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Query: UMLS:C0027819 (neuroblastoma)
 27,800 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

1. Acetylcholine (ACh) produces two membrane current changes when applied to NG108-15 mouse neuroblastoma x rat glioma hybrid cells transformed (by DNA transfection) to express m1 muscarinic receptors: it activates a Ca(2+)-dependent K+ conductance, producing an outward current, and it inhibits a voltage-dependent K+ conductance (the M conductance), thus diminishing the M-type voltage-dependent K+ current (IK(M)) and producing an inward current. The present experiments were undertaken to find out how far inhibition of IK(M) might be secondary to stimulation of phospholipase C, by recording membrane currents and intracellular Ca2+ changes with indo-1 using whole-cell patch-clamp methods. 2. Bath application of 100 microM ACh reversibly inhibited IK(M) by 47.3 +/- 3.2% (n = 23). Following pressure-application of 1 mM ACh, the mean latency to inhibition was 420 ms at 35 degrees C and 1.79 s at 23 degrees C. Latencies to inhibition by Ba2+ ions were 148 ms at 35 degrees C and 92 ms at 23 degrees C. 3. The involvement of a G-protein was tested by adding 0.5 mM GTP-gamma-S or 10 mM potassium fluoride to the pipette solution. These slowly reduced IK(M), with half-times of about 30 and 20 min respectively, and rendered the effect of superimposed ACh irreversible. Effects of ACh were not significantly changed after pretreatment for 24 h with 500 ng ml-1 pertussis toxin or on adding up to 10 mM GDP-beta-S to the pipette solution. 4. The role of phospholipase C and its products was tested using neomycin (to inhibit phospholipase C), inositol 1,4,5-trisphosphate (InsP3) and inositol 1,3,4,5-tetrakisphosphate (InsP4), heparin, and phorbol dibutyrate (PDBu) and staurosporin (to activate and inhibit protein kinase C respectively). Both neomycin (1 mM external) and InsP3 (100 microM intrapipette) inhibited the ACh-induced outward current and/or intracellular Ca2+ transient but did not block ACh-induced inhibition of IK(M). Intrapipette heparin (1 mM) blocked activation of IK(Ca) and reduced Ach-induced inhibitions of IK(M), but also reduced inhibition of ICa via endogeneous m4 receptors. PDBu (with or without intrapipette ATP) and staurosporin had no significant effects.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:On the mechanism of M-current inhibition by muscarinic m1 receptors in DNA-transfected rodent neuroblastoma x glioma cells. 827 Nov 96

Cholinergic neurons in PNS and CNS are identified by the presence of choline acetyltransferase and the accumulation of choline by a high-affinity, sodium-coupled choline transporter to be used for acetylcholine synthesis. It appears that expression of choline acetyltransferase can be altered by several physiological conditions, including hormones and trophic factors, but little is known about control of expression of the sodium-coupled choline carrier or whether these two phenotypic markers are regulated similarly. In the present study, the cholinergic human neuroblastoma LA-N-2 was used to investigate regulation of expression of choline acetyltransferase and choline uptake activity associated with differentiation and neurite extension. Cells grown in serum-containing basal medium maintained a relatively undifferentiated morphology, expressed low levels of choline acetyltransferase activity, and accumulated choline by a sodium-dependent process followed by conversion to acetylcholine. Transfer of cells to an enriched, serum-free defined medium resulted in morphological and neurochemical differentiation, with an enhancement of cholinergic phenotype. Hemicholinium-sensitive choline uptake activity was increased about sixfold over a 4-day period, with no change in choline acetyltransferase or acetylcholinesterase specific activity. Acetylcholine synthesis was increased in parallel with the changes in choline accumulation; choline metabolism in the differentiated cells differed significantly from that observed in the undifferentiated cells, with proportionally less converted to phosphorylcholine and proportionally more remaining as unmetabolized choline and converted to acetylcholine. The enhanced choline accumulation appeared to be mediated by an increased number of choline carriers, demonstrated by increased binding of the affinity ligand [3H]-choline mustard to the transporter and by an increased Vmax for the uptake process. The increased expression of the transport function appeared to be under transcriptional control, as the enhancement of uptake was blocked by the RNA polymerase II inhibitor alpha-amanitin as well as by the protein synthesis inhibitor cycloheximide. These results show that expression of sodium-coupled choline carriers and choline acetyltransferase may be regulated separately in the differentiating neuroblastoma LA-N-2 and that neurotransmitter synthesis is controlled by provision of precursor rather than at the level of the biosynthetic enzyme.
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PMID:Regulation of expression of cholinergic neuronal phenotypic markers in neuroblastoma LA-N-2. 837 93

Studies were conducted to ascertain whether cells of the IMR-32 human neuroblastoma clone express ligand binding sites and functional responsiveness attributable to ganglia-type nicotinic acetylcholine receptors (nAChR) and/or neuronal/nicotinic alpha-bungarotoxin binding sites. Some comparative studies were conducted using cells of other clonal lines, including BC3H-1 mouse muscle line cells that are known to express muscle-type nAChR. Two classes of specific binding sites for 3H-labeled acetylcholine ([3H]ACh) and a single class of high-affinity, specific 125I-labeled alpha-bungarotoxin binding sites are expressed on membrane fractions prepared from IMR-32 cells. Radioligand binding to these sites on IMR-32 cells is relatively insensitive to blockade by the muscle-type nAChR-selective inhibitors, succinyldicholine and decamethonium, indicating that they are distinct from muscle-type nAChR. [3H]ACh binding to its sites on IMR-32 cell membranes is insensitive to blockade by alpha-bungarotoxin, indicating that IMR-32 cell [3H]ACh and 125I-labeled alpha-bungarotoxin binding sites also can be distinguished. Functional nAChR ion channels of the ganglia-type are expressed by IMR-32 cells, as assessed by the abilities of nicotinic agonists to stimulate 86Rb+ efflux, the relatively higher sensitivity of those responses to blockade by mecamylamine than by d-tubocurarine, and the inability of alpha-bungarotoxin to antagonize nAChR function. These results are consistent with expression by IMR-32 cells of functional ganglia-type nAChR that correlate with high affinity [3H]ACh binding sites as well as expression of a distinct class of neuronal/nicotinic alpha-bungarotoxin binding sites that have a ganglia-type pharmacology.
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PMID:Expression of ganglia-type nicotinic acetylcholine receptors and nicotinic ligand binding sites by cells of the IMR-32 human neuroblastoma clonal line. 847 12

Lithium has a biphasic effect of the agonist-dependent accumulation of Ins(1,4,5)P3 in human neuroblastoma SH-SY5Y cells. These effects consist of a transient reduction, followed by a long-lasting increase in Ins(1,4,5)P3 as compared to controls. The Li+ effects are dose dependent, and were observed at concentrations used in the treatment of bipolar disorders, and thus may have therapeutic implications. The mechanism of the Li+ effect on Ins(1,4,5)P3 accumulation requires further investigation. The transient reduction of Ins(1,4,5)P3 was observed under conditions where Li+ causes only a moderate increase in the inositol mono- and bi-phosphates. Supplementation with exogenous inositol had no effect on the level of Ins(1,4,5)P3, indicating that the mechanism of the Li(+)-dependent reduction of Ins(1,4,5)P3 is not due to inositol depletion. Li+ did not interfere with degradation of Ins(1,4,5)P3 after receptor-blockage with atropine, suggesting that Li+ has no direct effect on the Ins(1,4,5)P3 metabolizing enzymes. A direct effect of Li+ on the phospholipase C is also unlikely. Entry of Ca2+ into the cells is an important factor, which affects agonist-stimulated accumulation of Ins(1,4,5)P3, as well as absolute values of Li(+)-dependent increase in Ins(1,4,5)P3; however, it is not essential for the manifestation of Li+ effects. Our results also show that manifestation of Li+ effects in human neuroblastoma cells requires the stimulation of muscarinic receptors and activation of PLCs, PKCs, and/or that other staurosporine/H-7/GF 109203X-sensitive protein kinases are involved in the regulation of Ins(1,4,5)P3 during the plateau phase of ACh-stimulation. We also suggest an important role for these enzymes in the Li(+)-dependent elevation of Ins(1,4,5)P3.
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PMID:Phosphoinositide signalling in human neuroblastoma cells: biphasic effect of Li+ on the level of the inositolphosphate second messengers. 886 50

In cultured mouse neuroblastoma N1E-115 cells, inorganic lead (Pb2+) affects inward currents induced by activation of neuronal type nicotinic acetylcholine receptors (nAChR) in a biphasic manner. At nanomolar concentrations a blocking action is observed, while at submillimolar concentrations this blocking effect is reversed, resulting in a U-shaped concentration-effect curve. Maximal block by 90% is observed at 1-3 microM Pb2+. The interactions of Pb2+ with nAChR were examined at the blocking concentration of 10 nM Pb2+ and at 10 microM Pb2+, presenting the reversal of block. The fitted Emax for nAChR receptor activation by ACh was attenuated at both high and low Pb2+ concentrations by 24% and 54%, respectively. The EC50 values of the activation curves were not significantly altered; amounting to 53 microM, 64 microM and 86 microM ACh in the control situation and in the presence of 10 nM and 10 microM Pb2+, respectively. Further, receptor desensitization and ion channel block by ACh were also not affected by Pb2+. The results indicate that Pb2+ affects nAChR in a dual manner that involves inhibition and potentiation, both by non-competitive interactions. Neuronal nAChR expressed in N1E-115 cells resemble a combination of alpha 4 and beta 2 subunits, that constitute the predominant subunits in the central nervous system. The differential inhibition and potentiation of nAChR, together with the high sensitivity, are of interest with respect to Pb2+ neurotoxicity.
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PMID:Dual, non-competitive interaction of lead with neuronal nicotinic acetylcholine receptors in N1E-115 neuroblastoma cells. 904 21

Functional effects of human alpha 5 nicotinic ACh receptor (AChR) subunits coassembled with alpha 3 and beta 2 or with alpha 3 and beta 4 subunits, were investigated in Xenopus oocytes. The presence of alpha 5 subunits altered some properties of both alpha 3 AChRs and differentially altered other properties of alpha 3 beta 2 AChRs vs. alpha 3 beta 4 AChRs. alpha 5 subunits increased desensitization and Ca++ permeability of all alpha 3 AChRs. The Ca++ permeabilities of both alpha 3 beta 2 alpha 5 and alpha 3 beta 4 alpha 5 AChRs were comparable to that of alpha 7 AChRs. As we have shown previously, alpha 5 subunits increased the ACh sensitivity of alpha 3 beta 2 AChRs 50-fold but had little effect on alpha 3 beta 4 AChRs. alpha 5 caused only subtle changes in the activation potencies of alpha 3 AChRs for nicotine, cytisine and 1,1-dimethyl-4-plenylpiperazinium (DMPP). However, alpha 5 increased the efficacies of nicotine and DMPP on alpha 3 beta 2 AChRs but decreased them on alpha 3 beta 4 AChRs. Immunoisolation of cloned human AChRs expressed in oocytes showed that alpha 5 efficiently coassembled with alpha 3 plus beta 2 and/or beta 4 subunits. As expected, human AChRs immunoisolated from SH-SY5Y neuroblastoma cells showed that AChRs containing alpha 3 and probably alpha 5 subunits were present, but alpha 4 AChRs were not. In brain, by contrast, alpha 4 beta 2 AChRs were shown to predominate over alpha 3 AChRs. Some of the brain alpha 4 beta 2 AChRs were found to contain alpha 5 subunits.
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PMID:alpha 5 Subunit alters desensitization, pharmacology, Ca++ permeability and Ca++ modulation of human neuronal alpha 3 nicotinic receptors. 965 74

The effects of the carbamate physostigmine and of the organophosphates (OPs) parathion, paraoxon and phenyl saligenin cyclic phosphate (PSP) were examined on different subtypes of neuronal nicotinic acetylcholine receptors (nAChR). Stimulation with 1 mM ACh induced transient nicotinic inward currents in mouse N1E-115 and human SH-SY5Y neuroblastoma and in locust thoracic ganglion cells. All four acetylcholinesterase (AChE) inhibitors reduced the nicotinic currents in a concentration-dependent manner. Parathion is about 50 times more potent in blocking nAChR, compared to its active AChE inhibiting metabolite paraoxon. The relative blocking potency of the different AChE inhibitors was the same in all cell types, and followed the order parathion > physostigmine > PSP > paraoxon. In N1E-115 cells the IC50 values of block amounted to 2 microM, 30 microM, 39 microM and 96 microM for parathion, physostigmine, PSP and paraoxon, respectively. In all cell types, the nicotinic currents were equally blocked by parathion. Human nAChR in SH-SY5Y cells appeared more sensitive to block by physostigmine, PSP and paraoxon, while these AChE inhibitors similarly inhibited nicotinic currents in insect cells and in mouse neuroblastoma cells. The observation that the concentration-dependence of block is different from that of AChE inhibition, indicates a distinct interaction of AChE inhibitors with nAChR. Only in locust cells physostigmine induced a non-desensitizing inward current, that appeared to originate from nAChR activation. Occasionally, the OPs were able to activate slow ionic currents in mouse, but not in human and locust cells. As the OP-induced agonistic activity in mouse cells was not associated with the blocking action, the target site appeared to be distinct from nAChR. These results show that AChE inhibitors block nAChR with different potencies, dependent on the compound and the receptor subtype, and may activate distinct ion currents in neuronal cells of different species origin.
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PMID:Differential effects of physostigmine and organophosphates on nicotinic receptors in neuronal cells of different species. 986 67

1. The role of cyclic ADP ribose and ryanodine receptors in the inhibition of the M-like current (IK(M,ng)) by acetylcholine was investigated in m1 muscarinic receptor-transformed mouse neuroblastoma-rat glioma hybrid (NG108-15) cells using patch-clamp techniques and calcium microfluorimetry. 2. Acetylcholine (1-100 microM) decreased IK(M,ng) by up to 55 %. Application, via the patch pipette, of the cyclic ADP ribose antagonists 8-amino-cyclic ADP ribose (10-100 microM) and 8-bromo-cyclic ADP ribose (100-1000 microM) reduced this inhibition of IK(M,ng) in a concentration-dependent manner. The half-maximal inhibition concentrations for 8-amino- cyclic ADP ribose and 8-bromo-cyclic ADP ribose were around 40 microM and 1 mM, respectively. 3. Neither of the cyclic ADP ribose antagonists altered the amplitude of IK(M,ng) per se, or the incidence of the concurrent Ca2+-activated K+ current (IIK(Ca)) activation, also mediated by acetylcholine. 4. The ryanodine receptor modulators ryanodine (1-10 microM) and Ruthenium Red (10 microM) did not alter IK(M,ng) amplitude or IK(M,ng) inhibition mediated by acetylcholine. There was a statistically significant increase in the proportion of cells showing outward currents in the presence of Ruthenium Red. 5. Intracellular calcium levels measured with fura-2 microfluorimetry were increased with low concentrations of ryanodine (1 microM), more consistently with caffeine (10 mM), and in almost every case with both bradykinin (300 nM) and acetylcholine (100 microM). Caffeine-, but not bradykinin-evoked responses were abolished by preincubation with ryanodine (10 microM). 6. The fast 'rundown rate' of the M-current recorded in rat superior cervical ganglion cells under whole-cell conditions precluded an investigation of the effects of intracellular dialysis of cyclic ADP ribose. However, when cyclic ADP ribose (5 microM) was applied directly to the cytoplasmic face of inside-out membrane patches excised from rat superior cervical ganglion cells containing M-channels, it had no effect on the main parameters of single channel activity (conductance, mean open time or frequency of opening). 7. These results indicate that cyclic ADP ribose acts on a specific intracellular site to mediate IK(M,ng) inhibition. However, unlike previously established effects of cyclic ADP ribose, the ryanodine receptor is not required, suggesting that another molecular target may be involved. Studies at the single channel level indicate that cyclic ADP ribose may not act directly on the M-channels in inside-out patches.
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PMID:The role of ryanodine receptors in the cyclic ADP ribose modulation of the M-like current in rodent m1 muscarinic receptor-transformed NG108-15 cells. 1043 36

Mechanisms of preferential loss of cholinergic neurons in the course of neurodegenerative diseases are unknown. Therefore, we investigated whether differentiation-evoked changes in acetyl-CoA and acetylcholine metabolism contribute to the susceptibility of cholinergic neuroblastoma to cytotoxic effects of Al. In SN56 cells differentiated with retinoic acid and dibutyryl cAMP (DC), pyruvate utilization and acetyl-CoA content were lower and acetylcholine level higher than in nondifferentiated cells (NC), respectively. In DC Al and Ca accumulations were 50% and 100%, respectively higher than in NC. Acute Al addition caused inhibition, whereas its chronic application had no effect on pyruvate utilization both in NC and in DC. On the other hand, in both experiments, Al evoked a greater decrease of acetyl-CoA level in DC than in NC. Acute addition of Al depressed acetylcholine release from DC to two times lower values than in NC. On the other hand, chronic addition of Al increased ACh release from DC over twofold, being without effect on its release from NC. These findings indicate that higher accumulation of Ca, along with low levels of acetyl-CoA, could make DC more susceptible to neurotoxic inputs than NC. Excessive acetylcholine release, evoked by Al, is likely to increase acetyl-CoA utilization for resynthesis of the neurotransmitter pool and cause deficit of this metabolite in DC. On the other hand, NC, owing to lower Ca accumulation, slower ACh metabolism, and higher level of acetyl-CoA, would be less prone to these harmful conditions.
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PMID:Acute and chronic effects of aluminum on acetyl-CoA and acetylcholine metabolism in differentiated and nondifferentiated SN56 cholinergic cells. 1107 May 6

We characterized the functional and molecular properties of nicotinic acetylcholine receptors (AChRs) expressed by IMR-32, a human neuroblastoma cell line, and compared them to human alpha3 AChRs expressed in stably transfected human embryonic kidney (HEK) cells. IMR-32 cells, like neurons of autonomic ganglia, have been shown to express alpha3, alpha5, alpha7, beta2, and beta4 AChR subunits. From these subunits, several types of alpha3 AChRs as well as homomeric alpha7 AChRs could be formed. However, as we show, the properties of functional AChRs in these cells overwhelmingly reflect alpha3beta4 AChRs. alpha7 AChR function was not detected, yet we estimate that there are 70% as many surface alpha7 AChRs in IMR-32 when compared with alpha3 AChRs. Agonist potencies (EC(50) values) followed the rank order of 1,1-dimethyl-4-phenylpiperazinium (DMPP; 16+/-1 microM) > nicotine (Nic; 48 +/- 7 microM) > or = cytisine (Cyt; 57 +/- 3 microM) = acetylcholine (ACh; 59 +/- 6 microM). All agonists exhibited efficacies of at least 80% relative to ACh. The currents showed strong inward rectification and desensitized at a rate of 3 s(-1) (300 microM ACh; -60 mV). Assays that used mAbs confirmed the predominance of alpha3- and beta4-containing AChRs in IMR-32 cells. Although 18% of total alpha3 AChRs contained beta2 subunits, no beta2 subunit was detected on the cell surface. Chronic Nic incubation increased the amount of total, but not surface alpha3beta2 AChRs in IMR-32 cells. Nic incubation and reduced culture temperature increased total and surface AChRs in alpha3beta2 transfected HEK cells. Characterization of various alpha3 AChRs expressed in HEK cell lines revealed that the functional properties of the alpha3beta4 cell line best matched those found for IMR-32 cells. The rank order of agonist potencies (EC(50) values) for this line was DMPP (14 +/- 1 microM) = Cyt (18 +/- 1 microM) > Nic (56 +/- 15 microM > ACh (79 +/- 8 microM). The efficacies of both Cyt and DMPP were approximately 80% when compared with ACh and the desensitization rate was 2 s(-1). These data show that even with the potential to express several human nicotinic AChR subtypes, the functional properties of AChRs expressed by IMR-32 are completely attributable to alpha3beta4 AChRs.
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PMID:Functional properties of human nicotinic AChRs expressed by IMR-32 neuroblastoma cells resemble those of alpha3beta4 AChRs expressed in permanently transfected HEK cells. 1169 12


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