Dysfunctional Presynaptic M2 Receptors in the Presence of Chronically High Acetylcholine Levels: Data from the PRiMA Knockout Mouse.

Franziska Mohr,Jochen Klein,Martina Zimmermann,Eric Krejci

doi:10.1371/journal.pone.0141136

Abstract

The muscarinic M2 receptor (M2R) acts as a negative feedback regulator in central cholinergic systems. Activation of the M2 receptor limits acetylcholine (ACh) release, especially when ACh levels are increased because acetylcholinesterase (AChE) activity is acutely inhibited. Chronically high ACh levels in the extracellular space, however, were reported to down-regulate M2R to various degrees. In the present study, we used the PRiMA knockout mouse which develops severely reduced AChE activity postnatally to investigate ACh release, and we used microdialysis to investigate whether the function of M2R to reduce ACh release in vivo was impaired in adult PRiMA knockout mice. We first show that striatal and hippocampal ACh levels, while strongly increased, still respond to AChE inhibitors. Infusion or injection of oxotremorine, a muscarinic M2 agonist, reduced ACh levels in wild-type mice but did not significantly affect ACh levels in PRiMA knockout mice or in wild-type mice in which ACh levels were artificially increased by infusion of neostigmine. Scopolamine, a muscarinic antagonist, increased ACh levels in wild-type mice receiving neostigmine, but not in wild-type mice or in PRiMA knockout mice. These results demonstrate that M2R are dysfunctional and do not affect ACh levels in PRiMA knockout mice, likely because of down-regulation and/or loss of receptor-effector coupling. Remarkably, this loss of function does not affect cognitive functions in PRiMA knockout mice. Our results are discussed in the context of AChE inhibitor therapy as used in dementia.

Highlights

G protein-coupled receptors (GPCR) are workhorses for neurotransmitter actions, mediating pre- as well as postsynaptic effects
The present study of adaptive processes in the central cholinergic system was motivated by the use of AChE inhibitors in human disease, primarily for the treatment of dementias such as Alzheimers disease (AD) [19,20]
The PRiMA knockout mouse is an excellent model for cholinergic hyperfunction, because (a.) residual AChE activity is less than 10% of that in WT mice [11]; (b.) striatal ACh levels in these mice were reportedly very high and (c.) these mice are phenotypically normal and seizure-free with only minor problems of motoric function [12]

Summary

Introduction

G protein-coupled receptors (GPCR) are workhorses for neurotransmitter actions, mediating pre- as well as postsynaptic effects. An interesting phenomenon in this respect is the observation that M2R are down-regulated when synaptic ACh levels remain high for extended periods of time This is the case in transgenic mouse models with reduced AChE activity. The PRiMA knockout mouse, AChE activity in the brain is strongly reduced but not absent [11]. These mice are phenotypically normal, and motor function shows only minor and difficult-to-detect changes, ACh levels in the brain are extremely high in these mice as well [12]. Muscarinic receptors, especially the M2 subtype, are down-regulated by 20–60% depending on the brain region investigated [12], and this down-regulation occurs in parallel with the development of the central cholinergic fibres postnatally [13]

Methods

Results

Conclusion