Cortical synapses of the world's smallest mammal: An FIB/SEM study in the Etruscan shrew

Alonso‐Nanclares, Lidia; Toledo-Rodriguez, Maria; Merchán-Pérez, Ángel; González‐Soriano, Juncal; Plaza-Alonso, Sergio; Cano‐Astorga, Nicolás; Naumann, Robert K.; Brecht, Michael; DeFelipe, Javier

doi:10.1002/cne.25432

Cited by 12 publications

(4 citation statements)

References 93 publications

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“…In L1 and L3, we found synaptic densities of 1.80 and 1.08 synapses/µm 3 , respectively, that is, clearly lower than in the SR. These data are in line with the synaptic densities described in the somatosensory cortex of the Etruscan shrew 53 . In the somatosensory cortex of the juvenile rat, the synaptic densities are lower, but the proportion of excitatory and inhibitory synapses is similar to the mouse data, including the fact that layer 1 has a higher percentage of inhibitory synapses than layer 3 54 , 55 .…”

Section: Discussionsupporting

confidence: 90%

Tracing nerve fibers with volume electron microscopy to quantitatively analyze brain connectivity

Turegano-Lopez,

de las Pozas,

Santuy

et al. 2024

Commun Biol

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The highly complex structure of the brain requires an approach that can unravel its connectivity. Using volume electron microscopy and a dedicated software we can trace and measure all nerve fibers present within different samples of brain tissue. With this software tool, individual dendrites and axons are traced, obtaining a simplified “skeleton” of each fiber, which is linked to its corresponding synaptic contacts. The result is an intricate meshwork of axons and dendrites interconnected by a cloud of synaptic junctions. To test this methodology, we apply it to the stratum radiatum of the hippocampus and layers 1 and 3 of the somatosensory cortex of the mouse. We find that nerve fibers are densely packed in the neuropil, reaching up to 9 kilometers per cubic mm. We obtain the number of synapses, the number and lengths of dendrites and axons, the linear densities of synapses established by dendrites and axons, and their location on dendritic spines and shafts. The quantitative data obtained through this method enable us to identify subtle traits and differences in the synaptic organization of the samples, which might have been overlooked in a qualitative analysis.

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Section: Discussionsupporting

confidence: 90%

Tracing nerve fibers with volume electron microscopy to quantitatively analyze brain connectivity

Turegano-Lopez,

de las Pozas,

Santuy

et al. 2024

Commun Biol

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“…We found that the majority of synapses had a macular shape (79–87%) and were the smallest synapses. These findings are in line with previous reports in other brain regions and species (Geinisman et al, 1987; Jones et al, 1991; Neuman et al, 2016; Hsu et al, 2017; Calì et al, 2018; Santuy et al, 2018a; Domínguez-Álvaro et al, 2019, 2021a; Montero-Crespo et al, 2020; Cano-Astorga et al, 2021, 2023; Turégano-López, 2022; Alonso-Nanclares et al, 2023). It has been reported that complex-shaped synapses possess more AMPA and NMDA receptors than macular synapses, characterizing them as a potentially ’powerful’ population associated with more enduring memory-related functionality than macular synapses (Geinisman et al, 1987, 1991, 1992a, 1992b, 1993; Lüscher et al, 2000; Toni et al, 2001; Ganeshina et al, 2004a, 2004b; Spruston 2008).…”

Section: Common Synaptic Characteristicssupporting

confidence: 93%

Volume Electron Microscopy Analysis of Synapses in Associative and Primary Regions of the Human Cerebral Cortex

Cano-Astorga,

Plaza-Alonso,

DeFelipe

et al. 2024

Preprint

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Functional and structural studies investigating macroscopic connectivity in the human cerebral cortex suggest that high-order associative regions exhibit greater connectivity compared to primary ones. However, the synaptic organization of these brain regions remains unexplored due to the difficulties involved in studying the human brain at the nanoscopic level. In the present work, we conducted volume electron microscopy to investigate the synaptic organization of the human brain obtained at autopsy. Specifically, we examined layer III of Brodmann areas 17, 3b, and 4, as representative areas of primary visual, somatosensorial, and motor cortex. Additionally, we conducted comparative analyses with our previous synaptic datasets of layer III from temporopolar and anterior cingulate associative cortical regions (Brodmann areas 24, 38, and 21). 9,690 synaptic junctions were 3D reconstructed, showing that certain synaptic characteristics appeared to be specific to particular cortical regions. The number of synapses per volume, the proportion of the postsynaptic targets, and the synaptic size may distinguish one region from another, regardless of whether they are associative or primary cortex. By contrast, other synaptic characteristics were common to all analyzed regions, such as the proportion of excitatory and inhibitory synapses, their shapes, their spatial distribution, and a higher proportion of synapses located on dendritic spines. These observations may be included within the general rules of synaptic organization of the human cerebral cortex. The present results on nanoscopic characteristics of synapses provide further insights into the structural design of the human cerebral cortex.

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“…However, this characteristic is often misinterpreted as implying that synapses on dendritic shafts are mostly SS (Motta et al, 2019;Karimi et al, 2020;Loomba et al, 2022). In fact, quantitative analyses of synapses in the neuropil have shown that most synapses on dendritic shafts are AS (∼80%), with relatively few being SS (∼20%) (Beaulieu et al, 1992;Peters et al, 2008;Hsu et al, 2017;Calì et al, 2018;Santuy et al, 2018b;Domínguez-Álvaro et al, 2019Domínguez-Álvaro et al, , 2021aYakoubi et al, 2019b;Montero-Crespo et al, 2020Cano-Astorga et al, 2021, 2023Alonso-Nanclares et al, 2023). Therefore, synaptic organization datasets with incorrect assumptions regarding SS identification could introduce an important source of bias.…”

Section: Discussionmentioning

confidence: 99%

“…The most noticeable distinction lies in the postsynaptic density: AS have a thick postsynaptic density, while SS have a thin postsynaptic density. In general, regarding all synapses in the neuropil of the cerebral cortex, AS outnumber SS approximately 95:5 (see Alonso-Nanclares et al, 2023, andCano-Astorga et al, 2023, and the references therein). Importantly, it has been demonstrated that, in the cerebral cortex, most AS are excitatory (glutamatergic), while SS are inhibitory (GABAergic) (Colonnier, 1968;Gray, 1969;Peters and Kaiserman-Abramof, 1969;Houser et al, 1984;Peters and Palay, 1996;Ascoli et al, 2008).…”

Section: Introductionmentioning

confidence: 99%

Unambiguous identification of asymmetric and symmetric synapses using volume electron microscopy

Cano-Astorga,

Plaza-Alonso,

Turegano-Lopez

et al. 2024

Front. Neuroanat.

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The brain contains thousands of millions of synapses, exhibiting diverse structural, molecular, and functional characteristics. However, synapses can be classified into two primary morphological types: Gray’s type I and type II, corresponding to Colonnier’s asymmetric (AS) and symmetric (SS) synapses, respectively. AS and SS have a thick and thin postsynaptic density, respectively. In the cerebral cortex, since most AS are excitatory (glutamatergic), and SS are inhibitory (GABAergic), determining the distribution, size, density, and proportion of the two major cortical types of synapses is critical, not only to better understand synaptic organization in terms of connectivity, but also from a functional perspective. However, several technical challenges complicate the study of synapses. Potassium ferrocyanide has been utilized in recent volume electron microscope studies to enhance electron density in cellular membranes. However, identifying synaptic junctions, especially SS, becomes more challenging as the postsynaptic densities become thinner with increasing concentrations of potassium ferrocyanide. Here we describe a protocol employing Focused Ion Beam Milling and Scanning Electron Microscopy for studying brain tissue. The focus is on the unequivocal identification of AS and SS types. To validate SS observed using this protocol as GABAergic, experiments with immunocytochemistry for the vesicular GABA transporter were conducted on fixed mouse brain tissue sections. This material was processed with different concentrations of potassium ferrocyanide, aiming to determine its optimal concentration. We demonstrate that using a low concentration of potassium ferrocyanide (0.1%) improves membrane visualization while allowing unequivocal identification of synapses as AS or SS.

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Cortical synapses of the world's smallest mammal: An FIB/SEM study in the Etruscan shrew

Cited by 12 publications

References 93 publications

Tracing nerve fibers with volume electron microscopy to quantitatively analyze brain connectivity

Tracing nerve fibers with volume electron microscopy to quantitatively analyze brain connectivity

Volume Electron Microscopy Analysis of Synapses in Associative and Primary Regions of the Human Cerebral Cortex

Unambiguous identification of asymmetric and symmetric synapses using volume electron microscopy

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