Age-related mitochondrial alterations in brain and skeletal muscle of the YAC128 model of Huntington disease.

Mitochondrial dysfunction and bioenergetics failure are common pathological hallmarks in Huntington's disease (HD) and aging. In the present study, we used the YAC128 murine model of HD to examine the effects of mutant huntingtin on mitochondrial parameters related to aging in brain and skeletal muscle. We have conducted a cross-sectional natural history study of mitochondrial DNA changes in the YAC128 mouse. Here, we first show that the mitochondrial volume fraction appears to increase in the axons and dendrite regions adjacent to the striatal neuron cell bodies in old mice. Mitochondrial DNA copy number (mtDNAcn) was used as a proxy measure for mitochondrial biogenesis and function. We observed that the mtDNAcn changes significantly with age and genotype in a tissue-specific manner. We found a positive correlation between aging and the mtDNAcn in striatum and skeletal muscle but not in cortex. Notably, the YAC128 mice had lower mtDNAcn in cortex and skeletal muscle. We further show that mtDNA deletions are present in striatal and skeletal muscle tissue in both young and aged YAC128 and WT mice. Tracking gene expression levels cross-sectionally in mice allowed us to identify contributions of age and genotype to transcriptional variance in mitochondria-related genes. These findings provide insights into the role of mitochondrial dynamics in HD pathogenesis in both brain and skeletal muscle, and suggest that mtDNAcn in skeletal muscle tissue may be a potential biomarker that should be investigated further in human HD.

Neuronal Cell Cycle Re-Entry Enhances Neuropathological Features in AppNLF Knock-In Mice.

BACKGROUND: Aberrant cell cycle re-entry is a well-documented process occurring early in Alzheimer's disease (AD). This is an early feature of the disease and may contribute to disease pathogenesis. OBJECTIVE: To assess the effect of forced neuronal cell cycle re-entry in mice expressing humanized Aß, we crossed our neuronal cell cycle re-entry mouse model with AppNLF knock-in (KI) mice. METHODS: Our neuronal cell cycle re-entry (NCCR) mouse model is bitransgenic mice heterozygous for both Camk2a-tTA and TRE-SV40T. The NCCR mice were crossed with AppNLF KI mice to generate NCCR-AppNLF animals. Using this tet-off system, we triggered NCCR in our animals via neuronal expression of SV40T starting at 1 month of age. The animals were examined at the following time points: 9, 12, and 18 months of age. Various neuropathological features in our mice were evaluated by image analysis and stereology on brain sections stained using either immunofluorescence or immunohistochemistry. RESULTS: We show that neuronal cell cycle re-entry in humanized Aß plaque producing AppNLF KI mice results in the development of additional AD-related pathologies, namely, pathological tau, neuroinflammation, brain leukocyte infiltration, DNA damage response, and neurodegeneration. CONCLUSION: Our findings show that neuronal cell cycle re-entry enhances AD-related neuropathological features in AppNLF mice and highlight our unique AD mouse model for studying the pathogenic role of aberrant cell cycle re-entry in AD.

Gliosis Precedes Amyloid-ß Deposition and Pathological Tau Accumulation in the Neuronal Cell Cycle Re-Entry Mouse Model of Alzheimer's Disease.

BACKGROUND: The presence of cell cycle markers in postmortem Alzheimer's disease (AD) brains suggest a potential role of cell cycle activation in AD. It was shown that cell cycle activation in postmitotic neurons in mice produces Aß and tau pathologies from endogenous mouse proteins in the absence of AßPP or tau mutations. OBJECTIVE: In this study, we examined the microglial and astrocytic responses in these mice since neuroinflammation is another key pathological feature in AD. METHODS: Our neuronal cell cycle re-entry (NCCR) mouse model are bitransgenic mice heterozygous for both Camk2a-tTA and TRE-SV40T. Using this tet-off system, we triggered NCCR in our animals via neuronal expression of SV40T starting at 1 month of age. TRE-SV40T Tg mice were used as SV40T transgene controls. The animals were examined at following time points: 2, 3, 4, 6, and 12 months of age. The microglia and astrocyte responses in our mice were determined by image analysis and stereology on brain sections immunofluorescently labeled using the following antibodies: Iba1, CD45, CD68, MHCII, and GFAP. Cellular senescent marker p16 was also used in this study. RESULTS: Our NCCR mice demonstrate early and persistent activation of microglia and astrocytes. Additionally, proinflammatory and senescent microglia phenotype and brain leukocyte infiltration is present at 12 months of age. CONCLUSION: In the absence of FAD gene mutations, our NCCR mice simultaneously display many of the pathological changes associated with AD, such as ectopic neuronal cell cycle re-entry, Aß and tau pathologies, neuroinflammation, and neurodegeneration. These animals represent a promising alternative AD mouse model.

p35 Hemizygous Deletion in 5xFAD Mice Increases Aß Plaque Load in Males but Not in Females.

Increased amyloid beta (Aß) deposition is implicated in early stages of Alzheimer's disease (AD). Although aberrant Cdk5 activity mediated by Cdk5/p25 is suggested to promote Aß plaque deposition, the effects of Cdk5 inhibition on Aß plaque loads in AD mouse models have been equivocal, possibly due to the fact that Cdk5 can be activated by p35 or p39 and their cleaved products. Here we evaluated the effect of p35 knockdown on Aß plaque formation by constitutively knocking out a single p35 allele in 5xFAD mice. Surprisingly, our results show that the simultaneous reduction in the levels of p35 and p25 increases cortical Aß plaque loads in male 5xFAD mice, but not in females. This change is associated with male specific decrease in pSer9 GSK3ß levels. Furthermore, p35 hemizygous deletion has sexually dimorphic effects on Iba1 and GFAP protein levels. Our findings demonstrate sex differences in the effects of p35 reduction on biochemical pathways relevant to the modulation of Aß plaque deposition and confirm the importance of examining both sexes in preclinical AD research.

p35 hemizygosity activates Akt but does not improve motor function in the YAC128 mouse model of Huntington's disease.

Huntington's disease (HD) is a hereditary neurodegenerative disorder resulting from N-terminal polyglutamine expansion in the huntingtin protein. A relatively selective and early loss of medium spiny neurons in the striatum is a hallmark of HD neuropathology. Although the exact mechanism of mutant huntingtin-mediated neurodegeneration is unclear, recent evidence suggests that NMDA-receptor-mediated excitotoxicity is involved. Our previously published findings show that decreasing levels of the cdk5 activators, p35 and p25, reduces NMDA receptor-mediated excitotoxicity in striatal neurons in vivo. In this study we directly examined the effect of reducing levels of p35 and p25 in the context of mutant huntingtin toxicity, using the B6 YAC128 mouse model of HD. Our findings demonstrate that deletion of a single allele of p35 in the B6 YAC128 mice results in an upregulation of Akt activity, and increases phosphorylation of mutant huntingtin at Ser421. Longitudinal behavioral analysis showed that this 50% reduction in p35 and p25 levels did not improve accelerating Rotarod performance in these YAC128 mice. However, a complete deletion of p35 normalized the accelerating Rotarod performance relative to their non-transgenic littermates at four months of age.

Mechanisms of Muscle Denervation in Aging: Insights from a Mouse Model of Amyotrophic Lateral Sclerosis.

Muscle denervation at the neuromuscular junction (NMJ) is thought to be a contributing factor in age-related muscle weakness. Therefore, understanding the mechanisms that modulate NMJ innervation is a key to developing therapies to combat age-related muscle weakness affecting the elderly. Two mouse models, one lacking the Cu/Zn superoxide dismutase (SOD1) gene and another harboring the transgenic mutant human SOD1 gene, display progressive changes at the NMJ, including muscle endplate fragmentation, nerve terminal sprouting, and denervation. These changes at the NMJ share many of the common features observed in the NMJs of aged mice. In this review, research findings demonstrating the effects of PGC-1α, IGF-1, GDNF, MyoD, myogenin, and miR-206 on NMJ innervation patterns in the G93A SOD1 mice will be highlighted in the context of age-related muscle denervation.

Postnatal muscle modification by myogenic factors modulates neuropathology and survival in an ALS mouse model.

MyoD and myogenin are myogenic transcription factors preferentially expressed in adult fast and slow muscles, respectively. Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder in which motor neuron loss is accompanied by muscle denervation and paralysis. Studies suggest that muscle phenotype may influence ALS disease progression. Here we demonstrate that myogenin gene transfer into muscle supports spinal cord motor neuron survival and muscle endplate innervation in the G93A SOD1 fALS mice. On the other hand, MyoD gene transfer decreases survival and enhances motor neuron degeneration and muscle denervation. Although an increase in motor neuron count is associated with increased succinic dehydrogenase staining in the muscle, muscle overexpression of PGC-1α does not improve survival or motor function. Our study suggests that postnatal muscle modification influences disease progression and demonstrates that the muscle expression of myogenic and metabolic regulators differentially impacts neuropathology associated with disease progression in the G93A SOD1 fALS mouse model.

Decreasing Levels of the cdk5 Activators, p25 and p35, Reduces Excitotoxicity in Striatal Neurons.

Huntington's disease (HD) is a progressive neurodegenerative disorder caused by an expanded CAG trinucleotide repeat sequence in the huntingtin gene. The resulting poly-glutamine expansion in the huntingtin protein imparts a novel toxic gain of function causing selective loss of medium spiny neurons (MSNs) in the striatum. Although the exact mechanism of cell death is unclear, recent evidence suggests involvement of NMDA-receptor mediated excitotoxicity and aberrant cyclin dependent kinase 5 (cdk5) activity in striatal cells undergoing neurodegeneration. In this study we directly tested the effect of reduced levels of p25 and p35, two proteins required for cdk5 activation, on striatal neurodegeneration using mice with targeted deletion of p35. Quinolinic acid (QA) injected into the striatum of mice causes NMDA-receptor mediated cell death, and these QA-induced striatal lesions were examined in p35 hemizygous null (p35+/-) and wildtype (WT) mice. Striatal QA lesion volumes were 30% smaller in p35+/- mice than in WT mice. Furthermore, primary neuronal cultures of MSNs from P0 p35+/- pups displayed 33% less apoptotic neurons following NMDA treatment than those from WT pups. Examination of YAC128 mouse model of HD showed elevated p25 levels in striatum following intrastriatal QA injection. Our findings provide direct evidence for p25 and p35 involvement in excitotoxic neurodegeneration of MSNs and suggest a role for the cdk5 pathway in HD striatal neurodegeneration.

Presymptomatic biochemical changes in hindlimb muscle of G93A human Cu/Zn superoxide dismutase 1 transgenic mouse model of amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is primarily a motor neuron disorder. Intriguingly, early muscle denervation preceding motor neuron loss is observed in mouse models of ALS. Enhanced muscle vulnerability to denervation process has been suggested by accelerated muscle deterioration following peripheral nerve injury in an ALS mouse model. Here we provide evidence of biochemical changes in the hindlimb muscle of young, presymptomatic G93A hSOD1 transgenic mice. In this report, we demonstrate that cdk5 activity is reduced in hindlimb muscle of 27-day-old G93A hSOD1 transgenic mice. In vitro analysis revealed mutant hSOD1-mediated suppression of cdk5 activity. Furthermore, the decrease in muscle cdk5 activity was accompanied by a significant reduction in MyoD and cyclin D1 levels. These early muscle changes raise the possibility that the progressive deterioration of muscle function is potentiated by altered muscle biochemistry in these mice at a very young, presymptomatic age.

Conditional neuronal simian virus 40 T antigen expression induces Alzheimer-like tau and amyloid pathology in mice.

A large body of evidence has shown the activation of a cohort of cell cycle regulators and the duplication of DNA in degenerating neurons of Alzheimer's disease (AD) brain. Activation of these regulators and duplication of chromosomes precede neurodegeneration and formation of neurofibrillary tangles (NFTs), one of the diagnostic lesions of AD. These findings, in combination with evidence for cell cycle regulation of amyloid precursor protein and tau, has led to the hypothesis that reentry into the cell cycle underlies AD pathogenesis. To test this hypothesis directly, we have created transgenic mice with forced cell cycle activation in postmitotic neurons via conditional expression of the simian virus 40 large T antigen (TAg) oncogene. We show that TAg mice recapitulate the cell cycle changes seen in AD and display a neurodegenerative phenotype accompanied by tau pathology and NFT-like profiles. Moreover, plaque-like amyloid deposits, similar to those seen in AD, are also observed in the brains of TAg mice. These data provide support for an essential role of ectopic cell cycle activation in the generation of the characteristic pathological hallmarks of AD. Furthermore, our TAg mice are the first model to develop NFTs and amyloid pathology simultaneously and in the absence of any human transgenes. These mice will be useful for further defining the nongenetic mechanisms in AD pathogenesis and for the development of cell cycle-based therapies for AD.

Seasonal-like plasticity of spontaneous firing rate in a songbird pre-motor nucleus.

Many animals exhibit seasonal changes in behavior and its underlying neural substrates. In seasonally breeding songbirds, the brain nuclei that control song learning and production undergo substantial structural changes at the onset of each breeding season, in association with changes in song behavior. These changes are largely mediated by photoperiod-dependent changes in circulating concentrations of gonadal steroid hormones. Little is known, however, about whether changes in the electrophysiological activity of neurons accompany the dramatic morphological changes in the song nuclei. Here we induced seasonal-like changes in the song systems of adult white-crowned sparrows and used extracellular recording in acute brain slices from those individuals to study physiological properties of neurons in the robust nucleus of the arcopallium (RA), a pre-motor nucleus necessary for song production. We report that: RA neurons from birds in breeding condition show a more than twofold increase in spontaneous firing rate compared to those from nonbreeding condition; this change appears to require both androgenic and estrogenic actions; and this change is intrinsic to the RA neurons. Thus, neurons in the song circuit exhibit both morphological and physiological adult seasonal plasticity.

Influence of restraint and acute isolation on the selectivity of the adult zebra finch zenk gene response to acoustic stimuli.

Zebra finches respond to certain auditory stimuli with the activation of the immediate early gene zenk. It has been shown that the amount of sound-mediated zenk gene expression varies in the zebra finch caudomedial neostriatum (NCM), apparently correlated with stimulus type (conspecific>heterospecific>noise>tones) and familiarity. Here we tested the impact of two additional factors-song-specific acoustical properties and testing conditions-on the specificity of the sound-mediated zenk response, as assessed by in situ hybridization. A variant of a normal conspecific song was first produced by randomizing the spectral content while retaining the amplitude envelope ('song-enveloped noise'). This stimulus and related controls were presented to birds which were either free in cages or restrained in a stereotaxic instrument, after isolation either overnight or for only 1 h prior to testing. We confirmed prior results that unrestrained birds show a greater zenk response to normal conspecific song than to other acoustic stimuli. However, under restraint, birds showed little or no selectivity for conspecific song compared to matched stimuli lacking a song organization. Thus the specificity of the zenk response to song is not determined simply by the acoustic structure and familiarity of the stimulus. We conclude that the intrinsic selectivity of sensory responses measured in the CNS may be influenced by factors associated with attention, arousal or vigilance, and may be significantly altered by experimental conditions that involve physical restraint.