Neuronal Acetylcholinesterase Research Articles

Lack of inhibition of human plasma cholinesterase and red cell acetylcholinesterase by antimony compounds including stibine.

1. The toxic gas hypothesis proposes exposure to stibine (antimony trihydride) generated from microbial contamination of cot mattress materials as a possible cause of unexplained death in infancy (SIDS) as a consequence of cholinesterase inhibition. We have measured the direct effects of antimony compounds including stibine on the activity of plasma cholinesterase, red blood cell acetylcholinesterase (AChE) and mouse neuronal AChE in vitro. 2. Colorimetric assays for the different esterases with potassium antimonyl tartrate or antimony trichloride at concentrations up to 10(-3) M in the presence of substrate concentrations sufficient to produce 80% of the maximum reaction rate produced no inhibition of enzyme activity. Exposure of enzyme preparations to stibine gas at concentrations sufficient to cause denaturation of red cell haemogloblin caused no measurable inhibition of esterase activity. 3. We conclude that stibine gas or soluble antimony compounds are not capable of inhibiting cholinesterase activity at toxicologically relevant concentrations.

CSF cholinesterase activity in demented and non-demented subjects.

Samples of cerebrospinal fluid (CSF) were examined, looking mainly at total cholinesterase (ChE) and acetyl-cholinesterase (AChE) levels from 139 living subjects. At the completion of the study, 35 of the 139 patients had died and pathological confirmation of the presence of dementia had been obtained. These results, together with results from other laboratories, provide evidence that a low CSF ChE level presenting in demented patients may indicate a depletion of the brain AChE system, and this may confirm a clinical diagnosis of AD as well as other types of dementia which are associated with an alteration of the brain AChE system. The overlap in the levels of CSF biochemical markers between demented and non-demented subjects which has led to many conflicting reports has always disappointed investigators. It is suggested that some 'control' subjects with CSF ChE activity indistinguishable from that in AD patients may have an abnormal ageing process in their brains (brain at risk), although the symptoms of dementia have not yet been detected. Recognition of a pre-clinical or incubation period is very beneficial for explaining discrepancies in biochemistry and pathology in the literature, and must be considered for both the treatment and the prevention of dementia. The long used treatment, which was designed to inhibit AChE, should no longer be used: treatment must be designed to enhance the activity of the neuronal AChE system, or slow its degeneration.

Donepezil: an anticholinesterase inhibitor for Alzheimer's disease.

The pharmacology, pharmacokinetics, clinical efficacy, adverse effects, drug interactions, and dosage and administration of donepezil are reviewed. Donepezil is a synthetic noncovalent reversible inhibitor of acetylcholinesterase (AChE) for the treatment of mild to moderate dementia associated with Alzheimer's disease. In contrast to tacrine hydrochloride, the only comparable agent currently approved by FDA, donepezil exhibits a relatively high degree of selectivity for neuronal AChE as opposed to butyrylcholinesterase. It has a half-life of 60 hours in young adults and 104 hours in elderly patients. In clinical trials, donepezil has been associated with significant improvements in Alzheimer's Disease Assessment Scale-cognitive subscale and Clinical Interview-Based Impression of Change scores. The most common adverse effects associated with donepezil are nausea, diarrhea, anorexia, and vomiting, which are most likely to occur during dose initiation or adjustment. Hepatotoxicity, a dose-limiting adverse effect that sometimes requires discontinuation of tacrine, has not been reported with donepezil. Donepezil does not appear to interact with theophylline, cimetidine, warfarin, or digoxin. Ketoconazole and quinidine inhibit the metabolism of donepezil in vitro, but there is a lack of clinical data showing that these drugs decrease the clearance of donepezil. The initial recommended dosage is 5 mg daily before bedtime, with a dosage increase to 10 mg after four to six weeks according to the patient's response and tolerance. Donepezil appears to be preferable to tacrine as the initial agent for patients with mild to moderate dementia associated with Alzheimer's disease.

Acetylcholinesterase-transgenic mice display embryonic modulations in spinal cord choline acetyltransferase and neurexin Ibeta gene expression followed by late-onset neuromotor deterioration.

To explore the possibility that overproduction of neuronal acetylcholinesterase (AChE) confers changes in both cholinergic and morphogenic intercellular interactions, we studied developmental responses to neuronal AChE overexpression in motoneurons and neuromuscular junctions of AChE-transgenic mice. Perikarya of spinal cord motoneurons were consistently enlarged from embryonic through adult stages in AChE-transgenic mice. Atypical motoneuron development was accompanied by premature enhancement in the embryonic spinal cord expression of choline acetyltransferase mRNA, encoding the acetylcholine-synthesizing enzyme choline acetyltransferase. In contrast, the mRNA encoding for neurexin-Ibeta, the heterophilic ligand of the AChE-homologous neuronal cell surface protein neuroligin, was drastically lower in embryonic transgenic spinal cord than in controls. Postnatal cessation of these dual transcriptional responses was followed by late-onset deterioration in neuromotor performance that was associated with gross aberrations in neuromuscular ultrastructure and with pronounced amyotrophy. These findings demonstrate embryonic feedback mechanisms to neuronal AChE overexpression that are attributable to both cholinergic and cell-cell interaction pathways, suggesting that embryonic neurexin Ibeta expression is concerted in vivo with AChE levels and indicating that postnatal changes in neuronal AChE-associated proteins may be involved in late-onset neuromotor pathologies.

The significance of the activity of CSF cholinesterases in dementias

This article reviews the significance of changes in the level of cerebrospinal fluid acetylcholinesterase or cholinesterase in patients with Alzheimer's disease or other dementias. Evidence has shown that the methodology of assaying cerebrospinal fluid acetylcholinesterase or cholinesterase is reliable and the activity of the enzyme is stable. Low acetylcholinesterase or cholinesterase levels presenting in cerebrospinal fluid of a demented individual may confirm the clinical diagnosis of Alzheimer's disease or other organic dementia. A low activity of acetylcholinesterase or cholinesterase existing in cerebrospinal fluid of a non-demented individual may indicate a brain at risk, or that the person is in the preclinical stage of dementia. Recognition of the presence of the preclinical stage may be very beneficial for explaining the real meaning of the ‘overlap’ in the biochemistry and pathology between dementia and non-dementia, and also very important for prevention and treatment. Therefore, the strategy of prevention and of treatment should no longer be designed to inhibit acetylcholinesterase activity. In contrast, it should be designed to enhance the neuronal acetylcholinesterase activity or to delay the degeneration of brain acetylcholinesterase system.

Comparative effects of the metabolic inhibitors 2,4-dinitrophenol and iodoacetate on mouse neuroblastoma cells in vitro

The toxic effects of two metabolic inhibitors, dinitrophenol and iodoacetic acid, were compared. Mouse neuroblastoma cell cultures (Neuro-2a) were exposed to different concentrations of the toxic compounds for 24, 48 and 72 h to study basal toxicity effects (cell proliferation by quantification of total protein content (PR) and relative neutral red uptake (RNRU) by lysosomes). The following biochemical indicators assessed in the in vitro test system were: cytosolic phosphofructokinase (PFK) and enolase (ENL) activities in glycolysis; mitochondrial succinate dehydrogenase (SDH) activity in the citric acid cycle; lysosomal β-galactosidase (GAL) activity; and neuronal acetylcholinesterase (AChE) activity. The effects of the two metabolic inhibitors on the various indicators differed. Iodoacetic acid was found to be far more toxic than dinitrophenol to neuroblastoma cell proliferation at 24 h exposure. Though 2,4-dinitrophenol and iodoacetic acid both inhibited cell proliferation of the neuroblastoma cells, their effects on the other endpoints were opposite. Dinitrophenol was a general activator of the metabolism, particularly affecting lysosomal function. Iodoacetic acid did not significantly alter general metabolism, but considerably modified lysosomal function and AChE activity. The modification of lysosomal function of Neuro-2a cells by the two compounds was quite different: dinitrophenol increased RNRU and GAL activity, and iodoacetic acid decreased both parameters.

Transgenic expression of human acetylcholinesterase induces progressive cognitive deterioration in mice.

Cognitive deterioration is a characteristic symptom of Alzheimer's disease. This deterioration is notably associated with structural changes and subsequent cell death which occur, primarily, in acetylcholine-producing neurons, progressively damaging cholinergic neurotransmission. We have reported previously that excess acetylcholinesterase (AChE) alters structural features of neuromuscular junctions in transgenic Xenopus tadpoles. However, the potential of cholinergic imbalance to induce progressive decline of memory and learning in mammals has not been explored. To approach the molecular mechanisms underlying the progressive memory deficiencies associated with impaired cholinergic neurotransmission, we created transgenic mice that express human AChE in brain neurons. With enzyme levels up to two-fold higher than in control mice, transgenic mice displayed an age-independent resistance to the hypothermic effects of the AChE inhibitor, paraoxon. In addition to this improved scavenging capacity for anti-AChEs, however, these transgenic mice also resisted muscarinic, nicotinic and serotonergic agonists, indicating that secondary pharmacological changes had occurred. The transgenic mice also developed progressive learning and memory impairments, although their locomotor activities and open-field behaviour remained similar to those of matched control mice. By six months of age, transgenic mice lost their ability to respond to training in a spatial learning water maze test, whereas they performed normally in this test at the age of four weeks. This animal model is therefore suitable for investigating the transcriptional changes associated with cognitive deterioration and for testing drugs that may attenuate progressive damage. We conclude that upsetting cholinergic balance may by itself cause progressive memory decline in mammals, suggesting that congenital and/or acquired changes in this vulnerable balance may contribute to the physiopathology of Alzheimer's disease.

Developmentally transient expression of acetylcholinesterase within cortical pyramidal neurons of the rat brain

Using a histochemical method for the visualization of cholinesterase activity in neurons, we have observed developmentally transient expression of acetylcholinesterase (AChE) in cortical pyramidal neurons of the rat brain. Depending on the extent of the deposition of AChE reaction product, several types of cortical neurons could be visualized. We designated neurons with moderate-to-high staining intensity as AChE H and neurons with relatively lower staining intensity as AChE L. At birth (P0), very little AChE activity was found within cortical neurons. Between P1–P4, there was a gradual emergence of AChE-stained cortical neurons. At this stage, the majority of these neurons were of the AChE L type. At P5–P7 we observed an abrupt increase in AChE-stained cortical neurons. The number and the staining intensity of these neurons was at a peak at P8–P10. At this age range, the majority of these neurons were of the AChE H variety and displayed morphological characteristics of cortical pyramidal neurons. At P11–P15, there was an abrupt decrease in the number of AChE H neurons. After P15, the density and staining intensity of cortical AChE-positive (cholinergic) axons gradually increased. Nevertheless, AChE L pyramidal neurons were detected through these fibers up to P21. At P21, a dense plexus of AChE-positive axons was observed in all cortical areas while very little AChE reaction product was visible in pyramidal neurons, and this pattern continued into adult life. When the adult cortex was denervated from its AChE-positive axons by lesions of the nucleus basalis magnocellularis, many AChE L pyramidal neurons were uncovered. However, virtually no AChE H cortical pyramidal neurons were observed in such preparations. Developmentally transient AChE activity within these cortical pyramidal neurons might contribute to their maturation and establishment of synaptic connections.

Acetylcholinesterase activity and post-lesional plasticity in the hippocampus of young and aged rats

Applying quantitative microscopic histochemistry, the activity of acetylcholinesterase was determined in the various layers of the rat hippocampus at three different levels along the rostrocaudal extent. Two age groups of animals were examined: young adults (two to three months old) and aged subjects (26 months old). Young adults were divided into controls, and animals killed eight and 35 days following bilateral ibotenate lesioning of the medial septum-diagonal band complex. Aged rats were divided into controls and animals 35 days post-lesion. Analysis of variance revealed that the mean acetylcholinesterase activities of the entire hippocampus of individuals were not significantly different between young and aged rats when averaged across controls and 35 days post-lesion. There was a significant decrease of acetylcholinesterase activity (−52%) in young adults eight days post-lesion as compared to controls, but a significant increase (+63%) took place until 35 days post-lesion as compared to eight days post-lesion. Significantly lower activities existed, however, in young (−22%) and aged rats (−18%) 35 days post-lesion as compared to controls. This decrease in mean activity was not age dependent. As acetylcholinesterase is considered to be a good cholinergic indicator in the hippocampus, the results suggest a homotypic collateral sprouting from spared cholinergic afferents following ibotenate lesion of the medial septum-diagonal band complex in young and aged rats. Based on the data obtained, it is reasonable to assume that there was no difference in the post-lesional plasticity of neuronal acetylcholinesterase between young adult and aged rats.

Tonic activation of locus coeruleus neurons by systemic or intracoerulear microinjection of an irreversible acetylcholinesterase inhibitor: Increased discharge rate and induction of c-fos

Recent studies in this laboratory have demonstrated that intramuscular injection of the irreversible acetylcholinesterase (AChE) inhibitor, soman (pinacolylmethylphosphonofluoridate), produces a rapid (1–2 h) and profound depletion (70% of control) of norepinephrine (NE) in the olfactory bulb and forebrain. NE is decreased only in convulsing animals. As NE-containing locus coeruleus (LC) neurons provide the only NE input to the olfactory bulb and the major NE innervation of the forebrain, the reduction of NE suggests that soman may cause tonic activation of LC neurons leading to rapid depletion of NE. Activation of LC may result from: (i) facilitation of cholinergic transmission in LC; (ii) soman-induced activation of excitatory inputs to LC; or (iii) generalized activation of LC neurons due to seizures. The present experiments were designed to assess these alternatives. We examined whether LC neuronal activity, c-fos expression, and AChE staining are altered after peripheral (systemic) or direct intracoerulear injection of soman in anesthetized rats. Both modes of soman administration rapidly and potently increase the spontaneous discharge rate of LC neurons. This activation was associated with a desynchronization of the electroencephalogram, but not with seizures. The discharge of LC neurons remained elevated at all postsoman intervals examined (up to 2 h) and was rapidly and completely reversed by systemic injection of the muscarinic receptor antagonist scopolamine hydrochloride, but not by the nicotinic receptor antagonist mecamylamine. Both systemic and intracoerulear soman administration completely inhibited AChE staining in LC and rapidly induced the expression of c-fos in LC neurons. These results demonstrate that soman potently and tonically activates LC neurons. This effect appears to be mediated by direct inhibition of AChE in LC leading to a rapid accumulation of ACh. Unhydrolyzed ACh tonically activates LC neurons via muscarinic receptors. Soman-induced activation of LC neurons does not require seizures. We conclude that depletion of forebrain and olfactory bulb NE after systemic administration of soman results from tonic hypercholinergic stimulation of LC.

Effect of intrauterine exposure to cocaine on acetylcholinesterase in primary cultures of fetal mouse brain cells

The effect of in utero exposure to cocaine on the developmental pattern of acetylcholinesterase (AChE), a major regulator of the central nervous system neurotransmitter acetylcholine, was studied in fetal brain cell cultures collected from mice on gestational day 15 after maternal exposure to cocaine from gestational days 6–14. A significant decrease in total and specific activity (expressed per mg of protein) of acetylcholinesterase was seen throughout the culture period in the cells grown from the cocaine-exposed animals as compared to controls ( p < .002). Similarly, the total protein content of the brain cells grown from the cocaine-exposed animals was significantly decreased as compared to controls ( p < .03). Utilizing AChE as a biochemical marker, these studies have shown that in utero exposure to cocaine has an adverse effect on the normal developmental pattern of the production of acetylcholinesterase in cholinergic neurons in the fetal mouse brain.

Acetylcholinesterase-containing neurons, substance P and enkephalin fibers in the ventral horns of developing human embryos and fetuses

The presence of the acetylcholinesterase neurons and substance P-like and enkephalin-like fibers in the various nuclear columns of the ventral horns of the spinal cords was studied in the developing human by acetylcholinesterase histochemistry and substance P and enkephalin immunohistochemistry. Acetylcholinesterase-positive neurons initially appeared in the lateral neuronal columns and eventually were also observed in the medial columns as well as the median columns at various levels of the spinal cord by 10 weeks' gestation. Acetylcholinesterase-positive neurons in the lower sacral levels were not detected until 11–12 weeks' gestation. Diffused substance P- and enkephalin-like fibers were demonstrated as early as 10 weeks' gestation but did not align with any particular nuclear column until after 15 weeks' gestation. These fibers further increased in length and adopted reticular branching patterns and many of these tended to surround the cell bodies of the nuclear columns. Possible interaction of acetylcholinesterase neurons and substance P and enkephalin fibers would commence by 15 weeks' gestation.

A one-step cobalt-ferrocyanide method for histochemical demonstration of acetylcholinesterase activity in central nervous system tissue.

We introduce a one-step histochemical method with cobalt as the precipitating agent for ferrocyanide for the light microscopic demonstration of acetylcholinesterase activity. This method was used to demonstrate acetylcholinesterase in normal cortical fibers and neurons, as well as pathological elements such as plaques and tangles. This procedure can also be easily combined with immunohistochemical methods that use diaminobenzidine as a chromogen.

Electron microscopic localization of cholinesterase activity in Alzheimer brain tissue

Acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) activity was localized by electron microscopic enzyme cytochemistry in cortex from Alzheimer brains and brains from non-demented cases. In the tangle-rich medial temporal cortex of the Alzheimer brain, most of the neuronal AChE was associated with neurofibrillary tangles. These structures also contained BChE activity. In normal neurons AChE activity was found in the rough endoplasmic reticulum, nuclear envelope and Golgi apparatus. Little BChE activity was noted in normal cortex. In neuritic plaques, AChE and BChE activity was associated mostly with the amyloid, but also with the neuritic component.

Neuronal Acetylcholinesterase Levels in Cerebrospinal Fluid and Serum Determined by a Specific and Sensitive Immunoassay

Acetylcholinesterase levels were determined in cerebrospinal fluid and serum of 272 patients with various neurological disorders. The patients were ordered in 13 diagnostic groups. The assay employed was an antigen capture assay based on an immobilized monoclonal antibody selective for neuronal acetylcholinesterase (Rasmussen et al., Clin. Chim. Acta 166 (1987) 17-25 (1)). In 100% of the cases the acetylcholinesterase levels in cerebrospinal fluid were 500-10,000 fold higher than expected for a regular serum-derived protein (0.1-0.5%) at the intact blood-brain barrier. In both compartments the test values showed wide variations, which were much greater in serum than in cerebrospinal fluid. No specific relationship was discernible between the values obtained and any of the diagnostic groups, although decreased levels of acetylcholinesterase were found in bacterial and viral meningitis, and elevated levels were found in groups with more chronic diseases. A possible transfer of acetylcholinesterase from cerebrospinal fluid to blood is discussed.

Quantitative distribution of AChE-positive neurons in the hippocampus of young and aged rats

Acetylcholinesterase (AChE)-positive neurons were counted in the different layers of the rostral (septal) third, the middle third and the caudal (temporal) third of the hippocampus from 3 month (young) and 27 month old rats (aged) using AChE stained cryostat sections. The rats were treated with 3 and 2.5 mg of diisopropylphosphofluoridate/kg body weight, respectively 3 h before sacrifice. The study showed -1) a high numerical density of AChE-positive neurons (10.9 to 18.9 perikarya/mm2) in the hilus (fascia dentata), the str. oriens/pyramidale of CA1 and the subiculum, a particularly low density (less than 0.1 perikarya/mm2) in the str. granulosum and moleculare of the dentate area; -2) a significant (p less than 0.05) linear increase of the numerical density in most of the hippocampal layers from the rostral to the caudal pole; -3) no significant differences between young and aged animals; - and 4) a higher sensitivity to DFP-treatment in aged than in young animals. The distribution of AChE-positive neurons corresponds with the distribution of somatostatin-immunoreactive neurons described in the literature. A modulatory effect on neurotransmission is discussed as a possible function of the AChE in peptidergic neurons.

Localization of acetylcholinesterase positive neurons and substance P and enkephalin positive fibers by histochemistry and immunohistochemistry in the sympathetic intermediate zone of the developing human spinal cord

Localization of acetylcholinesterase positive neurons and substance P and enkephalin fibers were studied by histochemistry and immunohistochemistry in the intermediate sympathetic zone of the spinal cords of 39 human embryos/fetuses from gestation ages five to 40 weeks. Acetylcholinesterase positive neurons were observed in the nucleus intermediolateralis pars principalis as early as the fifth week of gestation. By the ninth to 13th weeks of gestation, positive neurons were also seen in the nuclei intermedialis pars funicularis, intercalatus spinalis and intercalatus pars paraependymalis. Increase in amount of these positive acetylcholinesterase neurons was demonstrated till term. Substance P and enkephalin fibers were initially observed by the eighth gestation week in the intermediolaterlis pars principalis nucleus and positive fibers were then detected in the nucleus intermedialis pars funicularis as well as the nucleus intercalatus spinalis by the 14th week of gestation. By the 26th week of gestation, all the major nuclei intermediolateralis par principalis, intermedialis pars funicularis, intercalatus spinalis and intercalatus pars paraependymalis has substance P and enkephalin fibers. Initial demonstration of acetylcholinesterase positive neurons appeared to be at an earlier stage than that of our substance P and enkephalin positive fibers.

The postnatal expression of acetylcholinesterase in somatostatin-positive cells of mouse hippocampus

The neuroanatomical distributions of acetylcholinesterase (AChE) staining and somatostatin-like immunoreactivity (SOMLI) of neurons intrinsic to the mouse hippocampal formation have been evaluated during postnatal development. Besides the progressive development of neuropil staining for AChE, as a consequence of the septohippocampal innervation, intense AChE staining was also expressed in a subpopulation of neurons intrinsic to the stratum oriens and the hilus of dentate gyrus. In the stratum oriens, the number of AChE-positive cells increased between postnatal day (PND) 3 and PND 10 and declined slightly after PND 21. In the hilus of the dentate gyrus, the number of AChE-stained cell bodies increased progressively until PND 21 when the adult complement was achieved. The AChE-positive neurons of strata radiatum and lacunosum-moleculare, which were few and scattered, increased progressively from PND 7 until adulthood. SOMLI-positive neurons were present in the hippocampal formation by PND 3, and their density showed initial increases followed by decreases in the second to third postnatal week. SOMLI cell distribution on the other hand did not change remarkably during subsequent maturation. Because of the similar developmental time course and localization of AChE and SOMLI neurons, co-localization was assessed by a double-staining method. A large percentage of the neurons staining for one of these markers also stained for the other. In the stratum oriens, from PND 3 to PND 10, the number of SOMLI neurons expressing AChE was increased while a slight decrease from the PND 21 to adulthood was evident. Virtually all SOMLI-positive neurons in the dentate gyrus stained for AChE from PND 7 through adulthood, although the intensity of AChE reactivity declined with maturation.

Ventral and dorsal horn acetylcholinesterase neurons are maintained in organotypic cultures of postnatal rat spinal cord explants

Transverse sections of postnatal rat spinal cord have been cultured using the organotypic roller tube method. These explant cultures retain identifiable anatomical landmarks, allow identification of individual neurons, can be maintained for up to 8 weeks, and undergo maturational changes in vitro. Putative ventral horn motoneurons were identified in these cultures by localization to ventral horn regions analogous to those of motoneurons in vivo and by staining for choline acetyltransferase (ChAT) immunoreactivity and acetylcholinesterase (AChE) activity. Morphometric studies of the photomicrographic areas of cell bodies of these ventralhorn neurons in intact cultures show a range of sizes up to 1635 μm 2 with the average size being245 ± 7 μm 2 ( n = 724) (average ± S.E.M.). The size ranges are roughly comparable to cross-sectional areas determined previously for ventral horn motoneurons in vivo. Dorsal horn regions of these cultures also development AChE activity that was absent at explantation. Biochemical analysis of ChAT and AChE activity in pooled samples of whole cultures showed ChAT activity to be0.48 ± 0.08 ( n = 7) μmol/min/g protein and AChE activity to be2.2 ± 2.0 ( n = 7) μmol/min/g protein at 37°C (averages ± S.E.M.). These values are comparable to previously reported values for neonatal rat spinal cord in situ. Organotypic roller tube cultures of postnatal rat spinal cord provide an attractive system for studies of survival, morphology, growth and differentiation of mammalian ventral horn neurons in vitro.

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 acteylcholinesterase 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.