PiB Fails to Map Amyloid Deposits in Cerebral Cortex of Aged Dogs with Canine Cognitive Dysfunction

Abstract

Dogs with Canine Cognitive Dysfunction (CCD) accumulate amyloid beta (Aβ) in the brain. As the cognitive decline and neuropathology of these old dogs share features with Alzheimer's disease (AD), the relation between Aβ and cognitive decline in animal models of cognitive decline is of interest to the understanding of AD. However, the sensitivity of the biomarker Pittsburgh Compound B (PiB) to the presence of Aβ in humans and in other mammalian species is in doubt. To test the sensitivity and assess the distribution of Aβ in dog brain, we mapped the brains of dogs with signs of CCD (n = 16) and a control group (n = 4) of healthy dogs with radioactively labeled PiB ([(11)C]PiB). Structural magnetic resonance imaging brain scans were obtained from each dog. Tracer washout analysis yielded parametric maps of PiB retention in brain. In the CCD group, dogs had significant retention of [(11)C]PiB in the cerebellum, compared to the cerebral cortex. Retention in the cerebellum is at variance with evidence from brains of humans with AD. To confirm the lack of sensitivity, we stained two dog brains with the immunohistochemical marker 6E10, which is sensitive to the presence of both Aβ and Aβ precursor protein (AβPP). The 6E10 stain revealed intracellular material positive for Aβ or AβPP, or both, in Purkinje cells. The brains of the two groups of dogs did not have significantly different patterns of [(11)C]PiB binding, suggesting that the material detected with 6E10 is AβPP rather than Aβ. As the comparison with the histological images revealed no correlation between the [(11)C]PiB and Aβ and AβPP deposits in post-mortem brain, the marked intracellular staining implies intracellular involvement of amyloid processing in the dog brain. We conclude that PET maps of [(11)C]PiB retention in brain of dogs with CCD fundamentally differ from the images obtained in most humans with AD.

Keywords: 6E10 immunohistochemistry; Alzheimer’s disease; Pittsburgh compound B; beta-amyloid; canine cognitive dysfunction; dog.

Figure 1

Showing different sections of a dog brain with the ROI as color coded areas outlined by the authors based on a stereotaxic atlas by Dua-Sharma, in order to extract PET image values for relevant regional brain areas (Dua-Sharma et al., 1970). (A) Sagittal mid section of a dog brain with identification of ROI. (B) Dorsal section of dog brain at the level of the third ventricle and caudate nucleus with identification of ROI. (C) Transverse section of dog brain at the level of the third and lateral ventricles and the habenular nucleus with identification of ROI. Color code identifies the gray matter for each ROI used. ROI, regions of interest.

Figure 2

Magnetic resonance imaging of the brain of CCD dogs with reference region of max average theta value (red/dark red) identical with an area of maximal washout characteristics of [11C]PiB highlighted in red and therefore minimum binding of [11C]PiB. (A) Mid-sagittal section of dog brain showing an area of max theta value (red/dark red) and therefore minimum [11C]PiB binding in the thalamic adhesion and the cerebellum. (B) Dorsal section of dog brain at the level of the third ventricle and caudate nucleus. Showing areas of max theta value (red/dark red) in the parietal and temporal lobes. (C) Transverse section of dog brain at the level of the third and lateral ventricles and the habenular nuclei. Identifying areas of maximum theta (red/dark red) and therefore minimum [11C]PiB binding in the temporal lobes and the thalamic adhesion.

Figure 3

Correlation of MRI [¹¹C]PiB PET imaging and histopathology of a transverse brain section at the level of the rostral commissure and the corpus striatum. The brain is from a 17-year old small mixed breed male dog with clinical diagnosis of CCD. (A) Transverse MRI of brain from CCD dog at the level of rostral commissure and the corpus striatum (B) MRI from picture 1 and corresponding [¹¹C]PiB PET imaging. [¹¹C]PiB binding is indicated by a color scale with red indicating high retention and black indicating low retention. Calculated using the Hypotime method. (C) Corresponding section to 1 and 2 immunostained with amyloid (Aβ) monoclonal antibody 6E10. Dark areas identifies positive areas of immunostaining using 6E10 indicating and thereby either Aβ and AβPP deposits or both (Marked with arrows). MRI, magnetic resonance imaging; PiB, Pittsburgh compound B; PET, positron emission tomography; CCD, canine cognitive dysfunction.

Figure 4

Histological sagittal sections of the frontal and temporal lobes including the hippocampus. Sections are immunostained with amyloid (Aβ) monoclonal antibody 6E10 (A,B) are histological section from the frontal and temporal lobe, respectively. Obtained from a 17-year old small mixed breed with signs of cognitive dysfunction. The sections exhibit diffuse staining of cortical gray matter in the frontal and temporal cortex and marked staining of the perforant pathway in the hippocampus. (C,D) are histological section from the frontal and temporal lobe, respectively. Obtained from a 13-year old collie with signs of cognitive dysfunction. The sections exhibit diffuse staining of the cortical gray matter in the frontal and temporal cortex. The perforant pathway in the hippocampus only demonstrate marked staining in a very small area compared to (B).

Figure 5

Histological horizontal sagittal sections through the anterior lobe of the cerebellum. Obtained from a 13-year old female Border Collie with clinical signs of canine cognitive dysfunction. (A,B) Low magnification view immunostained with amyloid (Aβ) monoclonal antibody 6E10 showing the presence of marked immunostaining of Purkinje cells (black arrows), but also exhibiting diffuse staining of the granular and molecular layer (white arrows). (C,D) High magnification view of Purkinje cells in (A,B) showing intracellular accumulation of Aβ positive material. CCD, canine cognitive dysfunction.