scholarly journals Quantitative mapping of dense microtubule arrays in mammalian neurons

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Eugene A Katrukha ◽  
Daphne Jurriens ◽  
Desiree M Salas Pastene ◽  
Lukas C Kapitein
Keyword(s):  
Hippocampal Neurons ◽  
Intracellular Transport ◽  
Motor Proteins ◽  
Three Dimensional ◽  
Super Resolution ◽  
Absolute Number ◽  
Microtubule Network ◽  
Spatial Regulation ◽  
Quantitative Mapping ◽  
Dendritic Shaft

The neuronal microtubule cytoskeleton underlies the polarization and proper functioning of neurons, amongst others by providing tracks for motor proteins that drive intracellular transport. Different subsets of neuronal microtubules, varying in composition, stability and motor preference, are known to exist, but the high density of microtubules has so far precluded mapping their relative abundance and three-dimensional organization. Here we use different super-resolution techniques (STED, Expansion Microscopy) to explore the nanoscale organization of the neuronal microtubule network in rat hippocampal neurons. This revealed that in dendrites acetylated microtubules are enriched in the core of the dendritic shaft, while tyrosinated microtubules are enriched near the plasma membrane, thus forming a shell around the acetylated microtubules. Moreover, using a novel analysis pipeline we quantified the absolute number of acetylated and tyrosinated microtubules within dendrites and found that they account for 65-75% and ~20-30% of all microtubules, respectively, leaving only few microtubules that do not fall in either category. Because these different microtubule subtypes facilitate different motor proteins, these novel insights help to understand the spatial regulation of intracellular transport.

scholarly journals Quantitative mapping of dense microtubule arrays in mammalian neurons

2021 ◽  
Author(s):  
Eugene A. Katrukha ◽  
Daphne Jurriens ◽  
Desiree Salas Pastene ◽  
Lukas C. Kapitein
Keyword(s):  
Intracellular Transport ◽  
Motor Proteins ◽  
Three Dimensional ◽  
Super Resolution ◽  
Absolute Number ◽  
Analysis Pipeline ◽  
Microtubule Network ◽  
Spatial Regulation ◽  
Quantitative Mapping ◽  
Dendritic Shaft

ABSTRACTThe neuronal microtubule cytoskeleton underlies the polarization and proper functioning of neurons, amongst others by providing tracks for motor proteins that drive intracellular transport. Different subsets of neuronal microtubules, varying in composition, stability and motor preference, are known to exist, but the high density of microtubules has so far precluded mapping their relative abundance and three-dimensional organization. Here we use different super-resolution techniques (STED, Expansion Microscopy) to explore the nanoscale organization of the neuronal microtubule network. This revealed that in dendrites stable, acetylated microtubules are enriched in the core of the dendritic shaft, while dynamic, tyrosinated microtubules enrich near the plasma membrane, thus forming a shell around the stable microtubules. Moreover, using a novel analysis pipeline we quantified the absolute number of acetylated and tyrosinated microtubules within dendrites and found that they account for 65-75% and ∼20-30% of all microtubules, respectively, leaving only few microtubules that do not fall in either category. Because these different microtubule subtypes facilitate different motor proteins, these novel insights help to understand the spatial regulation of intracellular transport.

Three-Dimensional Super-Resolution Imaging of the Cytoskeleton in Hippocampal Neurons Using Selective Plane Illumination

2020 ◽  
pp. 261-293
Author(s):  
Frances Camille M. Wu ◽  
Feby Wijaya Pratiwi ◽  
Chin-Yi Chen ◽  
Chieh-Han Lu ◽  
Wei-Chun Tang ◽  
...  
Keyword(s):  
Hippocampal Neurons ◽  
Three Dimensional ◽  
Super Resolution ◽  
Resolution Imaging

scholarly journals Neuronal Aβ42 is enriched in small vesicles at the presynaptic side of synapses

2018 ◽  
Vol 1 (3) ◽  
pp. e201800028 ◽  
Author(s):  
Yang Yu ◽  
Daniel C Jans ◽  
Bengt Winblad ◽  
Lars O Tjernberg ◽  
Sophia Schedin-Weiss
Keyword(s):  
Hippocampal Neurons ◽  
Memory Function ◽  
Three Dimensional ◽  
Amyloid Β ◽  
Super Resolution ◽  
Long Term Potentiation ◽  
Synaptic Function ◽  
Amyloid Β Peptide ◽  
Term Potentiation ◽  
Super Resolution Microscopy

The amyloid β-peptide (Aβ) is a physiological ubiquitously expressed peptide suggested to be involved in synaptic function, long-term potentiation, and memory function. The 42 amino acid-long variant (Aβ42) forms neurotoxic oligomers and amyloid plaques and plays a key role in the loss of synapses and other pathogenic events of Alzheimer disease. Still, the exact localization of Aβ42 in neurons and at synapses has not been reported. Here, we used super-resolution microscopy and show that Aβ42 was present in small vesicles in presynaptic compartments, but not in postsynaptic compartments, in the neurites of hippocampal neurons. Some of these vesicles appeared to lack synaptophysin, indicating that they differ from the synaptic vesicles responsible for neurotransmitter release. The Aβ42-containing vesicles existed in presynapses connected to stubby spines and mushroom spines, and were also present in immature presynapses. These vesicles were scarce in other parts of the neurites, where Aβ42 was instead present in large, around 200–600 nm, vesicular structures. Three-dimensional super-resolution microscopy confirmed that Aβ42 was present in the presynapse and absent in the postsynapse.

scholarly journals A Super-Resolved View of the Alzheimer’s Disease-Related Amyloidogenic Pathway in Hippocampal Neurons

2021 ◽  
pp. 1-20
Author(s):  
Yang Yu ◽  
Yang Gao ◽  
Bengt Winblad ◽  
Lars Tjernberg ◽  
Sophia Schedin Weiss
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Hippocampal Neurons ◽  
Three Dimensional ◽  
Amyloid Β ◽  
Stimulated Emission ◽  
Amyloid Β Peptide ◽  
Primary Neurons ◽  
Amyloid Β Protein ◽  
Early Endosomes

Background: Processing of the amyloid-β protein precursor (AβPP) is neurophysiologically important due to the resulting fragments that regulate synapse biology, as well as potentially harmful due to generation of the 42 amino acid long amyloid β-peptide (Aβ 42), which is a key player in Alzheimer’s disease. Objective: Our aim was to clarify the subcellular locations of the amyloidogenic AβPP processing in primary neurons, including the intracellular pools of the immediate substrate, AβPP C-terminal fragment (APP-CTF) and the product (Aβ 42). To overcome the difficulties of resolving these compartments due to their small size, we used super-resolution microscopy. Methods: Mouse primary hippocampal neurons were immunolabelled and imaged by stimulated emission depletion (STED) microscopy, including three-dimensional, three-channel imaging and image analyses. Results: The first (β-secretase) and second (γ-secretase) cleavages of AβPP were localized to functionally and distally distinct compartments. The β-secretase cleavage was observed in early endosomes, where we were able to show that the liberated N- and C-terminal fragments were sorted into distinct vesicles budding from the early endosomes in soma. Lack of colocalization of Aβ 42 and APP-CTF in soma suggested that γ-secretase cleavage occurs in neurites. Indeed, APP-CTF was, in line with Aβ 42 in our previous study, enriched in the presynapse but absent from the postsynapse. In contrast, full-length AβPP was not detected in either the pre- or the postsynaptic side of the synapse. Furthermore, we observed that endogenously produced and endocytosed Aβ 42 were localized in different compartments. Conclusion: These findings provide critical super-resolved insight into amyloidogenic AβPP processing in primary neurons.

scholarly journals Three-dimensional super-resolution imaging for fluorescence emission difference microscopy

AIP Advances ◽  
2015 ◽  
Vol 5 (8) ◽  
pp. 084901 ◽  
Author(s):  
Shangting You ◽  
Cuifang Kuang ◽  
Shuai Li ◽  
Xu Liu ◽  
Zhihua Ding
Keyword(s):  
Three Dimensional ◽  
Fluorescence Emission ◽  
Super Resolution ◽  
Resolution Imaging

Super-resolution imaging using a three-dimensional metamaterials nanolens

2010 ◽  
Vol 96 (2) ◽  
pp. 023114 ◽  
Author(s):  
B. D. F. Casse ◽  
W. T. Lu ◽  
Y. J. Huang ◽  
E. Gultepe ◽  
L. Menon ◽  
...  
Keyword(s):  
Three Dimensional ◽  
Super Resolution ◽  
Resolution Imaging
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