated. This causes an activation of the adenylate cyclase and an increased formation of cAMP, the intracellular second messenger. The same principles hold good for other hormones coupled to adenylate cyclase. The sensitivity of the adenylate cyclase may be altered in different clinical and experimental conditions. An increased sensitivity is seen in hyperthyroidism in man and in the rat, and during starvation in rats. A decreased sensitivity is seen in hypothyroidism, in patients with pheochromocytoma, pseudohypoparathyroidism type I or multiple symmetric lipomatosis. Several reasons for the altered sensitivity have been suggested. The number of hormone receptors, the coupling between receptor and N-protein, the amount or function of the N-protein or the PDE activity may all vary in different conditions.

Topics: Adenylyl Cyclases; Adrenal Gland Neoplasms; Animals; Catecholamines; Cell Membrane; Cyclic AMP; Enzyme Activation; GTP-Binding Proteins; Guanosine Triphosphate; Humans; Lipolysis; Lipomatosis; Pheochromocytoma; Pituitary Hormones, Anterior; Pseudohypoparathyroidism; Rats; Receptors, Adrenergic, beta; Receptors, Cell Surface; Thyroid Diseases; Thyroid Hormones

PC12 cells, a nerve growth factor-responsive clone of rat pheochromocytoma, contain a membrane-bound adenylate cyclase, which can be activated by adenosine analogs. The characteristics of the cyclase response indicate the presence of stimulatory adenosine receptors. Adenosine analogs also produce a marked increase in the ornithine decarboxylase levels of the cells, and the characteristics of this response suggest that it is linked to the adenylate cyclase-stimulatory adenosine receptors. The ornithine decarboxylase response elicited by 5'-N-ethylcarboxamideadenosine (NECA), a potent stimulatory adenosine analog, is synergistic with that produced by nerve growth factor. Differentiation of the cells with nerve growth factor, however, does not substantially alter either the response of cyclase to the adenosine analog or the magnitude of the adenosine-evoked ornithine decarboxylase response. Treatment of the cells with NECA produces an increase in the phosphorylation of a specific non-histone nuclear protein. While causing little or no morphological alteration by itself, NECA is synergistic with nerve growth factor in producing neurite outgrowth in PC12 cells. NECA does not cause an induction of acetylcholinesterase in the cells. NECA does not cause an induction of acetylcholinesterase in the cells, nor does it appear to affect the induction of this enzyme by nerve growth factor.

Topics: Acetylcholinesterase; Adenosine; Adenosine-5'-(N-ethylcarboxamide); Adenylyl Cyclases; Adrenal Gland Neoplasms; Animals; Carboxy-Lyases; Cell Line; Cell Membrane; Clone Cells; Guanosine Triphosphate; Guanylyl Imidodiphosphate; Kinetics; Nerve Growth Factors; Ornithine Decarboxylase; Pheochromocytoma; Rats