Dataset Information

Impaired NGF/TrkA Signaling Causes Early AD-Linked Presynaptic Dysfunction in Cholinergic Primary Neurons.

ABSTRACT: Alterations in NGF/TrkA signaling have been suggested to underlie the selective degeneration of the cholinergic basal forebrain neurons occurring in vivo in AD (Counts and Mufson, 2005; Mufson et al., 2008; Niewiadomska et al., 2011) and significant reduction of cognitive decline along with an improvement of cholinergic hypofunction have been found in phase I clinical trial in humans affected from mild AD following therapeutic NGF gene therapy (Tuszynski et al., 2005, 2015). Here, we show that the chronic (10-12 D.I.V.) in vitro treatment with NGF (100 ng/ml) under conditions of low supplementation (0.2%) with the culturing serum-substitute B27 selectively enriches the basal forebrain cholinergic neurons (+36.36%) at the expense of other non-cholinergic, mainly GABAergic (-38.45%) and glutamatergic (-56.25%), populations. By taking advantage of this newly-developed septo-hippocampal neuronal cultures, our biochemical and electrophysiological investigations demonstrate that the early failure in excitatory neurotransmission following NGF withdrawal is paralleled by concomitant and progressive loss in selected presynaptic and vesicles trafficking proteins including synapsin I, SNAP-25 and ?-synuclein. This rapid presynaptic dysfunction: (i) precedes the commitment to cell death and is reversible in a time-dependent manner, being suppressed by de novo external administration of NGF within 6 hr from its initial withdrawal; (ii) is specific because it is not accompanied by contextual changes in expression levels of non-synaptic proteins from other subcellular compartments; (ii) is not secondary to axonal degeneration because it is insensible to pharmacological treatment with known microtubule-stabilizing drug such paclitaxel; (iv) involves TrkA-dependent mechanisms because the effects of NGF reapplication are blocked by acute exposure to specific and cell-permeable inhibitor of its high-affinity receptor. Taken together, this study may have important clinical implications in the field of AD neurodegeneration because it: (i) provides new insights on the earliest molecular mechanisms underlying the loss of synaptic/trafficking proteins and, then, of synapes integrity which occurs in vulnerable basal forebrain population at preclinical stages of neuropathology; (ii) offers prime presynaptic-based molecular target to extend the therapeutic time-window of NGF action in the strategy of improving its neuroprotective in vivo intervention in affected patients.

SUBMITTER: Latina V

PROVIDER: S-EPMC5350152 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

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Impaired NGF/TrkA Signaling Causes Early AD-Linked Presynaptic Dysfunction in Cholinergic Primary Neurons.

Latina Valentina V Caioli Silvia S Zona Cristina C Ciotti Maria T MT Amadoro Giuseppina G Calissano Pietro P

Frontiers in cellular neuroscience 20170315

Alterations in NGF/TrkA signaling have been suggested to underlie the selective degeneration of the cholinergic basal forebrain neurons occurring <i>in vivo</i> in AD (Counts and Mufson, 2005; Mufson et al., 2008; Niewiadomska et al., 2011) and significant reduction of cognitive decline along with an improvement of cholinergic hypofunction have been found in phase I clinical trial in humans affected from mild AD following therapeutic NGF gene therapy (Tuszynski et al., 2005, 2015). Here, we show ...[more]

PMID: 28360840

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Project description:Basal forebrain cholinergic neurons (BFCNs) depend on nerve growth factor (NGF) for their survival/differentiation and innervate cortical and hippocampal regions involved in memory/learning processes. Cholinergic hypofunction and/or degeneration early occurs at prodromal stages of Alzheimer's disease (AD) neuropathology in correlation with synaptic damages, cognitive decline and behavioral disability. Alteration(s) in ubiquitin-proteasome system (UPS) is also a pivotal AD hallmark but whether it plays a causative, or only a secondary role, in early synaptic failure associated with disease onset remains unclear. We previously reported that impairment of NGF/TrkA signaling pathway in cholinergic-enriched septo-hippocampal primary neurons triggers "dying-back" degenerative processes which occur prior to cell death in concomitance with loss of specific vesicle trafficking proteins, including synapsin I, SNAP-25 and α-synuclein, and with deficit in presynaptic excitatory neurotransmission. Here, we show that in this in vitro neuronal model: (i) UPS stimulation early occurs following neurotrophin starvation (-1 h up to -6 h); (ii) NGF controls the steady-state levels of these three presynaptic proteins by acting on coordinate mechanism(s) of dynamic ubiquitin-C-terminal hydrolase 1 (UCHL-1)-dependent (mono)ubiquitin turnover and UPS-mediated protein degradation. Importantly, changes in miniature excitatory post-synaptic currents (mEPSCs) frequency detected in -6 h NGF-deprived primary neurons are strongly reverted by acute inhibition of UPS and UCHL-1, indicating that NGF tightly controls in vitro the presynaptic efficacy via ubiquitination-mediated pathway(s). Finally, changes in synaptic ubiquitin and selective reduction of presynaptic markers are also found in vivo in cholinergic nerve terminals from hippocampi of transgenic Tg2576 AD mice, even from presymptomatic stages of neuropathology (1-month-old). By demonstrating a crucial role of UPS in the dysregulation of NGF/TrkA signaling on properties of cholinergic synapses, these findings from two well-established cellular and animal AD models provide novel therapeutic targets to contrast early cognitive and synaptic dysfunction associated to selective degeneration of BFCNs occurring in incipient early/middle-stage of disease.

Project description:BackgroundSeptal cholinergic neurons account for most of the cholinergic innervations of the hippocampus, playing a key role in the regulation of hippocampal synaptic activity. Disruption of the septo-hippocampal pathway by an experimental transection of the fimbria-fornix drastically reduces the target-derived trophic support received by cholinergic septal neurons, mainly nerve growth factor (NGF) from the hippocampus. Axotomy of cholinergic neurons induces a reduction in the number of neurons positive for cholinergic markers in the medial septum. In several studies, the reduction of cholinergic markers has been interpreted as analogous to the neurodegeneration of cholinergic cells, ruling out the possibility that neurons lose their cholinergic phenotype without dying. Understanding the mechanism of cholinergic neurodegeneration after axotomy is relevant, since this paradigm has been extensively explored as an animal model of the cholinergic impairment observed in neuropathologies such as Alzheimer's disease.The principal aim of this study was to evaluate, using modern quantitative confocal microscopy, neurodegenerative changes in septal cholinergic neurons after axotomy and to assess their response to delayed infusion of NGF in rats.ResultsWe found that there is a slow reduction of cholinergic cells labeled by ChAT and p75 after axotomy. However, this phenomenon is not accompanied by neurodegenerative changes or by a decrease in total neuronal number in the medial septum. Although the remaining axotomized-neurons appear healthy, they are unable to respond to delayed NGF infusion.ConclusionsOur results demonstrate that at 3 weeks, axotomized cholinergic neurons lose their cholinergic phenotype without dying and down-regulate their NGF-receptors, precluding the possibility of a response to NGF. Therefore, the physiological role of NGF in the adult septal cholinergic system is to support phenotypic differentiation and not survival of neurons. This evidence raises questions about the relationship between transcriptional regulation of the cholinergic phenotype by retrograde-derived trophic signaling and the transcriptional changes experienced when retrograde transport is impaired due to neuropathological conditions.

Dataset Information

Impaired NGF/TrkA Signaling Causes Early AD-Linked Presynaptic Dysfunction in Cholinergic Primary Neurons.

Publications

Impaired NGF/TrkA Signaling Causes Early AD-Linked Presynaptic Dysfunction in Cholinergic Primary Neurons.

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