Dysfunctional hippocampal inhibition in the Ts65Dn mouse model of Down syndrome

GABAergic dysfunction is implicated in hippocampal deficits of the Ts65Dn mouse model of Down syndrome (DS). Since Ts65Dn mice overexpress G-protein coupled inward-rectifying potassium (GIRK2) containing channels, we sought to evaluate whether increased GABAergic function disrupts the functioning of hippocampal circuitry. After confirming that GABA(B)/GIRK current density is significantly elevated in Ts65Dn CA1 pyramidal neurons, we compared monosynaptic inhibitory inputs in CA1 pyramidal neurons in response to proximal (stratum radiatum; SR) and distal (stratum lacunosum moleculare; SLM) stimulation of diploid and Ts65Dn acute hippocampal slices. Synaptic GABA(B) and GABA(A) mediated currents evoked by SR stimulation were generally unaffected in Ts65Dn CA1 neurons. However, the GABA(B)/GABA(A) ratios evoked by stimulation within the SLM of Ts65Dn hippocampus were significantly larger in magnitude, consistent with increased GABA(B)/GIRK currents after SLM stimulation. These results indicate that GIRK overexpression in Ts65Dn has functional consequences which affect the balance between GABA(B) and GABA(A) inhibition of CA1 pyramidal neurons, most likely in a pathway specific manner, and may contribute to cognitive deficits reported in these mice.     

Luteolin induces hippocampal neurogenesis in the Ts65Dn mouse model of Down syndrome

Studies have shown that the natural flavonoid luteolin has neurotrophic activity. In this study, we investigated the effect of luteolin in a mouse model of Down syndrome. Ts65 Dn mice, which are frequently used as a model of Down syndrome, were intraperitoneally injected with 10 mg/kg luteolin for 4 consecutive weeks starting at 12 weeks of age. The Morris water maze test was used to evaluate learning and memory abilities, and the novel object recognition test was used to assess recognition memory. Immunohistochemistry was performed for the neural stem cell marker nestin, the astrocyte marker glial fibrillary acidic protein, the immature neuron marker DCX, the mature neuron marker NeuN, and the cell proliferation marker Ki67 in the hippocampal dentate gyrus. Nissl staining was used to observe changes in morphology and to quantify cells in the dentate gyrus. Western blot assay was used to analyze the protein levels of brain-derived neurotrophic factor(BDNF) and phospho-extracellular signal-regulated kinase 1/2(p-ERK1/2) in the hippocampus. Luteolin improved learning and memory abilities as well as novel object recognition ability, and enhanced the proliferation of neurons in the hippocampal dentate gyrus. Furthermore, luteolin increased expression of nestin and glial fibrillary acidic protein, increased the number of DCX~+ neurons in the granular layer and NeuN~+ neurons in the subgranular region of the dentate gyrus, and increased the protein levels of BDNF and p-ERK1/2 in the hippocampus. Our findings show that luteolin improves behavioral performance and promotes hippocampal neurogenesis in Ts65 Dn mice. Moreover, these effects might be associated with the activation of the BDNF/ERK1/2 pathway.     

Working memory in the Ts65Dn mouse, a model for Down syndrome

This study used a matching-to-position schedule of reinforcement to examine working memory in Ts65Dn and littermate control mice. Initially there appeared to be a memory deficit in the Ts65Dn mice, which disappeared with extended practice. Thus, what appeared as a memory deficit may actually be the result of a delay in learning the concept of matching. These results suggest that delayed learning may be an important factor in other procedures examining working memory in Ts65Dn mice and have important implications for clinical treatment of Down syndrome patients.     

Episodic-like memory in Ts65Dn, a mouse model of Down syndrome

Ts65Dn mice, like individuals with Down syndrome (DS), demonstrate a functional dissociation between explicit and implicit forms of memory, showing selective impairment in explicit or declarative learning tasks. Here, we explored Ts65Dn explicit memory deficits further by evaluating the ability of these mice to assimilate the temporal and spatial contexts under which previously novel objects had been encountered. We found that Ts65Dn mice could in fact form contextual representations of objects over the course of a few hours, contrary to their inability to discriminate object novelty over a more prolonged period of 24h. These results suggest that Ts65Dn mice might have particular difficulties in declarative tasks requiring long-term memory, presenting an especially important putative therapeutic target for pre-clinical and clinical DS research.     

Working memory in the aged Ts65Dn mouse, a model for Down syndrome

The Ts65Dn mouse displays several phenotypic abnormalities that parallel characteristics found in Down syndrome. One important characteristic associated with Down syndrome is an increased incidence of early-onset Alzheimer's disease. Since Alzheimer's disease is characterized largely by progressive memory loss, it is of interest to study working memory in the Ts65Dn mouse. Previous research in our lab using a titrating, delayed matching-to-position schedule of reinforcement has demonstrated that young, adult male Ts65Dn mice do not display a working memory deficit when compared to age-matched littermate controls. However, there have been no studies examining the working memory of these mice as they age. Due to the correlation between Down syndrome and Alzheimer's disease, and as part of a larger effort to further characterize the phenotype of the Ts65Dn mouse, the purpose of this study was to determine whether aged Ts65Dn mice possess a working memory deficit when compared to age-matched littermate controls. In order to study working memory, two groups of mice were trained under a titrating, delayed matching-to-position schedule of reinforcement. The first group was trained beginning at 3 months of age, and the second group began training at 15 months of age. Both groups were studied to 24 months of age. Initially, both groups of Ts65Dn mice performed at a lower level of accuracy than the control mice; however, this difference disappeared with further practice. The results from these lifespan studies indicate that the aged Ts65Dn mouse does not possess a working memory deficit when compared to age-matched controls.     

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.     

Differential effects of environmental enrichment on behavior and learning of male and female Ts65Dn mice,a model for Down syndrome

We have assessed the effects of enriched environment (EE) upon behavioral and cognitive performances of partially trisomic Ts65Dn (TS) mice and their control (CO) littermates. Enriched environment was applied to pups for 7 weeks after weaning. Circadian spontaneous activity (actimetry), exploratory behavior (hole board), activity in the open field and spatial memory (Morris Water Maze, repeated acquisition and cued paradigms) were analyzed in 86 female and 75 male mice, starting 15 days after completing enrichment. For each gender, mice were distributed in non-enriched and enriched control and trisomic groups. Enriched environment reduced in trisomic females and enhanced in trisomic males' circadian activity. Exploratory behavior was increased by enrichment in all groups, regardless of gender or presence of trisomy. In the Morris Water Maze, a significant improvement of the spatial memory was observed in enriched-control females, but not in enriched-control male mice, as assessed by distances traveled. Performances in the four groups of control animals were also consistently and significantly better than those of matching trisomic mice. In the acquisition trials, enrichment improved performance in trisomic female animals, but deteriorated in trisomic male mice. In all groups, changes in escape latencies and distances induced by enrichment were accounted for by changes in the total time spent in the periphery of the pool, indicating changes in learning strategy. Working memory was the function more affected by enrichment. It is concluded that enriched environment induces behavioral and learning changes in trisomic mice, although gender plays a significant modulatory role.     

Abnormal APP, cholinergic and cognitive function in Ts65Dn Down's model mice

We evaluated Ts65Dn Down's syndrome mice and their littermates (LM) at 1-2, 4, and 12 months of age to determine amyloid precursor protein (APP)-related cellular and biochemical changes associated with cognitive deficits. Ts65Dn mice showed cognitive deficits in the Morris water maze compared to LM mice at 4 and 12 months of age. Ts65Dn, but not LM mice, developed a septohippocampal cholinergic neuronal degeneration of choline acetyltransferase (ChAT)-positive neurons at 12 months of age. These cellular changes were compensated by increases in ChAT enzyme activity of remaining cholinergic terminals in the hippocampus. By 12 months of age, Ts65Dn mice had elevations of APP protein levels in the hippocampus compared to their LM. At this age, both Ts65Dn mice and their LM abnormally responded to cholinergic muscarinic M1 agonist treatment in terms of hippocampal APP, nerve growth factor (NGF), and brain-derived neurotrophic factor (BDNF) levels compared to young adult C57BL/6 mice. In summary, the Ts65Dn mice show developmental and progressive age-related behavioral deficits, hippocampal APP, and cholinergic pathology. The relatively better cognitive spatial performance in LM compared to Ts65Dn mice suggests that high APP levels combined with progressive degeneration of the cholinergic system are critical to the pathology and cognitive deficits seen in Ts65Dn mice.     

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.     

Behavioral, cognitive and biochemical responses to different environmental conditions in male Ts65Dn mice, a model of Down syndrome

Ts65Dn mouse is the most widely accepted model for Down syndrome. We previously showed that environmental enrichment improved spatial learning in female but deteriorated it in male Ts65Dn mice. This study analyzed the factors contributing to the disturbed cognition of male Ts65Dn mice after enriched housing, by allocating male control and Ts65Dn mice in four conditions after weaning: small (n = 2-3) and large group (n = 8-10) housing, and enriched housing in small (2-3) and large groups (8-10). Learning, aggressive behavior, anxiety-like behavior and biochemical correlates of stress were evaluated when Ts65Dn and control mice were 4-5 months old. Environmental enrichment in large mixed colonies of Ts65Dn and diploid littermates disturbed behavioral and learning skills of Ts65Dn mice in the Morris water maze. ACTH and testosterone levels were not modified in any group of mice. Ts65Dn and control mice subjected to enriched housing in large groups and Ts65Dn mice housed in large groups showed higher corticosterone levels. Aggressive behavior was evaluated by measuring the number of attacks performed in the presence of an intruder. Ts65Dn mice performed less attacks than controls in all conditions, especially after enriched housing, indicating subordination. In the plus maze, cognitive aspects (i.e. risk assessment) and motor components (open arm avoidance) of anxiety behavior were evaluated; no difference in any condition was found. It is suggested that an excess of social and/or physical stimulation in Ts65Dn mice may affect cognition by disturbing the emotional and behavioral components of the learning process.     

Hypothermic responses to 8-OH-DPAT in the Ts65Dn mouse model of Down syndrome

Recently, we have demonstrated that potassium channels containing G-protein-activated potassium channel 2 (GIRK2) subunits play a significant role in hypothermia induced by several neurotransmitter receptor agonists, including the serotonin (5-HT)1A/5-HT7 receptor agonist 8-OH-DPAT [R-(+)-8-hydroxy-2-(di-n-propylamino) tetralin]. The GIRK2 gene is located in human chromosome 21 (its mouse ortholog, Girk2, is in mouse chromosome 16). Down syndrome is produced by the trisomy of chromosome 21. Here, we used quantitative radiotelemetry to investigatehypothermic responses to 8-OH-DPAT in the Down syndrome mouse model Ts65Dn (which carries an extra chromosomal 16 segment containing Girk2). Our results indicate that, in relation to euploid controls, Ts65Dn mice display significantly increased hypothermic responses to 8-OH-DPAT. This finding may be relevant to the understanding of previously reported differences in serotoninergic neurotransmission in persons with Down syndrome.     

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.     

Synaptic structural abnormalities in the Ts65Dn mouse model of Down Syndrome

The Ts65Dn mouse is a genetic model for Down syndrome. Although this mouse shows abnormalities in cognitive function that implicate hippocampus as well as marked deficits in hippocampal long-term potentiation, the structure of the hippocampus has been little studied. We characterized synaptic structure in Ts65Dn and control (2N) mice, studying the hippocampus (fascia dentata, CA1) as well as the motor and somatosensory cortex, entorhinal cortex, and medial septum. Confocal microscopy was used to examine immunostained presynaptic boutons and to detail the structure of dendrites after Lucifer yellow microinjection. Both presynaptic and postsynaptic elements were significantly enlarged in Ts65Dn in all regions examined. The changes were detected at the youngest age examined (postnatal day 21) and in adults. In studies detailing the changes in fascia dentata and motor cortex, the enlargement of spines affected the entire population, resulting in the presence of spines whose volume was greatly increased. Electron microscopy confirmed that boutons and spines were enlarged and demonstrated abnormalities in the internal membranes of both. In addition, spine density was decreased on the dendrites of dentate granule cells, and there was reorganization of inhibitory inputs, with a relative decrease in inputs to dendrite shafts and an increase in inputs to the necks of spines. Taken together, the findings document widespread abnormalities of synaptic structure that recapitulate important features seen in Down syndrome. They establish the Ts65Dn mouse as a model for abnormal synapse structure and function in Down syndrome and point to the importance of studies to elucidate the mechanisms responsible for synapse enlargement.     

G-protein-associated signal transduction processes are restored after postweaning environmental enrichment in Ts65Dn, a Down syndrome mouse model

Individuals with Down syndrome (DS) present cognitive deficits that can be improved by early implementation of special care programs. However, they showed limited and temporary cognitive effects. We previously demonstrated that postnatal environmental enrichment (EE) improved clearly, though temporarily, the execution of visuospatial memory tasks in Ts65Dn mice, a DS model bearing a partial trisomy of murine chromosome 16; but in contrast to wild-type littermates, there was a lack of structural plasticity in pyramidal cell structure in the trisomic cerebral cortex. In the present study, we have investigated the impact of EE on the function of adenylyl cyclase and phospholipase C as a possible mechanism underlying the time-limited improvements observed. Basal production of cyclic adenosine monophosphate (cAMP) was not affected, but responses to GTPγS, isoprenaline, noradrenaline, SKF 38393 and forskolin were depressed in the Ts65Dn hippocampus. In EE conditions, cAMP accumulation was not significantly modified in control animals with respect to nonenriched controls. However, EE had a marked effect in Ts65Dn mice, in which cAMP production was significantly increased. Similarly, EE increased phospholipase C activity in Ts65Dn mice, in response to carbachol and calcium. We conclude that EE restores the G-protein-associated signal transduction systems that are altered in Ts65Dn mice.     

Reactivity to object and spatial novelty is normal in older Ts65Dn mice that model Down syndrome and Alzheimer's disease

Ts65Dn mice, a model for Down syndrome and Alzheimer's disease, have a spontaneous age-related reduction of cholinergic markers in medial septal neurons, hippocampal abnormalities, and an age-related learning deficit in a task that requires an intact hippocampus. Others have shown that when normal rodents explored an open field with objects, they detected the displacement of some of the familiar objects within the arena (spatial novelty) and the presence of a new object (object novelty); whereas rodents with hippocampal, fornix, or neonatal selective basal forebrain cholinergic lesions were impaired in detecting spatial, but not object, novelty. In this study, both control and Ts65Dn mice responded to both the spatial and object changes. This unexpected finding could have several explanations. One may be related to recent studies that suggest that only rats with neonatal, but not adult, basal forebrain cholinergic 192 IgG-saporin lesions are impaired in reacting to spatial novelty.     

Altered expression of neuropeptides in the primary somatosensory cortex of the Down syndrome model Ts65Dn

Down syndrome is the most common genetic disorder associated with mental retardation. Subjects and mice models for Down syndrome (such as Ts65Dn) show defects in the formation of neuronal networks in both the hippocampus and the cerebral cortex. The principal neurons display alterations in the morphology, density and distribution of dendritic spines in the cortex as well as in the hippocampus. Several evidences point to the possibility that the atrophy observed in principal neurons could be mediated by changes in their inhibitory inputs and, in fact, an imbalance between excitation and inhibition has been observed in Ts65Dn mice in these regions, which are crucial for learning and information processing. These animals have an increased density of interneurons in the primary somatosensory cortex, especially of those expressing calretinin and calbindin D-28k. Here, we have analysed the expression and distribution of several neuropeptides in the primary somatosensory cortex of Ts65Dn mice in order to investigate whether these subpopulations of interneurons are affected. We have observed an increase in the total density of somatostatin expressing interneurons and of those expressing VIP in layer IV in Ts65Dn mice. The typology of the somatostatin and VIP interneurons was unaltered as attested by the pattern of co-expression with other markers. Somatostatin immunoreactive neurons co-express mainly D-28k calbindin and VIP expressing interneurons maintain its pattern of co-expression with calcium binding proteins. These alterations, in case they were also present in subjects with Down syndrome, could be related to their impairment in cognitive profile and could be involved in the neurological defects observed in this disorder.     

Effects of chronic administration of SGS-111 during adulthood and during the pre- and post-natal periods on the cognitive deficits of Ts65Dn mice, a model of Down syndrome

The Ts65Dn mouse is the most commonly used model of Down syndrome. This mouse shows many phenotypic characteristics present in people with Down syndrome, including behavioral and cognitive deficits. SGS-111 is a novel analogue of the nootropic piracetam, which prevents oxidative damage and apoptosis in both normal and Down syndrome human cortical neurons. In this work we tested the ability of chronic administration of SGS-111 to adult Ts65Dn mice to reverse the cognitive deficit found in these mice. Moreover, since oxidative stress has been reported as early as the fetal stage, SGS-111 was also administered to pregnant Ts65Dn females from the day of conception throughout the pregnancy and to Ts65Dn pups during their entire life (5 months), from birth to the end of the behavioral testing period. A characterization of the effects of SGS-111 treatment on Ts65Dn and control mice sensorimotor abilities, motor coordination, spontaneous activity, activity in the open field, exploration, anxiety and spatial and non-spatial short- and long-term learning and memory was performed. The behavioral characterization showed that chronic administration of the antioxidant SGS-111 reduced the hyperactivity shown by Ts65Dn mice in their home cage, in the open field and in the hole board test. SGS-111 administration during adulthood improved performance in the first session in the Morris water maze in control mice, and when administered during the pre- and post-natal periods, improved spatial learning in the control mice but not in Ts65Dn mice. Chronic SGS-111 administration failed to affect behavior and cognition in Ts65Dn mice.     

Environmental enrichment rescues postnatal neurogenesis defect in the male and female Ts65Dn mouse model of Down syndrome

Down syndrome (DS), the most frequent genetic cause of intellectual disability and developmental delay, results from impaired neural stem cell proliferation and differentiation. Impaired neurogenesis in the neocortex, hippocampus and cerebellum is believed to be the underlying cause of learning and behavioral deficits in the Ts65Dn mouse model of DS. Aggressive sensorimotor and cognitive therapies have shown promise in mitigating the cognitive disabilities in DS but these behavioral therapies have not yet been investigated at the cellular level. Here, using the Ts65Dn mouse model of DS, we demonstrate that a combination of environmental enrichment and physical exercise starting in juvenile mice (postnatal day 18) markedly increases cell proliferation, neurogenesis and gliogenesis in the hippocampal dentate gyrus (DG) and the forebrain subventricular zone (SVZ) of both male and female mice. Enrichment and exercise increased the rate of Ts65Dn DG neurogenesis to be comparable to that of the nonenriched euploid group, while the effect on SVZ neurogenesis was reduced and seen only after prolonged exposure. These results clearly indicate that in a comprehensive stimulatory environment, the postnatal DS brain has the intrinsic capability of improving neurogenesis and gliogenesis to the levels of normal matched controls and that this cellular response underlies the cognitive improvement seen following behavioral therapies.     

Increased efficiency of the GABAA and GABAB receptor-mediated neurotransmission in the Ts65Dn mouse model of Down syndrome

Cognitive impairment in Down syndrome (DS) involves the hippocampus. In the Ts65Dn mouse model of DS, deficits in hippocampus-dependent learning and synaptic plasticity were linked to enhanced inhibition. However, the mechanistic basis of changes in inhibitory efficiency remains largely unexplored, and efficiency of the GABAergic synaptic neurotransmission has not yet been investigated in direct electrophysiological experiments. To investigate this important feature of neurobiology of DS, we examined synaptic and molecular properties of the GABAergic system in the dentate gyrus (DG) of adult Ts65Dn mice. Both GABAA and GABAB receptor-mediated components of evoked inhibitory postsynaptic currents (IPSCs) were significantly increased in Ts65Dn vs. control (2N) DG granule cells. These changes were unaccompanied by alterations in hippocampal levels of GABAA (α1, α2, α3, α5 and γ2) or GABAB (Gbr1a and Gbr1b) receptor subunits. Immunoreactivity for GAD65, a marker for GABAergic terminals, was also unchanged. In contrast, there was a marked change in functional parameters of GABAergic synapses. Paired stimulations showed reduced paired-pulse ratios of both GABAA and GABAB receptor-mediated IPSC components (IPSC2/IPSC1), suggesting an increase in presynaptic release of GABA. Consistent with increased gene dose, the level of the Kir3.2 subunit of potassium channels, effectors for postsynaptic GABAB receptors, was increased. This change was associated with enhanced postsynaptic GABAB/Kir3.2 signaling following application of the GABAB receptor agonist baclofen. Thus, both GABAA and GABAB receptor-mediated synaptic efficiency is increased in the Ts65Dn DG, thus likely contributing to deficient synaptic plasticity and poor learning in DS.