Related Experiment Video
Updated: Aug 1, 2026
Assessment of Ultrastructural Neuroplasticity Parameters After In Utero Transduction of the Developing Mouse Brain and Spinal Cord
Published on: February 26, 2019
Field emission scanning electron microscopy and freeze-fracture transmission electron microscopy of mouse cerebellar
O J Castejón1, R P Apkarian, H V Castejón
1Institute of Biological Investigations Dres. Orlando J. Castejon y Haydee Viloria de Castejón, Faculty of Medicine, University of Zulia, Maracaibo, Venezuela. ocastejo@cantv.net
Abstract:
Samples of albino mice were processed by the cryofracture method for scanning electron microscopy and examined with the field emission scanning electron microscope (FESEM). Freeze-etching direct replicas of mice cerebellar cortex were also studied with the transmission electron microscope (FFTEM), as a complementary technique for obtaining higher resolution, three-dimensional correlative images of cerebellar synaptic contacts. At the granular, Purkinje cells and molecular layers, the cryofracture method for FESEM selectively removed the neuroglial cell investment, facilitating the visualization of the outer and inner surfaces of cerebellar synaptic contacts. In addition, FFTEM showed the real extension of perisynaptic neuroglial investment. The outer surface of mossy fiber rosettes and their digitiform processes were seen at the granular layer, making flat and invaginated synaptic contacts with the granule cell dendrites. At the molecular layer, the longitudinal traject of parallel fibers or nonsynaptic segments and their synaptic varicosities were characterized. These latter established synaptic contacts with Purkinje dendritic spines. Fractured parallel fiber endings showed the SE-I images of clustered spheroidal synaptic vesicles and mitochondria and the surrounding cotton-like appearance of Bergmann glial cell cytoplasm. Climbing fibers showed a characteristic crossing-over bifurcation pattern in the white matter and in the three-layer structure of cerebellar cortex, formation of tendril collaterals in the granular layer, topographical relationship with Purkinje cell soma and retrograde collaterals in the molecular layer. The climbing fiber synaptic relationship with Purkinje dendritic spines was characterized, by means of FFTEM, by the presence of large synaptic endings and aggregation of intramembrane particles at the P and E faces of presynaptic endings, characteristic of excitatory synapses.
Insights
Cryofracture and freeze-etching electron microscopy revealed detailed cerebellar synaptic structures in mice. This technique visualized synaptic contacts between neurons and glial cells, offering new insights into cerebellar circuitry.
Area of Science:
- Neuroscience
- Cell Biology
- Microscopy
Background:
- Understanding the intricate three-dimensional structure of cerebellar synaptic contacts is crucial for deciphering neural circuit function.
- Previous imaging techniques have limitations in resolving the fine details of synaptic architecture and glial-neuronal interactions.
Purpose of the Study:
- To visualize and characterize cerebellar synaptic contacts in albino mice using cryofracture for field emission scanning electron microscopy (FESEM) and freeze-etching transmission electron microscopy (FFTEM).
- To obtain higher resolution, three-dimensional correlative images of synaptic structures and their surrounding neuroglial investment.
Main Methods:
- Cryofracture method applied to albino mouse cerebellar samples for FESEM.
- Freeze-etching direct replicas of cerebellar cortex for FFTEM.
- Correlative imaging to combine high-resolution 3D structural information.
Main Results:
- Cryofracture FESEM facilitated visualization of synaptic contact surfaces by removing neuroglial cells.
- FFTEM revealed the extent of perisynaptic neuroglial investment and detailed synaptic structures, including mossy fiber rosettes, parallel fibers, and climbing fibers.
- Specific synaptic relationships were characterized, such as parallel fibers with Purkinje dendritic spines and climbing fibers with Purkinje dendritic spines, noting features of excitatory synapses.
Conclusions:
- The combined cryofracture-FESEM and freeze-etching FFTEM techniques provide unprecedented detail of cerebellar synaptic organization.
- These methods allow for the clear visualization of neuronal and glial components at synaptic sites, enhancing our understanding of cerebellar circuitry.
