Fluoxetine rescues deficient neurogenesis in hippocampus of the Ts65Dn mouse model for Down syndrome

The Ts65Dn mouse, an adult model of Down syndrome displays behavioral deficits consistent with a dysfunctional hippocampus, similar to that seen with DS. In looking for mechanisms underlying these performance deficits, we have assessed adult neurogenesis in the dentate gyrus of Ts65Dn. Under untreated conditions, Ts65Dn mice (2-5 months old) showed markedly fewer BrdU-labeled cells than euploid animals. Chronic antidepressant treatment for over 3 weeks with the serotonin selective reuptake inhibitor, fluoxetine, increased neurogenesis in the Ts65Dn to comparable levels seen in the euploid by augmenting both proliferation and survival of BrdU-labeled cells in the subgranular layer and granule cell layer of the hippocampus, respectively.     

Alterations of central noradrenergic transmission in Ts65Dn mouse, a model for Down syndrome

Mice with segmental trisomy 16 (Ts65Dn) which have triplication of a region of mouse chromosome 16 homologous to the Down syndrome critical region in human chromosome 21, are used as a model for Down syndrome. Functioning of the central β-noradrenergic transmission was studied in Ts65Dn mice. Binding analysis in cerebral cortex revealed no change in the number of β-adrenoceptors and a slight reduction of affinity. The β-adrenoceptor transduction was assessed by analyzing cAMP formation in the cerebral cortex, hippocampus and cerebellar cortex under basal conditions and after stimulation with isoprenaline and forskolin. Basal production of cAMP was significantly reduced in hippocampus and cerebellar cortex of Ts65Dn mice compared to control, but not in cerebellum. After phosphodiesterase inhibition, net increments in cAMP accumulation were similar in both groups of mice. Stimulation of cAMP production by isoprenaline (10 μM) and forskolin (10 μM) was much higher in hippocampus than in cerebral cortex of either group. In both areas, but not in cerebellum, the stimulatory responses were consistently and significantly smaller in Ts65Dn than in control mice. Concentration–response curves for isoprenaline and forskolin were generated in the cerebral cortex. E_max responses were lower in trisomic than in control mice; however, in Ts65Dn mice the slope of the response curve to isoprenaline was markedly depressed whereas that to forskolin was similar to control. It is concluded that Ts65Dn mice show severe deficiencies in the synaptic transmission of the central β-noradrenergic system, which are selective for specific brain areas.     

The Ts65Dn mouse model of Down syndrome shows reduced expression of the Bcl-XL antiapoptotic protein in the hippocampus not accompanied by changes in molecular or cellular markers of cell death

The Ts65Dn (TS) mouse, the most widely used model of Down syndrome (DS), has a partial trisomy of a segment of chromosome 16 that is homologous to the distal part of human chromosome 21. This mouse shares many phenotypic characteristics with people with DS including neuromorphological, neurochemical, and cognitive disturbances. Both TS and DS brains show earlier aging and neurodegeneration. Since fibroblast cultures from TS mice and human DS hippocampal regions show increased apoptotic cell death it has been suggested that alterations in cerebral apoptosis might be implicated in the cognitive deficits found in TS mice and in people with DS. In the present study we have evaluated brain expression levels of several proapoptotic and antiapoptotic proteins from the mitochondrial (Bcl-2, Bcl-X_L, Bax and Bad) and the extrinsic (Fas-R and Fas-L) apoptotic pathways as well as the final executioner caspase-3, in the cortex and hippocampus of TS mice. No significant alterations in the expression levels of the proapoptotic Bad and Bax or the antiapoptotic Bcl-2 proteins in the cortex or hippocampus were found in TS mice. However, TS mice showed downregulation of Bcl-X_L in the hippocampus. In the extrinsic pathway we found unchanged levels of Fas-L in both structures and also in the expression levels of Fas-R in the hippocampus. Although Bcl-X_L downregulation suggests that the hippocampus of TS mice is less protected against programmed cell death, we did not find any evidence for increased apoptosis in TS mice since neither TUNEL-positive cells nor active caspase-3 expression were found in cortex or hippocampus of TS or CO mice.     

Three-dimensional synaptic ultrastructure in the dentate gyrus and hippocampal area CA3 in the Ts65Dn mouse model of Down syndrome

Down syndrome (DS) results from trisomy of human chromosome 21. Ts65Dn mice are an established model for DS and show several phenotypes similar to those in people with DS. However, there is little data on the structural plasticity of synapses in the trisynaptic pathway in the hippocampus. Here we investigate 3D ultrastructure of synapses in the hippocampus of age-matched control (2N) and Ts65Dn male mice. Serial ultrathin sections and 3D reconstructions characterize synapses in the middle molecular layer (MML) of dentate gyrus and in thorny excrescences (TEs) in proximal portions of apical dendrites of CA3 pyramidal neurons. 3D analysis of synapses shows phenotypes that distinguish Ts65Dn from 2N mice. For the MML, synapse density was reduced by 15% in Ts65Dn vs. 2N mice (P < 0.05). Comparative 3D analyses demonstrate a significant decrease in the number of thorns per TE in CA3 in Ts65Dn vs. 2N mice (by ≈45%, P = 0.01). Individual thorn volume was 3 times smaller in Ts65Dn vs. 2N mice (P = 0.02). A significant decrease in the number of thorn projections per TE in Ts65Dn vs. 2N mice was accompanied by a decrease of filopodium-like protrusions on the surface of TEs (P = 0.02). However, the volume of postsynaptic densities in CA3 Ts65Dn and 2N mice was unchanged (P = 0.78). Our findings suggest that the high degree of plasticity of CA3 thorns may be connected with their filopodial origin. Alterations of 3D synaptic structure in Ts65Dn mice may further contribute to the diminished plasticity in DS.     

Neonatal mice of the down syndrome model, Ts65Dn, exhibit upregulated VIP measures and reduced responsiveness of cortical astrocytes to VIP stimulation

The Ts65Dn segmental mouse model of Down syndrome (DS) possesses a triplication of the section of chromosome 16 that is most homologous to the human chromosome 21 that is trisomic in DS. This model exhibits many of the characteristics of DS including small size, developmental delays, and a decline of cholinergic systems and cognitive function with age. Recent studies have shown that vasoactive intestinal peptide (VIP) systems are upregulated in aged Ts65Dn mice and that VIP dysregulation during embryogenesis is followed by the hypotonia and developmental delays as seen in both DS and in Ts65Dn mice. Additionally, astrocytes from aged Ts65Dn brains do not respond to VIP stimulation to release survival-promoting substances. To determine if VIP dys-regulation is age-related in Ts65Dn mice, the current study examined VIP and VIP receptors (VPAC-1 and VPAC-2) in postnatal day 8 Ts65Dn mice. VIP and VPAC-1 expression was significantly increased in the brains of trisomic mice compared with wild-type mice. VIP-binding sites were also significantly increased in several brain areas of young Ts65Dn mice, especially in the cortex, caudate/putamen, and hippocampus. Further, in vitro treatment of normal neurons with conditioned medium from VIP-stimulated Ts65Dn astrocytes from neonatal mice did not enhance neuronal survival. This study indicates that VIP anomalies are present in neonatal Ts65Dn mice, a defect occurs in the signal transduction mechanism of the VPAC-1 VIP receptor, cortical astrocytes from neonatal brains are dysfunctional, and further, that VIP dysregulation may play a significant role in DS.     

Cell lines derived from hippocampal neurons of the normal and trisomy 16 mouse fetus (a model for Down syndrome) exhibit neuronal markers,cholinergic function,and functional neurotransmitter receptors

We have established hippocampal cell lines from normal and trisomy 16 fetal mice, a model of human trisomy 21. Both cell lines, named H1b (derived from a normal animal) and HTk (trisomic) possess neuronal markers by immunohistochemistry (enolase, synaptophysin, microtubule associated protein-2, and choline acetyltransferase) and lack glial markers (glial fibrillary acidic protein and S-100). Also, we evaluated intracellular Ca(2+) levels ([Ca(2+)](i)) in response to neurotransmitter agonists, in cells loaded with the fluorescent Ca(2+) indicators Indo-1 and Fluo-3. Both cell lines responded to glutamatergic stimuli induced by glutamate, N-methyl-D-aspartate, I-amino-2,3-dihydro-5-methyl-3-oxo-4-isoxazole propanoic acid or kainate. Glutamate responses were only partially prevented by addition of 5 mM EGTA and the metabotropic glutamate receptor agonist, trans-(1S,3R)-1-amino-1,3-cyclopentanedicarboxylic acid (ACPD), increased [Ca(2+)](i) in both cell types. These results confirm the presence of glutamatergic metabotropic receptors. In glutamate-induced responses, HTk cells exhibited slower time-dependent decay kinetics than H1b cells. Cholinergic agonists (nicotine and muscarine) induced a rapid, transient increase in [Ca(2+)](i) in both cell types. Furthermore, some cells were sensitive to histamine and norepinephrine. All responses to the aforementioned agonists were prevented by addition of specific antagonists. We also studied incorporation and release of [(3)H]choline in the cells, and observed no differences in uptake parameters. However, release induced by K(+) and nicotine depolarization was greatly reduced in HTk cells. The results show that H1b and HTk cells retain neuronal characteristics and respond to specific neurotransmitter stimuli. The HTk differences could be related to neuronal pathophysiology in Down syndrome.