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PubMed2000

Donepezil dose-dependently inhibits acetylcholinesterase activity in various areas and in the presynaptic cholinergic and the postsynaptic cholinoceptive enzyme-positive structures in the human and rat brain.

In the symptomatic treatment of mild to moderately severe dementia associated with Alzheimer's disease, donepezil (E2020) has been introduced for the inhibition of acetylcholinesterase activity in the human brain. However, there is no morphological evidence as to how this chemical agent affects the acetylcholinesterase-positive structures in the various areas of the human and the rat CNS. This study demonstrates by histochemical means that donepezil exerts a dose-dependent inhibitory effect in vitro on acetylcholinesterase activity. The most sensitive areas were the cortex and the hippocampal formation. Within the different layers of the cortex, the cholinoceptive acetylcholinesterase-positive postsynaptic pyramidal cell bodies were more sensitive than the presynaptic cholinergic axonal processes. In the cortex, the cell body staining was already abolished by even 2 x 10(-8)M donepezil, whereas the axonal staining could be eliminated only by at least 5 x 10(-8)M donepezil. In the hippocampus, the axonal acetylcholinesterase reaction end-product was eliminated by 5 x 10(-7)M donepezil. The most resistant region was the putamen, where the staining intensity was moderately reduced by 1 x 10(-6)M donepezil. In the rat brain, the postsynaptic cholinoceptive and presynaptic cholinergic structures were inhibited by nearly the same dose of donepezil as in the human brain. These histochemical results provide the first morphological evidence that, under in vitro circumstances, donepezil is not a general acetylcholinesterase inhibitor in the CNS, but rather selectively affects the different brain areas and, within these, the cholinoceptive and cholinergic structures. The acetylcholinesterase staining in the nerve fibers (innervating the intracerebral blood vessels of the human brain and the extracerebral blood vessels of the rat brain) and at the neuromuscular junction in the diaphragm and gastrocnemius muscle of rat, was also inhibited dose dependently by donepezil. It is concluded that donepezil may be a valuable tool with which to influence both the pre- and the postsynaptic acetylcholinesterase-positive structures in the human and rat central and peripheral nervous systems.

Acetylcholinesterase↗

PubMed2005

Ortho effects for inhibition mechanisms of butyrylcholinesterase by o-substituted phenyl N-butyl carbamates and comparison with acetylcholinesterase, cholesterol esterase, and lipase.

Phenyl carbamates are used to treat Alzheimer's disease. These compounds inhibit acetylcholinesterase and butyrylcholinesterase. The goal of this work was to determine the chemical characteristics of ortho substituents that make some carbamates better inhibitors of butyrylcholinesterase than of acetylcholinesterase, cholesterol esterase, and lipase. The inhibition constants, Ki, Ki', kc, and ki were measured for nine different carbamates. The values were plotted according to Hammett, Taft-Kutter-Hansch, and Swan-Lupton to obtain constants that correlated the chemical nature of the substituents with inhibition potency. It was found that the negative charges of tetrahedral intermediates were more stabilized by ortho electron-withdrawing substituents of the inhibitors in butyrylcholinesterase than in acetylcholinesterase. This result confirmed formation of 3-pronged hydrogen bonds for the oxyanion hole of butyrylcholinesterase and 2-pronged hydrogen bonds for the oxyanion hole of acetylcholinesterase. Furthermore, it was found that ortho electron-donating substituents of the inhibitors accelerated inhibition of butyrylcholinesterase by ortho polar effects. Conformations of enzyme-inhibitor tetrahedral intermediates for butyrylcholinesterase were different from those for acetylcholinesterase and cholesterol esterase; ortho substituents in the tetrahedral intermediates were located far from the negatively charged carbonyl oxygens in butyrylcholinesterase, but close to the negatively charged carbonyl oxygens in acetylcholinesterase and cholesterol esterase. In conclusion, electron-donating substituents in the ortho position were better inhibitors of butyrylcholinesterase than acetylcholinesterase, while electron-withdrawing substituents were better inhibitors of acetylcholinesterase.

Acetylcholinesterase↗

PubMed2000

Changes of acetylcholinesterase activity in brain areas and liver of sucrose- and ethanol-fed rats.

The effects of chronic ethanol or sucrose administration to rats on acetylcholinesterase from brain and liver were investigated. Membrane-bound and soluble acetylcholinesterase activities were determined in fractions prepared by centrifugation. The thermal stability and the effects of temperature and different types of alcohols on acetylcholinesterase activity were also studied. Membrane-bound acetylcholinesterase activity increased (p < 0.01) in the liver after chronic ethanol administration, whereas no differences among groups in the encephalic areas, except in the brain stem soluble form, were found. Membrane-bound acetylcholinesterase from the ethanol- and sucrose-treated groups was more stable at the different temperatures assayed between 10 and 50 degrees C than that corresponding to the control group. Non-linear Arrhenius plots were obtained with preparations of membrane-bound acetylcholinesterase from rat liver, with discontinuities at 30 degrees C (control or sucrose groups) or 34-35 degrees C (alcohol group). Assays made with membrane-bound or soluble enzyme from brain showed linear Arrhenius plots in all groups studied. The inhibitory effects of increasing concentrations of ethanol, n-propanol and n-butanol on acetylcholinesterase preparations from forebrain, cerebellum, brain stem and liver of the three experimental groups (control, sucrose-fed and ethanol-fed) were very similar. However, n-butanol displayed a biphasic action on particulate or soluble preparations of rat forebrain. n-butanol inhibited (competitive inhibition) at higher concentrations (250-500 mM), while at lower concentrations (10-25 mM), the alcohol inhibited at low substrate concentrations but activated at high substrate concentration. These results suggest that the liver is more affected by ethanol than the brain. Moreover, the lipid composition of membranes is probably modified by ethanol or sucrose ingestion and this would affect membrane fluidity and consequently the behaviour of acetylcholinesterase.

1-Butanol↗

PubMed2001

Interactions of rat brain acetylcholinesterase with the detergent Triton X-100 and the organophosphate paraoxon.

Inhibition of the critical enzyme acetylcholinesterase (E.C. 3.1.1.7) with subsequent cholinergic crisis is the mechanism of acute toxicity of the organophosphorus insecticides (B. E. Mileson et al., 1998, Toxicol. Sci.41, 8-20). Consequently, measurement of acetylcholinesterase activity is important for evaluating the mammalian toxicity of this commonly used class of insecticides. While mammalian acetylcholinesterase activity has often been determined in tissue homogenates in the presence of the nondenaturing detergent Triton X-100 at a concentration of 1%, the potential actions of this detergent on the activity of this critical enzyme are not understood. In the current study, homogenization of rat brain in buffer containing 1% Triton X-100 slightly elevated the (app)V(max) for hydrolysis of acetylthiocholine, without affecting the (app)K(m) or the (app)K(ss). However, the presence of both 1% Triton X-100 and paraoxon (at concentrations of 5 nM-100 nM) resulted in complex kinetic interactions with acetylcholinesterase, as evidenced by a curvilinear secondary plot for determination of the (app)k(i). These results suggest that measurement of acetylcholinesterase activity in the presence of up to 1% Triton X-100, but in the absence of oxon, should pose no problems with regard to data interpretation, provided it is recognized that the detergent slightly elevates activity. However, measurement of acetylcholinesterase activity after enzyme was exposed simultaneously to Triton X-100 and oxon could be problematic. Caution is warranted when interpreting data where acetylcholinesterase activity was determined under such conditions since in the presence of 1% Triton X-100, the capacity of oxon to inhibit acetylcholinesterase might change as a function of oxon levels.

Acetylcholine↗

PubMed1989

Different membrane-bound forms of acetylcholinesterase are present at the cell surface of hepatocytes.

In the present study we have determinated the acetylcholinesterase molecular forms present in rat liver hepatocytes; we have also studied the association of acetylcholinesterase with the cell surface of the hepatocytes. Subcellular fractionation indicated that rough endoplasmic reticulum and plasma-membrane-enriched fractions contains G4 and G2 acetylcholinesterase forms bound to membranes. Hepatocytes incubated with phosphatidylinositol-specific phospholipase C released about 70% of the surface acetylcholinesterase. Sedimentation analysis showed that all the solubilized acetylcholinesterase activity comes exclusively from a G2 dimer. The G4 hydrophobic form of acetylcholinesterase accounts for the additional cell-surface activity. The existence of these two forms of acetylcholinesterase on the surface of hepatocytes was further established by analyzing the phosphatidylinositol-specific phospholipase C sensitivity of the acetylcholinesterase molecular forms present in isolated rat liver plasma membranes.

Acetylcholinesterase↗

PubMed1983

Isolation of the secretory form of soluble acetylcholinesterase by using affinity chromatography on edrophonium-Sepharose.

A single molecular form of soluble acetylcholinesterase was isolated from a variety of mammalian tissues by use of a novel affinity matrix. This matrix was synthesised by coupling the reversible cholinesterase inhibitor, edrophonium chloride, to epoxy-activated Sepharose. This simple synthesis produced a matrix which was exceptionally stable and had the novel property of selectively binding only one molecular form of acetylcholinesterase. Soluble proteins from a variety of mammalian tissues, including brain, adrenal glands, cerebrospinal fluid, and blood, were separated by centrifugation. These contained combinations of acetylcholinesterase (EC 3.1.1.7) and cholinesterase (EC 3.1.1.8), varying from a single form of acetylcholinesterase to multiple forms of both acetylcholinesterase and cholinesterase. The edrophonium-Sepharose matrix bound only one form of acetylcholinesterase. This form of acetylcholinesterase corresponded in molecular size and electrophoretic mobility to the unique form found in cerebrospinal fluid, i.e. secretory acetylcholinesterase. Cholinesterase was not bound to the matrix.

Acetylcholinesterase↗

PubMed1989

Secretion of acetylcholinesterase and butyrylcholinesterase from the guinea-pig isolated ileum.

1. Strips of longitudinal muscle from guinea-pig ileum, retaining Auerbach's plexus, were superfused with oxygenated Krebs solution. Addition of 50 mM KCl led to a pronounced Ca2+-dependent increase in the activities of both acetylcholinesterase and non-specific cholinesterase (butyrylcholinesterase) in the perfusate but with no change in lactate dehydrogenase activity. 2. No release of acetylcholinesterase, either spontaneous or K+-evoked was observed in tissue freed of the nerve plexus, although release of butyrylcholinesterase still occurred. 3. Carbachol induced a marked Ca2+-dependent increase in the release of acetylcholinesterase but had no effect on the release of butyrylcholinesterase or lactate dehydrogenase. This carbachol-evoked increase in acetylcholinesterase release was blocked by hexamethonium but not by atropine. 4. Four readily soluble molecular forms of acetylcholinesterase and three soluble molecular forms of butyrylcholinesterase were present in innervated longitudinal muscle strips, but insignificant amounts of acetylcholinesterase were detected in denervated strips of muscle. Only one of the four molecular forms of acetylcholinesterase was recovered in the perfusates. 5. It is concluded that acetylcholinesterase is secreted from the nerves of Auerbach's plexus in response to depolarizing stimuli or to nicotinic cholinergic stimulation, while butyrylcholinesterase is secreted from non-neural elements, possibly the longitudinal muscle cells, of guinea-pig ileum in response to a depolarizing stimulus.

Acetylcholinesterase↗

PubMedDate not supplied

Localization of a novel adhesion-promoting site on acetylcholinesterase using catalytic antiacetylcholinesterase antibodies displaying cholinesterase-like activity.

A monoclonal antibody (MAb) raised against human acetylcholinesterase was found to have catalytic activity. A similar phenomenon was observed in a polyclonal antibody raised against the same antigen. The antibodies were demonstrated to be pure, and no contamination with either acetylcholinesterase or butyrylcholinesterase was found. Both antibodies hydrolyzed acetylthiocholine, an acetylcholinesterase substrate, and the MAb followed Michaelis-Menten kinetics. Six other MAbs and one other polyclonal antibody showed no evidence of catalytic activity. Acetylcholinesterase is a key component in the transmission of the nerve impulse, and is also expressed nonsynaptically during embryonic development, and abnormalities in expression are seen in neural tumors and degenerative disorders. This unusual expression is believed to be associated with a novel function of the enzyme related to differentiation and cell adhesion. Autoantibodies to acetylcholinesterase have been observed in a variety of neurologic, muscular, and autoimmune disorders. In an investigation of the possible role of acetylcholinesterase in cell adhesion, we showed that the enzyme promoted neurite outgrowth in neuroblastoma cell lines, and conversely, that certain antiacetylcolinesterase antibodies abrogated cell-substrate adhesion. Interestingly, the antibodies most effective in this regard were catalytic. Preliminary epitope analysis indicated a conformational epitope in the N-terminal domain. This domain contains the active site within a deep gorge and the peripheral anionic site at the rim of the gorge. Peripheral-site inhibitors, but not active-site inhibitors, also interfered with adhesion, and competed with the catalytic monoclonal binding to acetylcholinesterase, indicating that the epitope recognized is associated with the peripheral anionic site. The inhibitor data also support the supposition that catalysis in these antibodies may have arisen from stable complexation of acetylcholinesterase with an inhibitor. We conclude that the catalytic antiacetylcholinesterase antibody interacts with structures associated with the peripheral anionic site, thus defining a novel site on the molecule involved in cell adhesion. This finding has implications for our understanding of the potential importance of this peripheral site in a variety of congenital, neoplastic, and degenerative conditions.

Acetylcholinesterase↗

PubMed1985

Fluorescent phosphonate labels for serine hydrolases. Kinetic and spectroscopic properties of (7-nitrobenz-2-oxa-1,3-diazole)aminoalkyl methylphosphonofluoridates and their conjugates with acetylcholinesterase molecular forms.

The synthesis, kinetic, and spectral characterization of (7-nitrobenz-2-oxa-1,3-diazole)aminoethyl and (7-nitrobenz-2-oxa-1,3-diazole)aminopentyl methylphosphonofluoridate are described. These homologous organophosphorous agents contain the environmentally sensitive 7-nitrobenz-2-oxa-1,3-diazole chromophore. They inhibit acetylcholinesterase from Torpedo at rates exceeding 10(7) M-1 min-1 to form long-lived conjugates with one chromophore/80-kilodalton subunit. The intensity, position, and line width of the absorption spectra of the conjugates and reactivation kinetics in the presence and absence of the bisquaternary oxime 1,1'-trimethylene-bis(4-formylpyridinium bromide) dioxime indicate that these agents form conjugates in which the NBD-aminoalkyl moieties experience distinctive microscopic environments within the active center. NBD-aminoethyl methylphosphono-acetylcholinesterase undergoes oxime-induced as well as spontaneous reactivation at rates that are 3.6 and 35 times faster, respectively, than the corresponding rates measured for the NBD-aminopentyl conjugate. Hence, reactivation exhibits a marked dependence on structure of the methylphosphonate. Fluorescence emission at wavelengths greater than 520 nm is highly quenched and exhibits quantum efficiencies of less than 5%. Absorption maxima for the covalent NBD-aminoethyl methylphosphono-acetylcholinesterase appear at 475-480 nm while those for the corresponding NBD-aminopentyl methylphosphono-acetylcholinesterase appear at 485-490 nm. Bandwidths of the absorption maxima are substantially broader for the acetylcholinesterase adduct with NBD-aminoethyl methylphosphonofluoridate (3870 cm-1) than for the enzyme adduct with NBD-aminopentyl methylphosphonofluoridate (2870 cm-1). The CD spectrum of NBD-aminopentyl methylphosphono-acetylcholinesterase shows optical activity coincident with the shape and position of the absorption spectrum. In contrast, in addition to optically active transitions at the absorption maxima, the CD spectrum of NBD-aminoethyl methylphosphono-acetylcholinesterase shows intense optical activity at 430 nm, a wavelength region coincident with the region of spectral broadening. The spectral properties of alpha-chymotrypsin conjugates formed by reaction with the two probes are different, and the respective spectra differ also from those observed for the acetylcholinesterase conjugates. These results indicate that there is a reciprocal relationship between the structure of the probe and the structure of the active center.(ABSTRACT TRUNCATED AT 400 WORDS)

Acetylcholinesterase↗

PubMed1994

Mutation His322Asn in human acetylcholinesterase does not alter electrophoretic and catalytic properties of the erythrocyte enzyme.

The YT blood group antigen is located on human red blood cell (RBC) acetylcholinesterase. Wild-type acetylcholinesterase, YT1, has histidine at codon 322, whereas the genetic variant of acetylcholinesterase, YT2, has asparagine. This mutation is located within exon 2 of the ACHE gene, an exon that is present in all alternatively spliced forms of acetylcholinesterase. Therefore, acetylcholinesterase in brain and muscle has the same mutation as RBC acetylcholinesterase. We compared the electrophoretic and kinetic properties of RBC acetylcholinesterases having His 322 or Asn 322. We found no differences in the isoelectric point, mobility on non-denaturing gel electrophoresis, affinity for acetylthiocholine, activity per milligram of RBC ghost protein, substrate inhibition constants, binding to the peripheral site ligand (propidium), and binding to active site ligands (tetrahydroaminoacridine and edrophonium). Thus, although the point mutation elicits antibody production in nonmatching blood transfusion recipients, it has no effect on the enzymatic properties of acetylcholinesterase.

Acetylcholinesterase↗

PubMed1994

Differential inhibition of soluble and membrane-bound acetylcholinesterase forms from mouse brain by choline esters with an acyl moiety of an intermediate size.

Differential inhibitions of soluble and membrane-bound acetylcholinesterase forms purified from mouse brain were examined by the comparison of kinetic constants such as a Km value, a Kss value (substrate inhibition constant), and IC50 values of active site-selective ligands including choline esters. Membrane-bound acetylcholinesterase form (solubilized only in the presence of detergent) showed lower Km and Kss values than soluble acetylcholinesterase form (easily solubilized without detergent). Edrophonium expressed a slightly but significantly (p < 0.01) higher inhibition of detergent-soluble acetylcholinesterase form than aqueous-soluble acetylcholinesterase form, while physostigmine inhibited both forms with a similar potency. A remarkable difference in inhibition was observed using choline esters; although choline esters with acyl chain of a short size (acetyl- to butyrylcholine) or a long size (heptanoyl- to decanoylcholine) showed a similar inhibitory potency for two forms of acetylcholinesterase, pentanoylcholine and hexanoylcholine inhibited more strongly aqueous-soluble acetylcholinesterase than detergent-soluble acetylcholinesterase. Thus, it is suggested that the two forms of AChE may be distinguished kinetically by pentanoyl- or hexanoylcholine.

Acridines↗

PubMed1982

Selective inhibition of 'motor endplate-specific' acetylcholinesterase by beta-endorphin and related peptides.

The effects of beta-endorphin (lipotropin 61-91) and related naturally-occurring peptides upon acetylcholinesterase activity in rat hind-limb muscles was investigated. beta-endorphin weakly inhibited the activity in a plasma membrane-enriched fraction. The inhibition by beta-endorphin of the membrane-associated acetylcholinesterase was less marked when the fractions were prepared from muscles which had been denervated 4-6 days previously. The membrane-associated acetylcholinesterase was solubilised from normal muscle preparations and separated by sucrose density gradient centrifugation into three major peaks (16S, 10S and 4S). beta-Endorphin inhibited the activity in the 16S peak but not that in the 10S and 4S peaks, whilst tensilon, a competitive inhibitor of acetylcholinesterase, inhibited the activity of all three peaks. beta-Endorphin inhibited the activity in the 16S peak but not that in the 10S and 4S peaks, whilst tensilon, a competitive inhibitor of acetylcholinesterase, inhibited the activity of all three peaks. beta-Endorphin inhibited the 16S activity in a concentration-dependent manner and its action was partly prevented if naloxone was added simultaneously. Purified natural porcine and bovine beta-endorphin were equipotent in terms of effective concentration range but the maximum inhibition was greater with the bovine peptide. beta-Lipotropin was approximately 4 times less potent than beta-endorphin, whilst C-fragment (lipotropin 61-87) was 100 times less potent. Prolonged treatment with collagenase did not reduce the catalytic activity of 16S acetylcholinesterase, but it was no longer susceptible to the inhibitory action of beta-endorphin. Kinetic studies indicated a complex type of inhibition by beta-endorphin (hyperbolic Lineweaver-Burke plot). Methionine enkephalin inhibited acetylcholinesterase in a weakly non-competitive manner and its action was not abolished if the enzyme was predigested with collagenase. beta-Endorphin produces a novel form of inhibition of acetylcholinesterase, acting only on the 16S (A12 or 'motor endplate-specific') form of the enzyme. The findings are discussed in the light of evidence that beta-endorphin-related immunoreactivity is expressed in motor nerve axons in the immature rat.

Animals↗

PubMed1979

Relationship between inhibition of acetylcholinesterase and response of the rat phrenic nerve-diaphragm preparation to indirect stimulation at higher frequencies.

1 Rat isolated diaphragm preparations were stimulated indirectly either intermittently at 20, 50 or 100 Hz or continuously at 0.2 Hz.2 Addition of 1.8 muM paraoxon (which inhibits acetylcholinesterase by forming a phosphorylated enzyme which undergoes slow spontaneous reactivation) for 5 min to the organ bath produced a failure of the muscle to maintain tetanic tension (tetanic fade, Wedensky inhibition) and potentiated the neuromuscular blocking activity of exogenous acetylcholine. The rates of recovery from both these effects were recorded.3 In a series of experiments with dyflos (which inhibits acetylcholinesterase by forming a phosphorylated enzyme which does not undergo spontaneous reactivation) the relationship between functional acetylcholinesterase activity and neuromuscular blocking activity of exogenous acetylcholine was also determined.4 From the data obtained, the relationship between functional acetylcholinesterase activity and tetanic fade was calculated. These calculations show that (i) a considerable reduction in functional acetylcholinesterase activity is required before the diaphragm loses its ability to respond with a sustained tetanus to indirect stimulation at higher frequencies, (ii) the minimum (critical) level of functional acetylcholinesterase activity required for a normal tetanic response is directly related to the frequency of stimulation and (iii) once functional acetylcholinesterase activity has been reduced to the critical level, a very small further reduction leads to a complete tetanic fade.5 The meaning of functional acetylcholinesterase assays and of conclusions which can be drawn from them, is discussed.

Animals↗

PubMed1994

Expression of dimeric and tetrameric acetylcholinesterase isoforms on the surface of cultured bovine adrenal chromaffin cells.

Acetylcholinesterase is a highly polymorphic enzyme, which can be anchored to the cell surface through several different mechanisms. Dimeric (G2) acetylcholinesterase isoforms are attached by a glycosyl-phosphatidyl-inositol (GPI) linkage, whereas tetrameric (G4) forms are linked through a 20 kilodalton hydrophobic subunit. Although cells of haemopoietic origin contain large amounts of G2 GPI-linked acetylcholinesterase, most tissues express only trace amounts of this isoform. We examined the expression of acetylcholinesterase isoforms in cultured bovine adrenal medullary chromaffin cells. Two major isoforms (G2 and G4) were identified on the cell surface. The G2 isoform, which accounted for approximately half the cell-surface enzyme activity, was linked to the membrane through a GPI anchor. After treatment with diisopropylfluorophosphate to completely inhibit cellular acetylcholinesterase, the G4 isoform was found to be resynthesised and transported to the cell surface more rapidly than the G2 isoform. As the addition of GPI anchors is known to be a very rapid step, this finding suggested that the G2 and G4 isoforms might be transported to the cell surface by two different mechanisms. This conclusion was supported by results from subcellular fractionation experiments. The ratio of G4/G2 membrane-bound acetylcholinesterase varied between different subcellular fractions. The membrane-bound G2 isoform was greatly enriched in a high-speed "microsomal" fraction. G4 acetylcholinesterase is known to be actively secreted by chromaffin cells in culture. Although the G4 isoform was present on the cell surface, most of the secreted enzyme was derived from an intracellular pool. Thus, it is unlikely that the cell-surface G4 isoform contributes significantly to the pool of secreted enzyme. Instead, the expression of two different membrane-bound isoforms may provide a means by which chromaffin cells can target the enzyme to different locations on the cell surface.

Acetylcholinesterase↗

PubMed1984

A non-cholinergic function for acetylcholinesterase in the substantia nigra: behavioural evidence.

Acetylcholinesterase is released from substantia nigra neurons, independently of cholinergic transmission. In an attempt to discover the functional significance of this phenomenon, the behavioural effects of injecting acetylcholinesterase into one substantia nigra of the rat were investigated. Following a single injection of the enzyme, intraperitoneal amphetamine evoked circling behaviour in a direction away from the side of injection. Purified acetylcholinesterase with a similar electrophoretic mobility to the endogenous secreted form, was far more potent in eliciting circling than much higher activities of commercial enzyme, consisting of several molecular species of acetylcholinesterase. Similar infusions of butyrylcholinesterase did not induce circling. Depending upon the amount of enzyme initially given, the behavioural effects of a single injection of acetylcholinesterase persisted for up to thirty days. During this period apomorphine, administered systemically, induced transient circling towards the acetylcholinesterase-treated side. It is concluded that secreted acetylcholinesterase has a functional significance within the substantia nigra, independent of cholinergic transmission. This released enzyme could exert long-term changes in the activity of the nigrostriatal system, involving modification of dopamine striatal receptors.

Acetylcholine↗

PubMed1980

Differential behaviour of eel and erythrocyte acetylcholinesterase on N-methylacridine affinity columns. Importance of ligand affinity and concentration.

The retention and elution of acetylcholinesterase from bovine erythrocytes and electric eel on N-methylacridinium affinity columns have been compared at various ligand concentrations. A soluble 7.7 S dimeric form of bovine erythrocyte acetylcholinesterase required a ligand concentration of 2.0-2.8 mumol/ml in 0.1 M NaCl for retention, compared to 0.44 mumol/ml for various forms of the electric eel acetylcholinesterase. The difference in the retention of acetylcholinesterase from these two sources could not be explained by differences in their oligomeric structure. The affinity of bovine erythrocyte acetylcholinesterase for N-methylacridinium was 13-fold or more lower than the electric eel acetylcholinesterase at similar ionic strengths. N-Methylacridinium appeared to react selectively with the catalytic anionic site of both enzymes. It was concluded that the affinity of the side arm ligand was the major determinant of the differences in the retention properties of the eel and erythrocyte acetylcholinesterase. The difference in affinity for N-methylacridinium probably reflects differences in the organic cation binding region of the two enzymes.

Acetylcholinesterase↗

PubMed1982

HI-6: reactivation of central and peripheral acetylcholinesterase following inhibition by soman, sarin and tabun in vivo in the rat.

HI-6, ([[[(4-aminocarbonyl)pyridino]methoxy]methyl]-2-] (hydroxyimino)methyl]-pyridinium dichloride), is an oxime which, when combined with atropine, is an extremely effective therapy against organophosphate poisoning. It was found that, following soman (287 micrograms/kg) poisoning, HI-6 reactivated acetylcholinesterase in the diaphragm and intercostal muscles but not in the brain. At a lower dose of soman (110 micrograms/kg), HI-6 reactivated sarin-inhibited acetylcholinesterase in the brain and in the respiratory musculature but did not reactivate tabun-inhibited acetylcholinesterase. It was also found that soman produced a differential inhibition of diaphragm and intercostal muscle acetylcholinesterase in vivo, whereas the in vitro I50 for soman was the same in both areas. HI-6 was capable of reactivating soman-inhibited acetylcholinesterase when administered up to 30 min post-soman, indicating that the rate of aging of the soman-acetylcholinesterase complex is slower than previously reported. The above results suggest that, in severe soman poisoning, the primary lesion occurs in peripheral acetylcholinesterase in the respiratory musculature (specifically the diaphragm).

Acetylcholinesterase↗

PubMed1984

Stereospecific reactivation by some Hagedorn-oximes of acetylcholinesterases from various species including man, inhibited by soman.

Reactivation by bispyridinium mono-oximes (Hagedorn-oximes) and some classical oximes (0.03 or 1mM) was studied in vitro of rat, bovine and human erythrocyte acetylcholinesterase and of electric eel acetylcholinesterase inhibited by soman. Relative reactivating potencies of the oximes are similar for the three inhibited erythrocyte enzymes. In general, Hagedorn-oximes are more potent than the classical oximes. Among the Hagedorn-oximes, HI-6 is the most potent reactivator for the three inhibited enzymes. Relative reactivating potencies for the inhibited erythrocyte acetylcholinesterases and electric eel acetylcholinesterase, however, clearly differ. Since the reactivation experiments were carried out with racemic soman, a mixture of the two inhibited enzymes may be formed, which may cause additional problems in the comparison of various results. In order to get more detailed information on differences between human erythrocyte and electric eel acetylcholinesterase, reactivation of these enzymes inhibited with the P(-)-isomers of C(+)- and C(-)-soman were studied separately. Reactivation appeared to be dependent on the chirality of the alpha-carbon atom in the pinacolyl group. HI-6 is by far the most potent reactivator for the human enzyme inhibited by the two P(-)-isomers. It is suggested that electric eel acetylcholinesterase is not a reliable model for in vitro testing of therapeutic potencies of oximes against soman intoxication in mammals. Rate constants of aging of the four acetylcholinesterases inhibited with racemic soman and of the human and eel enzyme inhibited by the P(-)-isomers of C(+)- and C(-)-soman were also determined. The aging of the inhibited rat enzymes proceeds remarkably slowly (t1/2 = 21 min). The rate of aging is not affected by the chirality on the alpha-carbon atom in the pinacolyl group. Consequences of the present results are discussed in view of extrapolation of reactivation data of a series of reactivators to their relative therapeutic effect, ultimately in man. It is speculated that the more rapid aging of the human inhibited enzyme may hamper oxime-therapy in man more seriously than in rat.

Acetylcholinesterase↗