Periodic Reporting for period 1 - Hibernating_Synapses (Synaptic resilience in Tau-induced neurodegeneration)
Periodo di rendicontazione: 2023-01-01 al 2025-06-30
There are approx. 55 million people living with dementia and this number will keep rising due to the increase in live expectancy and the lack of effective treatments. Patients suffering from Alzheimer’s disease (the most common form of dementia) lose their memories because of the loss of synaptic connections between the neurons in their brain, a process that slowly and irreversibly progresses during decades and that correlates with the hyperphosphorylation of Tau protein. Interestingly, seminal work showed that synapse loss and Tau hyperphosphorylation also occur simultaneously in the brain of certain animal species when they hibernate. Hibernation is an energy-saving strategy associated to extended periods of adverse external conditions, and the brain in general and synapses in particular are highly energy-demanding structures. Thus, when animals enter into hibernation (torpor) synapses are lost in different brain regions (notably the hippocampus, the brain region primarily involved in memory formation and consolidation) and Tau becomes hyperphosphorylated. Afterwards, when the periods of inactivity and lowered bodily function are finished (arousal), Tau dephosphorylation and synapse recovery occur within a matter of hours. This remarkable synaptic resilience is unique and could be potentially used for the development of novel therapeutic strategies against dementia. However, the molecular mechanisms that control this process are largely unknown and the connection with Alzheimer’s disease phenotypes remains circumstantial.
The overall goal of my project is to elucidate the molecular mechanisms of synaptic remodelling during hibernation and use this knowledge to counteract Tau-induced cognitive decline. In the first part, we will first define hibernation-induced molecular changes (both at the proteomic and transcriptional level) in the brain of the golden hamster (a hibernation model). Afterwards, we will perform several mechanistic studies to determine which of those changes (genes, proteins, pathways and cellular processes) are driving synapse loss and recovery across the hibernation cycle. In the second part of the project, we will use CRISPRi/a screening to identify “hibernation targets” (genes that are up- or down-regulated in arousal) that rescue Tau-induced synaptic dysfunction in different screens. The findings of this project will significantly impact dementia research and translation by opening largely unexplored therapeutic inroads to combat these disorders, but they will also help us understand a unique and extreme form of synaptic plasticity found in the brain of hibernating animals.
The overall goal of my project is to elucidate the molecular mechanisms of synaptic remodelling during hibernation and use this knowledge to counteract Tau-induced cognitive decline. In the first part, we will first define hibernation-induced molecular changes (both at the proteomic and transcriptional level) in the brain of the golden hamster (a hibernation model). Afterwards, we will perform several mechanistic studies to determine which of those changes (genes, proteins, pathways and cellular processes) are driving synapse loss and recovery across the hibernation cycle. In the second part of the project, we will use CRISPRi/a screening to identify “hibernation targets” (genes that are up- or down-regulated in arousal) that rescue Tau-induced synaptic dysfunction in different screens. The findings of this project will significantly impact dementia research and translation by opening largely unexplored therapeutic inroads to combat these disorders, but they will also help us understand a unique and extreme form of synaptic plasticity found in the brain of hibernating animals.
The Hibernating_Synapses project is successfully moving forward and most of the mid-term milestones that were set at the start have been achieved. These are our main achievements:
- We conducted single nucleus RNA sequencing on the hippocampus of euthermic, torpor and arousal hamsters and generated the first single-cell resolution atlas of the hibernating brain, starting to unveil the list of genes that are up- and down-regulated in each individual cell type at the different stages of the hibernation cycle. We validated several of these genes using RNAscope.
- We used advanced bioinformatic tools (SCENIC) and showed that several transcriptional networks (“regulons”) are up- or down-regulated in all cell types during torpor and arousal, revealing the existence of common coordinated transcriptional changes across different cell types during hibernation.
- We compared the transcriptional profile of different cell types from hibernating hamsters and from AD patients and found remarkable similarities between the response of specific cells during hibernation and cell states described in Alzheimer’s disease. This is an important achievement beyond the state-of-the-art in the field, because we show mechanistic similarities (and differences) between hibernation and Alzheimer’s disease.
- We conducted preliminary mechanistic studies and showed that Tau and microglia are functionally connected to the loss of synapses during hibernation. This is also an important advancement in the field because previous evidence was not causal.
- We developed novel tools and techniques to study synapse remodelling during hibernation, such as sparse labelling of cells and we started using in vivo two-photon imaging. This will allow us to conduct in vivo longitudinal monitoring of synapse remodelling across the hibernation cycle and to address gaps in the field, such as whether after hibernation dendritic spines grow back in the exact same location as they were before.
- We conducted single nucleus RNA sequencing on the hippocampus of euthermic, torpor and arousal hamsters and generated the first single-cell resolution atlas of the hibernating brain, starting to unveil the list of genes that are up- and down-regulated in each individual cell type at the different stages of the hibernation cycle. We validated several of these genes using RNAscope.
- We used advanced bioinformatic tools (SCENIC) and showed that several transcriptional networks (“regulons”) are up- or down-regulated in all cell types during torpor and arousal, revealing the existence of common coordinated transcriptional changes across different cell types during hibernation.
- We compared the transcriptional profile of different cell types from hibernating hamsters and from AD patients and found remarkable similarities between the response of specific cells during hibernation and cell states described in Alzheimer’s disease. This is an important achievement beyond the state-of-the-art in the field, because we show mechanistic similarities (and differences) between hibernation and Alzheimer’s disease.
- We conducted preliminary mechanistic studies and showed that Tau and microglia are functionally connected to the loss of synapses during hibernation. This is also an important advancement in the field because previous evidence was not causal.
- We developed novel tools and techniques to study synapse remodelling during hibernation, such as sparse labelling of cells and we started using in vivo two-photon imaging. This will allow us to conduct in vivo longitudinal monitoring of synapse remodelling across the hibernation cycle and to address gaps in the field, such as whether after hibernation dendritic spines grow back in the exact same location as they were before.
We have generated the first single-cell resolution atlas of the brain of the golden hamster. Using a combination of automated and manual methods based on the expression of cell type-specific markers, we annotated 34 clusters that unequivocally represent different cell types. Although we observe a high degree of similarity with the cell types previously described in the mouse and human atlases, the depth and quality of our analysis allowed us to identify rare neuronal populations that have not been previously resolved (to our knowledge) in other single-cell atlases. When fully analysed we will make the atlas publicly available, and this will be very useful for the research community studying the golden hamster as well as other organisms.
Additionally, have unveiled the full list of genes that are up- or down-regulated at the different stages of the hibernation cycle. Within this project, we are specifically looking at genes and pathways that are involved in synapse remodelling and that can be used to rescue Tau pathology. However, there are numerous other hibernation genes that could be potentially relevant for other groups and research fields. For instance, we found several ribosomal and mitochondrial genes upregulated across different cell types. It would be very interesting to further investigate what are the consequences of these changes for metabolism and protein synthesis, as well as for overall neuronal (or glial) function and survival during stress periods. Moreover, since metabolic defects and protein homeostasis defects are also found in Alzheimer’s disease (as well as in other neurodegenerative disorders), these phenotypes could be potentially interconnected with synapse loss and Tau hyperphosphorylation.
Finally, we are currently developing several methods and tools that will impact the hibernation research field. We have implanted a cranial window in the cortex of the hamster, injected AAVs to sparsely label different cells, and performed two-photon live imaging to study the dynamics of synapse remodelling during the hibernation cycle. This methodology represents a critical breakthrough because for the first time it is possible to follow individual neurons and synapses in the same animal at the different stages of hibernation. This will help to answer key and unresolved questions, such as whether dendritic spines regenerate in the exact same position they occupied before hibernation, whether different types of spines are more prone to degenerate and regenerate, and what the precise kinetics of synaptic spine de/regeneration are.
Additionally, have unveiled the full list of genes that are up- or down-regulated at the different stages of the hibernation cycle. Within this project, we are specifically looking at genes and pathways that are involved in synapse remodelling and that can be used to rescue Tau pathology. However, there are numerous other hibernation genes that could be potentially relevant for other groups and research fields. For instance, we found several ribosomal and mitochondrial genes upregulated across different cell types. It would be very interesting to further investigate what are the consequences of these changes for metabolism and protein synthesis, as well as for overall neuronal (or glial) function and survival during stress periods. Moreover, since metabolic defects and protein homeostasis defects are also found in Alzheimer’s disease (as well as in other neurodegenerative disorders), these phenotypes could be potentially interconnected with synapse loss and Tau hyperphosphorylation.
Finally, we are currently developing several methods and tools that will impact the hibernation research field. We have implanted a cranial window in the cortex of the hamster, injected AAVs to sparsely label different cells, and performed two-photon live imaging to study the dynamics of synapse remodelling during the hibernation cycle. This methodology represents a critical breakthrough because for the first time it is possible to follow individual neurons and synapses in the same animal at the different stages of hibernation. This will help to answer key and unresolved questions, such as whether dendritic spines regenerate in the exact same position they occupied before hibernation, whether different types of spines are more prone to degenerate and regenerate, and what the precise kinetics of synaptic spine de/regeneration are.