Moshe Shani

Transgenic expression of human acetylcholinesterase induces progressive cognitive deterioration in mice

BACKGROUND Cognitive deterioration is a characteristic symptom of Alzheimers disease. This deterioration is notably associated with structural changes and subsequent cell death which occur, primarily, in acetylcholine-producing neurons, progressively damaging cholinergic neurotransmission. We have reported previously that excess acetylcholinesterase (AChE) alters structural features of neuromuscular junctions in transgenic Xenopus tadpoles. However, the potential of cholinergic imbalance to induce progressive decline of memory and learning in mammals has not been explored. RESULTS To approach the molecular mechanisms underlying the progressive memory deficiencies associated with impaired cholinergic neurotransmission, we created transgenic mice that express human AChE in brain neurons. With enzyme levels up to two-fold higher than in control mice, transgenic mice displayed an age-independent resistance to the hypothermic effects of the AChE inhibitor, paraoxon. In addition to this improved scavenging capacity for anti-AChEs, however, these transgenic mice also resisted muscarinic, nicotinic and serotonergic agonists, indicating that secondary pharmacological changes had occurred. The transgenic mice also developed progressive learning and memory impairments, although their locomotor activities and open-field behaviour remained similar to those of matched control mice. By six months of age, transgenic mice lost their ability to respond to training in a spatial learning water maze test, whereas they performed normally in this test at the age of four weeks. This animal model is therefore suitable for investigating the transcriptional changes associated with cognitive deterioration and for testing drugs that may attenuate progressive damage. CONCLUSION We conclude that upsetting cholinergic balance may by itself cause progressive memory decline in mammals, suggesting that congenital and/or acquired changes in this vulnerable balance may contribute to the physiopathology of Alzheimers disease.

Manipulations of ACHE gene expression suggest non-catalytic involvement of acetylcholinesterase in the functioning of mammalian photoreceptors but not in retinal degeneration

To explore role(s) of acetylcholinesterase (AChE) in functioning and diseased photoreceptors, we studied normal (rd/+) and degenerating (rd/rd) murine retinas. All retinal neurons, expressed AChEmRNA throughout fetal development. AChE and c-Fos mRNAs peaked at post-natal days 10-12, when apoptosis of rd/rd photoreceptors begins. Moreover, c-Fos and AChEmRNA were co-overexpressed in rd/rd mice producing transgenic human (h), and host (m) AChE, but not in rd/+ mice. However, mAChE overexpression also occurred in transgenics expressing human serum albumin. Drastic variations in AChE catalytic activity were ineffective during development. Neither transgenic excess nor diisopropylfluorophosphonate (DFP) inhibition (80%) affected the rd phenotype; nor did DFP exposure induce photoreceptor degeneration or affect other key cholinergic proteins in rd/+ mice, unlike reports of adult mice and despite massive induction under DFP of c-Fos70 years). Therefore, the extreme retinal sensitivity to AChE modulation may reflect non-catalytic function(s) of AChE in adult photoreceptors. These findings exclude AChE as causing the rd phenotype, suggest that its primary function(s) in mammalian retinal development are non-catalytic ones and indicate special role(s) for the AChE protein in adult photoreceptors.

Cholinergic drug resistance and impaired spatial learning in transgenic mice overexpressing human brain acetylcholinesterase

Publisher Summary This chapter focuses on the cholinergic drug resistance and impaired spatial learning in transgenic mice overexpressing human brain acetylcholinesterase. Appropriate functioning of the cholinergic synapse requires a precise balance of its elements. Important contributors toward this balance are the acetylcholine (ACh) synthesizing enzyme choline acetyltransferase, the hydrolyzing enzyme acetylcholinesterase (AChE), and ACh receptors. Balanced cholinergic neurotransmission is disrupted in several pathological situations. Expression of human AChE under control of the authentic human promoter and first intron is observed in central nervous system neurons of transgenic mice. This expression changed responses to hypothermia-inducing drugs acting on cholinergic and probably serotonergic receptors. In addition, it created a progressive spatial learning and memory impairment. In contrast, the open field behavior of these transgenic animals remained normal. These findings suggest that subtle alterations in the cholinergic balance may cause physiologically observable changes and contribute by itself to the memory deterioration in at least a part of the patients with cholinergic deficits.

Testicular Gene Amplification and Impaired BCHE Transcription Induced in Transgenic Mice by the Human BCHE Coding Sequence

Multiple findings implicate acetylcholine with sperm functioning 1,2 and acetyl-and butyrylcholinesterase activities (ACHE, BCHE) were observed in mammalian sperm cells and during oocyte development 1–3. In vivo amplification of the human BCHE gene was first found in a father and son exposed to cholinesterase inhibitors 4, but it remained unclear whether the amplified DNA was transmitted as such from father to son or whether the amplification phenomenon re-occurred in germ cells, particularly during male meiosis or sperm differentiation.

Modulating Cholinergic Neurotransmission Through Transgenic Overexpression of Human Cholinesterases

Impaired cholinergic metabolism has been associated with several neurodegenerative disorders of the central (Wurtman, 1992) and peripheral nervous system (Harding, 1992), suggesting that experimental modulation of cholinergic neurotransmission can serve to study the relationship between cholinergic deficits and neuropathology. To this end, we established transgenic models for overexpressing human cholinesterases (ChEs) (Soreq and Zakut, 1993) in cholinergic synapses of Xenopus laevis tadpoles and mice. Cholinesterase over-expression should create a local deficit of acetylcholine at the “engineered” synapses and initiate feedback responses from which one may learn about the role of cholinergic neurotransmission in regulating synaptic structure and function.

54 Neuromuscular deterioration in ache-transgenic mice

The long-term contribution of balanced cholincrgic neurotransmission toward neuromuscular properties was studied in transgcnic mice b) expressing human acctylcholincsterasc (hAChE) in motoneurons, but not muscle. Spinal cord Cholinergic axodcndritic synapses in transgcnic mice wverc morphologically normal despite 7-fold higher AChE activity staining as compared with control synapses In contrast, although muscle extracts included only ca. 6%) of hAChE from motoneuron origin, transgcnic neuromuscular junctions were 60% larger than control ones and displayed either exaggerated or degenerated post-synaptic folds Neuromuscular impairment uas evident in grip tests at the age of 1 weeks, worsened with age and was accompanied by progressive amyotrophy and abnormal electromyogmphic potentials, rcflccting cnlargcd motor units and junctional dysfunction. The vulnerability of vertebrate neuromuscular .junctions to alterations in cholinergic ncurotransmission, highlights the morphogenic role of AChE Human AChE-expressing mice can hence be used for dissecting tllC molecular mechanisms underlying neuromuscular proprrties. FUNCTIONAL STUDIES OF THE SYNAPTIC VESICLE PROTEINS