Changes In Acetylcholinesterase Activity Research Articles

Effects of physostigmine on the cardiopulmonary system of conscious pigs

Physostigmine, as a pretreatment candidate for nerve agent poisoning, was examined for cardiopulmonary side effects. Cardiovascular and pulmonary parameters were monitored in unanesthetized domestic pigs which received pulmonary arterial infusion of 5 μg/kg/min physostigmine salicylate for 2 hr. A level of 74% inhibition of red blood cell (RBC) acetylcholinesterase (AChE) activity was attained in 45 min, and this level of carbamylation increased only slightly during the remaining infusion period. In addition to this large change in AChE activity, minor changes were observed in hematocrit, heart rate, body temperature, mean aortic pressure, pulmonary arterial wedge pressure, and pulmonary artery pressure. Typically, these parameters showed a trend toward elevated levels. Blood gases, pH, respiratory rate, tidal and minute volume, cardiac output, nonelastic resistance, and dynamic compliance were not significantly different from baseline values. The unanesthetized pig responds to physostigmine in a manner similar to that reported for other species and appears to be a suitable model for evaluating cardiopulmonary effects of cholinesterase inhibitors.

Effects of Physostigmine on the Cardiopulmonary System of Conscious Pigs

Effects of Physostigmine on the Cardiopulmonary System of Conscious Pigs. STEMLER, F. W., CORCORAN, K. D., PARRISH, J. H., HURT, H. H., TEZAK-REID, T. M., KAMINSKJS, A., AND JAEGER, J. J. (1990). Fundam. Appl. Toxicol. 14, 96–103. Physostigmine, as a pretreatment candidate for nerve agent poisoning, was examined for cardiopulmonary side effects. Cardiovascular and pulmonary parameters were monitored in unanesthetized domestic pigs which received pulmonary arterial infusion of 5 μg/kg/min physostigmine salicylate for 2 hr. A level of 74% inhibition of red blood cell (RBC) acetylcholinesterase (AChE) activity was attained in 45 min, and this level of carbamylation increased only slightly during the remaining infusion period. In addition to this large change in AChE activity, minor changes were observed in hemato-crit, heart rate, body temperature, mean aortic pressure, pulmonary arterial wedge pressure, and pulmonary artery pressure. Typically, these parameters showed a trend toward elevated levels. Blood gases, pH, respiratory rate, tidal and minute volume, cardiac output, nonelastic resistance, and dynamic compliance were not significantly different from baseline values. The unanesthetized pig responds to physostigmine in a manner similar to that reported for other species and appears to be a suitable model for evaluating cardiopulmonary effects of cholinesterase inhibitors.

Changes in the properties of synaptic channels opened by acetylcholine in denervated frog muscle

Acetylcholine (ACh)-activated channels in end-plates of frog sartorius muscle were studied at various times after denervation. Mean open times of the synaptic membrane channels were derived from the time constant of decay of miniature end-plate currents ( τ MEPC) evoked by ACh quanta released from Schwann cells, which replace the motor nerve terminals after these degenerate. Membrane current noise, elicited by iontophoretic application of ACh to voltage-clamped end-plates, was also used to determine mean open time ( τ noise) and conductance of the ion channels. About 1 week after denervation, soon after Schwann cell MEPCs appeared. they had a τ similar to that of the neural MEPC in innervated end-plates. However, 5–6 weeks after denervation τ MEPC was increased by a factor of about 5. Circa 4 weeks after denervation, cholinesterase activity of the denervated muscle decreased to 76% of that in the contralateral, innervated muscle, and even 4 months after the operation it was still 64%. Thus, it is unlikely that a change in acetylcholinesterase activity is the main factor responsible for the increase in τ of Schwann cell MEPC. About 1 week after denervation τ noise was close to that in innervated end-plates (about 2 ms). Twelve to 24 days after denervation the average channel open time was 4.5 ± 1.0 ms, with some end-plates still showing normal ‘fast’ channels. However, in muscles denervated for 47–113 days the open time was 12.9 ± 1.9 ms. In the early and intermediate periods, ACh-induced noise spectra with two components were obtained from many end-plates, indicating the simultaneous activation of two different types of channels. At some end-plates during the early and intermediate periods after denervation, but not after about 5 weeks, neostigmine caused the appearance of a component, which was as fast as that of normal end-plate channels. In other experiments small doses of α-bungarotoxin were applied in order to predominantly block extra-junctional receptors. In the early period of denervation, when two components were present in the noise spectra, α-bungarotoxin eliminated the slow component leaving channels as fast as in innervated end-plates. After prolonged denervation, a component with τ of about 5.5 ms was occasionally disclosed by application of α-bungarotoxin. τ noise and τ MEPC from the same end-plate closely agreed. Our results indicate that at frog end-plates the open time of the majority of the synaptic channels opened by ACh becomes longer with increasing time after denervation.

Time dependence of Li+ action on acetylcholinesterase activity in correlation with spontaneous quantal release of acetylcholine in rat diaphragm.

The mean miniature end-plate potential (m.e.p.p.) frequency and the acetylcholinesterase (AChE) activity have been examined in the rat diaphragm, in relation to the time of Li+ action. Substitution of LiCl for NaCl caused increase in m.e.p.p. frequency, the final level of which depended on Li+ concentration in the bath solution. Incubation of diaphragm homogenate with NaCl caused initially a slight increase of AChE activity, reaching a plateau after 20 min. Substitution of LiCl for NaCl produced an increase in AChE activity to a maximum, followed by a decrease to a minimum. The activity consequently returned to a steady level. During complete substitution of LiCl for NaCl, the maximum value of AChE activity coincided with the maximum m.e.p.p. frequency (Fmax) induced by Li+; the minimum value correlated with the precise instant that the frequency reached a plateau. The addition of prostigmine caused: a) blockade of AChE activity, and b) prostigmine concentration-dependent decrease of Fmax and/or disappearance of m.e.p.p.s from neuromuscular junctions treated with Li+. These results are discussed in terms of correlation between changes of AChE activity induced by Li+ and variations of spontaneous quantal release of the transmitter from nerve terminals of the rat diaphragm treated with Li+.

Ultrastructural changes in acetylcholinesterase activity in the deafferented spinal trigeminal nucleus.

Acetylcholinesterase (AChE) activity was investigated in synaptic areas of the cat spinal trigeminal nucleus (pars interpolaris and pars caudalis) ipsilateral and contralateral to complete retrogasserian rhizotomy. Vibratome sections of tissue taken from animals of 1, 3, 6, 14, and 21 days survival were examined by electron microscopy following a histochemical reaction for AChE activity employing a method based on the Karnovsky-Roots technique for demonstrating reaction product. As degeneration progressed with survival time, enzymatic activity was initially reduced in synaptic clefts of injured afferent terminals and subsequently was enhanced throughout the extracellular space, including within synaptic clefts of possibly reinnervated sites. These changes in enzymatic activity with primary deafferentation are discussed in relation to the process of reinnervation, the development of neuronal hyperactivity, and possible noncholinergic functions of AChE.

Changes in acethylcholinesterase activity and total protein in the developing fetal brain.

Pre-natal changes in the physiological development of the porcine conceptus indexed by acetylcholinesterase (AChE) activity and total protein content of the fetal brain and amniotic fluid were determined from 4 to 12 weeks of gestation at intervals of two weeks. Marked brain and body development was observed between four and six weeks of gestation. AChE activity in the amniotic fluid declined non-significantly with gestation length while fetal brain AChE activity increased with advancing gestation. Total protein levels in both the amniotic fluid and fetal brain were relatively steady and no significant changes were observed. Changes in AChE activity of the fetal brain may therefore be related to growth changes in the fetus.

Effects of ketamine anesthesia on rat-brain membranes: fluidity changes and kinetics of acetylcholinesterase.

This investigation shows that the effects of general anesthetics previously observed in vitro on membrane fluidity and on enzymic activities and occurring at concentrations calculated to be clinically relevant can be reproduced in vivo in anesthetized animals. Anesthesia with 2-chlorophenyl-2-methylaminocyclohexanone (ketamine) induces a more fluid state of rat-brain synaptic and mitochondrial membranes, as shown by the rotational correlation times of the spin labels 16-doxylstearate and 5-doxylstearate. Changes in acetylcholinesterase activity, with a decrease in Vmax and no change in the Km for acetylcholine, closely follow the fluidity increase.

Acetylcholinesterase activity in regions of the rat brain following a convulsion.

The effects of electroconvulsive shock on the levels of acetylcholinesterase in several brain regions of the rat were studied. Hippocampus, mesencephalon, cortex, and striatum exhibited rapid changes in acetylcholinesterase activity during the first few minutes following the convulsion, whereas brainstem and basal forebrain levels remained unchanged. In both hippocampus and midbrain there was a sustained decrease in activity: the total acetylcholinesterase activity was decreased by up to 40% within 2 min of the convulsion and did not return to control values for another 3 h. Thirty minutes after a flurothyl-induced convulsion there was a similar fall in acetylcholinesterase activity in both these regions, whereas a subconvulsive electric shock produced no change. It is concluded that a convulsion produces significant short-term decreases in acetylcholinesterase activity in areas of the rat brain that are involved in the generation and propagation of seizures, and the question is raised of whether this is related to the increase in seizure threshold that follows a convulsion.

Effects of ketamine anesthesia on rat-brain membranes: fluidity changes and kinetics of acetylcholinesterase

This investigation shows that the effects of general anesthetics previously observed in vitro on membrane fluidity and on enzymic activities and occurring at concentrations calculated to be clinically relebant can be reproduced in vivo in anesthetized animals. Anesthesia with 2-chlorophenyl-2-methylaminocyclohexanone (ketamine) induces a more fluid state of rat-brain synaptic and mitochondrial membranes, as shown by the rotational correlation times of the spin labels 16-doxylstearate and 5-doxylstearate. Changes in acetylcholinesterase activity, with a decrease in V max and no change in the K m for acetylcholine, closely follow the fluidity increase.

Observations on erythrocytic acetylcholinesterase activity in experimentally induced anaplasmosis in calves

Erythrocytic acetylcholinesterase (AChE) activity was assessed in splnectomized bovine calves inoculated with Anaplasma marginale. Other parameters, i.e., haematological values (haemoglobin, packed-cell volume, and total erythrocytic count), percentage parasitaemia and the ensuing host reactions during anaplasma infection were also studied in relation to AChE activity. There was significant inhibition of red blood cell-AChE activity commencing soon after inoculation of infected blood; this preceded a substantial decrease in haematological values along with high parasitaemia. The changes in AChE activity may possibly be associated with increased cell membrane permeability, thus triggering the entry of A. marginale organisms into red blood cells.

Effect of limb immobilization on skeletal muscle.

The immobilization of limbs resulted in atrophy of those muscles that are fixed either at or less than resting length. The loss in protein in these muscles can be described by a first-order equation. Decreases in protein synthesis rate in muscles of immobilized limbs occur during the first 6 h of immobilization, and this decrease probably played a role in initiating muscular atrophy. After weeks of immobilization, muscles composed predominately of slow-twitch fibers took on properties characteristic of fast twitch muscles. The EMG activity of muscles in immobilized limbs was reduced to 5--15% of control levels. Insulin responsiveness for 2-deoxyglucose uptake into the soleus muscle of a limb is decreased at the 24th h of limb immobilization. Muscles of immobilized limbs have either no change or a decrease in resting membrane potential, an increase in extrajunctional acetylcholine receptors of lesser magnitude than the increase that occurred in denervated muscle, and no change in acetylcholinesterase activity in neuromuscular junctions. Immobilizing muscles at stretched lengths prevented the decrease in nerve afterhyperpolarization that was seen in muscles immobilized at shortened positions. These observations suggested that metabolic changes in muscles have a retrograde trophic influence on motor nerves. The model of limb immobilization permits the study of many fundamental problems concerned with mechanisms by which a muscle adapts so that it can meet the requirements of the external environment.

Methylmercury-induced changes in the activities of neurotransmitter enzymes in nervous tissues of the rat

The activities of neurotransmitter-metabolizing enzymes were determined in the nervous tissues of rats treated with methylmercury chloride (10 mg/kg/day, for 7 days). The activity of choline acetyltransferase was lowered consistently in the cytosol and synaptosomal fractions of the brain in methylmercury-treated rats, while changes in acetylcholinesterase activity in the brain subcellular fractions were small. In peripheral nerves, decreases in activities of choline acetyltransferase in the sciatic nerve and of acetylcholinesterase in the dorsal root were most profound. Decreases in these enzyme activities started at an early phase and increased markedly with the progress of intoxication. The activity of acetylcholinesterase in the dorsal root ganglion and in the sciatic nerve was also inhibited significantly at the latent period and more profoundly at the symptomatic period at which time crossing of hind limbs, a typical sign of organomercurial poisoning, was observed in the animals. Activities of tyrosine hydroxylase and monoamine oxidase were elevated in the brain homogenate and especially in the synaptosomal fraction with respect to the former enzyme after methylmercury treatment. Effects of methylmercury in vitro on the activities of these enzymes revealed that a much higher amount of methylmercury was required to produce in vitro an inhibitory action equivalent to that observed in vivo. These results suggest that the neurotoxic action of methylmercury could be mediated, at least in part, through the level of neurotransmitter enzymes.

Effect of nembutal anesthesia, electric shock, and shock avoidance conditioning on acetylcholinesterase activity and protein content in various regions of the rat brain

Nembutal [pentobarbital sodium] was found to increase the protein content in some regions of the rat brain (posterior cortex, caudate nucleus) and to decrease the protein content in the ventral cortex. Acetylcholinesterase (AChE) activity, expressed in terms of wet tissue weight, increased in the cerebellum and medulla, but decreased in the medial cortex, hippocampus, thalamus, and the caudate nucleus. These changes in AChE activity were not explicable in terms of a direct effect of the anesthetic on the enzyme. A decrease in the protein content was observed in the frontal cortex, ventral cortex, hippocampus, and the caudate nucleus after electric shock is applied. Following shock avoidance conditioning, a decrease in protein content was observed in the medial cortex, posterior cortex, cerebellum, and the ventral cortex; the protein content increased in the thalamus. An increase in AChE activity was observed in the frontal cortex and in the medulla after shock, but its content decreased in the caudate nucleus, hypothalamus, thalamus, and the olfactory bulb. Following shock avoidance conditioning AChE activity increased in the posterior cortex, hippocampus, thalamus, and hypothalamus, but decreased in the ventral cortex.

Epidemiologic study of physiological effects in usual and volunteer citrus workers from organophosphate pesticide residues at reentry.

Biological parameters associated with organophosphate (OP) pesticide exposure were evaluated in a study of citrus harvesters. Changes in these parameters related to environmental residues of cholinesterase (ChE) inhibiting pesticides were studied. Further, it was determined whether usual and volunteer workers differed in their biochemical parameters after exposure to pesticide residues during field operations. Urine metabolite findings for usual farm workers showed that some exposure to OP pesticides had occurred. Too few workers were available during the reentry stage of the field study to evaluate the effect of this exposure on blood ChE. Baseline (nonexposure) acetylcholinesterase (AChE) and plasma cholinesterase (PChE) activities were significantly higher in usual (Mexican American) workers than in volunteer workers (student volunteers). Student volunteers in the test citrus grove showed statistically significant declines in PChE during the exposure period, yet changes in AChE activity, urine metabolites, and residue levels were very small. The implications of these findings are discussed in light of current reentry standards as well as future epidemiologic studies on reentry research.

Correlation between retention of defensive conditioned reflexes and brain acetylcholinesterase activity in rats.

1. Acetylcholinesterase activity in the motor and visual cortex and in the hippocampus of rats 1, 2, or 4 weeks after the formation of a defensive conditioned two-way avoidance reflex in the animals does not differ from the corresponding values obtained by studying the brain of animals exposed to the uncomblned action of the stimuli (active control). 2. Testing preservation of conditioned reflexes by their repeated formation 1 or 2 weeks after initial training was not accompanied by any change in acetylcholinesterase activity of the brain structures examined compared with the active control. When retention of conditioned reflexes was tested 4 weeks after their formation, a considerable decrease in acetylcholinesterase activity was found in the motor and visual cortex of the trained animals. 3. The dynamics of retention of conditioned reflexes is evidently due to changes in reactivity of the cholinergic systems of the brain with respect to stimuli used for conditioning.

Localization of α-bungarotoxin binding sites to the goldfish retinotectal projection

The optic tectum of the goldfish Carassius auratus is a rich source of α-bungarotoxin (α-Btx) binding protein. In order to determine whether some fraction of these receptors is present at retinotectal synapses, we have compared the histological distribution of receptors revealed by the use of [ 125Iα-Btx radioautography to the distribution of optic nerve terminals revealed by the use of cobalt and horseradish peroxidase (HRP) techniques. The majority of α-Btx binding is concentrated in those tectal layers containing primary retinotectal synapses. The same layers contain high concentrations of acetylcholinesterase (AChE), revealed histochemically. Following enucleation of one eye, there is a loss of α-Btx binding in the contralateral tectum, observed both by radioautography and by a quantitative binding assay of α-Btx binding. Approximately 40% of the α-Btx binding sites are lost within two weeks following enucleation. By contrast, no significant change in AChE activity could be demonstrated up to 6 months enucleation. These results are discussed in light of recent studies which show that the α-Btx binding protein and the nicotinic acetylcholine receptor are probably identical in goldfish tectum. We conclude that the 3 main classes of retinal ganglion cells projecting to the goldfish tectum are nicotinic cholinergic and that little or no postdenervation hypersensitivity due to receptor proliferation occurs in tectal neurons following denervation of the retinal input.

Neural regulation of cholinesterase in newt skeletal muscle. II. The effects of denervation and of culture in vitro

AbstractThe effect of denervation in vivo on the acetylcholinesterase (AChE) of newt (Notophthalrnw uiridescens) triceps muscle was compared with the effect of culturing explanted muscles in organ culture. Two weeks after denervation, the AChE activity of triceps muscles falls to approximately 60% of that in innervated muscles. The initial rate of loss of AChE is more rapid during summer than during winter, but no seasonal differences were observed in the specific activity of AChE in innervated muscles or in muscles two weeks after denervation. Changes in AChE activity when the triceps muscles were maintained in organ culture were similar in rate and in extent to those that occurred after denervation in vivo.The effect of denervation in vivo and culturing upon the individual AChE species separated by velocity sedimentation was also determined. Denervation and organ culture both resulted in a marked and rapid loss of two forms of AChE with sedimentation coefficients of 16.5S and 12.8S on sucrose density gradients. The former is associated with the motor endplates and the latter with the musculotendinous junctions. The activity of another form of AChE, with a sedimentation coefficient of 9.8S, transiently increased after denervation, but after one week returned to the initial level. The activity of a major form of AChE, with a sedimentation coefficient of 5,5S, remained constant after denervation, but that of the slowest sedimenting form (3.7S) declined slightly after one week. Similar alterations in the activities of these forms were observed in cultured muscles. Co‐culture of the muscles with dorsal root (sensory) ganglia from newts prevented both the loss of total AChE activity and the alterations in molecular forms of AChE in the muscles. This innervation‐like effect of the ganglia is apparently mediated by a diffusible factor(s) since ganglia separated from the muscles by a Millipore filter (0.22 μm pore size) also prevented alterations in AChE activity and molecular forms in the cultured muscles.These results demonstrate that the response of AChE to the absence or presence of nervous tissue, apparently mediated by a neurotrophic factor, is identical to that observed in vivo, at least in the case of the characteristics examined.

Effect of electric shock and diazepam on of Na, K-ATPase activity in brain membranes

The phenomenon of inactivation of Na,K-ATPase of the unpurified synaptosomal fraction from rat cerebral cortex in electric shock may be connected with “modification” of the potassium combining site of the enzyme. The anticonvulsant diazepam, if injected intramuscularly, also inhibits the Na,K-ATPase of the brain membranes, but diazepam followed by electrical stimulation of the brain in vivo activates Na,K-ATPase by comparison with the control. Diazepam also abolishes clonic convulsions following electrical stimulation of the brain. Meanwhile diazepam does not abolish the compensatory changes in acetylcholinesterase activity of brain and spinal cord synaptosomes of rats with electric shock. The results are discussed in connection with the view that inhibition of Na,K-ATPase activity of the nerve ending membranes may be the pathogenetic mechanism of seizure activity.

Acetylcholinesterase activity in specific regions of rabbit brain: Effect of pentobarbitone anesthesia

Acetylcholinesterase activity was determined in the cerebral cortex, cerebellum, hypothalamus, midbrain and medulla oblongata of a) control mature male rabbits, and b) male rabbits injected i.p. with an anesthetic dose of sodium pentobarbitone (60 mg/kg b.w.). Enzyme activity was highest in the midbrain and lowest in the cerebral cortex in controls. Pentobarbitone anesthesia increased enzyme activity in the cerebral cortex. No significant change in acetylcholinesterase activity occurred in the other brain areas studied following pentobarbitone administration. These findings are compared with the results of previous studies in the rat.

Effect of hydration and dehydration of animals on bioenergetics and mediator processes in neurosecretory cells of the anterior hypothalamus

Histochemical methods were used to study the activity of oxidoreductases and enzymes inactivating mediators in the neurosecretory cells of the anterior hypothalamus during hydration and dehydration in rabbits. Enzymes of the Krebs cycle and of the electron transport system were shown to respond by increased activity to dehydration and by reduced activity to hydration. Activity of α-glycerophosphate dehydrogenase and glucose-6-phosphate dehydrogenase was increased compared with the control in both cases. Monoamine oxidase activity was reduced during dehydration but increased during hydration; changes in acetylcholinesterase activity were in the opposite direction.