Jenna C. Carroll

Pathological α-Synuclein Transmission Initiates Parkinson-like Neurodegeneration in Nontransgenic Mice

Synthetic Parkinsons Parkinsons disease (PD) and related α-synucleinopathies are defined by the accumulation of α-synuclein (α-Syn)–containing intraneuronal inclusions—Lewy bodies (LBs) and Lewy neurites (LNs)—in association with the loss of dopaminergic neurons in the substantia nigra pars compacta (SNpc) and other brain regions. However, a cause-and-effect relationship between LB/LN formation and neurodegeneration remains unclear. Indeed, whether LB/LNs are toxic or represent a neuroprotective response has been contentious. Luk et al. (p. 949) injected α-Syn fibrils generated from recombinant mouse α-Syn protein into the dorsal striatum of wild-type mice and found that misfolded α-Syn caused the formation of PD-like LB/LNs and subsequent cell-to-cell transmission of pathologic α-Syn to anatomically interconnected regions, including the SNpc. Furthermore, the formation of LB/LNs and their accumulation in SNpc resulted in the progressive loss of these dopaminergic neurons, reduced dopamine innervations to the dorsal striatum, and culminated in motor deficits similar to PD. Thus, a synthetic misfolded wild-type protein (that is, α-Syn) was able to elicit and transmit disease pathology and neurodegeneration in healthy nontransgenic mice. Intracerebral inoculation of synthetic misfolded α-synuclein mimics Parkinson’s disease in wild-type mice. Parkinson’s disease is characterized by abundant α-synuclein (α-Syn) neuronal inclusions, known as Lewy bodies and Lewy neurites, and the massive loss of midbrain dopamine neurons. However, a cause-and-effect relationship between Lewy inclusion formation and neurodegeneration remains unclear. Here, we found that in wild-type nontransgenic mice, a single intrastriatal inoculation of synthetic α-Syn fibrils led to the cell-to-cell transmission of pathologic α-Syn and Parkinson’s-like Lewy pathology in anatomically interconnected regions. Lewy pathology accumulation resulted in progressive loss of dopamine neurons in the substantia nigra pars compacta, but not in the adjacent ventral tegmental area, and was accompanied by reduced dopamine levels culminating in motor deficits. This recapitulation of a neurodegenerative cascade thus establishes a mechanistic link between transmission of pathologic α-Syn and the cardinal features of Parkinson’s disease.

Protective actions of sex steroid hormones in Alzheimer’s disease

Risk for Alzheimers disease (AD) is associated with age-related loss of sex steroid hormones in both women and men. In post-menopausal women, the precipitous depletion of estrogens and progestogens is hypothesized to increase susceptibility to AD pathogenesis, a concept largely supported by epidemiological evidence but refuted by some clinical findings. Experimental evidence suggests that estrogens have numerous neuroprotective actions relevant to prevention of AD, in particular promotion of neuron viability and reduction of beta-amyloid accumulation, a critical factor in the initiation and progression of AD. Recent findings suggest neural responsiveness to estrogen can diminish with age, reducing neuroprotective actions of estrogen and, consequently, potentially limiting the utility of hormone therapies in aged women. In addition, estrogen neuroprotective actions are also modulated by progestogens. Specifically, continuous progestogen exposure is associated with inhibition of estrogen actions whereas cyclic delivery of progestogens may enhance neural benefits of estrogen. In recent years, emerging literature has begun to elucidate a parallel relationship of sex steroid hormones and AD risk in men. Normal age-related testosterone loss in men is associated with increased risk to several diseases including AD. Like estrogen, testosterone has been established as an endogenous neuroprotective factor that not only increases neuronal resilience against AD-related insults, but also reduces beta-amyloid accumulation. Androgen neuroprotective effects are mediated both directly by activation of androgen pathways and indirectly by aromatization to estradiol and initiation of protective estrogen signaling mechanisms. The successful use of hormone therapies in aging men and women to delay, prevent, and or treat AD will require additional research to optimize key parameters of hormone therapy and may benefit from the continuing development of selective estrogen and androgen receptor modulators.

Progesterone and Estrogen Regulate Alzheimer-Like Neuropathology in Female 3xTg-AD Mice

Estrogen depletion in postmenopausal women is a significant risk factor for the development of Alzheimers disease (AD), and estrogen-based hormone therapy may reduce this risk. However, the effects of progesterone both alone and in combination with estrogen on AD neuropathology remain unknown. In this study, we used the triple transgenic mouse model of AD (3xTg-AD) to investigate the individual and combined effects of estrogen and progesterone on β-amyloid (Aβ) accumulation, tau hyperphosphorylation, and hippocampal-dependent behavioral impairments. In gonadally intact female 3xTg-AD mice, AD-like neuropathology was apparent by 3 months of age and progressively increased through age 12 months, a time course that was paralleled by behavioral impairment. Ovariectomy-induced depletion of sex steroid hormones in adult female 3xTg-AD mice significantly increased Aβ accumulation and worsened memory performance. Treatment of ovariectomized 3xTg-AD mice with estrogen, but not progesterone, prevented these effects. When estrogen and progesterone were administered in combination, progesterone blocked the beneficial effect of estrogen on Aβ accumulation but not on behavioral performance. Interestingly, progesterone significantly reduced tau hyperphosphorylation when administered both alone and in combination with estrogen. These results demonstrate that estrogen and progesterone independently and interactively regulate AD-like neuropathology and suggest that an optimized hormone therapy may be useful in reducing the risk of AD in postmenopausal women.

The Microtubule-Stabilizing Agent, Epothilone D, Reduces Axonal Dysfunction, Neurotoxicity, Cognitive Deficits, and Alzheimer-Like Pathology in an Interventional Study with Aged Tau Transgenic Mice

Neurodegenerative tauopathies, such as Alzheimers disease (AD), are characterized by insoluble deposits of hyperphosphorylated tau protein within brain neurons. Increased phosphorylation and decreased solubility has been proposed to diminish normal tau stabilization of microtubules (MTs), thereby leading to neuronal dysfunction. Earlier studies have provided evidence that small molecule MT-stabilizing drugs that are used in the treatment of cancer may have utility in the treatment of tauopathies. However, it has not been established whether treatment with a small molecule MT-stabilizing compound will provide benefit in a transgenic model with pre-existing tau pathology, as would be seen in human patients with clinical symptoms. Accordingly, we describe here an interventional study of the brain-penetrant MT-stabilizing agent, epothilone D (EpoD), in aged PS19 mice with existing tau pathology and related behavioral deficits. EpoD treatment reduced axonal dystrophy and increased axonal MT density in the aged PS19 mice, which led to improved fast axonal transport and cognitive performance. Moreover, the EpoD-treated PS19 mice had less forebrain tau pathology and increased hippocampal neuronal integrity, with no dose-limiting side effects. These data reveal that brain-penetrant MT-stabilizing drugs hold promise for the treatment of AD and related tauopathies, and that EpoD could be a candidate for clinical testing.

Epothilone D Improves Microtubule Density, Axonal Integrity and Cognition in a Transgenic Mouse Model of Tauopathy

Neurons in the brains of those with Alzheimers disease (AD) and many frontotemporal dementias (FTDs) contain neurofibrillary tangles comprised of hyperphosphorylated tau protein. Tau normally stabilizes microtubules (MTs), and tau misfolding could lead to a loss of this function with consequent MT destabilization and neuronal dysfunction. Accordingly, a possible therapeutic strategy for AD and related “tauopathies” is treatment with a MT-stabilizing anti-cancer drug such as paclitaxel. However, paclitaxel and related taxanes have poor blood–brain barrier permeability and thus are unsuitable for diseases of the brain. We demonstrate here that the MT-stabilizing agent, epothilone D (EpoD), is brain-penetrant and we subsequently evaluated whether EpoD can compensate for tau loss-of-function in PS19 tau transgenic mice that develop forebrain tau inclusions, axonal degeneration and MT deficits. Treatment of 3-month-old male PS19 mice with low doses of EpoD once weekly for a 3 month period significantly improved CNS MT density and axonal integrity without inducing notable side-effects. Moreover, EpoD treatment reduced cognitive deficits that were observed in the PS19 mice. These results suggest that certain brain-penetrant MT-stabilizing agents might provide a viable therapeutic strategy for the treatment of AD and FTDs.

Androgens Regulate the Development of Neuropathology in a Triple Transgenic Mouse Model of Alzheimer's Disease

Normal age-related testosterone depletion in men is a recently identified risk factor for Alzheimers disease (AD), but how androgen loss affects the development of AD is unclear. To investigate the relationship between androgen depletion and AD, we compared how androgen status affects the progression of neuropathology in the triple transgenic mouse model of AD (3xTg-AD). Adult male 3xTg-AD mice were sham gonadectomized (GDX) or GDX to deplete endogenous androgens and then exposed for 4 months to either the androgen dihydrotestosterone (DHT) or to placebo. In comparison to gonadally intact 3xTg-AD mice, GDX mice exhibited robust increases in the accumulation of β-amyloid (Aβ), the protein implicated as the primary causal factor in AD pathogenesis, in both hippocampus and amygdala. In parallel to elevated levels of Aβ, GDX mice exhibited significantly impaired spontaneous alternation behavior, indicating deficits in hippocampal function. Importantly, DHT treatment of GDX 3xTg-AD mice attenuated both Aβ accumulation and behavioral deficits. These data demonstrate that androgen depletion accelerates the development of AD-like neuropathology, suggesting that a similar mechanism may underlie the increased risk for AD in men with low testosterone. In addition, our finding that DHT protects against acceleration of AD-like neuropathology predicts that androgen-based hormone therapy may be a useful strategy for the prevention and treatment of AD in aging men.

Chronic Stress Exacerbates Tau Pathology, Neurodegeneration, and Cognitive Performance through a Corticotropin-Releasing Factor Receptor-Dependent Mechanism in a Transgenic Mouse Model of Tauopathy

Because overactivation of the hypothalamic–pituitary–adrenal (HPA) axis occurs in Alzheimers disease (AD), dysregulation of stress neuromediators may play a mechanistic role in the pathophysiology of AD. However, the effects of stress on tau phosphorylation are poorly understood, and the relationship between corticosterone and corticotropin-releasing factor (CRF) on both β-amyloid (Aβ) and tau pathology remain unclear. Therefore, we first established a model of chronic stress, which exacerbates Aβ accumulation in Tg2576 mice and then extended this stress paradigm to a tau transgenic mouse model with the P301S mutation (PS19) that displays tau hyperphosphorylation, insoluble tau inclusions and neurodegeneration. We show for the first time that both Tg2576 and PS19 mice demonstrate a heightened HPA stress profile in the unstressed state. In Tg2576 mice, 1 month of restraint/isolation (RI) stress increased Aβ levels, suppressed microglial activation, and worsened spatial and fear memory compared with nonstressed mice. In PS19 mice, RI stress promoted tau hyperphosphorylation, insoluble tau aggregation, neurodegeneration, and fear–memory impairments. These effects were not mimicked by chronic corticosterone administration but were prevented by pre-stress administration of a CRF receptor type 1 (CRF1) antagonist. The role for a CRF1-dependent mechanism was further supported by the finding that mice overexpressing CRF had increased hyperphosphorylated tau compared with wild-type littermates. Together, these results implicate HPA dysregulation in AD neuropathogenesis and suggest that prolonged stress may increase Aβ and tau hyperphosphorylation. These studies also implicate CRF in AD pathophysiology and suggest that pharmacological manipulation of this neuropeptide may be a potential therapeutic strategy for AD.

Sex differences in β-amyloid accumulation in 3xTg-AD mice: Role of neonatal sex steroid hormone exposure

The risk of Alzheimers disease (AD) is higher in women than in men, a sex difference that likely results from the effects of sex steroid hormones. To investigate this relationship, we first compared progression of β-amyloid (Aβ) pathology in male and female triple transgenic (3xTg-AD) mice. We found that female 3xTg-AD mice exhibit significantly greater Aβ burden and larger behavioral deficits than age-matched males. Next, we evaluated how the organizational effects of sex steroid hormones during postnatal development may affect adult vulnerability to Aβ pathology. We observed that male 3xTg-AD mice demasculinized during early development exhibit significantly increased Aβ accumulation in adulthood. In contrast, female mice defeminized during early development exhibit a more male-like pattern of Aβ pathology in adulthood. Taken together, these results demonstrate significant sex differences in pathology in 3xTg-AD mice and suggest that these differences may be mediated by organizational actions of sex steroid hormones during development.

Selectively Silencing GSK-3 Isoforms Reduces Plaques and Tangles in Mouse Models of Alzheimer's Disease

Glycogen synthase kinase-3 (GSK-3) is linked to the pathogenesis of Alzheimers disease (AD), senile plaques (SPs), and neurofibrillary tangles (NFTs), but the specific contributions of each of the GSK-3 α and β isoforms to mechanisms of AD have not been clarified. In this study, we sought to elucidate the role of each GSK-3α and GSK-3β using novel viral and genetic approaches. First, we developed recombinant adeno-associated virus 2/1 short hairpin RNA constructs which specifically reduced expression and activity of GSK-3α or GSK-3β. These constructs were injected intraventricularly in newborn AD transgenic (tg) mouse models of SPs (PDAPP+/−), both SPs and NFTs (PDAPP+/−;PS19+/−), or wild-type controls. We found that knockdown (KD) of GSK-3α, but not GSK-3β, reduced SP formation in PDAPP+/− and PS19+/−;PDAPP+/− tg mice. Moreover, both GSK-3α and GSK-3β KD reduced tau phosphorylation and tau misfolding in PS19+/−;PDAPP+/− mice. Next, we generated triple tg mice using the CaMKIIα-Cre (α-calcium/calmodulin-dependent protein kinase II-Cre) system to KD GSK-3α in PDAPP+/− mice for further study of the effects of GSK-3α reduction on SP formation. GSK-3α KD showed a significant effect on reducing SPs and ameliorating memory deficits in PDAPP+/− mice. Together, the data from both approaches suggest that GSK-3α contributes to both SP and NFT pathogenesis while GSK-3β only modulates NFT formation, suggesting common but also different targets for both isoforms. These findings highlight the potential importance of GSK-3α as a possible therapeutic target for ameliorating behavioral impairments linked to AD SPs and NFTs.

Testosterone regulation of Alzheimer-like neuropathology in male 3xTg-AD mice involves both estrogen and androgen pathways.

Normal, age-related depletion of the androgen testosterone is a risk factor for Alzheimers disease (AD) in men. Previously, we reported that experimental androgen depletion significantly accelerates development of AD-like neuropathology in the 3xTg-AD triple-transgenic mouse model of AD, an effect prevented by androgen treatment. Because testosterone is metabolized in brain into both the androgen dihydrotestosterone (DHT) and the estrogen 17β-estradiol (E2), testosterone can mediate its effects through androgen and or estrogen pathways. To define the role of androgen and estrogen pathways in regulation of AD-like neuropathology, we compared the effects of testosterone (T) and its metabolites DHT and E2 in male 3xTg-AD mice depleted of endogenous sex steroid hormones by gonadectomy (GDX). Male 3xTg-AD mice were sham GDX or GDX, immediately treated with vehicle, T, DHT, or E2, and 4 months later evaluated for two indices of AD-like neuropathology, β-amyloid (Aβ) accumulation and tau hyperphosphorylation. In comparison to sham GDX mice, we observed a significant increase in Aβ accumulation in GDX mice in subiculum, hippocampus, and amygdala. Treatment of GDX mice with T prevented the increased Aβ accumulation in all three brain regions. DHT treatment yielded similar results, significantly reducing Aβ accumulation across brain regions. Interestingly, E2 prevented Aβ accumulation in hippocampus but exerted only partial effects in subiculum and amygdala. Levels of tau hyperphosphorylation in sham GDX male 3xTg-AD mice were modest and only slightly increased by GDX. Treatment of GDX mice with T or E2 but not DHT reduced tau hyperphosphorylation to levels lower than observed in sham animals. These data suggest that testosterone regulates Aβ pathology through androgen and estrogen pathways and reduces tau pathology largely through estrogen pathways. These findings further define hormone pathways involved in regulation of AD-related pathology, information that is important for understanding disease etiology and developing pathway-specific hormone interventions.