Appearance Of Acetylcholinesterase Research Articles

Acetylcholinesterase and responses to acetylcholine in the embryonic chicken heart.

Three and 4 day old embryonic chicken hearts were examined for their responsiveness to acetylcholine and presence of acetylcholinesterase (AChE) to determine the role of the enzyme in the cardiac effects of the transmitter. The effects of acetylcholine on rate and contractility of 3 day old hearts were indistinguishable from those on 4 day old hearts. The effects were readily blocked by atropine at both stages of development. In 3 day old hearts the responses to acetylcholine were not affected by the AChE inhibitor physostigmine but in 4 day old hearts they were considerably potentiated. The effect of acetylcholine on the rates of 4 day old hearts is of short duration (5 min or less). In 3 day old hearts it persists for a much longer time. Thus, the appearance of AChE in the embryonic heart of the chicken does not seem to modify the responsiveness of the cholinergic receptor to the transmitter.

Early appearance of acetylcholinergic, serotoninergic, and peptidergic neurons and fibers in the developing human central nervous system.

Animal experiments have already shown that neurotransmitters and neuropeptides are not only important for normal functioning of the adult central nervous system (CNS) but are also crucial to its development. However, information on the spatio-temporal distribution of these endogenous substances in the developing human CNS is still scarce. With the use of immunocytochemical staining and a constant supply of properly fixed human abortuses from southern China, an early appearance of acetylcholinesterase, enkephalin, and substance P immunoreactivities was detected first in the spinal cord (weeks 5 to 7 of gestation), then in the brainstem nuclei (weeks 11 to 12). Their overlapping localizations in many regions of the CNS suggest possible interactions among neurons containing these substances, which are in turn important for the proper establishment of the neuronal circuitry. Immunoreactivity for neuropeptide Y appeared initially in the lateral region of upper segments of the spinal cord at week 12 of gestation, then spread latero-medially and cranio-caudally to the sacral region. In the hippocampus, neuropeptide Y neurons appeared from week 15 onwards. Serotoninergic neurons were found in the dorsal raphe nucleus at week 10 and then decreased in number as the fetus grew older. Somatostatin releasing inhibitory factor, vasopressin, and oxytocin were detected in the hypothalamus from weeks 12 to 14 onwards, and monoamine oxidase, succinic dehydrogenase, parvalbumin, calbindin D28K, and vasoactive intestinal peptide were found in the visual cortex at midgestation. The early appearance and the abundance of the neurotransmitters and neuropeptides in the developing CNS indicate that they may play a key role in neuronal differentiation.

Early appearance of acetylcholinergic, serotoninergic, and peptidergic neurons and fibers in the developing human central nervous system

Animal experiments have already shown that neurotransmitters and neuropeptides are not only important for normal functioning of the adult central nervous system (CNS) but are also crucial to its development. However, information on the spatio-temporal distribution of these endogenous substances in the developing human CNS is still scarce. With the use of immunocytochemical staining and a constant supply of properly fixed human abortuses from southern China, an early appearance of acetylcholinesterase, enkephalin, and substance P immunoreactivities was detected first in the spinal cord (weeks 5 to 7 of gestation), then in the brainstem nuclei (weeks 11 to 12). Their overlapping localizations in many regions of the CNS suggest possible interactions among neurons containing these substances, which are in turn important for the proper establishment of the neuronal circuitry. Immunoreactivity for neuropeptide Y appeared initially in the lateral region of upper segments of the spinal cord at week 12 of gestation, then spread latero-medially and cranio-caudally to the sacral region. In the hippocampus, neuropeptide Y neurons appeared from week 15 onwards. Serotoninergic neurons were found in the dorsal raphe nucleus at week 10 and then decreased in number as the fetus grew older. Somatostatin releasing inhibitory factor, vasopressin, and oxytocin were detected in the hypothalamus from weeks 12 to 14 onwards, and monoamine oxidase, succinic dehydrogenase, parvalbumin, calbindin D28K, and vasoactive intestinal peptide were found in the visual cortex at midgestation. The early appearance and the abundance of the neurotransmitters and neuropeptides in the developing CNS indicate that they may play a key role in neuronal differentiation. Microsc. Res. Tech. 45:389–400, 1999. © 1999 Wiley-Liss, Inc.

Acetylcholine as a Signaling System to Environmental Stimuli in Plants. I. Contribution of Ca2+ in heat-stressed Zea mays seedlings.

Plant growth is influenced by transport of water, ions and hormones under various environmental stresses. If acetylcholine (ACh) mediates the opening and/or closing of ion channels in plants just as in animal systems, it is important to detect such evidence. The effect of heat stress on the acetylcholinesterase (AChE) activity and Ca2+ as a trigger of ACh release can be determined in selected organs of dark grown Zea mays seedlings. The enzyme activity in the coleoptile node was 3-7 fold higher than that of other organs. By heat stress, the AChE activity increased about 20% in the node. Cytochemical evidence for AChE activity was found only in the node. This reaction appeared in the cortical cells around the vascular system. After heat stress, the localization of AChE was radicaly changed in the node. AChE activity was detected in all endodermal cells surrounding the vascular system. Further, fluorescent labeled Ca2+ in the node was identified in cortex cells around the vascular system, epidermis and adhering peripheral cortical cells with a laser scanning confocal microscope. Following heat stress, Ca2+ was found more in the cortical cells and whole endodermal cells between the cortex and stele. The appearance of AChE and Ca2+ in endodermal cells after heat stress seems to be correlated with ACh function controlling ion channels.

Distribution pattern of acetylcholinesterase in early embryonic chicken hearts

To study the developmental appearance of acetylcholinesterase in early embryonic hearts, an enzyme-histochemical study was carried out in chicken embryos ranging from cardiogenic plate to late tubular stages. Initially acetylcholinesterase is present in all cells of the (future) myocardium. When 13-14 pairs of somites have developed, i.e., shortly before blood propulsion starts, acetylcholinesterase selectively disappears from the ventral and lateral wall of the developing ventricle. Slightly later, when 18-19 pairs of somites have developed, acetylcholinesterase also disappears from the dorsal and anterior wall of the atrium. High concentrations of acetylcholinesterase remain present in the outflow tract and lower concentrations in a continuous tract along the lesser curvature of the heart, the atrial side of the atrioventricular canal, and the left wall of the atrium. In late tubular stages of heart development, acetylcholinesterase is reexpressed in the inner myocardial layer of the ventricle, i.e., in the developing trabeculae and the ventricular side of the atrioventricular canal, where it is continuous with the acetylcholinesterase-expressing cells of the atrial side of the atrioventricular canal. The expression pattern of acetylcholinesterase in early embryonic chick hearts coincides with that of areas that control the conduction of the impulse and may reveal a cholinergic signal transduction system that is responsible for a coordinated contraction pattern of the myocardium prior to the development of the definitive conductive system.

Development of cholinergic markers in mouse forebrain. I. Choline acetyltransferase enzyme activity and acetylcholinesterase histochemistry

Measurements of choline acetyltransferase (ChAT) activity were made during the development of the neocortical cholinergic innervation, and correlated with the development of the acetylcholinesterase (AChE) staining pattern in mouse cerebral cortex and several other areas of the forebrain between the time of initial onset and maturity. ChAT activity can first be measured on postnatal day 6 (P6). The enzyme reaches 40% of adult activity by P18 and adult values by 7 weeks postnatal. The onset of AChE staining varies for different regions of the forebrain and for various areas within the cerebral cortex. The earliest appearance of AChE is seen in several basal forebrain nuclei including the striatum, the ventromedial region of the globus pallidus and the hypothalamus on embryonic day 18 (E18). In neocortex and olfactory cortex, AChE-stained axons are seen in the white matter before birth, but do not enter cingulate cortex and hippocampus until P2. By P2, almost all areas of the basal forebrain and diencephalon have acquired some AChE staining pattern. The adult distribution of AChE staining is reached by 3 weeks postnatal in all areas of the forebrain. Adult cerebral cortex shows a characteristic pattern of alternating AChE dense and AChE sparse bands which vary in depth depending on the cortical area. The cortical banding pattern develops in an ‘inside-out’ fashion, starting in layer VI and gradually entering more superficial layers. In parallel with the AChE pattern of development in cortex, transient AChE staining can be observed in some thalamic nuclei and in some forebrain fiber systems. In the neostriatum patches of intense AChE staining first develop along the ventrolateral border, then spread throughout the whole nucleus and finally coalesce to a uniform high density over the entire neostriatum. We discuss the close spatial and temporal correspondence between AChE pattern development and reported data on synapse formation, and speculate on the role of the cortical cholinergic system in development.

Membrane-Bound Form of Acetylcholinesterase Activated during Postnatal Development of the Rat Somatosensory Cortex

We are interested in the expression of synapse-specific macromolecules and its correlation with the appearance of neuronal types and synaptogenesis during development of the mammalian brain. We report here studies showing that the appearance of acetylcholinesterase (AChE) at layer IV of the rat somatosensory cortex is correlated with the expression of a membrane-bound AChE. Both its electrophoretic mobility and its sedimentation coefficient remain unaltered during maturation; however, its kinetics parameters, the heat and fixative sensitivities and the substrate inhibition changed through development. Our results suggest that an adult form of membrane-bound AChE is expressed postnatally.

Factors influencing the cholinesterases of cerebrospinal fluid in the anaesthetized cat

Both acetylcholinesterase and non-specific cholinesterase are found in cerebrospinal fluid and blood plasma of the cat; the ratio of activities acetylcholinesterase/non-specific cholinesterase is about 1.5 in cerebrospinal fluid and 0.15 in plasma. A search was made for factors capable of influencing the concentration of the two cholinesterases in cerebrospinal fluid. Either the ventricular system was perfused with artificial cerebrospinal fluid from a lateral ventricle to the aqueduct, or the atlanto-occipital membrane was punctured and cerebrospinal fluid was collected continuously from the cisterna magna. Factors studied included: (a) procedures affecting the composition or formation of cerebrospinal fluid, such as changes in the ionic constituents of the perfusate, the inhibition of cerebrospinal fluid formation by acetazolamide or ouabain, or the rapid intra-carotid infusion of hypertonic urea; (b) arousal (noise or stimulation of the central ends of the sciatic nerves), or deepening of anaesthesia; (c) changes in blood pressure; (d) central stimulants and depressants, pyrogens, prostaglandins, antagonists of acetylcholine. Whereas most procedures or drugs tested increased the concentration of acetylcholinesterase, some central depressants (e.g. chlorpromazine) reduced, while another (ether) increased the appearance of acetylcholinesterase in the cerebrospinal fluid. The effect of ether was, in all probability, due to damage to the blood-brain barrier. A rise in acetylcholinesterase concentration was obtained upon stimulation of the central ends of the sciatic nerves; this was inhibited by atropine but not by N-methylatropine, indicating that the rise was due to increased nervous activity and not to the circulatory eflects of the stimulation, since the changes in blood pressure caused by the stimulation remained the same after atropine administration. Amphetamine or leptazol raised the levels of acetylcholinesterase but it was not possible to determine whether this was due only to increased central nervous activity, since there was invariably leakage through the blood-brain barrier which by itself would be sufficient to produce the effect. A rise in the level of acetylcholinesterase was seen after administration of pyrogen; this was apparently not a simple effect of warming the body, but due to the action of the pyrogen on centers concerned with temperature control, since warming the animal by external heat failed to produce a similar change. In general the concentration of non-specific cholinesterase in cerebrospinal fluid bore little relation to that of acetylcholinesterase: when changes in the latter were pronounced, movements of non-specific cholinesterase in the same direction were frequent, but nearly always of a smaller magnitude. The failure of hypertonic urea to raise the concentration of non-specific cholinesterase in the perfusates indicates that, in the cat, 2 osM urea is not a reliable means of opening the blood-brain barrier for molecules of this size. In a separate group of perfusion experiments, eserine (3 × 10 −5 M) was added to the artificial cerebrospinal fluid in order to measure the release of acetylcholine. There was the expected fall when anaesthesia was deepened, and a rise after d-amphetamine given i.v., both responses being rapid in contrast with the usual slow changes in the esterases. In general there was no constant relation between release of acetylcholine into, and the appearance of acetylcholinesterase in, the cerebrospinal fluid. In experiments on the desheathed vagi of rabbits there was appreciable leakage of acetylcholinesterase from the surface of the nerves, but electrical stimulation of their proximal ends did not increase the leakage. We conclude that increased acetylcholinesterase content of the cerebrospinal fluid is readily obtained by giving centrally acting drugs or by afferent nervous stimuli. Haemo- and hydrodynamic effects accompany the administration of many drugs and the use of afferent stimuli. It is, therefore, often difficult to decide whether the rises in concentration of acetylcholinesterase are due to increased nervous activity as such and/or to the accompanying circulatory and metabolic events.

Time of origin of the neuroblasts in the neural tube of the chick embryo determined by histochemical observation of acetyl - cholinesterase activity.

The time of origin of the neuroblasts in the neural tube of the chick embryo was determined by histochemical observation of acetylcholinesterase (AChE) activity.In the central nervous system, the neuroblasts appeared first in the caudal half of the rhombencephalon of the 1.5-day chick embryo (13-somite, stage 11). They were round medium-sized cells and located in both the ventral and the dorsal halves of the wall of the neural tube. In the cervical, thoracic and lumbosacral neural tubes, the neuroblasts first appeared at stage 12, 14 and 17 respectively. At these levels, they were large and located in the ventral half of the wall of the neural tube. Occasionally they provided an axon which extended to the ventrolateral edge of the neural tube.The time of origin of the neuroblasts in the neural tube of the chick embryo is more accurately determined by histochemical observation of AChE activity than by 3H-thymidine autoradiography. The time difference between the actual neuroblastic differentiation and the appearance of AChE moderately positive cells is much smaller than that between the actual neuroblastic differentiation and the differentiation determined by 3H-thymidine autoradiography.

Regulation of acetylcholinesterase appearance at neuromuscular junctions in vitro

The appearance of acetylcholinesterase (AChE) at newly formed nerve-muscle synapses depends on synaptic transmission. Synapses form when cultures are grown in the presence of acetylcholine receptor antagonists, but AChE does not accumulate at these synapses. The important component of transmission seems to be muscle activity. Treatment with dibutyryl cyclic GMP mimics muscle activity, directly inducing synaptic AChE appearance.

Cytochemical demonstration of the postnatal development of cholinesterases in the sympathetic ganglion of the rat

The appearance and localization of acetylcholinesterase and butyrylcholinesterase is studied during the development of nerve cells in the superior cervical ganglion, using cytochemical procedures. Electron microscopic evidence indicates a relationship between the intracellular appearance of acetylcholinesterase and the maturation of rat nerve cells.In the synaptic region, the first appearance of acetylcholinesterase activity, presumably of postsynaptic origin, precedes the appearance of presynaptic enzyme activity localized at the axolemma of presynaptic axons.The localization of butyrylcholinesterase activity during the postnatal development of ganglion cells is similar to that observed for acetylcholinesterase. The former enzyme, however, never seems to exhibit any significant activity either at the presynaptic axolemma of nerve endings or at postsynaptic junctional structures. Thus, it is most probably devoid of any synaptically localized function.

Appearance of acetylcholinesterase and creatine kinase in plasma of normal chickens after denervation

Evidence that acetylcholinesterase (AChE) activity is released from normal chick embryonic muscle fibers and from muscles of chickens with inherited muscular dystrophy suggested that denervated chick muscles, which have AChE properties similar to dystrophic muscles, would also release AChE. Bilateral denervation of the breast and wing muscles of normal chickens was followed by the appearance of AChE activity, distinguished from plasma cholinesterase by differential substrate hydrolysis, inhibitor sensitivity, and electrophoretic migration. Plasma creatine kinase (CK) activity was also elevated after denervation.

Effects of a snake alpha-neurotoxin on the development of innervated skeletal muscles in chick embryo.

The evolution of the cholinergic (nicotinic) receptor in chick muscles is monitored during embryonic development with a tritiated alpha-neurotoxin from Naja nigricollis and compared with the appearance of acetylcholinesterase. The specific activity of these two proteins reaches a maximum around the 12th day of incubation. By contrast, choline acetyltransferase reaches an early maximum of specific activity around the 7th day of development, and later continuously increases until hatching. Injection of alpha-toxin in the yolk sac at early stages of development causes an atrophy of skeletal and extrinsic ocular-muscles and of their innervation. In 16-day embryos treated by the alpha-toxin, the endplates revealed by the Koelle reaction are almost completely absent. The total content and specific activities of acetylcholinesterase and choline acetyltransferase in atrophic muscles are markedly reduced.

Subcellular distribution of sodium-potassium adenosine triphosphatase, acetylcholine and acetylcholinesterase in developing rat brain

Summary The normal pattern of appearance of acetylcholinesterase, acetylcholine and sodium-potassium adenosine triphosphatase has been determined in the synaptosomal and microsomal fractions during the development of rat brain. Changes in the concentrations of acetylcholinesterase and acetylcholine were found to be gradual with development whereas that of sodium-potassium adenosine triphosphatase increased dramatically around birth and levelled off during the first 2 weeks after birth. The pattern of appearance of both enzyme systems was different in both subcellular fractions and further studies on the effect of activators and inhibitors on the two enzymes showed no relationship between their activities. The possible interrelations between the synaptosomes and microsomes were discussed.

Acetylcholinesterase activity in the chick embryo spinal cords

Summary 1. The cholinesterase in the spinal cord of the developing chick embryo was found to be almost entirely the specific acetylcholinesterase (AChE). The enzyme was located primarily in the anterior horns of the gray matter. 2. AChE activity was highest in the lumbosacral region of the spinal cord and lower in the brachial and cervical regions. 3. AChE activity was measured from day 5 of development until hatching on day 21. Enzyme activity was low prior to day 7 of development, at which time it increased, reached a maximum level on day 10, after which the enzyme activity decreased and remained on a plateau for the remainder of development. 4. Chick embryo spinal cord was maintained in organ culture for 24 and 48 hours. The AChE activity was relatively high after 24 hours' incubation (79% of noncultured control value), but was only 45% of the control value by 48 hours. 5. The addition of acetylcholine (ACh) 0.02M to the medium reduced the loss of AChE activity after 24 and 48 hours' incubation. 6. The injection of ACh into the yolk sac of fertile eggs had no effect on AChE activity of the whole embryo after 72 or 96 hours' development. 7. The correlation between appearance of AChE in the central nervous system and the onset of neuromuscular function, as well as the implications of enzyme induction in ontogeny, are discussed.

Appearance of biochemical components related to acetylcholine metabolism during the embryonic development of chick brain

The normal pattern of appearance of acetylcholinesterase, acetylcholine, choline acetylase, acetothiokinase and choline during the development of chick embryo brain has been determined. During the early period of development, no dramatic changes occur in the concentration of these components of the acetylcholine metabolizing system. During the period from 13 days to hatching, when nervous function is initiated in the major portions of the chick brain, marked increases occur in the concentrations of acetylcholinesterase, choline acetylase, choline and protein and the acetylcholine concentration increases slightly. The concentration of acetothiokinase remains constant throughout the course of embryonic development. The possible interrelations of these changes are discussed.

Some Physiological Aspects of Organogenesis in the Insect Embryo

AbstractIn the embryo, as in the postembryonic insect, morphological development is accompanied by an increase or change in enzyme activity. The appearance of certain enzymatically active proteins in a particular tissue or organ may indicate when that tissue or organ has become functional. The time and place of appearance of several types of esterase has been determined in the tissues and organs of the developing embryo of the large milkweed bug,Oncopeltus fasciatus(Dall.). The appearance of acetylcholinesterase in the neuropile of the nervous system reflects the morphological development of this system; it may also determine the onset of functional activity. Similarly, the appearance of non-specific esterase in many tissues and organs at a definite time during their morphological development suggests a relationship between their functional differentiation and the appearance of the enzyme. Evidence for such a relationship will remain circumstantial until the physiological significance of these esterases is understood.

Development of the cholinergic system in insect eggs

Precise evidence has been obtained for the presence of acetylcholine, acetylcholinesterase, and choline acetylase in developing eggs of Musca domestica (L.) and Oncopeltus fasciatus (Dall.). During embryogenesis choline acetylase appears first; acetylcholine was detected only after the appearance of acetylcholinesterase which appears late in development. The amounts of substrate and enzymes increase progressively with the development of embryo until the time of hatching.