Evidence that increased Kcnj6 gene dose is necessary for deficits in behavior and dentate gyrus synaptic plasticity in the Ts65Dn mouse model of Down syndrome
Introduction
Down syndrome (DS), due to triplication of human chromosome 21 (HSA21) (Lejeune et al., 1959), results in a number of significant neurobiological and somatic phenotypes. The most salient are cognitive and behavioral impairments in children and the emergence of Alzheimer's disease in the elderly (Belichenko et al., 2016, Dierssen, 2012, Dykens, 2007, Kleschevnikov et al., 2012c, Lott, 2012, Nadel, 2003, Roizen and Patterson, 2003, Roper and Reeves, 2006, Sabbagh and Edgin, 2016). The gene-dosage hypothesis proposes that all DS-related phenotypes are due to the presence in excess of one or more genes or regulatory sequences on HSA21 (Epstein et al., 1981). Tests of this hypothesis has in recent years resulted in insights into a number of the genes and mechanisms responsible for DS phenotypes (Belichenko et al., 2009, Dierssen et al., 2011, Fotaki et al., 2002, Kleschevnikov et al., 2004, Olson et al., 2004b, Salehi et al., 2006, Vesa et al., 2005).
Mouse genetic models of DS provide an opportunity to investigate the neurobiology of DS (Belichenko et al., 2015, Das and Reeves, 2011, Edgin et al., 2012, Gardiner et al., 2003, Mojabi et al., 2016, Reeves et al., 1995, Rueda et al., 2012, Yu et al., 2010) and to explore the role of individual genes in DS phenotypes (Altafaj et al., 2013, Salehi et al., 2006). Similar to DS, mouse genetic models of DS exhibit deficient hippocampus-dependent long-term memory, working memory, and other physiological and behavioral changes consistent with deficits in cognition (Fernandez et al., 2007). Mouse models that contain different as well as overlapping sets of triplicated genes have been genetically engineered and examined to explore phenotype-genotype relationship (Davisson et al., 1990, Jiang et al., 2015, O'Doherty et al., 2005, Olson et al., 2004a, Pereira et al., 2009, Sago et al., 1998, Yu et al., 2010). The most intensively investigated mouse is the Ts65Dn model, whose creation 26 years ago was instrumental in advancing this field of study (Davisson et al., 1990). Ts65Dn mice exhibit many phenotypes characteristic of DS including deficient cognition (Braudeau et al., 2011, Busciglio et al., 2013, Costa, 2011, Demas et al., 1998, Faizi et al., 2011, Fernandez et al., 2007, Gutierrez-Castellanos et al., 2013, Martinez-Cue et al., 2014, Smith et al., 2014).
Studies in humans partially trisomic for HSA21 identified a region whose presence in three copies was sufficient to create many DS phenotypes, including intellectual disability; it was thereby labeled the Down Syndrome Critical Region (DSCR) (Korenberg, 1990, Peterson et al., 1994, Rahmani et al., 1989, Yamamoto et al., 2011). Recent studies in mice carrying all or part of the homologous region of mouse Chromosome 16 have shown that many DS-like phenotypes are linked to genes in this region (Belichenko et al., 2009, Dierssen and de Lagran, 2006, Jiang et al., 2015, Reeves et al., 1995). Within the DSCR, Kcnj6 is a candidate for contributing through increased dose to cognitive deficits. Kcnj6 is present in 3 copies in both people with DS and Ts65Dn mice. This gene encodes the Kir3.2 (Girk2) subunit of inwardly rectifying potassium channels which serve as effectors for a number of postsynaptic metabotropic receptors (Luscher et al., 1997, Mark and Herlitze, 2000, Yamada et al., 1998). As predicted by increased dose, the Kir3.2 product of Kcnj6 is increased in Ts65Dn mice (Harashima et al., 2006, Kleschevnikov et al., 2012b, Kleschevnikov et al., 2005). Suggesting a physiologically meaningful contribution for increased Kir3.2 in these mice, there was increased signaling through postsynaptic GABAB receptors in both primary cultures of hippocampal neurons (Best et al., 2007) and acute hippocampal slices (Best et al., 2012, Kleschevnikov et al., 2012b). In addition, suppressing enhanced GABAB/Kir3.2 signaling by treating with selective GABAB receptor antagonists restored synaptic plasticity and long-term memory in Ts65Dn mice (Kleschevnikov et al., 2012a). Recently, cognitive assessment in a series of mouse genetic models bearing distinct sets of genes within DSCR pointed to a contribution of Kcnj6 (Jiang et al., 2015).
A direct test of the impact of increased dose of specific genes is essential for defining contribution(s) to phenotypes. To address the impact of Kcnj6 triplication on cognitive phenotypes in DS, we genetically deleted the third copy by producing Ts65Dn mice with 2 copies of Kcnj6 (i.e., Ts65Dn:Kcnj6 ++−). Littermate Ts65Dn mice with three copies of Kcnj6 (Ts65Dn:Kcnj6 +++) and normosomic mice (2N:Kcnj6 ++) served as controls. Reduction of the Kcnj6 gene dose restored to normal the level of Kir3.2, long term memory, and short- and long- term potentiation in the DG. Remarkably, pharmacologically inhibiting Kir3.2-containing channels also restored synaptic plasticity. The findings are evidence that increased expression of Kcnj6 is necessary for the significant cognitive impairment in this model of DS and suggests that strategies aimed at pharmacologically reducing channel function should be explored for enhancing cognition in DS.
Section snippets
Animals
Segmental trisomy 16 (Ts65Dn) mice were purchased from the Jackson Laboratory, Bar Harbor, ME, stock #001924. Heterozygous Kcnj6 +/− mice (129/sv–C57BL/6 hybrid crosses) were a gift from Dr. L. Jan (UCSF). These mice were characterized in a number of previous studies (Blednov et al., 2001, Mitrovic et al., 2003, Signorini et al., 1997). Ts65Dn:Kcnj6 ++− mice were the result of crossing female Ts65Dn with male Kcnj6 +/− mice. Diploid (2N:Kcnj6 ++) and trisomic (Ts65Dn:Kcnj6 +++) mice served as
Kcnj6 dose influences body weight during aging
Body weight is a measure of systemic health. Children with DS show reduced body weight (Cronk, 1978), while adult individuals with DS often have increased body weight due to obesity (de Asua et al., 2014). To assess for an effect of Kcnj6 gene dose, the body weights of mice of each genotype were examined from 1 to 12 months of age (Fig. 1). Body weight was significantly reduced in Ts65Dn:Kcnj6 +++ vs. 2N:Kcnj6 ++ mice at all ages (Fig. 1, red vs. blue bar respectively; Table 1). Interestingly,
Discussion
Herein we examined the effects of the Kcnj6 gene dose on behavioral and cognitive phenotypes in the Ts65Dn mouse model of DS. Genetically normalizing Kcnj6 gene dose reduced to 2N levels the Kir3.2 protein subunit of inwardly-rectifying potassium channels, restored synaptic plasticity in the DG, and improved long-term memory. Our findings are evidence that increased gene dose for Kcnj6 is necessary for deficits in hippocampal synaptic plasticity and hippocampally-mediated tests of long-term
Acknowledgements
The study supported by The Jerome Lejeune Foundation (Grant # 1483), Cure Alzheimer's Foundation, LuMind /RDS Foundation, and NIH (R01NS55371 and R01NS66072).
References (104)
Normalization of Dyrk1A expression by AAV2/1-shDyrk1A attenuates hippocampal-dependent defects in the Ts65Dn mouse model of Down syndrome
Neurobiol. Dis.
(2013)Dysfunctional hippocampal inhibition in the Ts65Dn mouse model of Down syndrome
Exp. Neurol.
(2012)Frontal cortex BDNF levels correlate with working memory in an animal model of Down syndrome
Behav. Brain Res.
(2003)GIRK2 deficient mice. Evidence for hyperactivity and reduced anxiety
Physiol Behav.
(2001)Long-term memory in mental retardation: evidence for a specific impairment in subjects with Down's syndrome
Neuropsychologia
(1997)GABAB-GIRK2-mediated signaling in Down syndrome
Adv. Pharmacol.
(2010)Impaired spatial working and reference memory in segmental trisomy (Ts65Dn) mice
Behav. Brain Res.
(1998)- et al.
Working memory in the Ts65Dn mouse, a model for down syndrome
Behav. Brain Res.
(2006) Human and mouse model cognitive phenotypes in Down syndrome: implications for assessment
Prog. Brain Res.
(2012)Impaired short- and long-term memory in Ts65Dn mice, a model for Down syndrome
Neurosci. Lett.
(1998)
Comprehensive behavioral phenotyping of Ts65Dn mouse model of Down syndrome: activation of beta1-adrenergic receptor by xamoterol as a potential cognitive enhancer
Neurobiol. Dis.
Mouse models of Down syndrome: how useful can they be? Comparison of the gene content of human chromosome 21 with orthologous mouse genomic regions
Gene
Increased efficiency of the GABAA and GABAB receptor-mediated neurotransmission in the Ts65Dn mouse model of Down syndrome
Neurobiol. Dis.
Discoveries in Down syndrome: moving basic science to clinical care
Prog. Brain Res.
Neurological phenotypes for Down syndrome across the life span
Prog. Brain Res.
G protein-coupled inwardly rectifying K + channels (GIRKs) mediate postsynaptic but not presynaptic transmitter actions in hippocampal neurons
Neuron
Keppen-Lubinsky syndrome is caused by mutations in the inwardly rectifying K + channel encoded by KCNJ6
Am. J. Hum. Genet.
An immunocytochemical study on the distribution of two G-protein-gated inward rectifier potassium channels (GIRK2 and GIRK4) in the adult rat brain
Neuroscience
My close encounter with GABA(B) receptors
Biochem. Pharmacol.
Down's syndrome
Lancet
Inhibitory modulation of long-term potentiation via the 5-HT1A receptor in slices of the rat hippocampal dentate gyrus
Brain Res.
Increased App expression in a mouse model of Down's syndrome disrupts NGF transport and causes cholinergic neuron degeneration
Neuron
Thigmotaxis as an index of anxiety in mice. Influence of dopaminergic transmissions
Behav Brain Res.
Excitation/inhibition balance and learning are modified by Dyrk1a gene dosage
Neurobiol. Dis.
Engineering DYRK1A overdosage yields Down syndrome-characteristic cortical splicing aberrations
Neurobiol. Dis.
Molecular and cellular characterization of the Down syndrome critical region protein 2
Biochem. Biophys. Res. Commun.
Implicit and explicit memory: a functional dissociation in persons with Down syndrome
Neuropsychologia
Working memory in the aged Ts65Dn mouse, a model for Down syndrome
Behav. Brain Res.
Working memory and Down syndrome
J. Intellect. Disabil. Res.
Keppen-Lubinsky syndrome: expanding the phenotype
Am. J. Med. Genet. A
The "Down syndrome critical region" is sufficient in the mouse model to confer behavioral, neurophysiological, and synaptic phenotypes characteristic of Down syndrome
J. Neurosci.
Down syndrome cognitive phenotypes modeled in mice trisomic for all HSA 21 homologues
PLoS One
An anti-beta-amyloid vaccine for treating cognitive deficits in a mouse model of Down syndrome
PLoS One
Ts65Dn, a mouse model of Down syndrome, exhibits increased GABAB-induced potassium current
J. Neurophysiol.
Object recognition in rats and mice: a one-trial non-matching-to-sample learning task to study 'recognition memory'
Nat. Protoc.
Early pharmacotherapy restores neurogenesis and cognitive performance in the Ts65Dn mouse model for Down syndrome
J. Neurosci.
Localization of a retroviral element within the rd gene coding for the beta subunit of cGMP phosphodiesterase
Proc. Natl. Acad. Sci. U. S. A.
Specific targeting of the GABA-A receptor alpha5 subtype by a selective inverse agonist restores cognitive deficits in Down syndrome mice
J. Psychopharmacol.
Down syndrome: genes, model systems, and progress towards pharmacotherapies and clinical trials for cognitive deficits
Cytogenet Genome Res.
Trisomy of the G protein-coupled K + channel gene, Kcnj6, affects reward mechanisms, cognitive functions, and synaptic plasticity in mice
Proc. Natl. Acad. Sci. U. S. A.
Trisomy for synaptojanin1 in Down syndrome is functionally linked to the enlargement of early endosomes
Hum. Mol. Genet.
On the promise of pharmacotherapies targeted at cognitive and neurodegenerative components of Down syndrome
Dev. Neurosci.
Growth of children with Down's syndrome: birth to age 3 years
Pediatrics
The use of mouse models to understand and improve cognitive deficits in Down syndrome
Dis. Model. Mech.
Segmental trisomy of murine chromosome 16: a new model system for studying Down syndrome
Prog. Clin. Biol. Res.
Evaluation of the impact of abdominal obesity on glucose and lipid metabolism disorders in adults with Down syndrome
Res. Dev. Disabil.
Reversing excitatory GABA(A)R signaling restores synaptic plasticity and memory in a mouse model of Down syndrome
Nat. Med.
Down syndrome: the brain in trisomic mode
Nat. Rev. Neurosci.
DYRK1A (dual-specificity tyrosine-phosphorylated and -regulated kinase 1A): a gene with dosage effect during development and neurogenesis
ScientificWorldJournal
Behavioral characterization of a mouse model overexpressing DSCR1/RCAN1
PLoS One
Cited by (33)
Identifying the joint signature of brain atrophy and gene variant scores in Alzheimer's Disease
2024, Journal of Biomedical InformaticsOverexpression of miR-99a in hippocampus leads to impairment of reversal learning in mice
2022, Behavioural Brain ResearchCitation Excerpt :DS individuals present cognitive dysfunction characterized by deficits in problem-solving, reasoning, learning, memory, as well as deficiencies in conceptual, social, and practical skills across the life span [2,3]. Neurological abnormalities in DS included reduced brain mass, impaired hippocampal synaptic plasticity, altered dendritic spine morphology, and synaptic density [1,4–6]. Accumulating evidences suggest that the altered expression of genes encoded on human chromosome 21 (HSA21), more than 400 genes including 5 microRNA (miRNA) genes (miR-99a, let-7c, miR-125b-2, miR-155, and miR-802) [7,8], are involved in the phenotypic features in DS [9,10].
Down syndrome: a model for chromosome abnormalities
2022, Neurobiology of Brain Disorders: Biological Basis of Neurological and Psychiatric Disorders, Second EditionRestoring neuronal chloride homeostasis with anti-NKCC1 gene therapy rescues cognitive deficits in a mouse model of Down syndrome
2021, Molecular TherapyCitation Excerpt :Most of these studies have been performed in Ts65Dn mice, the most widely used trisomic model of DS carrying a freely segregating extra chromosome that contains ∼55% of the mouse ortholog genes to HSA21.40 Interestingly, much of the work on HSA21 genes has focused on a small set of triplicated orthologs in Ts65Dn mice: the amyloid-precursor protein (App),41–43 the G protein-coupled inwardly rectifying potassium channel 2 (Girk2/Kcnj6),37,44 the dual-specificity tyrosine-phosphorylation-regulated kinase 1A (Dyrk1A)45–47, the basic helix-loop-helix (bHLH) transcription factors Olig1/Olig2,48 and the phosphoinositide phosphatase synaptojanin 1 (Synj1).49 These studies have clearly shown that specific DS-related phenotypes critically arise from the triplication of different HSA21 dosage-sensitive genes in Ts65Dn mice.
Therapeutic potential of targeting G protein-gated inwardly rectifying potassium (GIRK) channels in the central nervous system
2021, Pharmacology and TherapeuticsCholinergic neurodegeneration in Alzheimer disease mouse models
2021, Handbook of Clinical Neurology