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Mapping Synaptic Pathology within Cerebral Cortical Circuits in Subjects with Schizophrenia.

Sweet RA, Fish KN, Lewis DA - Front Hum Neurosci (2010)

Bottom Line: Efforts to localize these alterations in brain tissue from subjects with schizophrenia have frequently been limited to the quantification of structures that are non-selectively identified (e.g., dendritic spines labeled in Golgi preparations, axon boutons labeled with synaptophysin), or to quantification of proteins using methods unable to resolve relevant cellular compartments.An important adaptation required for studies of human disease is coupling this approach to stereologic methods for systematic random sampling of relevant brain regions.In this context, we provide examples of the examination of pre- and post-synaptic structures within excitatory and inhibitory circuits of the cerebral cortex.

View Article: PubMed Central - PubMed

Affiliation: Translational Neuroscience Program, Department of Psychiatry, University of Pittsburgh Pittsburgh, PA, USA.

ABSTRACT
Converging lines of evidence indicate that schizophrenia is characterized by impairments of synaptic machinery within cerebral cortical circuits. Efforts to localize these alterations in brain tissue from subjects with schizophrenia have frequently been limited to the quantification of structures that are non-selectively identified (e.g., dendritic spines labeled in Golgi preparations, axon boutons labeled with synaptophysin), or to quantification of proteins using methods unable to resolve relevant cellular compartments. Multiple label fluorescence confocal microscopy represents a means to circumvent many of these limitations, by concurrently extracting information regarding the number, morphology, and relative protein content of synaptic structures. An important adaptation required for studies of human disease is coupling this approach to stereologic methods for systematic random sampling of relevant brain regions. In this review article we consider the application of multiple label fluorescence confocal microscopy to the mapping of synaptic alterations in subjects with schizophrenia and describe the application of a novel, readily automated, iterative intensity/morphological segmentation algorithm for the extraction of information regarding synaptic structure number, size, and relative protein level from tissue sections obtained using unbiased stereological principles of sampling. In this context, we provide examples of the examination of pre- and post-synaptic structures within excitatory and inhibitory circuits of the cerebral cortex.

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Related in: MedlinePlus

Parvalbumin-positive basket cell boutons apposing GABAA α1 subunits in the plasma membrane of neurons. (A) In a single confocal z-plane GABAA α1 (green) is seen in the plasma membrane of a PV (red) neuron in a tissue section. (B) Color segmentation masks of the image in (A) which represents the intracellular PV (blue), PV boutons (red) and GABAA α1 clusters (green). The different masks were made using different parameters for our iterative approach. Only the red and green object masks that partially overlap, and thus correspond to PV puncta and α1 clusters that are directly apposed, are shown. The arrowheads point to α1 clusters that are within the PV neuron and are directly apposed by PV puncta. Note that in (A) several PV puncta appear to juxtapose the soma near the open arrowhead. However, in this z-plane only one is directly apposed to an α1 cluster (open arrowhead). (C) A single confocal z-plane of a tissue section was labeled for GABAA α1 subunit (green), GAD65 (red), PV (blue), and Nissl substance (gray). Note GAD65 and PV dual-labeled (purple) boutons (white arrows) and GAD65 labeled boutons (black arrows) in apposition to GABAA α1 subunits in the plasma membrane of a pyramidal cell. The open arrowhead indicates an α1 cluster for which the apposed bouton is below the plane of focus. All Bars = 5 μm.
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Figure 6: Parvalbumin-positive basket cell boutons apposing GABAA α1 subunits in the plasma membrane of neurons. (A) In a single confocal z-plane GABAA α1 (green) is seen in the plasma membrane of a PV (red) neuron in a tissue section. (B) Color segmentation masks of the image in (A) which represents the intracellular PV (blue), PV boutons (red) and GABAA α1 clusters (green). The different masks were made using different parameters for our iterative approach. Only the red and green object masks that partially overlap, and thus correspond to PV puncta and α1 clusters that are directly apposed, are shown. The arrowheads point to α1 clusters that are within the PV neuron and are directly apposed by PV puncta. Note that in (A) several PV puncta appear to juxtapose the soma near the open arrowhead. However, in this z-plane only one is directly apposed to an α1 cluster (open arrowhead). (C) A single confocal z-plane of a tissue section was labeled for GABAA α1 subunit (green), GAD65 (red), PV (blue), and Nissl substance (gray). Note GAD65 and PV dual-labeled (purple) boutons (white arrows) and GAD65 labeled boutons (black arrows) in apposition to GABAA α1 subunits in the plasma membrane of a pyramidal cell. The open arrowhead indicates an α1 cluster for which the apposed bouton is below the plane of focus. All Bars = 5 μm.

Mentions: As summarized in Section “Dorsolateral Prefrontal Cortex”, multiple lines of evidence indicate abnormalities of the PV+ class of GABA neurons in the DLPFC of subjects with schizophrenia. Importantly, the PV+ GABA neurons comprise two functionally and morphologically distinct subgroups: chandelier neurons whose axon boutons form distinctive arrays (cartridges) surrounding the axon initial segment of pyramidal cells and basket cells, whose boutons surround neuron cell bodies (Figure 5A). Because both chandelier cell cartridge boutons and PV+ basket cell boutons can be readily masked using iterative segmentation (Figures 5B–D) quantitative comparisons of relative fluorescence of proteins (e.g., GAD67) in these PV+ GABA bouton types in normal individuals, and in subjects with schizophrenia, can be accomplished. Moreover, within the PV+ basket cell subtype, boutons may target either pyramidal cell bodies, or the cell bodies of other inhibitory neurons. These can be distinguished via multiple label immunofluorescence (Figure 6), and the pre- and post-synaptic components masked for quantification of number and/or relative fluorescence intensity.


Mapping Synaptic Pathology within Cerebral Cortical Circuits in Subjects with Schizophrenia.

Sweet RA, Fish KN, Lewis DA - Front Hum Neurosci (2010)

Parvalbumin-positive basket cell boutons apposing GABAA α1 subunits in the plasma membrane of neurons. (A) In a single confocal z-plane GABAA α1 (green) is seen in the plasma membrane of a PV (red) neuron in a tissue section. (B) Color segmentation masks of the image in (A) which represents the intracellular PV (blue), PV boutons (red) and GABAA α1 clusters (green). The different masks were made using different parameters for our iterative approach. Only the red and green object masks that partially overlap, and thus correspond to PV puncta and α1 clusters that are directly apposed, are shown. The arrowheads point to α1 clusters that are within the PV neuron and are directly apposed by PV puncta. Note that in (A) several PV puncta appear to juxtapose the soma near the open arrowhead. However, in this z-plane only one is directly apposed to an α1 cluster (open arrowhead). (C) A single confocal z-plane of a tissue section was labeled for GABAA α1 subunit (green), GAD65 (red), PV (blue), and Nissl substance (gray). Note GAD65 and PV dual-labeled (purple) boutons (white arrows) and GAD65 labeled boutons (black arrows) in apposition to GABAA α1 subunits in the plasma membrane of a pyramidal cell. The open arrowhead indicates an α1 cluster for which the apposed bouton is below the plane of focus. All Bars = 5 μm.
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Figure 6: Parvalbumin-positive basket cell boutons apposing GABAA α1 subunits in the plasma membrane of neurons. (A) In a single confocal z-plane GABAA α1 (green) is seen in the plasma membrane of a PV (red) neuron in a tissue section. (B) Color segmentation masks of the image in (A) which represents the intracellular PV (blue), PV boutons (red) and GABAA α1 clusters (green). The different masks were made using different parameters for our iterative approach. Only the red and green object masks that partially overlap, and thus correspond to PV puncta and α1 clusters that are directly apposed, are shown. The arrowheads point to α1 clusters that are within the PV neuron and are directly apposed by PV puncta. Note that in (A) several PV puncta appear to juxtapose the soma near the open arrowhead. However, in this z-plane only one is directly apposed to an α1 cluster (open arrowhead). (C) A single confocal z-plane of a tissue section was labeled for GABAA α1 subunit (green), GAD65 (red), PV (blue), and Nissl substance (gray). Note GAD65 and PV dual-labeled (purple) boutons (white arrows) and GAD65 labeled boutons (black arrows) in apposition to GABAA α1 subunits in the plasma membrane of a pyramidal cell. The open arrowhead indicates an α1 cluster for which the apposed bouton is below the plane of focus. All Bars = 5 μm.
Mentions: As summarized in Section “Dorsolateral Prefrontal Cortex”, multiple lines of evidence indicate abnormalities of the PV+ class of GABA neurons in the DLPFC of subjects with schizophrenia. Importantly, the PV+ GABA neurons comprise two functionally and morphologically distinct subgroups: chandelier neurons whose axon boutons form distinctive arrays (cartridges) surrounding the axon initial segment of pyramidal cells and basket cells, whose boutons surround neuron cell bodies (Figure 5A). Because both chandelier cell cartridge boutons and PV+ basket cell boutons can be readily masked using iterative segmentation (Figures 5B–D) quantitative comparisons of relative fluorescence of proteins (e.g., GAD67) in these PV+ GABA bouton types in normal individuals, and in subjects with schizophrenia, can be accomplished. Moreover, within the PV+ basket cell subtype, boutons may target either pyramidal cell bodies, or the cell bodies of other inhibitory neurons. These can be distinguished via multiple label immunofluorescence (Figure 6), and the pre- and post-synaptic components masked for quantification of number and/or relative fluorescence intensity.

Bottom Line: Efforts to localize these alterations in brain tissue from subjects with schizophrenia have frequently been limited to the quantification of structures that are non-selectively identified (e.g., dendritic spines labeled in Golgi preparations, axon boutons labeled with synaptophysin), or to quantification of proteins using methods unable to resolve relevant cellular compartments.An important adaptation required for studies of human disease is coupling this approach to stereologic methods for systematic random sampling of relevant brain regions.In this context, we provide examples of the examination of pre- and post-synaptic structures within excitatory and inhibitory circuits of the cerebral cortex.

View Article: PubMed Central - PubMed

Affiliation: Translational Neuroscience Program, Department of Psychiatry, University of Pittsburgh Pittsburgh, PA, USA.

ABSTRACT
Converging lines of evidence indicate that schizophrenia is characterized by impairments of synaptic machinery within cerebral cortical circuits. Efforts to localize these alterations in brain tissue from subjects with schizophrenia have frequently been limited to the quantification of structures that are non-selectively identified (e.g., dendritic spines labeled in Golgi preparations, axon boutons labeled with synaptophysin), or to quantification of proteins using methods unable to resolve relevant cellular compartments. Multiple label fluorescence confocal microscopy represents a means to circumvent many of these limitations, by concurrently extracting information regarding the number, morphology, and relative protein content of synaptic structures. An important adaptation required for studies of human disease is coupling this approach to stereologic methods for systematic random sampling of relevant brain regions. In this review article we consider the application of multiple label fluorescence confocal microscopy to the mapping of synaptic alterations in subjects with schizophrenia and describe the application of a novel, readily automated, iterative intensity/morphological segmentation algorithm for the extraction of information regarding synaptic structure number, size, and relative protein level from tissue sections obtained using unbiased stereological principles of sampling. In this context, we provide examples of the examination of pre- and post-synaptic structures within excitatory and inhibitory circuits of the cerebral cortex.

No MeSH data available.


Related in: MedlinePlus