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Long COVID, the Hippocampus and the Spike Protein: Microstrokes vs Spike Protein Administration
Microstrokes disrupt the structural network of the hippocampus – and experimental spike protein exposure can disrupt hippocampal network function through inflammatory and vascular mechanisms.
WALTER M CHESNUT
FEB 2

长新冠、海马体与刺突蛋白：微小中风 vs 刺突蛋白给药 二甲双胍似乎可以改善大脑中刺突蛋白的一些有害影响

Long COVID, the Hippocampus, and the Spike Protein: Microstrokes vs. Spike Protein Administration
Metformin appears to mitigate some of the harmful effects of the spike protein in the brain

微小中风会破坏海马体的结构网络——而实验性刺突蛋白暴露也可通过炎症和血管机制扰乱海马体网络功能。

沃尔特·M·切斯纳特（Walter M. Chesnut）

长新冠病毒、海马体和尖峰蛋白：微中风与尖峰蛋白管理
微中风会破坏海马体的结构网络，而实验性刺突蛋白暴露会通过炎症和血管机制破坏海马体网络功能。
沃尔特·M·切斯纳特
2月2日

摘要：二甲双胍似乎可以改善大脑中刺突蛋白的一些有害影响。

https://open.substack.com/pub/wmcresearch/p/long-covid-the-hippocampus-and-the

(A) Intranasally injected S1 (His-tagged) in the olfactory bulb, striatum, and hippocampus was examined at 3 hr post S1 administration (0.5 μg/per animal) using immunoblotting. Representative images are presented. (B) Schematic of behavior tests. (C-G) Episodic-like memory task. (C) The episodic-like memory task consists of two sample phases and a test phase. In each sample phase, rats encountered two sets of four identical novel objects (old objects, sample 1; recent objects, sample 2). In the test phase, objects were mixed together. One of the old familiar objects was placed in new location (b: old familiar displaced). The other three objects were placed in same location as in sample phases (a: old familiar stationary, c: recent familiar stationary). (D) Distribution of exploratory time for ‘what-when’ (old familiar objects vs. recent familiar objects). (E) Recency discrimination of exploratory times for ‘what-when’ (old stationary objects vs. recent stationary objects) (F) Distribution of exploratory time for ‘what-where’ (old familiar displaced vs. recent familiar stationary). (G) Total exploratory distance for each group in the test phase. (H-M) Water maze test. (H) Representative swimming paths of the control and S1 injected group during the probe trial. (I-J) The escape latencies in the two groups over five consecutive training days (I) and on the 5th day of the spatial acquisition session (J). (K-L) The percentage of time spent and distance traveled in the target quadrant during the probe trial (K), the average crossing number over the platform-site, and the latency of the first target-site crossover (L). (M) The average swimming speed of two groups. (O-R) Open field test. (O) Representative track sheets of the control and S1 injected group during the test trial. (P-R) Graphs showing alterations in time spent in central zone (P), distance traveled in central zone (Q), and overall distance traveled (R). Values are presented as means±SEMs. p < 0.05, * p < 0.01, ***p < 0.001 vs. control. Con, control group (n = 12); S1, S1 injected group (n = 12).
If one recalls the trash compactor scene in Star Wars, we may use that scene as an analogy for what occurs to the hippocampus in the presence of the Spike Protein; We have a two-pronged, crushing assault. On the one hand, we have a microvascular attack impairing the hippocampus, functionally analogous to microstrokes in downstream network effects. And, on the other hand, we have a neuroinflammatory attack induced by the presence of the Spike Protein.
First, let’s look at what microstrokes do to the hippocampus. (A)在S1施用(0.5μg/每只动物)后3小时，使用免疫印迹检查嗅球、纹状体和海马中鼻内注射的S1(His标记)。 呈现了代表性图像。 (B) 行为测试示意图。 (C-G) 情景式记忆任务。 (C) 情景式记忆任务由两个样本阶段和一个测试阶段组成。 在每个样本阶段，老鼠遇到两组四个相同的新物体（旧物体，样本 1；最近的物体，样本 2）。 在测试阶段，对象被混合在一起。 旧熟悉物体之一被放置在新位置（b：旧熟悉物体移位）。 其他三个物体被放置在与样本阶段相同的位置（a：旧的熟悉的固定物，c：最近熟悉的固定物）。 (D)“什么-什么时候”的探索时间分布（旧熟悉的物体与最近熟悉的物体）。 （E）“什么时候”的探索时间的新近度辨别（旧的静止物体与最近的静止物体）（F）“什么地方”的探索时间分布（旧的熟悉的移位与最近熟悉的静止物体）。 (G) 测试阶段每组的总探索距离。 (H-M)水迷宫测试。 (H) 探针试验期间对照组和 S1 注射组的代表性游泳路径。 (I-J) 两组在连续五个训练日 (I) 和空间获取会话第五天 (J) 中的逃避潜伏期。 (K-L) 探测试验期间在目标象限中花费的时间和行进距离的百分比 (K)、平台站点上的平均交叉次数以及第一次目标站点交叉的延迟 (L)。 (M) 两组的平均游泳速度。 (O-R) 旷场测试。 (O) 试验期间对照组和 S1 注射组的代表性跟踪表。 (P-R) 图表显示在中心区域花费的时间 (P)、在中心区域行驶的距离 (Q) 以及总行驶距离 (R) 的变化。 值表示为平均值±SEM。 p < 0.05，* p < 0.01，***p < 0.001 与对照相比。 Con，对照组（n = 12）； S1，S1注射组（n = 12）。
如果人们回想起《星球大战》中垃圾压实机的场景，我们可以用这个场景来类比海马体在刺突蛋白存在时发生的情况； 我们正面临着双管齐下的毁灭性攻击。 一方面，微血管攻击损害了海马体，其功能类似于下游网络效应中的微中风。 另一方面，我们会因刺突蛋白的存在而诱发神经炎症发作。
首先，让我们看看微抚对海马体有何影响。

Cognitive deficits affect over 70% of stroke survivors, yet the mechanisms by which multiple small ischemic events contribute to cognitive decline remain poorly understood. In this study, we employed chronic two-photon calcium imaging to longitudinally track the fate of individual neurons in the hippocampus of mice navigating a virtual reality environment, both before and after inducing brain-wide microstrokes. Our findings reveal that, under normal conditions, hippocampal neurons exhibit varying degrees of stability in their spatial memory coding. However, microstrokes disrupted this functional network architecture, leading to cognitive impairments. Notably, the preservation of stable coding place cells, along with the stability, precision, and persistence of the hippocampal network, was strongly predictive of cognitive outcomes. Mice with more synchronously active place cells near important locations demonstrated recovery from cognitive impairment. This study uncovers critical cellular responses and network alterations following brain injury, providing a foundation for novel therapeutic strategies preventing cognitive decline.

认知缺陷影响了超过 70% 的卒中幸存者，但多个微小缺血事件如何导致认知功能下降，其机制仍不清楚。在本研究中，我们采用慢性双光子钙成像技术，对在虚拟现实环境中导航的小鼠海马体内单个神经元的命运进行了纵向追踪，观察其在全脑微梗死诱导前后的变化。研究结果显示，在正常条件下，海马神经元在空间记忆编码的稳定性方面存在差异。然而，微梗死破坏了这一功能性网络结构，进而导致认知功能受损。值得注意的是，稳定编码位置细胞的保留情况，以及海马网络在稳定性、精确性和持续性方面的表现，与认知结局高度相关。在关键位置附近具有更高同步活动的位置细胞的小鼠，表现出从认知障碍中的恢复。本研究揭示了脑损伤后关键的细胞反应和网络层面的改变，为预防认知功能衰退的全新治疗策略奠定了基础。

Brain-wide microstrokes affect the stability of memory circuits in the hippocampus
https://www.nature.com/articles/s41467-025-58688-4
Yet when one observes the interaction of the Spike Protein with the cerebral vasculature, we find a parallel effect. 全脑微中风影响海马体记忆回路的稳定性
https://www.nature.com/articles/s41467-025-58688-4
然而，当观察刺突蛋白与脑血管系统的相互作用时，我们发现了一种平行的效应。

Methods: S-protein was either injected intravenously or directly into the hippocampus of K-18 (h ACE2 in epithelial cells) or global h-ACE2 knock-in (h ACE2 KI) mice or wild-type mice. Cognitive functions were assessed by Y-maze and Barnes maze. Cerebrovascular density was determined using confocal 3-D image reconstruction. Human brain microvascular endothelial cells (HBMVEC) were treated with S-protein and assessed for apoptosis and inflammatory markers using immunoblotting and RT-PCR. K-18 and h-ACE2 KI mice received intraocular injections of S-protein; retinas were evaluated for vascular cell death and inflammation.
Results: S-protein injections caused significant deterioration in memory and learning function of K-18 and h-ACE2 KI mice but not in the wild-type mice (P<0.05). S-protein significantly increased inflammatory mediators, cytokine production, and apoptosis in the brains and HBMVECs (P<0.05). Significant cerebrovascular rarefaction was detected only in K-18 and h-ACE2 KI mice compared to wild-type mice (P<0.05). Retinal vascular cell death and inflammation were significantly increased after S-protein injection. (P<0.05)
Conclusions: COVID-19 spike protein decreases cognitive function via increased endothelial cell inflammation, apoptosis, and cerebrovascular rarefaction. Humanized ACE2 animal models are excellent and reliable for investigating the neurological symptoms of COVID-19. 方法：将S蛋白静脉注射或直接注射到K-18（上皮细胞中的h ACE2）或整体h-ACE2敲入（h ACE2 KI）小鼠或野生型小鼠的海马中。 通过Y迷宫和Barnes迷宫评估认知功能。 使用共焦 3-D 图像重建确定脑血管密度。 用 S 蛋白处理人脑微血管内皮细胞 (HBMVEC)，并使用免疫印迹和 RT-PCR 评估细胞凋亡和炎症标记物。 K-18和h-ACE2 KI小鼠接受S-蛋白眼内注射； 评估视网膜的血管细胞死亡和炎症。
结果：S蛋白注射导致K-18和h-ACE2 KI小鼠的记忆和学习功能显着恶化，但野生型小鼠则没有这种情况（P<0.05）。 S蛋白显着增加大脑和HBMVEC中的炎症介质、细胞因子的产生和细胞凋亡（P<0.05）。 与野生型小鼠相比，仅在 K-18 和 h-ACE2 KI 小鼠中检测到显着的脑血管稀疏（P<0.05）。 注射S-蛋白后，视网膜血管细胞死亡和炎症显着增加。 (P<0.05)
结论：COVID-19 刺突蛋白通过增加内皮细胞炎症、细胞凋亡和脑血管稀疏而降低认知功能。 人源化 ACE2 动物模型对于研究 COVID-19 的神经系统症状非常出色且可靠。

Abstract 53: Covid-19 Spike-protein Causes Cerebrovascular Rarefaction And Deteriorates Cognitive Functions In A Mouse Model Of Humanized ACE2
https://www.ahajournals.org/doi/10.1161/str.53.suppl_1.53
So, we observe inflammatory-mediated injury to the hippocampus following spike protein exposure in experimental models. There is also evidence of spike protein–associated neuroinflammatory effects and neuronal dysfunction within the hippocampus. 摘要 53：Covid-19 刺突蛋白导致人源化 ACE2 小鼠模型脑血管稀疏并恶化认知功能
https://www.ahajournals.org/doi/10.1161/str.53.suppl_1.53
因此，我们在实验模型中观察到刺突蛋白暴露后炎症介导的海马损伤。 还有证据表明刺突蛋白相关的神经炎症作用和海马内的神经元功能障碍。

We found that the spike protein alone induced a broad spectrum of proteome changes in the mouse skull marrow, meninges, and brain that are similar to those observed in COVID-19 patients and that the isolated effects of the spike protein on the nervous system caused anxiety-like behavior without memory deficits. While previous studies reported anxiety and cognitive impairment induced by direct administration of spike protein into the hippocampus or ventricles,27,96 the differences between our and previous studies are likely caused by the higher local concentration of spike protein in the hippocampus in those studies. The worsened outcomes after MCAo and TBI indicated that the spike protein may render the brain more vulnerable to subsequent insults. No significant difference was observed in the TBI model during the acute phase. However, a more pronounced injury attributable to spike treatment was detected in the chronic phase. Our MCAo assessment focused solely on the acute phase, and additional long-term spike protein-induced changes may not have been observed. While these findings do not support the direct induction of clinical neurological symptoms by the spike protein, they suggest the long-term consequences of spike protein-induced inflammation and dysfunctional signaling in the brain. 我们发现，刺突蛋白单独诱导了小鼠颅骨、骨髓、脑膜和大脑中广泛的蛋白质组变化，这与在 COVID-19 患者中观察到的相似，并且刺突蛋白对神经系统的孤立影响导致了焦虑样行为，但没有记忆缺陷。 虽然之前的研究报道了直接将刺突蛋白注射到海马体或心室会引起焦虑和认知障碍，27,96但我们和之前的研究之间的差异可能是由于这些研究中海马刺突蛋白的局部浓度较高造成的。 MCAo 和 TBI 后恶化的结果表明，刺突蛋白可能使大脑更容易受到随后的损伤。 TBI模型在急性期没有观察到显着差异。 然而，在慢性期发现了由尖峰治疗引起的更明显的损伤。 我们的 MCAo 评估仅关注急性期，可能没有观察到额外的长期刺突蛋白诱导的变化。 虽然这些发现并不支持刺突蛋白直接诱导临床神经症状，但它们表明刺突蛋白诱导的炎症和大脑中信号传导功能障碍的长期后果。

Persistence of spike protein at the skull-meninges-brain axis may contribute to the neurological sequelae of COVID-19
https://www.sciencedirect.com/science/article/pii/S1931312824004384 头骨-脑膜-大脑轴上刺突蛋白的持续存在可能会导致 COVID-19 的神经系统后遗症
https://www.sciencedirect.com/science/article/pii/S1931312824004384

We demonstrate that intranasally administered S1 quickly entered the hippocampus and was associated with cognitive impairment by 6 weeks post-injection. Transcriptomic analysis of hippocampal tissue revealed early alterations in gene expression associated with synaptic function. We observed that the expression of hypoxia-responsive genes was also altered, suggesting the involvement of HIF-1α signaling. Further analysis confirmed that S1 stabilized the HIF-1α protein in a hypoxia-independent manner, and siRNA-mediated knockdown of HIF-1α restored synaptic gene expression, including GRIN2A, SHANK1, and JPH3. By 6 weeks post-injection, hippocampal neuronal loss was accompanied by the accumulation of phosphorylated tau (p-tau) and aggregated α-synuclein.
In this study, after administering spike protein intranasally, there was a notable accumulation of tau and α-synuclein proteins, accompanied by a corresponding decline in cognitive function within the hippocampal region. These results suggest that direct delivery of spike protein to the central nervous system can induce neuropathological changes via neuroinflammation and synapse loss. Indeed, previous studies have shown that spike protein accumulation in the hippocampus is associated with increased inflammatory cytokines, and neuronal loss, leading to cognitive decline [14,61]. 我们证明，鼻内注射的 S1 快速进入海马体，并在注射后 6 周与认知障碍相关。 海马组织的转录组分析揭示了与突触功能相关的基因表达的早期变化。 我们观察到缺氧反应基因的表达也发生了改变，表明 HIF-1α 信号传导参与其中。 进一步分析证实，S1 以不依赖缺氧的方式稳定 HIF-1α 蛋白，并且 siRNA 介导的 HIF-1α 敲低恢复了突触基因表达，包括 GRIN2A、SHANK1 和 JPH3。 注射后 6 周，海马神经元损失伴随着磷酸化 tau (p-tau) 和聚集的 α-突触核蛋白的积累。
在这项研究中，鼻内施用刺突蛋白后，tau 蛋白和 α-突触核蛋白明显积累，并伴随着海马区域认知功能的相应下降。 这些结果表明，将刺突蛋白直接递送至中枢神经系统可以通过神经炎症和突触损失诱导神经病理学变化。 事实上，之前的研究表明，海马中刺突蛋白的积累与炎症细胞因子的增加和神经元损失有关，从而导致认知能力下降[14,61]。

SARS-CoV-2 spike protein causes synaptic dysfunction and p-tau and α-synuclein aggregation leading cognitive impairment: The protective role of metformin
https://pmc.ncbi.nlm.nih.gov/articles/PMC12594341/
Experimental studies show that isolated spike protein exposure can induce neuroinflammatory and vascular changes in animal models, particularly affecting hippocampal signaling and behavior. Although the underlying mechanisms differ, these experimental models produce overlapping behavioral and network-level impairments relevant to post-COVID cognitive symptoms. Metformin seems to ameliorate some of these deleterious effects of the Spike Protein in the brain. We will investigate this further. I would like to give an immense Thank You to the subscriber who became a Founding Member over the weekend. Please have a blessed week. SARS-CoV-2 刺突蛋白导致突触功能障碍，p-tau 和 α-突触核蛋白聚集导致认知障碍：二甲双胍的保护作用
https://pmc.ncbi.nlm.nih.gov/articles/PMC12594341/
实验研究表明，孤立的刺突蛋白暴露可引起动物模型中的神经炎症和血管变化，特别是影响海马信号传导和行为。 尽管潜在机制不同，但这些实验模型产生了与新冠病毒后认知症状相关的重叠行为和网络水平损伤。 二甲双胍似乎可以改善大脑中刺突蛋白的一些有害影响。 我们将对此进行进一步调查。 我要向周末成为创始会员的订阅者表示衷心的感谢。 祝您度过愉快的一周。