On counting and counting errors

Güillery, R. W.

doi:10.1002/cne.10221

Cited by 359 publications

(365 citation statements)

References 63 publications

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“…1 C1-C3). Brain regions can be approximately recovered by spatial clustering of the ABA (26,28,(29)(30)(31)(32), but estimating the density profile of cell types characterized by their transcription profile can provide a data-driven definition of some brain regions, on which there is no universal agreement (26,28,29).…”

Section: Resultsmentioning

confidence: 99%

Cell-type–based model explaining coexpression patterns of genes in the brain

Grange

Bohland

Okaty

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

125

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Spatial patterns of gene expression in the vertebrate brain are not independent, as pairs of genes can exhibit complex patterns of coexpression. Two genes may be similarly expressed in one region, but differentially expressed in other regions. These correlations have been studied quantitatively, particularly for the Allen Atlas of the adult mouse brain, but their biological meaning remains obscure. We propose a simple model of the coexpression patterns in terms of spatial distributions of underlying cell types and establish its plausibility using independently measured cell-typespecific transcriptomes. The model allows us to predict the spatial distribution of cell types in the mouse brain.neuroscience | bioinformatics | neuroanatomy B rain-wide and genome-wide maps of gene expression are now available (1, 2), due to the development of high-throughput neuroanatomical methods (3-6). This has enabled analysis of the spatial correlation structure of gene expression (7-12). In the Allen Brain Atlas (ABA) of the adult mouse, the brain is divided up into cubic voxels of size 200 μm. The expression energies of up to 20,000 genes in the adult C57BL/6J mouse are given by automatic processing of in situ hybridized (ISH) brain sections, coregistered to the Allen Reference Atlas (ARA) (13). Coexpression of two genes in a voxel may arise from two sources: ðiÞ both genes are expressed within the same cell type or ðiiÞ the two genes are expressed in two different cell types, both present in the voxel. These two possibilities cannot be disentangled solely on the basis of the ABA. Ideally, gene expression profiles should be experimentally obtained for each cell type in the brain, and indeed such transcriptome profiles are now available (14-21). This cell-based approach to the study of gene expression is complementary to the gene-based approach of the ABA. In ref. 22, the data of ref. 19 were used to extract neuron-specific genes, astrocyte-specific genes, and oligodendrocyte-specific genes, which resulted in three combinations of brain-wide maps from the ABA, whose clustering showed anatomical signatures of major brain subdivisions (see also ref. 23 for estimates of neuron-specific and oligodendrocyte-specific expression patterns, both in the mouse and in the human brain). The present paper goes beyond the broad classification of cell types into three classes and attempts to estimate density profiles of every cell type known by its transcriptome profile. To study the genes collectively we use a voxel-by-gene data matrix E, corresponding to V = 49;742 cubic voxels, and 3,041 genes, as in refs. 24-26. The entry Eðv; gÞ is the expression energy of the gene labeled g in the voxel labeled v [a measure representing the level of mRNA in situ hybridization (1, 10)]. We combine the ABA with cell-type-specific transcriptome profiles, to gain biological understanding of the coexpression patterns of the genes. Our model is based on G = 2;131 genes found in all these datasets and in the coronal ABA. The model proposed to estimate the brain-w...

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Section: Resultsmentioning

confidence: 99%

Cell-type–based model explaining coexpression patterns of genes in the brain

Grange

Bohland

Okaty

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

125

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“…The analysis was performed as described previously (Polgár et al, 2004) by using a modification of the optical disector method (Williams and Rakic, 1988;Coggeshall, 1992;Bjugn and Gundersen, 1993;West, 1999;Guillery, 2002) to provide unbiased counts of neuronal nuclei in laminas I-III. Briefly, merged images representing NeuN and propidium iodide from the z-series obtained with the 60ϫ objective were viewed with Neurolucida for Confocal (MicroBrightField, Colchester, VT).…”

Section: Methodsmentioning

confidence: 99%

Loss of Neurons from Laminas I-III of the Spinal Dorsal Horn Is Not Required for Development of Tactile Allodynia in the Spared Nerve Injury Model of Neuropathic Pain

Polgár¹,

Hughes²,

Arham³

et al. 2005

J. Neurosci.

128

108

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It has been proposed that death of inhibitory interneurons in the dorsal horn contributes to the neuropathic pain that follows partial nerve injury. In this study, we have used two approaches to test whether there is neuronal death in the dorsal horn in the spared nerve injury (SNI) model. We performed a stereological analysis of the packing density of neurons in laminas I-III 4 weeks after operation and found no reduction on the ipsilateral side compared with that seen on the contralateral side or in sham-operated or naive rats. In addition, we used two markers of apoptosis, terminal deoxynucleotidyl transferase-mediated biotinylated UTP nick end labeling (TUNEL) staining and immunocytochemical detection of cleaved (activated) caspase-3. Neither of these methods demonstrated apoptotic neurons in the dorsal spinal cord 1 week after operation. Although TUNEL-positive cells were present throughout the gray and white matter at this stage, they were virtually all labeled with antibody against ionized calcium-binding adapter molecule 1, a marker for microglia. All animals that underwent SNI showed clear signs of tactile allodynia affecting the ipsilateral hindpaw. These results suggest that a significant loss of neurons from the dorsal horn is not necessary for the development of tactile allodynia in the SNI model.

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“…Although counting all objects within a circumscribed area is a valid and efficient method, it is prone to over-counting. To minimize over-counting, we used the correction factor first computed by Abercrombie (Abercrombie, 1946;Duale et al, 2009;Guillery, 2002;Hou et al, 2008). The photomicrographs in all figures were optimized for final production by adjusting only the brightness and contrast using Adobe Photoshop 7.0 (Adobe Systems Inc., San Jose, CA).…”

Section: Quantification Of Prv-152-positive Cells In the Spinal Cord mentioning

confidence: 99%

Noxious Colorectal Distention in Spinalized Rats Reduces Pseudorabies Virus Labeling of Sympathetic Neurons

Duale

Lyttle²,

Smith³

et al. 2010

Journal of Neurotrauma

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The retrograde transsynaptic tracer pseudorabies virus (PRV) has been widely used as a marker for synaptic connectivity in the spinal cord. Notably, the PRV-152 construct expresses enhanced green fluorescent protein (EGFP). We recently reported a significant attenuation of PRV-152 labeling of the intermediolateral cell column (IML) and celiac ganglia after complete T4 spinal cord transection versus sham injury in rats at 96 h after PRV-152 inoculation of the left kidney. Here we found a significant increase in noxious colorectal distention (CRD)-evoked c-Fos expression in spinal cords of injured versus sham rats without PRV infection. In order to assess whether enhancing neuronal activity in spinalized rats might increase PRV-152 labeling, we subjected awake spinalized rats to 1.5 h of intermittent noxious CRD either: (1) just prior to inoculation, or (2) 96 h after inoculation (n ¼ 3/group). Equal numbers of spinalized rats in both groups received PRV-152 inoculations without CRD (non-stimulated; n ¼ 3/group). At 96 h post-inoculation fixed spinal cords and left celiac ganglionic tissues were assessed for the distribution and quantification of EGFP-labeled cells. The injured cohort that received CRD just prior to PRV injection showed a significant reduction in EGFP-labeled cells in both the IML and left celiac ganglion compared to non-stimulated injured rats. In contrast, the injured cohort that received CRD 96 h after PRV-152 inoculation showed no differences in EGFP-labeled cell numbers in the IML or celiac ganglia versus non-stimulated injured rats. Interestingly, microglia near c-Fos-positive cells after acute CRD appeared more reactive compared to non-stimulated spinalized rats, and activated microglial cells markedly reduce viral transduction and progression following PRV inoculation of the CNS. Hence our results imply that increased CRD-induced c-Fos expression in the injured paradigm, prior to but not after PRV injection, further attenuates PRV-152 uptake, perhaps through changes in neuronal activity and/or innate neuro-immune responses.

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On counting and counting errors

Cited by 359 publications

References 63 publications

Cell-type–based model explaining coexpression patterns of genes in the brain

Cell-type–based model explaining coexpression patterns of genes in the brain

Loss of Neurons from Laminas I-III of the Spinal Dorsal Horn Is Not Required for Development of Tactile Allodynia in the Spared Nerve Injury Model of Neuropathic Pain

Noxious Colorectal Distention in Spinalized Rats Reduces Pseudorabies Virus Labeling of Sympathetic Neurons

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