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Advances in imaging ultrastructure yield new insights into presynaptic biology.

Bruckner JJ, Zhan H, O'Connor-Giles KM - Front Cell Neurosci (2015)

Bottom Line: At presynaptic terminals, neurotransmitter-filled synaptic vesicles are released in response to calcium influx through voltage-gated calcium channels activated by the arrival of an action potential.Decades of electrophysiological, biochemical, and genetic studies have contributed to a growing understanding of presynaptic biology.The development of techniques for rapid immobilization and preservation of neuronal tissues for electron microscopy (EM) has led to a new renaissance in ultrastructural imaging that is rapidly advancing our understanding of synapse structure and function.

View Article: PubMed Central - PubMed

Affiliation: Cell and Molecular Biology Training Program, University of Wisconsin-Madison Madison, WI, USA.

ABSTRACT
Synapses are the fundamental functional units of neural circuits, and their dysregulation has been implicated in diverse neurological disorders. At presynaptic terminals, neurotransmitter-filled synaptic vesicles are released in response to calcium influx through voltage-gated calcium channels activated by the arrival of an action potential. Decades of electrophysiological, biochemical, and genetic studies have contributed to a growing understanding of presynaptic biology. Imaging studies are yielding new insights into how synapses are organized to carry out their critical functions. The development of techniques for rapid immobilization and preservation of neuronal tissues for electron microscopy (EM) has led to a new renaissance in ultrastructural imaging that is rapidly advancing our understanding of synapse structure and function.

No MeSH data available.


Related in: MedlinePlus

Structure-function relationships of the presynaptic terminal. (A) Diverse presynaptic terminals have a number of common structural characteristics visible in electron micrographs. The active zone (AZ) membrane is delineated by its electron-dense lipid bilayer. Complex cytoskeletal filaments project from the AZ membrane into the presynaptic cytoplasm and are often visible as an electron-dense projection. SVs are 40–60 nm in diameter and organized into three functionally defined pools: the reserve pool (purple), recycling pool (blue), and readily releasable pool (RRP; red). SVs of the reserve and recycling pools are typically linked to one another by 2–3 thin proteinaceous tethers 30–40 nm in length, and occasionally linked to the AZ membrane by longer filaments of roughly 60 nm in length. The reserve and recycling pools are morphologically intermixed and therefore defined primarily by their mobility in functional assays. RRP vesicles are tethered or docked at the membrane in close proximity to clusters of voltage-gated calcium channels at the base of the dense projection. Three modes of endocytosis are hypothesized for recovery of SVs following exocytosis: clathrin-mediated endocytosis (CME), kiss-and-run, and ultrafast endocytosis. The newly described ultrafast endocytosis involves the formation of 80-nm diameter vesicular intermediates within 50–100 ms after stimulus that fuse with early endosomal compartments within 1 s after stimulus. SVs are then reformed from the early endosome in a clathrin-dependent manner 3–5 s post stimulus. (B) The RRP includes SVs tethered to the AZ membrane by short filaments 5–25 nm in length and SVs in direct contact with the membrane. Although the exact molecular composition of SV tethers is unknown, some of the AZ proteins responsible for regulating SV tethering and docking/priming are known. (C) Cryopreservation of synapses reveals the morphological intricacies of dense projection structure previously masked by chemical fixation and dehydration. Although the unique functional requirements of distinct synapses within and between species likely underlie observed differences in morphology, most Dense projections (DPs) comprise a central core and radiating filaments of varying lengths that contact of distinct functional pools.
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Figure 1: Structure-function relationships of the presynaptic terminal. (A) Diverse presynaptic terminals have a number of common structural characteristics visible in electron micrographs. The active zone (AZ) membrane is delineated by its electron-dense lipid bilayer. Complex cytoskeletal filaments project from the AZ membrane into the presynaptic cytoplasm and are often visible as an electron-dense projection. SVs are 40–60 nm in diameter and organized into three functionally defined pools: the reserve pool (purple), recycling pool (blue), and readily releasable pool (RRP; red). SVs of the reserve and recycling pools are typically linked to one another by 2–3 thin proteinaceous tethers 30–40 nm in length, and occasionally linked to the AZ membrane by longer filaments of roughly 60 nm in length. The reserve and recycling pools are morphologically intermixed and therefore defined primarily by their mobility in functional assays. RRP vesicles are tethered or docked at the membrane in close proximity to clusters of voltage-gated calcium channels at the base of the dense projection. Three modes of endocytosis are hypothesized for recovery of SVs following exocytosis: clathrin-mediated endocytosis (CME), kiss-and-run, and ultrafast endocytosis. The newly described ultrafast endocytosis involves the formation of 80-nm diameter vesicular intermediates within 50–100 ms after stimulus that fuse with early endosomal compartments within 1 s after stimulus. SVs are then reformed from the early endosome in a clathrin-dependent manner 3–5 s post stimulus. (B) The RRP includes SVs tethered to the AZ membrane by short filaments 5–25 nm in length and SVs in direct contact with the membrane. Although the exact molecular composition of SV tethers is unknown, some of the AZ proteins responsible for regulating SV tethering and docking/priming are known. (C) Cryopreservation of synapses reveals the morphological intricacies of dense projection structure previously masked by chemical fixation and dehydration. Although the unique functional requirements of distinct synapses within and between species likely underlie observed differences in morphology, most Dense projections (DPs) comprise a central core and radiating filaments of varying lengths that contact of distinct functional pools.

Mentions: Early electron microscopy (EM) studies first identified many of the now well-known features of synapses, including SVs and post-synaptic densities (De Robertis and Bennett, 1955; Palay and Palade, 1955). Although synaptic ultrastructure varies across species and neuronal subtype, the general functions described above are shared among all synapses and generate commonalities in synaptic architecture (Zhai and Bellen, 2004). Neurotransmitter release is accomplished through a complex network of molecules at the presynaptic active zone (AZ; Couteaux and Pécot-Dechavassine, 1970; Südhof, 2012). The AZ provides both a structural and molecular foundation for synaptic activity, and is identifiable in all synapses as a strip of electron-dense presynaptic plasma membrane tightly apposed to a postsynaptic electron density comprising neurotransmitter receptors and associated cytoskeletal proteins. The presynaptic terminal also contains large clusters of SVs sorted into distinct pools, commonly subdivided into the following: (1) the reserve pool that maintains release during prolonged activity, (2) the recycling pool; and (3) the readily-releasable pool (RRP), which can be further subdivided into SVs “tethered” to the AZ membrane by short filaments and “docked” SVs in direct contact with the AZ membrane absent any visible tethers. The RRP is defined as SVs that are released upon mechanical stimulation with hypertonic sucrose and thought to represent the population accessed upon Ca2+ influx during normal physiological activity (Figures 1A,B; Rosenmund and Stevens, 1996).


Advances in imaging ultrastructure yield new insights into presynaptic biology.

Bruckner JJ, Zhan H, O'Connor-Giles KM - Front Cell Neurosci (2015)

Structure-function relationships of the presynaptic terminal. (A) Diverse presynaptic terminals have a number of common structural characteristics visible in electron micrographs. The active zone (AZ) membrane is delineated by its electron-dense lipid bilayer. Complex cytoskeletal filaments project from the AZ membrane into the presynaptic cytoplasm and are often visible as an electron-dense projection. SVs are 40–60 nm in diameter and organized into three functionally defined pools: the reserve pool (purple), recycling pool (blue), and readily releasable pool (RRP; red). SVs of the reserve and recycling pools are typically linked to one another by 2–3 thin proteinaceous tethers 30–40 nm in length, and occasionally linked to the AZ membrane by longer filaments of roughly 60 nm in length. The reserve and recycling pools are morphologically intermixed and therefore defined primarily by their mobility in functional assays. RRP vesicles are tethered or docked at the membrane in close proximity to clusters of voltage-gated calcium channels at the base of the dense projection. Three modes of endocytosis are hypothesized for recovery of SVs following exocytosis: clathrin-mediated endocytosis (CME), kiss-and-run, and ultrafast endocytosis. The newly described ultrafast endocytosis involves the formation of 80-nm diameter vesicular intermediates within 50–100 ms after stimulus that fuse with early endosomal compartments within 1 s after stimulus. SVs are then reformed from the early endosome in a clathrin-dependent manner 3–5 s post stimulus. (B) The RRP includes SVs tethered to the AZ membrane by short filaments 5–25 nm in length and SVs in direct contact with the membrane. Although the exact molecular composition of SV tethers is unknown, some of the AZ proteins responsible for regulating SV tethering and docking/priming are known. (C) Cryopreservation of synapses reveals the morphological intricacies of dense projection structure previously masked by chemical fixation and dehydration. Although the unique functional requirements of distinct synapses within and between species likely underlie observed differences in morphology, most Dense projections (DPs) comprise a central core and radiating filaments of varying lengths that contact of distinct functional pools.
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Figure 1: Structure-function relationships of the presynaptic terminal. (A) Diverse presynaptic terminals have a number of common structural characteristics visible in electron micrographs. The active zone (AZ) membrane is delineated by its electron-dense lipid bilayer. Complex cytoskeletal filaments project from the AZ membrane into the presynaptic cytoplasm and are often visible as an electron-dense projection. SVs are 40–60 nm in diameter and organized into three functionally defined pools: the reserve pool (purple), recycling pool (blue), and readily releasable pool (RRP; red). SVs of the reserve and recycling pools are typically linked to one another by 2–3 thin proteinaceous tethers 30–40 nm in length, and occasionally linked to the AZ membrane by longer filaments of roughly 60 nm in length. The reserve and recycling pools are morphologically intermixed and therefore defined primarily by their mobility in functional assays. RRP vesicles are tethered or docked at the membrane in close proximity to clusters of voltage-gated calcium channels at the base of the dense projection. Three modes of endocytosis are hypothesized for recovery of SVs following exocytosis: clathrin-mediated endocytosis (CME), kiss-and-run, and ultrafast endocytosis. The newly described ultrafast endocytosis involves the formation of 80-nm diameter vesicular intermediates within 50–100 ms after stimulus that fuse with early endosomal compartments within 1 s after stimulus. SVs are then reformed from the early endosome in a clathrin-dependent manner 3–5 s post stimulus. (B) The RRP includes SVs tethered to the AZ membrane by short filaments 5–25 nm in length and SVs in direct contact with the membrane. Although the exact molecular composition of SV tethers is unknown, some of the AZ proteins responsible for regulating SV tethering and docking/priming are known. (C) Cryopreservation of synapses reveals the morphological intricacies of dense projection structure previously masked by chemical fixation and dehydration. Although the unique functional requirements of distinct synapses within and between species likely underlie observed differences in morphology, most Dense projections (DPs) comprise a central core and radiating filaments of varying lengths that contact of distinct functional pools.
Mentions: Early electron microscopy (EM) studies first identified many of the now well-known features of synapses, including SVs and post-synaptic densities (De Robertis and Bennett, 1955; Palay and Palade, 1955). Although synaptic ultrastructure varies across species and neuronal subtype, the general functions described above are shared among all synapses and generate commonalities in synaptic architecture (Zhai and Bellen, 2004). Neurotransmitter release is accomplished through a complex network of molecules at the presynaptic active zone (AZ; Couteaux and Pécot-Dechavassine, 1970; Südhof, 2012). The AZ provides both a structural and molecular foundation for synaptic activity, and is identifiable in all synapses as a strip of electron-dense presynaptic plasma membrane tightly apposed to a postsynaptic electron density comprising neurotransmitter receptors and associated cytoskeletal proteins. The presynaptic terminal also contains large clusters of SVs sorted into distinct pools, commonly subdivided into the following: (1) the reserve pool that maintains release during prolonged activity, (2) the recycling pool; and (3) the readily-releasable pool (RRP), which can be further subdivided into SVs “tethered” to the AZ membrane by short filaments and “docked” SVs in direct contact with the AZ membrane absent any visible tethers. The RRP is defined as SVs that are released upon mechanical stimulation with hypertonic sucrose and thought to represent the population accessed upon Ca2+ influx during normal physiological activity (Figures 1A,B; Rosenmund and Stevens, 1996).

Bottom Line: At presynaptic terminals, neurotransmitter-filled synaptic vesicles are released in response to calcium influx through voltage-gated calcium channels activated by the arrival of an action potential.Decades of electrophysiological, biochemical, and genetic studies have contributed to a growing understanding of presynaptic biology.The development of techniques for rapid immobilization and preservation of neuronal tissues for electron microscopy (EM) has led to a new renaissance in ultrastructural imaging that is rapidly advancing our understanding of synapse structure and function.

View Article: PubMed Central - PubMed

Affiliation: Cell and Molecular Biology Training Program, University of Wisconsin-Madison Madison, WI, USA.

ABSTRACT
Synapses are the fundamental functional units of neural circuits, and their dysregulation has been implicated in diverse neurological disorders. At presynaptic terminals, neurotransmitter-filled synaptic vesicles are released in response to calcium influx through voltage-gated calcium channels activated by the arrival of an action potential. Decades of electrophysiological, biochemical, and genetic studies have contributed to a growing understanding of presynaptic biology. Imaging studies are yielding new insights into how synapses are organized to carry out their critical functions. The development of techniques for rapid immobilization and preservation of neuronal tissues for electron microscopy (EM) has led to a new renaissance in ultrastructural imaging that is rapidly advancing our understanding of synapse structure and function.

No MeSH data available.


Related in: MedlinePlus