Decrease In High-affinity Choline Uptake Research Articles

Altered Cholinergic Metabolism in Rat CNS Following Aluminum Exposure: Implications on Learning Performance

The effects of Al on the central cholinergic system have been studied. Al, at a dose of 10 mg/kg of body weight/day for 4 weeks, had a deleterious effect on the activities of biosynthetic (choline acetyltransferase) and hydrolytic (acetylcholinesterase) enzymes of the neurotransmitter acetylcholine. The levels of acetylcholine were also significantly lowered in different brain regions at the end of the dose regimen. There was a significant decrease in high-affinity choline uptake following Al exposure. Muscarinic acetylcholine receptor binding studies revealed a decreased number of binding sites (Bmax), with the maximum effects being manifested in the hippocampus. Exogenous addition of 10 microM desferrioxamine restored the muscarinic receptor binding completely. The impaired cholinergic functioning had severe effects on cognitive functions. Neurobehavioral deficits were manifested in terms of decreased active (52%) and passive (73.30%) avoidance tests. The results suggest that Al exerts its toxic effects by altering cholinergic transmission, which is ultimately reflected in neurobehavioral deficits.

Effects of iron-induced lipid peroxidation and of acidosis on choline uptake by synaptosomes

The effects of iron-induced lipid peroxidation and of lactic acidosis on [3H]choline uptake were investigated on crude synaptosomes prepared from rat cerebral cortices. Fe(2+)-induced lipid peroxidation as evidenced from the production of thiobarbituric acid reactives substances (TBARS) was correlated with a decrease in high-affinity choline uptake (HACU). Trolox C, a free radical scavenger, prevented both Fe(2+)-induced TBARS production and decrease in HACU. Lactic acidosis (pH 6.0 for 30 or 60 min) increased the TBARS production with concomitant decrease in HACU (-48%, -78%, respectively). The acidosis dependent decrease was not reversible following pH 7.4 readjustment after 60 min acidosis. It was not prevented by trolox C, although trolox C inhibited the acidosis-induced production of TBARS. The results suggest that the contribution of acidosis to peroxidative damages is probably of less importance in comparison to other cytotoxic mechanisms.

Effects of a 60 Hz magnetic field on central cholinergic systems of the rat.

We studied the effects of an acute (45 min) exposure to a 60 Hz magnetic field on sodium-dependent, high-affinity choline uptake in the brain of the rat. Decreases in uptake were observed in the frontal cortex and hippocampus after the animals were exposed to a magnetic field at flux densities > or = 0.75 mT. These effects of the magnetic field were blocked by pretreating the animals with the narcotic antagonist naltrexone, but not by the peripheral opioid antagonist, naloxone methiodide. These data indicate that the magnetic-field-induced decreases in high-affinity choline uptake in the rat brain were mediated by endogenous opioids in the central nervous systems.

Recovery of cholinergic phenotype in the injured rat neostriatum: roles for endogenous and exogenous nerve growth factor.

Polyclonal antibodies against recombinant human nerve growth factor (rhNGF) potently inhibited PC12 neurite outgrowth, blocked high-affinity 125I-rhNGF binding but not its receptor, and cross-reacted with rat, mouse, and human nerve growth factor (NGF) but not with brain-derived neurotrophic factor, neurotrophin-3, ciliary neurotrophic factor, insulin-like growth factor, epidermal growth factor, or activin A. Immunocytochemistry revealed many NGF-positive neurons in the rat neostriatum. The NGF-positive neurons disappeared by 3 days after mechanical injury to the neostriatum and were replaced by intensely NGF- and glial fibrillary acidic protein-positive astrocytes. Enzyme-linked immunosorbent assay measurements revealed that the NGF content of the injured striatum was elevated by eightfold 3 days postinjury and by twofold 2 weeks later. The high-affinity choline uptake (HACU) into cholinergic nerve terminals was decreased by 23% at 2 and 4 weeks postinjury, yet choline acetyltransferase (ChAT) activity in these neurons was unchanged at 2 weeks and decreased by 14% at 4 weeks. Daily infusion of 1 microgram of rhNGF into the injury area did not alter the loss of HACU. However, this treatment elevated ChAT activity by 23-29% above intact neostriatal levels and by 53-65% relative to HACU at both survival times. Thus, lesion-induced increases in NGF levels within astrocytes are associated with maintenance of striatal ChAT activity at normal levels following cholinergic injury, even with decreases in HACU. Pharmacologic doses of rhNGF can further augment ChAT activity in damaged cholinergic neurons, showing the usefulness of exogenous NGF even when endogenous NGF is elevated in response to injury.

Opioid receptor subtypes mediating the noise-induced decreases in high-affinity choline uptake in the rat brain

Acute (20 min) exposure to 100-dB white noise elicits a naltrexone-sensitive decrease in sodium-dependent high-affinity choline uptake in the frontal cortex and hippocampus of the rat. In the present study, the subtypes of opioid receptors involved were investigated by pretreating rats with microinjection of specific opioid-receptor antagonists into the lateral cerebroventricle before noise exposure. We found that the noise-induced decrease in high-affinity choline uptake in the hippocampus was blocked by pretreatment with either μ-, δ-, or κ-opioid-receptor antagonists, whereas the effect of noise on frontal cortical high-affinity choline uptake was blocked by a μ- and δ- but not by a κ-antagonist. These data further confirm the role of endogenous opioids in mediating the effects of noise on central cholinergic activity and indicate that different neural mechanisms are involved in the effects of noise on the frontal cortical and hippocampal cholinergic systems.

Inhibition of high affinity choline uptake in the rat brain by neurotoxins: Effect of monosialoganglioside GM1

Mustard derivatives of ethyl-choline and hemicholinium-3 have been suggested as possible specific cholinergic neurotoxins. In this study a structural analog of hemicholinium-3, a,a′-bis[di(2-chloroethyl)amino]-4,4′-2-biacetophenone (toxin 7), was added to synaptosomes prepared from the cortex, striatum or hippocampus of rat brain. Synaptosomal high affinity choline uptake (HACU) was significantly decreased in a dose-dependent manner by addition of toxin 7, while synaptosomal uptake of GABA or dopamine was not changed. Incubation of cortical synaptosomes with the monosialoganglioside GM1 prevented the decrease in HACU seen following administration of toxin 7. This preventative effect of GM1 was greater if GM1 was added prior to or concomitant with toxin 7, than if GM1 was added following toxin 7. Two newly synthesized hemicholinium-3 analogs, 4-[3′-di(2-chloroethyl)aminopropionyl] biphenyl (toxin 5) and 4-[3′-di(2-bromoethyl)aminopropionyl]biphenyl (toxin 6) caused a large decrease in HACU when added to cortical synaptosomes, this decrease was significantly greater than that seen with the same dose of toxin 7 or ethyl-choline aziridinium (AF64A). Ultrastructural changes in the synaptosomal membrane following incubation with toxin 7 or toxin 7 with GM1 were examined by electron microscopy. Development of a compound which is both a potent neurotoxin, and is specific for cholinergic neurons will allow new insights into the normal function of the cholinergic system in the CNS and provide animal models of disease states in which cholinergic degeneration is an important element.

Corticotropin-releasing factor antagonist blocks microwave-induced decreases in high-affinity choline uptake in the rat brain

Acute (45-min) irradiation with pulsed low-level microwaves (2450-MHz, 2 μsec pulses at 500 pps, average power density of 1 mW/cm 2, whole-body average specific absorption rate of 0.6 W/kg) decreased sodium-dependent high-affinity choline uptake (HACU) activity in the frontal cortex and hippocampus of the rat. These effects were blocked by pretreating the animals before exposure with intracerebroventricular injection of the specific corticotropin-releasing factor (CRF) receptor antagonist, α-helical-CRF 9–41 (25 μg). Similar injection of the antagonist had no significant effect on HACU in the brain of the sham-exposed rats. These data suggest that low-level microwave irradiation activates CRF in the brain, which in turn causes the changes in central HACU.

AF64A depletes hippocampal high-affinity choline uptake but does not alter the density of α-bungarotoxin binding sites or modify the effect of exogenous choline

Sodium-dependent, high-affinity choline uptake (HACU) and the density of α-bungarotoxin (BuTX) receptor-binding sites were measured in the hippocampus following the intraventricular infusion of ethylcholine aziridinium ion (AF64A), a neurotoxin that competes with choline at high-affinity choline transport sites and may result in the degeneration of cholinergic axons. Eight days after the infusion of AF64A into the lateral ventricles (2.5 nmol/side), HACU was depleted by 60% in the hippocampus of experimental animals in comparison with controls, but the density of BuTX-binding sites was not altered. The administration of 15 mg/ml of choline chloride in the drinking water increased the density of BuTX-binding sites, as previously reported by this laboratory 23,28. The administration of AF64A did not prevent the effect of exogenous choline on the density of binding sites, nor did choline treatment alter the effect of AF64A on HACU. These data indicate that the density of BuTX-binding sites in the hippocampus is not altered following a substantial decrease in HACU and presumed degeneration of cholinergic axons. Since the effect of exogenous choline was not prevented by AF64A treatment, the data are interpreted to support the hypothesis that the increase in the density of BuTX-binding sites following dietary choline supplementation is attributable to a direct effect of choline on receptor sites.

Interaction between nerve growth factor and GM 1 monosialoganglioside in preventing cortical choline acetyltransferase and high affinity choline uptake decrease after lesion of the nucleus basalis

Monosialoganglioside GM 1 and nerve growth factor (NGF) were administered alone or concomitantly to adult male rats with a unilateral ibotenic acid lesion of the nucleus basalis magnocellularis (NBM). High-affinity choline uptake (HACU) rate and choline acetyltransferase (ChAT) activity were measured, 4 and 21 days after surgery, respectively, in the frontal and parietal cortices of both hemispheres. A 33–34% decrease in HACU rate and a 43-39% decrease in ChAT activity was found in the ipsilateral cortices 4 and 21 days, respectively, after the lesion. If the lesioned rats received NGF (10 μg i.c.v.) twice a week or daily administrations of GM 1 (30 mg/kg, i.p.), beginning immediately after surgery the decrease in HACU rate and ChAT activity was smaller. If NGF and GM, were given concomitantly no decrease in HACU rate and ChAT activity was detected in the lesioned hemisphere and a slight increase occurred in the contralateral hemisphere. However, after the concurrent administration of NGF (10 μg i.c.v.) and the inactive dose of GM 1 10 mg/kg i.p. no decrease in HACU and ChAT activity was also found in the lesioned rats. The latter finding indicates a potentiation by GM 1 of NGF effects on the cholinergic neurons of the IBM. The two drugs may either antagonize the neurotoxic effects of ibotenic acid or stimulate a compensatory activity in the remaining neurons.

Effect of Single or Chronic Injection with a Carbamate, Propoxur, on the Brain Cholinergic System and Behavior of Mice

Propoxur, an anticholinesterase carbamate, was injected singly (10 mg/ kg, s.c.) or chronically (5 mg/kg/day, s.c., for 10 days) into mice. Animals were examined for effects on the cholinergic system in brain tissue and on behavior. Single injection caused an increase in brain acetylcholine (ACh) content at 10 and 60 min; and it caused decreases in acetylcholinesterase (AChE) activity at 10, 60 and 180 min, high-affinity choline uptake into synaptosomes at 10 and 60 min, and [3H]quinuclidinyl benzilate (QNB) binding at 10 min without causing any change in choline acetyltransferase (ChAT) activity. Open-field behavior and rotarod performance were depressed at 10 min and rectal temperature was decreased at 10, 60 and 120 min after a single injection. Chronic treatment caused decreases in high-affinity choline uptake and [3H]QNB binding 24 hr after the final injection without any changes in ACh content, AChE activity or ChAT activity. In behavioral tests conducted 10 min prior to daily administration, rotarod performance was slightly suppressed only during the period of injection. Although propoxur is considered to have effects similar to those of organophosphorus compounds, the changes observed in the present experiments seemed to be reversible.

Effect of single or chronic injection with a carbamate, propoxur, on the brain cholinergic system and behavior of mice.

Propoxur, an anticholinesterase carbamate, was injected singly (10 mg/kg, s.c.) or chronically (5 mg/kg/day, s.c., for 10 days) into mice. Animals were examined for effects on the cholinergic system in brain tissue and on behavior. Single injection caused an increase in brain acetylcholine (ACh) content at 10 and 60 min; and it caused decreases in acetylcholinesterase (AChE) activity at 10, 60 and 180 min, high-affinity choline uptake into synaptosomes at 10 and 60 min, and [3H]quinuclidinyl benzilate (QNB) binding at 10 min without causing any change in choline acetyltransferase (ChAT) activity. Open-field behavior and rotarod performance were depressed at 10 min and rectal temperature was decreased at 10, 60 and 120 min after a single injection. Chronic treatment caused decreases in high-affinity choline uptake and [3H]QNB binding 24 hr after the final injection without any changes in ACh content, AChE activity or ChAT activity. In behavioral tests conducted 10 min prior to daily administration, rotarod performance was slightly suppressed only during the period of injection. Although propoxur is considered to have effects similar to those of organophosphorus compounds, the changes observed in the present experiments seemed to be reversible.

Effects of training on a spatial memory task on high affinity choline uptake in hippocampus and cortex in young adult and aged rats

The relation of forebrain cholinergic function to learning and memory was explored by identification and characterization of a training-induced change in high-affinity choline uptake (HACU), an index of cholinergic activity. Young adult rats were trained to find an invisible escape platform in a water tank using environmental cues. After 4 d of this place-training (16 trials), hippocampal HACU was significantly reduced relative to that observed in rats trained to find a visible platform (cue-training), even when cue- and place-trained rats were yoked for swim time. These place- but not cue-trained rats showed significantly lower hippocampal HACU than did naive rats, and no effect of training was noted after only 1 d of training. Similar results were obtained in parietal cortex. These differential training effects on HACU correspond to previous reports that muscarinic blockade impairs place, but not cue, learning. A further experiment revealed that the decrease in HACU in hippocampus, but not in parietal cortex, occurred only during the acquisition phase of learning and was related to the rate of acquisition for individual animals. Hippocampal HACU in naive young and aged (24-27 months) rats did not differ, but the response of the septohippocampal cholinergic system to training was diminished in the aged rats. Old rats displayed impaired place learning and a corresponding dampening of the training-induced change in HACU. These results suggest that there is a task-specific engagement of cholinergic function in young animals that does not occur in behaviorally impaired aged animals, a finding that is consistent with a role for cholinergic dysfunction in memory impairments associated with aging.