Role of Acetylcholinesterase on the Structure and Function of Cholinergic Synapses: Insights Gained from Studies on Knockout Mice

Abstract

Electrophysiological and ultrastructural studies were performed on phrenic nerve-hemidiaphragm preparations isolated from wild-type and acetylcholinesterase (AChE) knockout (KO) mice to determine the compensatory mechanisms manifested by the neuromuscular junction to excess acetylcholine (ACh). The diaphragm was selected since it is the primary muscle of respiration, and it must adapt to allow for survival of the organism in the absence of AChE. Nerve-elicited muscle contractions, miniature endplate potentials (MEPPs) and evoked endplate potentials (EPPs) were recorded by conventional electrophysiological techniques from phrenic nerve-hemidiaphragm preparations isolated from 1.5- to 2-month-old wild-type (AChE(+/+)) or AChE KO (AChE(-/-)) mice. These recordings were chosen to provide a comprehensive assessment of functional alterations of the diaphragm muscle resulting from the absence of AChE. Tension measurements from AChE(-/-) mice revealed that the amplitude of twitch tensions was potentiated, but tetanic tensions underwent a use-dependent decline at frequencies below 70 Hz and above 100 Hz. MEPPs recorded from hemidiaphragms of AChE(-/-) mice showed a reduction in frequency and a prolongation in decay (37%) but no change in amplitude compared to values observed in age-matched wild-type littermates. In contrast, MEPPs recorded from hemidiaphragms of wild-type mice that were exposed for 30 min to the selective AChE inhibitor 5-bis(4-allyldimethyl-ammoniumphenyl)pentane-3-one (BW284C51) exhibited a pronounced increase in amplitude (42%) and a more marked prolongation in decay (76%). The difference between MEPP amplitudes and decays in AChE(-/-) hemidiaphragms and in wild-type hemidiaphragms treated with BW284C51 represents effective adaptation by the former to a high ACh environment. Electron microscopic examination revealed that diaphragm muscles of AChE(-/-) mice had smaller nerve terminals and diminished pre- and post-synaptic surface contacts relative to neuromuscular junctions of AChE(+/+) mice. The morphological changes are suggested to account, in part, for the ability of muscle from AChE(-/-) mice to function in the complete absence of AChE.

Skeletal Muscle Calpain Acts through Nitric Oxide and Neural miRNAs to Regulate Acetylcholine Release in Motor Nerve Terminals

“…The presynaptic changes in CEAD, however, were mysterious and explained as a possible “compensatory response” to the effect of excessive cholinergic activation (Engel et al, 2010). Similar changes in the nerve terminus have been seen in pharmacological models of cholinesterase inhibition (Roberts and Thesleff, 1969, Tiedt et al, 1978), and in genetic AChE knockout models (Adler et al, 2011). …”

Small molecule metalloprotease inhibitor with in vitro, ex vivo and in vivo efficacy against botulinum neurotoxin serotype A

“…Mice were housed in Association for Assessment and Accreditation of Laboratory Animal Care (AAALAC) International accredited facilities with food and water available ad libitum as described previously (Adler et al, 2011). To obtain isolated muscle preparations, the animals were euthanized by an overdose of isoflurane and decapitated.…”

“…In situ EDL muscle tension measurements were performed on male Sprague Dawley Crl:CD (SD)IGS BR rats (300–350 g) from Charles River housed under an approved AAALAC International program (Adler et al, 2011). Local intramuscular (i.m.)…”

New Pharmacological Approaches to the Cholinergic System: An Overview on Muscarinic Receptor Ligands and Cholinesterase Inhibitors

“…The development of knockout (−/−) mice for AChE and BuChE, separately, has substantially added to our broader understanding of the function of these enzymes in health and disease [177–178]. Interestingly, selective BuChE inhibitors similarly augment brain endogenous ACh concentrations [169] and, as confirmed in AChE −/− mice, achieve this via action on BuChE [170].…”

“…This is notable in light of the knowledge that rat, similar to healthy human brain, primarily comprises 90% AChE and 10% BuChE. Thus, a 39% inhibition of BuChE – comprising 10% of brain cholinesterase activity, elevated brain ACh levels to the same amount as a 27% inhibition of AChE comprising 90% of brain cholinesterase activity [172], and underlies that both AChE and BuChE co-regulate brain cholinergic function [169–170, 177–179]. …”