Focused ultrasound-induced blood-brain barrier opening improves adult hippocampal neurogenesis and cognitive function in a cholinergic degeneration dementia rat model

BACKGROUND: The persistence of adult hippocampal neurogenesis (AHN) is sharply decreased in Alzheimer's disease (AD). The neuropathologies of AD include the presence of amyloid-β deposition in plaques, tau hyperphosphorylation in neurofibrillary tangles, and cholinergic system degeneration. The focused ultrasound (FUS)-mediated blood-brain barrier opening modulates tau hyperphosphorylation, the accumulation of amyloid-β proteins, and increases in AHN. However, it remains unclear whether FUS can modulate AHN in cholinergic-deficient conditions. In this study, we investigated the effect of FUS on AHN in a cholinergic degeneration rat model of dementia. METHODS: Adult male Sprague-Dawley rats (n = 48; 200-250 g) were divided into control (phosphate-buffered saline injection), 192 IgG-saporin (SAP), and SAP+FUS groups; in the two latter groups, SAP was injected bilaterally into the lateral ventricle. We applied FUS to the bilateral hippocampus with microbubbles. Immunohistochemistry, enzyme-linked immunosorbent assay, immunoblotting, 5-bromo-2'-deoxyuridine labeling, an acetylcholinesterase assay, and the Morris water maze test were performed to assess choline acetyltransferase, acetylcholinesterase activity, brain-derived neurotrophic factor expression, neural proliferation, and spatial memory, respectively. Statistical significance of differences in between groups was calculated using one-way and two-way analyses of variance followed by Tukey's multiple comparison test to determine the individual and interactive effects of FUS on immunochemistry and behavioral analysis. P < 0.05 was considered significant. RESULTS: Cholinergic degeneration in rats significantly decreased the number of choline acetyltransferase neurons (P < 0.05) in the basal forebrain, as well as AHN and spatial memory function. Rats that underwent FUS-mediated brain-blood barrier opening exhibited significant increases in brain-derived neurotrophic factor (BDNF; P < 0.05), early growth response protein 1 (EGR1) (P < 0.01), AHN (P < 0.01), and acetylcholinesterase activity in the frontal cortex (P < 0.05) and hippocampus (P < 0.01) and crossing over (P < 0.01) the platform in the Morris water maze relative to the SAP group after sonication. CONCLUSIONS: FUS treatment increased AHN and improved spatial memory. This improvement was mediated by increased hippocampal BDNF and EGR1. FUS treatment may also restore AHN and protect against neurodegeneration, providing a potentially powerful therapeutic strategy for AD.ope

Combined Effects of Focused Ultrasound and Photodynamic Treatment for Malignant Brain Tumors Using C6 Glioma Rat Model

Purpose: Glioblastoma (GBM) is an intractable disease for which various treatments have been attempted, but with little effect. This study aimed to measure the effect of photodynamic therapy (PDT) and sonodynamic therapy (SDT), which are currently be ing used to treat brain tumors, as well as sono-photodynamic therapy (SPDT), which is the combination of these two. Materials and Methods: Four groups of Sprague-Dawley rats were injected with C6 glioma cells in a cortical region and treated with PDT, SDT, and SPDT. Gd-MRI was monitored weekly and 18F-FDG-PET the day before and 1 week after the treatment. The acoustic power used during sonication was 5.5 W/cm2 using a 0.5-MHz single-element transducer. The 633-nm laser was illumi nated at 100 J/cm2. Oxidative stress and apoptosis markers were evaluated 3 days after treatment using immunohistochemistry (IHC): 4-HNE, 8-OhdG, and Caspase-3. Results: A decrease in tumor volume was observed in MRI imaging 12 days after the treatment in the PDT group (p<0.05), but the SDT group showed a slight increase compared to the 5-Ala group. The high expression rates of reactive oxygen species-related factors, such as 8-OhdG (p<0.001) and Caspase-3 (p<0.001), were observed in the SPDT group compared to other groups in IHC. Conclusion: Our findings show that light with sensitizers can inhibit GBM growth, but not ultrasound. Although SPDT did not show the combined effect in MRI, high oxidative stress was observed in IHC. Further studies are needed to investigate the safety parameters to apply ultrasound in GBM.ope

Enhanced delivery of a low dose of aducanumab via FUS in 5×FAD mice, an AD model

Background: Aducanumab (Adu), which is a human IgG1 monoclonal antibody that targets oligomer and fibril forms of beta-amyloid, has been reported to reduce amyloid pathology and improve impaired cognition after administration of a high dose (10 mg/kg) of the drug in Alzheimer's disease (AD) clinical trials. The purpose of this study was to investigate the effects of a lower dose of Adu (3 mg/kg) with enhanced delivery via focused ultrasound (FUS) in an AD mouse model. Methods: The FUS with microbubbles opened the blood-brain barrier (BBB) of the hippocampus for the delivery of Adu. The combined therapy of FUS and Adu was performed three times in total and each treatment was performed biweekly. Y-maze test, Brdu labeling, and immunohistochemical experimental methods were employed in this study. In addition, RNA sequencing and ingenuity pathway analysis were employed to investigate gene expression profiles in the hippocampi of experimental animals. Results: The FUS-mediated BBB opening markedly increased the delivery of Adu into the brain by approximately 8.1 times in the brains. The combined treatment induced significantly less cognitive decline and decreased the level of amyloid plaques in the hippocampi of the 5×FAD mice compared with Adu or FUS alone. Combined treatment with FUS and Adu activated phagocytic microglia and increased the number of astrocytes associated with amyloid plaques in the hippocampi of 5×FAD mice. Furthermore, RNA sequencing identified that 4 enriched canonical pathways including phagosome formation, neuroinflammation signaling, CREB signaling and reelin signaling were altered in the hippocami of 5×FAD mice receiving the combined treatment. Conclusion: In conclusion, the enhanced delivery of a low dose of Adu (3 mg/kg) via FUS decreases amyloid deposits and attenuates cognitive function deficits. FUS-mediated BBB opening increases adult hippocampal neurogenesis as well as drug delivery. We present an AD treatment strategy through the synergistic effect of the combined therapy of FUS and Adu.ope

Long-lasting restoration of memory function and hippocampal synaptic plasticity by focused ultrasound in Alzheimer's disease

Background: Focused ultrasound (FUS) is a medical technology that non-invasively stimulates the brain and has been applied in thermal ablation, blood–brain barrier (BBB) opening, and neuromodulation. In recent years, numerous experiences and indications for the use of FUS in clinical and preclinical studies have rapidly expanded. Focused ultrasound-mediated BBB opening induces cognitive enhancement and neurogenesis; however, the underlying mechanisms have not been elucidated. Methods: Here, we investigate the effects of FUS-mediated BBB opening on hippocampal long-term potentiation (LTP) and cognitive function in a 5xFAD mouse model of Alzheimer's disease (AD). We applied FUS with microbubble to the hippocampus and LTP was measured 6 weeks after BBB opening using FUS. Field recordings were made with a concentric bipolar electrode positioned in the CA1 region using an extracellular glass pipette filled with artificial cerebrospinal fluid. Morris water maze and Y-maze was performed to test cognitive function. Results: Our results demonstrated that FUS-mediated BBB opening has a significant impact on increasing LTP at Schaffer collateral - CA1 synapses and rescues cognitive dysfunction and working memory. These effects persisted for up to 7 weeks post-treatment. Also, FUS-mediated BBB opening in the hippocampus increased PKA phosphorylation. Conclusion: Therefore, it could be a promising treatment for neurodegenerative diseases as it remarkably increases LTP, thereby improving working memory. © 2023 The Author(s)ope

알츠하이머 마우스 모델에서 집속초음파와 아두카누맙의 병합 처치를 통한 베타 아밀로이드 플라크 감소 및 신경발생 유도에 의한 치료효과 검증

Focused ultrasound (FUS) with microbubbles (MBs) can noninvasively open the blood–brain barrier (BBB) for drug delivery into the parenchyma. In preclinical studies, it is known that FUS-induced BBB opening not only enhances the penetration of therapeutic agents but also modulates neurogenesis and immunity. Recently, human clinical trials in Alzheimer’s disease (AD) with FUS treatment have been initiated. Beta-amyloid (Aβ) deposition is a typical pathological phenomenon of AD. Although its mechanism remains questionable and controversial as a target for AD treatment strategies, numerous studies have focused on targeting Aβ. Meanwhile, numerous studies have reported that FUS-mediated BBB opening reduces Aβ levels in the brain. As previously used drugs for AD are limited to relieve symptoms, studies are more inclined to develop drugs, and there is a trend to develop drugs that can solve the cause of the disease. Aducanumab is a human monoclonal antibody that reduces Aβ plaques in a dose-dependent manner. First, establishing FUS parameters to avoid blood vessel damage is an essential step to address safety issues and needs to be effective enough to open the BBB as well. Thus, the safety and efficacy of FUS-mediated BBB opening in the 5xFAD AD mouse model were examined, and whether FUS-mediated BBB opening contributes to the reduction of Aβ and improved cognitive function was investigated. As a result, a safe and efficient parameter of FUS was developed (1-Hz burst repetition frequency, 10-ms bursts, 120s in total, and average peak pressure 0.25 MPa). It did not only cause any hemorrhages or damage in the brain, but also proved its effectiveness. FUS-mediated BBB opening decreased Aβ in the hippocampus of 5xFAD mice and rescued cognitive function. Second, it was hypothesized that the combined therapy of FUS and aducanumab would be more effective than either FUS or aducanumab treatment alone. The effectiveness could be maximized depending on the number of treatments; FUS was targeted at four regions of the hippocampus where newly generated neurons occur, and aducanumab treatment was conducted three times biweekly and dose dependent. Combined therapy increased both proliferation and neurogenesis in the hippocampus. In line with this result, it was shown that the combined therapy significantly decreased Aβ plaques in the hippocampus and restored cognitive function. In addition, its effectiveness was more statistically significant when treated multiple times. In conclusion, the combined therapy of FUS and aducanumab for AD reduced amyloid aggregation, restored spatial memory, and increased neurogenesis. Furthermore, the remarkable aspect of this study is that the combined therapy was shown to be effective even with the minimal dose of the drug by increasing the BBB penetration rate with FUS, while excluding the possibility of side effects. This study provides a better insight into establishing a solid therapeutic strategy for the treatment of AD as well as other neurodegenerative diseases. 집속초음파는 미세기포와 함께 사용되어 비침습적으로 뇌혈관장벽을 제어하고 뇌 내로 약물을 전달할 수 있다. 집속초음파를 통한 뇌혈관 장벽의 제어는 뇌 내로 약물 전달이 가능하게 할 뿐만 아니라 신경발생이나 면역, 다양한 환경변화를 조절할 수 있다는 전임상 연구들이 보고되었으며, 최근 치매환자를 대상으로 집속초음파 임상시험이 시작되었다. 베타 아밀로이드 침착은 치매에서 나타나는 대표적인 병리학적 현상이다. 치매 치료 전략으로써 베타 아밀로이드가 적절한지에 대해서는 아직도 많은 논란과 의문이 남아있지만, 이를 타겟으로 많은 치료 연구들이 진행중에 있다. 최근 집속초음파를 통한 뇌혈관 장벽의 제어가 뇌 내 침착된 아밀로이드 플라크를 줄여준다는 보고가 있었다. 한편 치매치료제 개발에 있어 이전에는 증상을 완화시키고 지연시키는 목적에 국한되었다면, 최근에는 병의 근본적인 원인을 해결할 수 있는 치료제를 개발하는 추세이다. 아두카누맙은 인간단일클론항체 치료제로써 약물의 용량에 따라 치매 환자의 뇌 내 아밀로이드 플라크를 줄여주는 치료 효과가 확인되어 근본적인 치매 치료제로 여겨진다. 하지만 아두카누맙 역시 큰 분자량으로 뇌혈관장벽을 투과하기 어렵기 때문에 임상에서는 고용량을 사용하며 이는 다양한 부작용을 일으킬 수 있다. 본 연구의 최종 목적은 집속초음파와 아두카누맙의 병합 치료를 통해 신개념 치매 치료효과를 검증하는 것이다. 본 연구의 1부에서는 5xFAD 치매 마우스 모델을 이용하여 집속초음파를 통한 뇌혈관 장벽 제어의 안전성과 효율을 확인하였다. H&E 염색을 통해 뇌 내 미세손상이 발생하지 않는 안전한 파라미터를 구축하였다. 집속초음파에 의해 아밀로이드 플라크가 감소함을 면역조직화학염색을 통해 확인하고, 모리스 수중 미로 실험을 통해 인지기능이 개선됨을 확인함으로써 집속초음파가 치매 치료 수단으로서의 가능성을 확인하였다. 2부에서는 앞서 확립한 집속초음파 파라미터를 기반으로, 아두카누맙 치료제와 병합 치료에 대해 연구하였다. 연구를 위해 5xFAD, 5xFAD + 집속초음파, 5xFAD + 아두카누맙, 5xFAD + 집속초음파 + 아두카누맙, 대조군 총 5개 그룹으로 나누어 진행하였다. 집속초음파는 뇌 내 해마 영역의 4개 부위를 조사하였으며, 치료는 2주 간격으로 총 3번 실시하였다. 인지기능 회복을 조사하기위해 Y자 미로 테스트를 수행하였으며, 아밀로이드 플라크와 신경발생 변화를 조사하기 위해 면역조직화학염색을 수행하였다. 실험결과 집속초음파와 아두카누맙을 병합치료한 그룹에서 유의하게 인지기능이 회복되고, 아밀로이드 플라크가 감소하며, 신경발생이 유도되는 효과를 확인하였다. 또한 단회 치료보다 반복 치료했을 경우 인지기능 회복이 더 증가하였다. 본 연구에서는 집속초음파를 통해 뇌 내 투과율을 증가시킴으로써 치료제를 적은 용량으로 효과를 극대화 시킬 수 있게 하며, 부작용을 최소화할 수 있다는 장점을 제시한다. 또한 뇌혈관장벽에 의해 사용이 제한되는 항체치료제의 가능성을 보여주었다는 점에서 매우 의미 있는 연구 결과라고 생각한다. 본 연구에서 제시하는 새로운 치료전략이 알츠하이머병 치료에 크게 기여할 수 있기를 기대하며, 또 더 나아가 다양한 퇴행성 뇌질환 치료에 확대 적용할 수 있을 것이라 생각한다.open박

Optimization of Medial Forebrain Bundle Stimulation Parameters for Operant Conditioning of Rats

Background: The medial forebrain bundle (MFB) is involved in the integration of pleasure and reward. Previous studies have used various stimulation parameters for operant conditioning, though the effectiveness of these parameters has not been systematically studied. Objectives: The purpose of the present study was to investigate the optimal MFB stimulation parameters for controlling the conditioned behavior of rats. Methods: We evaluated four factors, including intensity, frequency, pulse duration, and train duration, to determine the effect of each on lever pressure applied by animals. We further compared burst and tonic stimulation in terms of learning and performance abilities. Results: The number of lever presses increased with each factor. Animals in the burst stimulation group exhibited more lever presses. Furthermore, the average speed in the maze among burst stimulation group subjects was higher. Conclusion: We determined the optimal parameters for movement control of animals in operant conditioning and locomotor tasks by adjusting various electrical stimulation parameters. Our results reveal that a burst stimulation is more effective than a tonic stimulation for increasing the moving speed and number of lever presses. The use of this stimulation technique also allowed us to minimize the training required to control animal behavior.restrictio