Gerhard Leinenga

Scanning ultrasound removes amyloid-β and restores memory in an Alzheimer’s disease mouse model

Repeated scanning ultrasound in the absence of a therapeutic agent removes amyloid-β and restores memory in an Alzheimer’s disease mouse model. Can ultrasound restore memory? Transgenic mice with increased amyloid-β (Aβ) production show several aspects of Alzheimer’s disease, including Aβ deposition and memory impairment. By repeatedly treating these Aβ-forming mice with scanning ultrasound, Leinenga and Götz now demonstrate that Aβ is removed and memory is restored as revealed by improvement in three memory tasks. These improvements were achieved without the use of any therapeutic agent, and the scanning ultrasound treatment did not induce any apparent damage to the mouse brain. The authors then showed that scanning ultrasound activated resident microglial cells that took up Aβ into their lysosomes. These findings suggest that repeated scanning ultrasound may be a noninvasive method with potential for treating Alzheimer’s disease. Amyloid-β (Aβ) peptide has been implicated in the pathogenesis of Alzheimer’s disease (AD). We present a nonpharmacological approach for removing Aβ and restoring memory function in a mouse model of AD in which Aβ is deposited in the brain. We used repeated scanning ultrasound (SUS) treatments of the mouse brain to remove Aβ, without the need for any additional therapeutic agent such as anti-Aβ antibody. Spinning disk confocal microscopy and high-resolution three-dimensional reconstruction revealed extensive internalization of Aβ into the lysosomes of activated microglia in mouse brains subjected to SUS, with no concomitant increase observed in the number of microglia. Plaque burden was reduced in SUS-treated AD mice compared to sham-treated animals, and cleared plaques were observed in 75% of SUS-treated mice. Treated AD mice also displayed improved performance on three memory tasks: the Y-maze, the novel object recognition test, and the active place avoidance task. Our findings suggest that repeated SUS is useful for removing Aβ in the mouse brain without causing overt damage, and should be explored further as a noninvasive method with therapeutic potential in AD.

What Renders TAU Toxic

TAU is a microtubule-associated protein that under pathological conditions such as Alzheimer’s disease (AD) forms insoluble, filamentous aggregates. When 20 years after TAU’s discovery the first TAU transgenic mouse models were established, one declared goal that was achieved was the modeling of authentic TAU aggregate formation in the form of neurofibrillary tangles. However, as we review here, it has become increasingly clear that TAU causes damage much before these filamentous aggregates develop. In fact, because TAU is a scaffolding protein, increased levels and an altered subcellular localization (due to an increased insolubility and impaired clearance) result in the interaction of TAU with cellular proteins with which it would otherwise either not interact or do so to a lesser degree, thereby impairing their physiological functions. We specifically discuss the non-axonal localization of TAU, the role phosphorylation has in TAU toxicity and how TAU impairs mitochondrial functions. A major emphasis is on what we have learned from the four available TAU knock-out models in mice, and the knock-out of the TAU/MAP2 homolog PTL-1 in worms. It has been proposed that in human pathological conditions such as AD, a rare toxic TAU species exists which needs to be specifically removed to abrogate TAU’s toxicity and restore neuronal functions. However, what is toxic in one context may not be in another, and simply reducing, but not fully abolishing TAU levels may be sufficient to abrogate TAU toxicity.

Ultrasound treatment of neurological diseases — current and emerging applications

Like cardiovascular disease and cancer, neurological disorders present an increasing challenge for an ageing population. Whereas nonpharmacological procedures are routine for eliminating cancer tissue or opening a blocked artery, the focus in neurological disease remains on pharmacological interventions. Setbacks in clinical trials and the obstacle of access to the brain for drug delivery and surgery have highlighted the potential for therapeutic use of ultrasound in neurological diseases, and the technology has proved useful for inducing focused lesions, clearing protein aggregates, facilitating drug uptake, and modulating neuronal function. In this Review, we discuss milestones in the development of therapeutic ultrasound, from the first steps in the 1950s to recent improvements in technology. We provide an overview of the principles of diagnostic and therapeutic ultrasound, for surgery and transient opening of the blood–brain barrier, and its application in clinical trials of stroke, Parkinson disease and chronic pain. We discuss the promising outcomes of safety and feasibility studies in preclinical models, including rodents, pigs and macaques, and efficacy studies in models of Alzheimer disease. We also consider the challenges faced on the road to clinical translation.

Combined effects of scanning ultrasound and a tau-specific single chain antibody in a tau transgenic mouse model

One of the greatest challenges for the treatment of neurodegenerative disease is crossing the blood-brain barrier. Nisbet et al. demonstrate that non-invasive scanning ultrasound increases the delivery of tau-specific single-chain antibody fragments across the blood-brain barrier and into neurons of tau transgenic mice, reducing anxiety-like behaviour and tau pathology.

Safety and Efficacy of Scanning Ultrasound Treatment of Aged APP23 Mice

Deposition of amyloid-β (Aβ) peptide leads to amyloid plaques that together with tau deposits characterize the brains of patients with Alzheimers disease (AD). In modeling this pathology, transgenic animals such as the APP23 strain, that expresses a mutant form of the amyloid precursor protein found in familial cases of AD, have been instrumental. In previous studies, we have shown that repeated treatments with ultrasound in a scanning mode (termed scanning ultrasound or SUS) were effective in removing Aβ and restoring memory functions, without the need for a therapeutic agent such as an Aβ antibody. Considering that age is the most important risk factor for AD, we extended this study in which the mice were only 12 months old at the time of treatment by assessing a cohort of 2 year-old mice. Interestingly, at this age, APP23 mice are characterized by cerebral amyloid angiopathy (CAA) and the presence of occasional microbleeds. We found that SUS in aged mice that have been exposed to four SUS sessions that were spread out over 8 weeks and analyzed 4 weeks later did not show evidence of increased CAA or microbleeds. Furthermore, amyloid was reduced as assessed by methoxy-XO4 fluorescence. In addition, plaque-associated microglia were more numerous in SUS treated mice. Together this adds to the notion that SUS may be a treatment modality for human neurodegenerative diseases.

Scanning ultrasound (SUS) causes no changes to neuronal excitability and prevents age-related reductions in hippocampal CA1 dendritic structure in wild-type mice

Scanning ultrasound (SUS) is a noninvasive approach that has recently been shown to ameliorate histopathological changes and restore memory functions in an Alzheimers disease mouse model. Although no overt neuronal damage was reported, the short- and long-term effects of SUS on neuronal excitability and dendritic tree morphology had not been investigated. To address this, we performed patch-clamp recordings from hippocampal CA1 pyramidal neurons in wild-type mice 2 and 24 hours after a single SUS treatment, and one week and 3 months after six weekly SUS treatments, including sham treatments as controls. In both treatment regimes, no changes in CA1 neuronal excitability were observed in SUS-treated neurons when compared to sham-treated neurons at any time-point. For the multiple treatment groups, we also determined the dendritic morphology and spine densities of the neurons from which we had recorded. The apical trees of sham-treated neurons were reduced at the 3 month time-point when compared to one week; however, surprisingly, no longitudinal change was detected in the apical dendritic trees of SUS-treated neurons. In contrast, the length and complexity of the basal dendritic trees were not affected by SUS treatment at either time-point. The apical dendritic spine densities were reduced, independent of the treatment group, at 3 months compared to one week. Collectively, these data suggest that ultrasound can be employed to prevent an age-associated loss of dendritic structure without impairing neuronal excitability.

Establishing sheep as an experimental species to validate ultrasound-mediated blood-brain barrier opening for potential therapeutic interventions

Rationale: Treating diseases of the brain such as Alzheimers disease (AD) is challenging as the blood-brain barrier (BBB) effectively restricts access of a large number of potentially useful drugs. A potential solution to this problem is presented by therapeutic ultrasound, a novel treatment modality that can achieve transient BBB opening in species including rodents, facilitated by biologically inert microbubbles that are routinely used in a clinical setting for contrast enhancement. However, in translating rodent studies to the human brain, the presence of a thick cancellous skull that both absorbs and distorts ultrasound presents a challenge. A larger animal model that is more similar to humans is therefore required in order to establish a suitable protocol and to test devices. Here we investigated whether sheep provide such a model. Methods: In a stepwise manner, we used a total of 12 sheep to establish a sonication protocol using a spherically focused transducer. This was assisted by ex vivo simulations based on CT scans to establish suitable sonication parameters. BBB opening was assessed by Evans blue staining and a range of histological tests. Results: Here we demonstrate noninvasive microbubble-mediated BBB opening through the intact sheep skull. Our non-recovery protocol allowed for BBB opening at the base of the brain, and in areas relevant for AD, including the cortex and hippocampus. Linear time-shift invariant analysis and finite element analysis simulations were used to optimize the position of the transducer and to predict the acoustic pressure and location of the focus. Conclusion: Our study establishes sheep as a novel animal model for ultrasound-mediated BBB opening and highlights opportunities and challenges in using this model. Moreover, as sheep develop an AD-like pathology with aging, they represent a large animal model that could potentially complement the use of non-human primates.

Ultrasound as a treatment modality for neurological diseases

ith an ageing population, neurological disorders present an increasing challenge to W our health care systems. Although antibodies are increasingly being explored for therapeutic intervention, the inefficiency of their uptake by the brain means that the estimated cost of a vaccine to treat neurodegenerative disorders such as Alzheimer disease (AD) will exceed US

Modeling ultrasound propagation through material of increasing geometrical complexity

25 000 per patient per year. Not only is this expected to challenge the health care systems of many countries, it also raises ethical issues associated with making these vaccines available to every patient.

ULTRASOUND AS A NOVEL TREATMENT MODALITY FOR THE MICROTUBULE-ASSOCIATED PROTEIN TAU THAT FORMS INTRANEURONAL PROTEIN AGGREGATES IN ALZHEIMER'S DISEASE

HighlightsUltrasound is increasingly being recognized as therapeutic tool for brain diseases.The acoustic properties of a set of simple bone‐modeling samples were analyzed.Wiener deconvolution predicts the Ultrasound Acoustic Response and attenuation.Finite Element Analysis observes scattering and refraction of wave propagation.Finite Element Analysis reveals differences depending on step size of models. ABSTRACT Ultrasound is increasingly being recognized as a neuromodulatory and therapeutic tool, inducing a broad range of bio‐effects in the tissue of experimental animals and humans. To achieve these effects in a predictable manner in the human brain, the thick cancellous skull presents a problem, causing attenuation. In order to overcome this challenge, as a first step, the acoustic properties of a set of simple bone‐modeling resin samples that displayed an increasing geometrical complexity (increasing step sizes) were analyzed. Using two Non‐Destructive Testing (NDT) transducers, we found that Wiener deconvolution predicted the Ultrasound Acoustic Response (UAR) and attenuation caused by the samples. However, whereas the UAR of samples with step sizes larger than the wavelength could be accurately estimated, the prediction was not accurate when the sample had a smaller step size. Furthermore, a Finite Element Analysis (FEA) performed in ANSYS determined that the scattering and refraction of sound waves was significantly higher in complex samples with smaller step sizes compared to simple samples with a larger step size. Together, this reveals an interaction of frequency and geometrical complexity in predicting the UAR and attenuation. These findings could in future be applied to poro‐visco‐elastic materials that better model the human skull.