Neurobiol Dis. 2015 Feb;74:204-18. doi: 10.1016/j.nbd.2014.12.005.

Long-term effects of neonatal treatment with fluoxetine on cognitive performance in Ts65Dn mice.

Stagni F1, Giacomini A1, Guidi S1, Ciani E1, Ragazzi E1, Filonzi M2, De Iasio R2, Rimondini R3, Bartesaghi R4.
  • 1Department of Biomedical and Neuromotor Sciences, University of Bologna, Italy.
  • 2Centralized Laboratory, S. Orsola-Malpighi University Hospital, Bologna, Italy.
  • 3Department of Medical and Surgical Sciences, University of Bologna, Italy.
  • 4Department of Biomedical and Neuromotor Sciences, University of Bologna, Italy. Electronic address: renata.bartesaghi@unibo.it.

 

Abstract

Individuals with Down syndrome (DS), a genetic condition caused by triplication of chromosome 21, are characterized by intellectual disability and are prone to develop Alzheimer’s disease (AD), due to triplication of the amyloid precursor protein (APP) gene. Recent evidence in the Ts65Dn mouse model of DS shows that enhancement of serotonergic transmission with fluoxetine during the perinatal period rescues neurogenesis, dendritic pathology and behavior, indicating that cognitive impairment can be pharmacologically restored. A crucial question is whether the short-term effects of early treatments with fluoxetine disappear at adult life stages. In the current study we found that hippocampal neurogenesis, dendritic pathology and hippocampus/amygdala-dependent memory remained in their restored state when Ts65Dn mice, which had been neonatally treated with fluoxetine, reached adulthood. Additionally, we found that the increased levels of the APP-derived βCTF peptide in adult Ts65Dn mice were normalized following neonatal treatment with fluoxetine. This effect was accompanied by restoration of endosomal abnormalities, a βCTF-dependent feature of DS and AD. While untreated adult Ts65Dn mice had reduced hippocampal levels of the 5-HT1A receptor (5-HT1A-R) and methyl-CpG-binding protein (MeCP2), a protein that promotes 5-HT1A-R transcription, in neonatally-treated mice both 5-HT1A-R and MeCP2 were normalized. In view of the crucial role of serotonin in brain development, these findings suggest that the enduring outcome of neonatal treatment with fluoxetine may be due to MeCP2-dependent restoration of the 5-HT1A-R. Taken together, results provide new hope for the therapy of DS, showing that early treatment with fluoxetine enduringly restores cognitive impairment and prevents early signs of AD-like pathology.

KEYWORDS: 5-HT1A receptor; Alzheimer’s disease; Cognitive impairment; Down syndrome; Pharmacotherapy

PMID: 25497735

 

SUPPLEMENTS:

Intellectual disability is the invariable hallmark and the most invalidating feature of Down syndrome (DS), a genetic disorder caused by triplication of chromosome 21. A unique characteristic of DS is the high risk for the onset of Alzheimer’s Disease (AD) in individuals over 35 years of age, with worsening of cognitive functions. AD neuropathology appears in virtually all adults with DS older than 40 years of age, with accumulation of beta-amyloid and hyperphosphorylation of tau proteins. This implies that trisomic genes play a paramount role in disease development.

 

Figure_1

 

Neuronal endosomal abnormalities are the earliest signs in sporadic AD and DS (1) and appear well before cognitive decline. Endosomal alterations are characterized by enlargement of early endosomes and increased immunoreactivity for endosome fusion (such as rab5) and recycling (rab4) markers. The endocytic pathway plays a crucial role in the internalization, recycling, and catabolic modulation of proteins relevant to AD, including amyloid precursor protein (APP), beta-amyloid peptide and apolipoprotein E. Thus, early endosomal alterations may be one of the mechanisms underlying neurodegenerative changes during AD progression.

App is one of the triplicated genes in DS thought to contribute not only to the progression of AD, through excessive formation of APP-derived beta-amyloid peptide, but also to development of early endosomal alterations. In fact, it has been shown that endosome dysfunction in DS is dependent on elevated levels of APP carbossiterminal fragment beta (βCTF) (3). This fragment derives from the amyloidogenic cleavage of APP by β secretase (BACE1) (Fig 1). Increased levels of BACE1 in the DS brain may additionally contribute to further enhance βCTF levels (Fig.1).

 

Figure_2

 

There are currently no therapies to prevent AD development in DS. Therefore in our study, we wondered whether it was possible to pharmacologically prevent early signs of AD exploiting the Ts65Dn mouse model of DS. Similarly to individuals with DS, this model is bound to develop AD with age. Noteworthy, the Ts65Dn mouse exhibits AD-like early endosomal abnormalities starting from 2 months of age, i.e. well before the appearance of cognitive deterioration. We treated mice with the antidepressant fluoxetine, a selective inhibitor of the serotonin transporter (SERT). This drug increases the amount of serotonin available to postsynaptic receptors, preventing serotonin from being taken back into the presynaptic neuron (Fig.2). We thought that treatment with fluoxetine may prevent AD development based on the following rationale: 1) serotonin signaling regulates APP processing (2) 2) the serotonergic system is altered in DS starting from early phases of brain development; 3) treatment with fluoxetine in an AD mouse model decreases beta amyloid levels similarly to infusion of serotonin into the hippocampus (2); 4) cognitively normal elderly individuals who were exposed to antidepressant drugs for five years have significantly less amyloid load in comparison with untreated individuals (2). We treated euploid and Ts65Dn mice during the first neonatal period with either saline or fluoxetine and we analyzed the outcome of treatment when mice reached adulthood.

As detailed below and shown in Fig. 2, we discovered very positive effects of fluoxetine on APP processing and endosomal alteration in treated Ts65Dn mice in comparison with their untreated counterparts. We found that adult Ts65Dn mice had higher bCTF and BACE1 levels in comparison with euploid mice. Importantly, in neonatally-treated Ts65Dn mice bCTF and BACE1 levels were fully normalized and became similar to those of euploid mice. We next evaluated the immunoreactivity of Rab5, a GTPase involved in early endosome fusion, in endosomes of individual cells in the septal complex and we found that untreated Ts65Dn mice exhibited abnormally high Rab5 levels. In parallel with normalization of bCTF levels after treatment, Rab5 levels in Ts65Dn mice underwent a reduction and became similar to those of euploid mice. Interestingly, the positive effects of early treatment in Ts65Dn mice were accompanied by full restoration of the expression levels of the 5-HT1A receptor, suggesting that restoration of serotonergic signaling may be involved in the observed restoration of APP processing and endosome alterations in adulthood.

These data indicate that an early therapy with fluoxetine can prevent early signs of AD in the Ts65Dn mouse model, highlighting the role of serotonin signaling in APP processing. The finding that early therapy with fluoxetine is able to induce long-lasting positive effects at adult life stages suggests that serotonin reuptake inhibitors may be a good strategy to prevent AD.

Importantly, the study of the effects of preventive treatments in a mouse model of DS, bound to develop AD, may benefit not only patients with DS but it may additionally provide important insights about the efficacy and timing of interventions targeting sporadic AD in the general population. Research efforts in discovering disease-modifying treatments against sporadic AD have focused on patients already diagnosed with AD, with scarce success. Since it is very likely that interventions initiated earlier in the disease process may be more beneficial, great efforts are currently made in order to detect biomarkers that may be predictive of sporadic AD in the general population, in order to correctly identifying individuals who will develop the pathology. Therefore once biomarkers alterations have been detected, information gained in DS may be exploited to devise appropriate therapeutic strategies to prevent AD in the general population.

 

References

  1. Cataldo AM, Petanceska S, Peterhoff CM, Terio NB, Epstein CJ, Villar A, Carlson EJ, Staufenbiel M, Nixon RA (2003) App gene dosage modulates endosomal abnormalities of Alzheimer’s disease in a segmental trisomy 16 mouse model of down syndrome. J Neurosci 23:6788-6792.
  2. Cirrito JR, Disabato BM, Restivo JL, Verges DK, Goebel WD, Sathyan A, Hayreh D, D’Angelo G, Benzinger T, Yoon H and others (2011) Serotonin signaling is associated with lower amyloid-beta levels and plaques in transgenic mice and humans. Proc Natl Acad Sci U S A 108:14968-14973.
  3. Jiang Y, Mullaney KA, Peterhoff CM, Che S, Schmidt SD, Boyer-Boiteau A, Ginsberg SD, Cataldo AM, Mathews PM, Nixon RA (2009) Alzheimer’s-related endosome dysfunction in Down syndrome is Abeta-independent but requires APP and is reversed by BACE-1 inhibition. Proc Natl Acad Sci U S A 107:1630-1635.

RB Figure_3

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