Automated measurement of acetylcholinesterase activity in rat peripheral tissues

Lassiter, T. Leon

doi:10.1016/s0300-483x(02)00752-7

Cited by 36 publications

(21 citation statements)

References 29 publications

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“…Homogenisation required between 2 and 5 min depending on the tissue; after every 30 s or so of homogenisation the mixture was rested for 10 s to allow cooling. The homogenate was then centrifuged and then decanted into an Eppendorf tubes at 9000 g for 5 min at 4°C [12, 15, 16]. However, the homogenates were thoroughly mixed and distributed into 16 equal portions in all animals (sheep, cattle, and pigs) and tissues (liver and muscle), representing storage temperatures (−80°C and −20°C) for immediate processing one-month intervals, over a period of eight months.…”

Section: Methodsmentioning

confidence: 99%

Comparison of Two Storage Methods for the Analysis of Cholinesterase Activities in Food Animals

2010

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Cholinesterases (ChE) are specialized carboxylic ester hydrolases that catalyse the hydrolysis of choline esters. They are classified into either acetylcholinesterase (AChE) or butyrylcholinesterase (BChE). Determination of ChE in the tissues is the appropriate tool for the diagnosis of organophosphorus and carbamate exposures. In general, a significant inhibition was seen in both AChE and BChE activities after 6 months of freezing at −80°C and after 3 months of freezing at −20°C. Linear regression of mean AChE and BChE was observed in all individual samples during the months of the two freezing methods. Bland and Altman plot of the ratios of the two freezing methods have showen the mean difference between the two freezing methods to be 8.8, and SD was 144.7 and −127.6 for upper and lower limits, respectively, for liver, while in muscle the mean difference was 1.5 and SD was 32.5 and −28.9 for upper and lower limits, respectively.

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Section: Methodsmentioning

confidence: 99%

Comparison of Two Storage Methods for the Analysis of Cholinesterase Activities in Food Animals

2010

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“…The brain was removed and the CA1 region and the perirhinal cortex were dissected out. AChE activity was determined using a standard spectophotometric method according to Ellman et al (1961) as modified by Lassiter et al (2003). Hydrolysis rates were measured at an acetylthiocholine concentration of 0.8 mM in 1 ml assay solutions with 30 mM phosphate buffer, pH 7.5, and 1.0 mM DTNB at 25 °C.…”

Section: Methodsmentioning

confidence: 99%

Early postnatal maternal deprivation in rats induces memory deficits in adult life that can be reversed by donepezil and galantamine

Benetti

Mello

Bonini

et al. 2008

Intl J of Devlp Neuroscience

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Early postnatal maternal deprivation is known to cause long-lasting neurobiological effects. Here, we investigated whether some of the cognitive aspects of these deficits might be related to a disruption of the cholinergic system. Pregnant Wistar rats were individually housed and maintained on a 12:12h light/dark cycle with food and water freely available. The mothers were separated from their pups for 3h per day from postnatal day 1 (PND-1) to PND-10. To do that, the dams were moved to a different cage and the pups maintained in the original home cage, which was transferred to a different room kept at 32 degrees C. After they reached 120-150 days of age, maternal-deprived and non-deprived animals were either sacrificed for brain acetylcholinesterase measurement, or trained and tested in an object recognition task and in a social recognition task as described by Rossato et al. (2007) [Rossato, J.I., Bevilaqua, L. R.M., Myskiw, J.C., Medina, J.H., Izquierdo, I., Cammarota, M. 2007. On the role hippocampal synthesis in the consolidation and reconsolidation of object recognition memory. Learn. Mem. 14, 36-46] and Lévy et al. (2003) [Lévy, F., Melo. A.I., Galef. B.G. Jr., Madden, M., Fleming. A.S. 2003. Complete maternal deprivation affects social, but not spatial, learning in adult rats. Dev. Psychobiol. 43, 177-191], respectively. There was increased acetylcholinesterase activity in hippocampus and perirhinal cortex of the deprived animals. In addition, they showed a clear impairment in memory of the two recognition tasks measured 24h after training. Oral administration of the acetylcholinesterase inhibitors, donepezil or galantamine (1mg/kg) 30min before training reversed the memory impairments caused by maternal deprivation. The findings suggest that maternal deprivation affects memory processing at adulthood through a change in brain cholinergic systems.

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“…In fact, the highest TNB -/hemoglobin ratio was determined at a wavelength of 462.5 nm. [33][34][35] Numerous studies investigated the effect of different experimental conditions on AChE activity and identified a number of determinants that have to be considered. Alternatively, hemoglobin-free AChE preparations are suitable for kinetic studies.…”

Section: Determination Of Ache Activitybasic Considerationsmentioning

confidence: 99%

Determination of acetylcholinesterase activity by the Ellman assay: A versatile tool for in vitro research on medical countermeasures against organophosphate poisoning

Worek¹,

Eyer

Thiermann³

2011

Drug Testing and Analysis

111

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Inhibition of acetylcholinesterase (AChE) is the main mechanism of action of organophosphorus compounds (OP), and AChE reactivators (oximes) are at present the only causal therapeutic approach. Being the key target of OP toxicity, AChE may serve as a valuable tool for diagnosis of OP exposure as well as for the investigation of the kinetics of interactions between OP and oximes. At present, the rapid, simple, and cheap spectrophotometric Ellman assay is widely used for diagnosis, therapeutic monitoring and in vitro kinetic investigations. Application of the assay for investigation of the interactions between AChE, inhibitors, and oximes requires the consideration of potential matrix effects (e.g. hemoglobin), side reactions (e.g. oximolysis of substrate) and other determinants (e.g. pH, temperature). By taking these factors into account, the Ellman assay allows the precise and reproducible determination of kinetic constants as a basis for the understanding of toxic OP effects and for the development of improved therapies against poisoning by OP. In addition, advanced applications of the Ellman assay, for example, in a dynamic in vitro model for the real-time activity determination of membrane-bound AChE, enables the proper investigation of relevant tissue, primarily respiratory muscle, and extends the applicability of this method.

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Automated measurement of acetylcholinesterase activity in rat peripheral tissues

Cited by 36 publications

References 29 publications

Comparison of Two Storage Methods for the Analysis of Cholinesterase Activities in Food Animals

Comparison of Two Storage Methods for the Analysis of Cholinesterase Activities in Food Animals

Early postnatal maternal deprivation in rats induces memory deficits in adult life that can be reversed by donepezil and galantamine

Determination of acetylcholinesterase activity by the Ellman assay: A versatile tool for in vitro research on medical countermeasures against organophosphate poisoning

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