Showing papers on "Acetylthiocholine published in 1997"

Research report In vitro phosphorylation of acetylcholinesterase at non-consensus protein kinase A sites enhances the rate of acetylcholine hydrolysis

Abstract: Here, we report that the catalytic subunit of cAMP-dependent protein kinase PKA but not casein kinase II or protein kinase C . phosphorylates recombinant human acetylcholinesterase AChE in vitro. This enhances acetylthiocholine hydrolysis up to 10-fold as compared to untreated AChE, while leaving unaffected the enzyme's affinity for this substrate and for various active and peripheral site inhibitors. Alkaline phosphatase treatment enhanced the electrophoretic migration, under denaturing conditions, of part of the AChE proteins isolated from various mammalian sources and raised the isoelectric point of some of the treated AChE molecules, indicating that part of the AChE molecules are also phosphorylated in vivo. Enhancement of acetylthiocholine hydrolysis also occurred with Torpedo . AChE, which has no consensus motif for PKA phosphorylation. Further, mutating the single PKA site in human AChE threonine-249 did not prevent this enhancement, suggesting that in both cases it was due to phosphorylation at non-consensus sites. In vivo suppression of the acetylcholine hydrolyzing activity of AChE and consequent impairment in cholinergic neurotransmission occur under exposure to both natural and pharmacological compounds, including organophosphate and carbamate insecticides and chemical warfare agents. Phosphorylation of AChE may possibly offer a rapid feedback mechanism that can compensate for impairments in cholinergic neurotransmission, modulating the hydrolytic activity of this enzyme and enabling acetylcholine hydrolysis to proceed under such challenges. q 1997 Elsevier Science B.V.

The effects of oximes in the assay of acetylcholinesterase activity in lysed erythrocytes in vitro

Abstract: Organophosphorus compounds are known to inhibit the esteratic site of acetylcholinesterase by phosphorylation. The phosphorylated esteratic site of acetylcholinesterase undergoes hydrolytic regeneration at a slow or negligible rate. Nucleophilic agents such as hydroxytamine, hydroxamic acids, and oximes reactivate the enzyme more erapidfy than does spontaneous hydrolysis. The red cell cholinesterose activity was assayed using dithio bis-2-nitrobenzoic acid (DTNB) commonly known as Ellman's reagent. The principle of this assay method is the rate of hydrolysis of acetylthiocholine (substrate) by a red celt suspension. Thiocholine that is produced, forms a yellow complex, when EUman's reagent (DTNB) is used in the assay. This was tested in vitro in lysed erythrocyte samples of 35 healthy persons who had no known exposure to cholinesterose inhibitors, after the observation of immediate increase in absorption of light at 440 nm. All of data were statistically analyzed using one-way ANOVA and student t-test. A value of p

Complete Staining of Nerve Fiber and Myoneural Junctions with Acetylthiocholine and Silver

Abstract: We describe a combined stain for simultaneous demonstration of the preterminal axons and cholinesterase activity at myoneural junctions of mammalian muscles. This technique employs acetylthiocholine iodide as the substrate for cholinesterase activity and silver nitrate impregnation of preterminal axons. The procedure is rapid, simple and uses fresh muscles. Intramuscular nerves, preterminal axons and myoneural junctions are stained simultaneously brown or black with minimal background staining of connective tissue and muscle fibers.