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PubMed1997

Measurement of acetylcholinesterase by positron emission tomography in the brains of healthy controls and patients with Alzheimer's disease.

BACKGROUND: Acetylcholinesterase activity, a marker for degeneration of the central cholinergic system, has consistently been reported, in necropsy brain studies, to be reduced in the cerebral cortex of patients with Alzheimer's disease. We have shown regional acetylcholinesterase activity in vivo in rodent and primate brains with radioactive acetylcholine analogues. In the present study, we used one of the analogues to map acetylcholinesterase activity in the brains of living people. METHODS: Positron emission tomography (PET) and a radiolabelled acetylcholine analogue with high hydrolytic specificity to acetylcholinesterase [11C]N-methyl-4-piperidyl acetate (MP4A), was used in eight elderly healthy controls and five patients with Alzheimer's disease who had mild dementia. All participants were given an intravenous injection of [11C]MP4A and then sequential patterns of radioactivity in various brain regions were obtained by PET. Time courses of [11C]MP4A concentration in arterial blood were also measured to obtain an input function. A three-compartment model was used to estimate regional acetylcholinesterase activity in the brain. FINDINGS: The estimated acetylcholinesterase distribution in the brain of the control participants agreed with the acetylcholinesterase distribution at necropsy. All patients with Alzheimer's disease had multiple cortical regions with a reduced estimated acetylcholinesterase activity in comparison with control participants. The reduction was more pronounced in the parietotemporal cortex, with an average reduction rate of 31% in temporal and 38% in parietal cortex, and less pronounced in other cortical lesions (19% in frontal, 24% in occipital, and 20% in sensorimotor cortex). Each patient was found to have at least two cortical regions with significantly reduced acetylcholinesterase activity. INTERPRETATION: The method we describe for non-invasive in-vivo detection of regional acetylcholinesterase changes in the living human brain that is feasible for biochemical assessment of Alzheimer's disease.

Acetylcholinesterase↗

PubMed1981

Direct demonstration of a correspondence between the dopamine islands and acetylcholinesterase patches in the developing striatum.

The distribution of dopamine-containing processes in the striatum of fetal and neonatal cats was studied by immunohistochemical and glyoxylic acid histofluorescence methods and compared to the distribution of acetylcholinesterase (acetylcholine acetylhydrolase, EC 3.1.1.7) observed by thiocholine histochemistry in the same or serially adjoining sections. Both methods for demonstrating the dopamine innervation revealed the characteristic patchwork of dopamine "islands" in the caudoputamen, in which catecholamine histofluorescence or tyrosine hydroxylase [tyrosine 3-monooxygenase; L-tyrosine, tetrahydropteridine:oxygen oxidoreductase (3-hydroxylating), EC 1.14.16.2]-like immunoreactivity was concentrated into 0.2- to 0.6-mm-wide patches. Both methods also demonstrated a high degree of patterning of the dopamine innervation in the ventral striatum, including the nucleus accumbens septi. A detailed and striking match was found between these configurations and the compartmental distribution of acetylcholinesterase observed in the caudoputamen and ventral striatum of the same brains. The correspondence between the dopamine and acetylcholinesterase figures was most obvious in the fetal brains, in which the background acetylcholinesterase staining was lightest, but matches between the dopamine islands and acetylcholinesterase patches could still be seen in the kittens. There was no clear alignment of striatal cell bodies stained for acetylcholinesterase with either the dopamine or the acetylcholinesterase-positive patches. Nor was there an obvious correspondence between dopamine and acetylcholinesterase in the striatal background matrix. We conclude that, at least during ontogenesis, it is the clustered arrangements of dopamine and acetylcholinesterase that are, in particular, tightly linked, and we suggest that information about the maturation of these clusters may be crucial in assessing the functions of striatal dopamine and acetylcholinesterase in the adult.

Acetylcholinesterase↗

PubMed2002

Serum acetylcholinesterase and prognosis of acute organophosphate poisoning.

OBJECTIVE: The aim of this study is to investigate the prognostic value of serum acetylcholinesterase levels and their relationship with neurological syndromes (Type 1 syndrome, intermediate syndrome, and delayed polyneuropathy) in acute organophosphate poisoning. MATERIALS AND METHODS: Thirty-two consecutive patients with acute organophosphate poisoning admitted to the Ondokuz Mayis University Emergency Department from June 1999 to January 2001 were evaluated. Patients were assessed according to admission time, symptoms, and results of clinical exams and their serum acetylcholinesterase levels were determined on days 1, 2, 3, 7, and the last day. RESULTS: There was no significant difference between the first-day serum acetylcholinesterase of the patients with severe poisoning (n = 22, 68.75%) and of the patients with mild poisoning (n = 10, 31.25%; NS). There was no discernible difference between the serum acetylcholinesterase obtained on days 1 and 3 after poisoning from the patients with intermediate syndrome (n = 5, 15.6%; means: 0.90 +/- 0.65 vs. 0.88 +/- 0.53, 19.35 vs. 18.92%; NS, sensitivity = 80%; specificity = 87.5%). There was a significant difference between the serum acetylcholinesterase obtained on days 1 and 3 from the patients with nonintermediate syndrome (n = 24, 75%; means: 1.05 +/- 0.24 vs. 1.68 +/- 0.29, 22.58 vs. 36.12%; p < 0.001). There was no discernible significant difference in serum acetylcholinesterase between the patients with organophosphorus-induced delayed polyneuropathy (n = 7, 21.8%) and nonorganophosphorus-induced delayed polyneuropathy. In the patients who died (n = 5, 15.6%), serum acetylcholinesterase showed no discernible increase day 1-the last day (means: 0.50 +/- 0.25 vs. 0.46 +/- 0.26, 10.75 vs. 9.89%; NS). There was a significant difference between the serum acetylcholinesterase levels obtained on days 1 and the last day from the patients who survived (n = 27, 84.3%; means: 1.14 +/- 0.25 vs. 2.32 +/- 0.26, 24.51 vs. 49.89%; p < 0.001). CONCLUSION: In the acute phase of organophosphate poisoning, low serum acetylcholinesterase (> 50% of minimum normal value) supports the diagnosis of organophosphate poisoning but it does not show a significant relationship to the severity of poisoning (NS). The serum acetylcholinesterase activity may be a useful parameter in following the acute prognosis of organophosphate poisoning.

Acetylcholinesterase↗

PubMed1984

Acetylcholinesterase release from mammalian erythrocytes by phosphatidylinositol-specific phospholipase C of Bacillus thuringiensis and characterization of the released enzyme.

The mode of acetylcholinesterase release from mammalian erythrocyte membranes by the action of phosphatidylinositol(PI)-specific phospholipase C of Bacillus thuringiensis was studied. As regards intact erythrocytes, a larger amount of acetylcholinesterase was released from sheep or bovine erythrocytes than from horse erythrocytes. From horse erythrocyte ghosts, acetylcholinesterase was more easily released than from intact cells. Bovine erythrocyte acetylcholinesterase released by PI-specific phospholipase C was purified by column chromatography on DEAE-cellulose, affinity gel and Sepharose 6B, to a homogeneous state, as indicated by polyacrylamide gel electrophoresis, with a recovery of 39%. Also, bovine erythrocyte acetylcholinesterase was solubilized by Triton X-100 and partially purified. The properties of these acetylcholinesterase preparations obtained by the action of PI-specific phospholipase C and/or Triton X-100 were studied in detail. On elution from the Sepharose 6B column, Triton X-100-solubilized acetylcholinesterase was eluted at the void volume while the enzyme obtained by further treatment with PI-specific phospholipase C was eluted in the region corresponding to M.W. 250,000. Furthermore, the heat stability of acetylcholinesterase purified after solubilization with PI-specific phospholipase C was higher than that of the Triton X-100-solubilized acetylcholinesterase. The close association and direct interaction of PI with acetylcholinesterase in the erythrocyte membrane was suggested by the above results.

Acetylcholinesterase↗

PubMed1995

Uptake of acetylcholinesterase by neurons in the substantia nigra.

It is known that acetylcholinesterase is secreted by the dopaminergic neurons of the substantia nigra and has a subsequent action independent of cholinergic transmission. Although non-cholinergic actions of this protein have been demonstrated, the subsequent fate of acetylcholinesterase is unknown. One possibility is that acetylcholinesterase is taken up following secretion into the extracellular space. This hypothesis has been tested in vivo, in both conscious and anaesthetized guinea-pigs. Exogenous acetylcholinesterase (2-20 pM) was infused via a push-pull cannula implanted into either the substantia nigra or the surrounding extranigral regions: the amount subsequently recovered in the perfusate was then compared with control values. Only when the push-pull cannulae were implanted in the substantia nigra was there a marked decrease in the amount of acetylcholinesterase recovered; this selective retention was abolished when the perfusion medium was cooled to 4 degrees C or when the experiment was performed post mortem. Direct visualization of immunocytochemically identified nigral dopaminergic cells revealed co-localized deposits of labelled, exogenous acetylcholinesterase. Moreover, when exogenous acetylcholinesterase was boiled to prevent detection by the assay system and to eliminate any classical enzymatic action, an enhancement in perfusate levels of endogenous acetylcholinesterase was observed from nigral but not from extranigral sites, indicating that endogenous and exogenous acetylcholinesterases were in competition. These results suggest that, within the substantia nigra, secreted acetylcholinesterase may be subject to a temperature- and energy-dependent uptake mechanism.

Acetylcholinesterase↗

PubMed1979

Molecular forms of chicken embryo acetylcholinesterase in vitro and in vivo. Isolation and characterization.

The four molecular forms of chick embryo leg muscle acetylcholinesterase have been isolated by velocity sedimentation; their apparent sedimentation coefficients are 19.5 S, 11.5 S, 7.1 S, and 5.4 S. All four forms are glycoproteins, exhibit the same Km for acetylcholine, and are inhibited to the same extent by specific inhibitors of acetyl- and buryrylcholinesterase. Treatment of the 19.5 S form of acetylcholinesterase with trypsin generates an array of molecular forms, several of which have sedimentation coefficients identical with the naturally occurring forms. Collagenase treatment of the 19.5 S acetylcholinesterase results in a somewhat different pattern of acetylcholinesterase forms including a novel 20.6 S form. Only the 19.5 S acetylcholinesterase is sensitive to collagenase treatment. Our results indicate that the several acetylcholinesterase forms share a common catalytic subunit, and suggest that the molecular forms of acetylcholinesterase in the chick represent different ensembles of a common monomer. In culture, the muscle cells contain only the 11.5 and 7.1 S acetylcholinesterase forms; however, they also secrete substantial amounts of enzyme into the medium. These secreted acetylcholinesterases have sedimentation coefficients of 9 S and 15 S. The relative abundance of the different acetylcholinesterase molecular forms changes during muscle development, both in vivo and in vitro, suggesting that the assembly and distribution of this family of membrane glycoproteins is developmentally regulated.

Acetylcholinesterase↗

PubMed1989

Electrophysiological estimation of the actions of acetylcholinesterase inhibitors on acetylcholine receptor and cholinesterase in physically isolated Aplysia neurones.

1. The actions of representative cholinesterase inhibitors on the acetylcholine responses of physically isolated single neurones from the pedal ganglion of Aplysia californica were studied, using electrophysiological techniques and rapid agonist application to analyse both the inhibitory actions on the acetylcholine receptor-channel complex and the degree of inhibition of acetylcholinesterase activity on the same neurone. The inhibitors used were physostigmine, edrophonium and diisopropylfluorophosphate (DFP). 2. When selected neurones were suddenly exposed to 50 microM acetylcholine by a 'concentration clamp' technique a large Na-dependent inward current was initiated, and decayed in the continued presence of acetylcholine without external perfusion. However, if perfusion of the acetylcholine solution was reinitiated the current increased somewhat, indicating that the decay of current was due to some combination of receptor desensitization and local depletion of acetylcholine at the membrane by acetylcholinesterase. 3. With simultaneous application of acetylcholine (50 microM) and physostigmine (0.1 to 100 microM) there was a dose-dependent reduction of peak amplitude of the acetylcholine response. However, physostigmine at low concentrations (0.1 to 10 microM) caused a time-dependent increase in the current amplitude alone with a time- and dose-dependent inhibition of acetylcholinesterase activity. At the highest concentration of physostigmine (100 microM) acetylcholinesterase activity was abolished but the current peak was very depressed. After removal of physostigmine from the bathing solution, the current amplitude decreased toward the control at the two lower concentrations as the inhibitory actions on acetylcholinesterase activity were almost reversible, while at the two higher concentrations (10 and 100 microM) the current increased and the inhibition of acethylcholinesterase remained. 4. When acetylcholine (50 microM) and edrophonium (0.1 to 10 microM) were applied simultaneously, edrophonium caused a dose-dependent increase in the peak amplitude that was correlated with a dose-dependent inhibition of acetylcholinesterase activity. Prolonged exposure to edrophonium did not change the peak amplitude and there was no time-dependent change in the inhibition of acetylcholinesterase activity. At the highest concentration of edrophonium used (100 microM), simultaneous application with acetylcholine augmented the peak amplitude relative to control, but to a lesser extent than 10 microM. Prolonged exposure to the highest concentration of edrophonium caused a time-dependent reduction in the peak amplitude. The effects of edrophonium were quickly reversible after the removal of the drug from the bathing solution. 5. DFP (1 and 10mM), similar to 1OO microM physostigmine, caused a dramatic reduction of the peak current on simultaneous application with ftetylcholine. During exposure to DFP the current amplitude and acetylcholinesterase activity were very depressed. After removing DFP from the bathing solution the current amplitude increased to more than the control level after 1 mm DFP, while it did not recover to the control level after 10mM DFP. The inhibition of acetylcholinesterase activity remained at both concentrations. 6. These results indicate that all three cholinesterase inhibitors have dose-dependent actions both at the acetylcholine receptor-channel complex and at acetylcholinesterase. The methods we have developed may be useful in the evaluation of various cholinesterase inhibitors.

Animals↗

PubMed1985

Effect of membrane fluidity upon binding of Electrophorus acetylcholinesterase to lipid vesicles.

A previous report (Watkins, M.S., Hitt, A.S. and Bulger, J.E. (1977) Biochem. Biophys. Res. Commun. 79, 640-647) has indicated that the asymmetric forms of Electrophorus acetylcholinesterase bind exclusively to sphingomyelin vesicles through interaction with the collagen-like 'tail' portion of the enzyme. We report here that acetylcholinesterase also binds to phosphatidylcholine vesicles containing saturated fatty acyl chains and to egg phosphatidylcholine vesicles containing cholesterol. This suggests preferential binding of acetylcholinesterase to membranes of lower fluidity. Surface charge of vesicles and density of zwitterionic lipid headgroups do not significantly affect binding of native acetylcholinesterase. The presence of chondroitin sulfate or hyaluronic acid slightly increases the binding of native acetylcholinesterase to sphingomyelin vesicles, while the presence of 1 M NaCl, bovine serum albumin, or tissue fractions enriched in basement membrane diminish binding. The dissociation constant for native acetylcholinesterase and sphingomyelin vesicles is (1.0-1.5) X 10(-7) M, as measured by a flotation binding assay. The globular, 11S form of acetylcholinesterase also binds to lipid vesicles, although not to the same degree as native acetylcholinesterase. This suggests that the collagen tail of the enzyme enhances binding, but is not essential for binding to occur. These results are consistent with the location of acetylcholinesterase on the surface of the postsynaptic plasma membrane in vivo.

Acetylcholinesterase↗

PubMed1991

Soman and sarin inhibition of molecular forms of acetylcholinesterase in mice. Time course of recovery and reactivation by the oxime HI-6.

The in vivo sensitivity of the molecular forms of the enzyme acetylcholinesterase to inhibition by either soman or sarin, reactivation by HI-6 and the time course of recovery following inhibition by soman were investigated in mice. Administration of HI-6 (50 mg/kg, i.p.) immediately after soman (100 micrograms/kg, s.c.) or sarin (150 micrograms/kg, s.c.) resulted in an apparent selective reactivation of the 10S and 16S molecular forms of acetylcholinesterase and no reactivation of the 4S form of diaphragm acetylcholinesterase. The apparent selectivity of the reactivation of the molecular forms of the acetylcholinesterase was probably due to the fact that the 10S and 16S forms of acetylcholinesterase are located primarily extracellularly and the 4S form intracellularly. The HI-6 was restricted primarily to the extracellular compartment due to its quaternary, hydrophilic nature. If the administration of HI-6 was delayed until 60 min following soman (100 micrograms/kg, s.c.) injection, no reactivation of any of the molecular forms of acetylcholinesterase could be found in the diaphragm. The soman-inhibited acetylcholinesterase had probably aged and, thus, was not susceptible to reactivation by HI-6. The time course of recovery of the molecular forms in the diaphragm occurred rather quickly with the smaller 4S and 10S forms recovering to control levels faster than the larger 16S form. It took between 8 and 16 days for the 16S form to recover to normal. In the brain, hypothalamic acetylcholinesterase molecular forms such as the 4S recovered faster than the 10S form which had not recovered to control 16 days after soman administration; the 16S form of acetylcholinesterase was not detected in the brain.

Acetylcholinesterase↗

PubMed1986

Release of acetylcholinesterase in the rat nigrostriatal pathway: relation to receptor activation and firing rate.

Acetylcholinesterase is released from both axon terminals and dendrites of nigrostriatal neurons. The relationship of this phenomenon to: (1) activation of receptors, and (2) firing rate, has been examined. In the first series of experiments, apomorphine or acetylcholine were infused into substantiae nigrae of urethane anaesthetized rats, via push-pull cannulae. In the ipsilateral striatum, the release of acetylcholinesterase was modified in a fashion reminiscent of the changes in firing rate induced by these drugs in nigrostriatal cells. Thus, application of acetylcholine into the substantia nigra induced an increase, while apomorphine induced a decrease in the release of acetylcholinesterase from the striatum. However, in the substantia nigra, the release of acetylcholinesterase did not follow this pattern: acetylcholine reduced local release, while apomorphine caused no change. In the second part of this study we studied the relationship between firing rate of nigrostriatal cells and the release of acetylcholinesterase. Two compounds known to block neuronal impulse flow were infused into the substantia nigra. These drugs were tetrodotoxin (a Na+ channel blocker), or gamma-hydroxybutyrate (a drug which blocks impulse flow specifically in dopaminergic cells). Both compounds reduced the release of acetylcholinesterase in the ipsilateral striatum. However, locally in the substantia nigra there was no decrease in release of the enzyme. In fact, following administration of gamma-hydroxybutyrate, there was a large Ca2+ dependent increase in release of acetylcholinesterase in the substantia nigra. These results suggest that release of acetylcholinesterase in the striatum may be linked to the discharge frequency of nigrostriatal neurons. On the other hand, release of acetylcholinesterase from the substantia nigra, which probably occurs from dendrites, is independent of Na+ mediated action potentials. This release may instead be associated with specific dendritic Ca2+ conductances.

Acetylcholine↗

PubMed1989

Acetylcholinesterase on the dendrites of central cholinergic neurons: an electron microscopical study in the ferret.

A combination of choline acetyltransferase immunohistochemistry and acetylcholinesterase histochemistry was used to characterize the ultrastructural distribution of acetylcholinesterase in identified cholinergic and non-cholinergic neurons in the ferret brain. Previous studies have shown that most of the cholinergic input to the brain arises from choline acetyltransferase-positive neurons found in the neostriatum, basal forebrain and dorsolateral pontine tegmentum. In all these cells intense staining for acetylcholinesterase was localized within the cisternae of the rough endoplasmic reticulum, in the nuclear envelope and Golgi apparatus, and along the plasma membranes of the soma and dendrites. In contrast, the distribution of acetylcholinesterase in non-cholinergic neurons was restricted mainly to the cisternae of the endoplasmic reticulum and the nuclear envelope. Since previous studies have associated high levels of acetylcholinesterase staining with non-cholinergic neurons in the locus coeruleus and substantia nigra zona compacta, these areas were examined as well. The ultrastructural localization of acetylcholinesterase in the principal locus coeruleus neurons was as observed in typical non-cholinergic neurons. On the other hand, the distribution of acetylcholinesterase in the principal cells of the zona compacta of the substantia nigra was more like that found in cholinergic neurons. In conclusion, the subcellular distribution of acetylcholinesterase in the principal cholinergic neurons of the brain follows a characteristic pattern which, with one exception, is different from that of acetylcholinesterase-positive non-cholinergic neurons.

Acetylcholinesterase↗

PubMed1989

Pressure ejection of acetylcholinesterase within the guinea-pig substantia nigra has non-classical actions on the pars compacta cells independent of selective receptor and ion channel blockade.

In the substantia nigra, acetylcholinesterase may have a non-classical function unrelated to cholinergic transmission. Acetylcholinesterase is released from the dendrites of dopamine-containing nigrostriatal neurons and has a subsequent action on these cells, independent of hydrolysis of acetylcholine. The aim of this study was to explore further the precise nature of this "non-cholinergic" action of acetylcholinesterase. Acetylcholinesterase was pressure-ejected in the vicinity of the dendrites of putative nigrostriatal neurons in vitro, in near-physiological amounts, and the effects of this treatment on neuronal membrane properties were investigated. It was found that acetylcholinesterase reversibly hyperpolarized the nigrostriatal cell membrane independent of sodium and calcium channel blockade. Acetylcholinesterase pretreated with an irreversible inhibitor (Soman) of its classical catalytic site produced the same hyperpolarizing effect: however, butyrylcholinesterase, which hydrolyses acetylcholine, was inefficacious. These effects persisted in the presence of the dopamine receptor antagonist sulpiride. It is suggested the acetylcholinesterase can facilitate the generation of a long-duration conductance, which enhances the firing of nigrostriatal cells if the neuron is first hyperpolarized. Hence the action of acetylcholinesterase would be to modulate inputs. These actions are independent of direct interaction with acetylcholine and dopamine systems. Hence, in the substantia nigra, acetylcholinesterase might serve as a "neuromodulatory" secretory protein.

Acetylcholinesterase↗

PubMed1990

Changes in the expression of acetylcholinesterase molecular forms during rat heart development.

The development of the molecular forms of acetylcholinesterase was studied in rat heart during the perinatal period. In this study, the activity of acetylcholinesterase increased both per unit wet weight and per unit protein from post-conception day 14 to day 42. Additionally, the activity of atrial acetylcholinesterase per unit wet weight increased more rapidly after birth than that of ventricular acetylcholinesterase. The percent contribution of the various molecular forms to the total acetylcholinesterase pool in heart changed dramatically from the fetal to the neonatal period. This switch primarily consisted of a decrease in the ratio of the asymmetric to globular forms. Thus, the specific activity of globular forms increased while that of the asymmetric forms remained relatively stable. When the atria and ventricles were examined separately at 19 days post-conception, the percent contribution of the individual molecular forms in the two cardiac areas was different. The atria contained a pool of acetylcholinesterase forms similar to postnatal heart while the ventricles contained a pool of acetylcholinesterase forms with a lower globular to asymmetric ratio. Finally, this study showed that greater than half of the acetylcholinesterase pool was inhibited by incubating hearts from fetal rats with echothiopate iodide, suggesting that a large portion of acetylcholinesterase catalytic sites are externalized in fetal heart.

Acetylcholinesterase↗

PubMed2004

Blood-cell-specific acetylcholinesterase splice variations under changing stimuli.

Developmental and trauma-induced mechanism(s) that modify inflammation and immune responses in blood cells were recently found to be regulated by acetylcholine. Here, we report corresponding blood cell-specific changes in acetylcholinesterase splice variants. Plasmon resonance and flow cytometry using acetylcholinesterase variant-specific antibody probes, revealed a progressive increase in myeloid cell fractions expressing the apoptosis-related acetylcholinesterase-S variant from newborns to adult controls and post-delivery mothers. Hematopoietic cell fractions positive for the myeloproliferative acetylcholinesterase-R variant, were similarly high in post-partum blood, both intracellular and on the cell surface. Moreover, intracellular acetylcholinesterase-S protein amounts as reflected by fluorescence intensity measurements remained unchanged in myeloid cells from post-partum mothers as compared with matched controls. Unlike brain neurons, which over-express intracellular acetylcholinesterase-R under stress, lymphocytes from post-partum mothers presented increased surface acetylcholinesterase-S and pronounced decreases in both the expression and contents of surface acetylcholinesterase-R. Peripheral stimuli-induced modulations in acetylcholine regulation may hence reflect blood cell lineage-dependent acetylcholinesterase splice variations.

Acetylcholine↗

PubMed2002

Catalytic antibodies with acetylcholinesterase activity.

We describe three catalytic cholinesterase-like catalytic antibodies (Ab1), as well as anti-idiotypic (Ab2) and idiotypic (Ab3) antibodies, to one of the Ab1s. The Ab1s were raised against the human erythrocyte acetylcholinesterase (AChE), and are unusual in that they both recognise and resemble acetylcholinesterase in their catalytic activity. No contamination of the antibody preparations with either acetylcholinesterase or butyrylcholinesterase (BChE) was found. None of the Ab2s showed catalytic activity, whereas four Ab3s did (an incidence of 1.26% of all Ab3s). Although there is considerable resemblance between Ab1s and Ab3s, there are significant differences between the two groups. All the antibodies were inhibited by phenylmethylsulphonyl fluoride (PMSF), indicating the presence of a serine residue in their active sites, and were inhibited by the cholinesterase active site inhibitors iso-OMPA and pyridostigmine, suggesting the similarity of the sites to those of cholinesterases. The Ab3s resemble the Ab1s in their ability to hydrolyse both acetyl and butyrylthiocholine (BTCh). However, the Ab3s appear to be better catalysts, having significantly reduced K(m) values (for acetyl, but not for butyrylthiocholine) and increased turnover numbers (K(cat)), rate enhancements (K(cat)/K(uncat)) and K(cat)/K(m) ratios, for both substrates, although these values by no means approach those of the natural enzymes. The Ab1s appear to have structures resembling the anionic sites of cholinesterases, as shown by their reaction with the anionic site inhibitors (edrophonium and tetramethylammonium). No such reactions were observed in the Ab3s. None of the antibodies show evidence of the sites resembling the peripheral anionic site (PAS) of acetylcholinesterase. All the antibodies recognise, to varying degrees, the peripheral anionic site of acetylcholinesterase. This was shown by their ability to inhibit acetylcholinesterase, to compete with peripheral site inhibitors, and to block acetylcholinesterase-mediated cell adhesion, a property of this site. The results indicate idiotypic mimicry of a catalytic antibody's active site, and suggest that the development of the catalytic activity in the anti-acetylcholinesterase antibodies may be related to the structural features of the peripheral anionic site of acetylcholinesterase.

Acetylcholine↗

PubMed1996

The spontaneous release of acetylcholinesterase in rat substantia nigra is altered by local changes in extracellular levels of dopamine.

Acetylcholinesterase release in the guinea-pig substantia nigra has been previously investigated 'on-line', using a sensitive chemiluminescent system. Since histological observations suggest that there is a difference in acetylcholinesterase distribution in the rat substantia nigra compared to that of the guinea-pig, the first aim of the present study was to use this chemiluminescent method to characterise acetylcholinesterase release in this brain region of the freely moving rat, and the second was explore the relationship between acetylcholinesterase release and dopamine systems in this region. Accordingly, acetylcholinesterase release in the rat substantia nigra was studied under basal conditions of spontaneous release and following the local administration of (a) elevated potassium ions (30, 45, 60'mM), (b) a stimulator of dopamine/acetylcholinesterase release-D-amphetamine (10(-7), 10(-6) and 10(-5) M), (c) an inhibitor of dopamine uptake-GBR12909 (10(-7), 10(-6) and 10(-5) M). Spontaneous release of acetylcholinesterase in this brain region of the rat appears to be comparable with that observed in the guinea-pig, despite the smaller number of acetylcholinesterase-containing neurones. Furthermore, not only elevated potassium ions, but D-amphetamine as well as GBR12909, all produced significant increases in the percentage spontaneous release of acetylcholinesterase. Thus, the release of acetylcholinesterase in this region may be triggered by levels of dopamine outside of the neurone.

Acetylcholinesterase↗

PubMed2003

Effect of thyroid hormones on acetylcholinesterase mRNA levels in the slow soleus and fast extensor digitorum longus muscles of the rat.

In the rat, the level of acetylcholinesterase messenger RNA in the typical slow soleus muscles is only about 20-30% of that in the fast extensor digitorum longus muscles. The expression of contractile proteins in muscles is influenced by thyroid hormones and hyperthyroidism makes the slow soleus muscle faster. The influence of thyroid hormones on the levels of acetylcholinesterase messenger RNA level in the slow soleus and fast extensor digitorum longus muscle of the rat was studied in order to examine the effect of thyroid hormones on muscle acetylcholinesterase expression. Hyperthyroidism was induced in rats by daily thyroid hormone injection or thyroid hormone releasing tablet implantation. Hind-limb suspension was applied to produce muscle unloading. Muscle denervation or reinnervation was achieved by sciatic nerve transection or crush. Acetylcholinesterase messenger RNA levels were analyzed by Northern blots and evaluated densitometrically. Hyperthyroidism increased the levels of acetylcholinesterase messenger RNA in the slow soleus muscles close to the levels in the fast extensor digitorum longus. The effect was the same in the unloaded soleus muscles. Acetylcholinesterase expression increased also in the absence of innervation (denervation), in the presence of changed nerve activation pattern (reinnervation), and under enhanced tonic neural activation of the soleus muscle (electrical stimulation). However, the changes were substantially smaller than those observed in the control soleus muscles. Enhancement of acetylcholinesterase expression in the soleus muscles by the thyroid hormones is, therefore, at last in part due to hormonal effect on the muscle itself. On the contrary, increased level of the thyroid hormones had no influence on acetylcholinesterase expression in the normal fast extensor digitorum longus muscles. However, some enhancing influence was apparent whenever the total number of nerve-induced muscle activations per day in the extensor digitorum longus muscle was increased. Thyroid hormones seem to be an independent extrinsic factor of acetylcholinesterase regulation in the slow soleus muscle.

Acetylcholinesterase↗

PubMed1997

Acetylcholinesterase promotes regeneration of neurites in cultured adult neurons of Aplysia.

Aplysia, a marine mollusc, has significant amounts of acetylcholinesterase in its hemolymph, reaching maximum levels in the adults with reproductive maturity [Srivatsan M., et al. (1992) J. comp. Physiol. 162, 29-37]. Since hemolymph of mature Aplysia is neurotrophic to Aplysia neurons in culture [Schacher S. and Proshanski E. (1983) J. Neurosci. 3, 2403-2413], we examined whether acetylcholinesterase is a hemolymph neurotrophic factor. Dopaminergic neurons from the pedal ganglia of young adult Aplysia were maintained in culture in defined medium or defined medium supplemented with hemolymph. After 24 h, neurons in defined medium supplemented with hemolymph were well attached to the substratum and exhibited multiple, long neurites. In contrast, neurons in defined medium alone attached poorly and exhibited one or two short neurites. When acetylcholinesterase was inhibited with a specific, membrane-impermeable inhibitor (1,5-bis(4-allyldimethylammoniumphenyl)-pentan-3-one dibromide) which binds to its catalytic and peripheral anionic sites, the neurotrophic effect of hemolymph was significantly reduced. However, inhibition of the catalytic site alone with membrane impermeable echothiophate still resulted in enhanced neurite growth. An analogue of acetylcholine, carbachol, which is not hydrolysed by acetylcholinesterase, did not interfere with neurite growth when added to the supplemented medium. Acetylcholinesterase isolated from the hemolymph and highly purified human acetylcholinesterase also promoted neurite growth in Aplysia neurons. These results show that i) acetylcholinesterase circulating in the hemolymph promotes neurite growth of adult neurons in culture; ii) the growth promoting action of acetylcholinesterase is independent of its function of hydrolysing acetylcholine and iii) the peripheral anionic site of acetylcholinesterase appears to be involved in neurite regeneration.

Acetylcholinesterase↗