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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

A highly conserved network of proteins organizes presynaptic function. Presynaptic proteins can be grouped into zones within the AZ based on studies of their location. Although the structural complexity of the presynapse is not illustrated (see Figure 1), it can be generally divided into the dense projection (orange), an AZ-proximal zone extending to approximately 50 nm from the membrane (green), an AZ-distal zone extending to approximately 100 nm from the membrane (blue), the peri-active zone beyond the electron dense AZ membrane (gray), and SVs that may transiently occupy any of the other zones. Presynaptic proteins discussed in this review are grouped according to their predominant localization and described in detail. For each molecule, conservation in multiple species is indicated with the gene name from humans, flies, and worms. Links between presynaptic proteins and human neurological disorders are summarized.
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Figure 2: A highly conserved network of proteins organizes presynaptic function. Presynaptic proteins can be grouped into zones within the AZ based on studies of their location. Although the structural complexity of the presynapse is not illustrated (see Figure 1), it can be generally divided into the dense projection (orange), an AZ-proximal zone extending to approximately 50 nm from the membrane (green), an AZ-distal zone extending to approximately 100 nm from the membrane (blue), the peri-active zone beyond the electron dense AZ membrane (gray), and SVs that may transiently occupy any of the other zones. Presynaptic proteins discussed in this review are grouped according to their predominant localization and described in detail. For each molecule, conservation in multiple species is indicated with the gene name from humans, flies, and worms. Links between presynaptic proteins and human neurological disorders are summarized.

Mentions: The precise organization and exact function of DPs remain enigmatic. However, a highly conserved network of interacting proteins comprising the CAZ has been well defined in many model organisms, including Piccolo/Aczonin, Bassoon, Rab3 interacting molecules (RIMs), Unc-13, RIM binding protein (RBP), liprin-α, and CAST/ELKS/ERC (Figure 2; Dresbach et al., 2001; Rosenmund et al., 2003; Zhai and Bellen, 2004; Jin and Garner, 2008; Bruckner et al., 2012). A large body of work has furthered our understanding of how each component contributes functionally to the behavior of the synapse and is reviewed elsewhere (Haucke et al., 2011; Gundelfinger and Fejtova, 2012). However, a detailed understanding of CAZ morphology has been hindered by the detection limits of conventional light microscopy where the diffraction limit of light restricts lateral resolution to between 200 and 300 nm and axial resolution to 500 nm (Abbe, 1873; Pawley, 2010; Bianchini et al., 2015). To bypass the diffraction limit and achieve nanometer resolution, both immuno-EM and super-resolution microscopy have been applied to the question of protein localization within the presynaptic CAZ. Stochastic optical reconstruction microscopy (STORM) and the related technique photo-activated light microscopy (PALM) take advantage of “on” and “off” states of photo-switchable fluorophores to temporally distribute localization information and computationally reconstruct the center of each diffraction spot (Betzig et al., 2006; Rust et al., 2006). STORM was used to establish the axial distribution of synaptic proteins within the presynaptic terminal in mouse olfactory bulb and cortex synapses, where RIM1 was found between 20 and 50 nm from the presynaptic membrane and the related Piccolo and Bassoon proteins were detected between 20 and 100 nm from the membrane (Dani et al., 2010). Combining immuno-gold labeling and HPF/FS EM has enabled localization of synaptic proteins with nanometer resolution in an ultrastructural context (Rostaing et al., 2006; Weimer et al., 2006). Such a study in cerebellar synapses found that RIM1 and Munc-13 localize closest to the plasma membrane at approximately 20 nm, while Piccolo adopts an L-shaped conformation at the tip of the DP where it extends to cover the distal AZ between 40 and 80 nm from the synaptic membrane (Limbach et al., 2011). In Drosophila, immuno-EM detecting two distinct epitopes of Bruchpilot demonstrated that it adopts an elongated conformation at the DP core (Fouquet et al., 2009). These data inform compelling hypotheses about the role of these proteins in synaptic function, where Piccolo and Bassoon regulate SV trafficking at the AZ perimeter; Munc13–1, RIM1, and CAST1 have a direct role in priming in support of SV release; and CAST/ELKS/ERC/Bruchpilot bridges these two compartments.


Advances in imaging ultrastructure yield new insights into presynaptic biology.

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

A highly conserved network of proteins organizes presynaptic function. Presynaptic proteins can be grouped into zones within the AZ based on studies of their location. Although the structural complexity of the presynapse is not illustrated (see Figure 1), it can be generally divided into the dense projection (orange), an AZ-proximal zone extending to approximately 50 nm from the membrane (green), an AZ-distal zone extending to approximately 100 nm from the membrane (blue), the peri-active zone beyond the electron dense AZ membrane (gray), and SVs that may transiently occupy any of the other zones. Presynaptic proteins discussed in this review are grouped according to their predominant localization and described in detail. For each molecule, conservation in multiple species is indicated with the gene name from humans, flies, and worms. Links between presynaptic proteins and human neurological disorders are summarized.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4440913&req=5

Figure 2: A highly conserved network of proteins organizes presynaptic function. Presynaptic proteins can be grouped into zones within the AZ based on studies of their location. Although the structural complexity of the presynapse is not illustrated (see Figure 1), it can be generally divided into the dense projection (orange), an AZ-proximal zone extending to approximately 50 nm from the membrane (green), an AZ-distal zone extending to approximately 100 nm from the membrane (blue), the peri-active zone beyond the electron dense AZ membrane (gray), and SVs that may transiently occupy any of the other zones. Presynaptic proteins discussed in this review are grouped according to their predominant localization and described in detail. For each molecule, conservation in multiple species is indicated with the gene name from humans, flies, and worms. Links between presynaptic proteins and human neurological disorders are summarized.
Mentions: The precise organization and exact function of DPs remain enigmatic. However, a highly conserved network of interacting proteins comprising the CAZ has been well defined in many model organisms, including Piccolo/Aczonin, Bassoon, Rab3 interacting molecules (RIMs), Unc-13, RIM binding protein (RBP), liprin-α, and CAST/ELKS/ERC (Figure 2; Dresbach et al., 2001; Rosenmund et al., 2003; Zhai and Bellen, 2004; Jin and Garner, 2008; Bruckner et al., 2012). A large body of work has furthered our understanding of how each component contributes functionally to the behavior of the synapse and is reviewed elsewhere (Haucke et al., 2011; Gundelfinger and Fejtova, 2012). However, a detailed understanding of CAZ morphology has been hindered by the detection limits of conventional light microscopy where the diffraction limit of light restricts lateral resolution to between 200 and 300 nm and axial resolution to 500 nm (Abbe, 1873; Pawley, 2010; Bianchini et al., 2015). To bypass the diffraction limit and achieve nanometer resolution, both immuno-EM and super-resolution microscopy have been applied to the question of protein localization within the presynaptic CAZ. Stochastic optical reconstruction microscopy (STORM) and the related technique photo-activated light microscopy (PALM) take advantage of “on” and “off” states of photo-switchable fluorophores to temporally distribute localization information and computationally reconstruct the center of each diffraction spot (Betzig et al., 2006; Rust et al., 2006). STORM was used to establish the axial distribution of synaptic proteins within the presynaptic terminal in mouse olfactory bulb and cortex synapses, where RIM1 was found between 20 and 50 nm from the presynaptic membrane and the related Piccolo and Bassoon proteins were detected between 20 and 100 nm from the membrane (Dani et al., 2010). Combining immuno-gold labeling and HPF/FS EM has enabled localization of synaptic proteins with nanometer resolution in an ultrastructural context (Rostaing et al., 2006; Weimer et al., 2006). Such a study in cerebellar synapses found that RIM1 and Munc-13 localize closest to the plasma membrane at approximately 20 nm, while Piccolo adopts an L-shaped conformation at the tip of the DP where it extends to cover the distal AZ between 40 and 80 nm from the synaptic membrane (Limbach et al., 2011). In Drosophila, immuno-EM detecting two distinct epitopes of Bruchpilot demonstrated that it adopts an elongated conformation at the DP core (Fouquet et al., 2009). These data inform compelling hypotheses about the role of these proteins in synaptic function, where Piccolo and Bassoon regulate SV trafficking at the AZ perimeter; Munc13–1, RIM1, and CAST1 have a direct role in priming in support of SV release; and CAST/ELKS/ERC/Bruchpilot bridges these two compartments.

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