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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.

No MeSH data available.


Related in: MedlinePlus

Low power confocal XY montage of human primary auditory cortex. (A) Immunoreactivity for VGlut1 demonstrates intense label of the entire gray matter, consistent with localization in boutons of cortical origin. (B) VGlut2 labels boutons in the thalamic termination zone in layer 4 and deep layer 3. Bar = 150 μm.
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Figure 7: Low power confocal XY montage of human primary auditory cortex. (A) Immunoreactivity for VGlut1 demonstrates intense label of the entire gray matter, consistent with localization in boutons of cortical origin. (B) VGlut2 labels boutons in the thalamic termination zone in layer 4 and deep layer 3. Bar = 150 μm.

Mentions: The prior observations of reduced bouton density in AI were based on identification of boutons using immunoreactivity for synaptophysin, a non-specific marker of most bouton types in cortex. Identifying whether these reductions reflect fewer boutons of intracortical or thalamocortical origin, and the extent to which excitatory and inhibitory boutons are reduced, is of fundamental importance in mapping the circuits involved in schizophrenia pathology. We have developed a multiple label fluorescent microscopic approach using antibodies directed against vesicular glutamate transporter 1 (VGlut1) which in rodent cortex has been shown to selectively label intracortical excitatory boutons, and VGlut2, which selectively labels thalamocortical boutons (Kaneko and Fujiyama, 2002). An example of a confocal micrograph in AI of a human subject labeled for VGlut1 and VGlut 2 and showing the predicted distribution of labeled boutons is shown in Figure 7. A higher power example from another subject is shown in Figure 8, triple labeled for VGlut1, VGlut2, and synaptophysin. It is readily apparent that VGlut1 and VGlut2 each label presynaptic boutons, indicated by the colocalization of each with synaptophysin (Figures 8A–D). An example of how the iterative segmentation procedure to generate object masks can be utilized to extract information on relative protein expression within boutons is shown in Figure 8E, for object masks created using VGlut1. It can be seen that this approach readily confirms both the lack of co-expressed VGlut2 within these boutons, and the range of synaptophysin relative expression within boutons. It is anticipated that this approach can be to map the expression of other presynaptic proteins which impact signaling characteristics within select populations of excitatory boutons in subjects with schizophrenia (Santos et al., 2009).


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

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

Low power confocal XY montage of human primary auditory cortex. (A) Immunoreactivity for VGlut1 demonstrates intense label of the entire gray matter, consistent with localization in boutons of cortical origin. (B) VGlut2 labels boutons in the thalamic termination zone in layer 4 and deep layer 3. Bar = 150 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2903233&req=5

Figure 7: Low power confocal XY montage of human primary auditory cortex. (A) Immunoreactivity for VGlut1 demonstrates intense label of the entire gray matter, consistent with localization in boutons of cortical origin. (B) VGlut2 labels boutons in the thalamic termination zone in layer 4 and deep layer 3. Bar = 150 μm.
Mentions: The prior observations of reduced bouton density in AI were based on identification of boutons using immunoreactivity for synaptophysin, a non-specific marker of most bouton types in cortex. Identifying whether these reductions reflect fewer boutons of intracortical or thalamocortical origin, and the extent to which excitatory and inhibitory boutons are reduced, is of fundamental importance in mapping the circuits involved in schizophrenia pathology. We have developed a multiple label fluorescent microscopic approach using antibodies directed against vesicular glutamate transporter 1 (VGlut1) which in rodent cortex has been shown to selectively label intracortical excitatory boutons, and VGlut2, which selectively labels thalamocortical boutons (Kaneko and Fujiyama, 2002). An example of a confocal micrograph in AI of a human subject labeled for VGlut1 and VGlut 2 and showing the predicted distribution of labeled boutons is shown in Figure 7. A higher power example from another subject is shown in Figure 8, triple labeled for VGlut1, VGlut2, and synaptophysin. It is readily apparent that VGlut1 and VGlut2 each label presynaptic boutons, indicated by the colocalization of each with synaptophysin (Figures 8A–D). An example of how the iterative segmentation procedure to generate object masks can be utilized to extract information on relative protein expression within boutons is shown in Figure 8E, for object masks created using VGlut1. It can be seen that this approach readily confirms both the lack of co-expressed VGlut2 within these boutons, and the range of synaptophysin relative expression within boutons. It is anticipated that this approach can be to map the expression of other presynaptic proteins which impact signaling characteristics within select populations of excitatory boutons in subjects with schizophrenia (Santos et al., 2009).

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