- Email: [email protected]
ALTERATIONS IN THE HIPPOCAMPAL MR CHARACTERISTICS OF A MOUSE MODEL OF AD
Poster Presentations: P3 with potent PDE10 inhibitors. This assay can be used to assess the receptor occupancy and correlate with efficacy in two different species. P3-012
ALTERATIONS IN THE HIPPOCAMPAL MR CHARACTERISTICS OF A MOUSE MODEL OF AD
Palamadai N. Venkatasubramanian1, Kaushik Govindaraju2, Benjamin Banks2, Frances Lendacki1, Jason Pych1, George Iordanescu1, Alice Wyrwicz1, 1NorthShore University HealthSystem, Evanston, Illinois, United States; 2Northwestern University, Evanston, Illinois, United States. Contact e-mail: [email protected] Background: High resolution MRI has been used previously to examine cortical and hippocampal atrophy in mouse models of AD. We report here a novel alteration in MR contrast specifically in the CA1 pyramidal cell layer (CA1py) in a transgenic mouse that coexpresses five FAD mutations. Methods: 5xFAD mice (Tg) coexpressing 3 APP and 2 PS1 FAD mutations and littermate wildtype (WT) controls were used. Fixed brains from 2, 4 and 10M old mice were imaged at14.1T. T 2 relaxation time maps were obtained using TR 6000ms, TE 7.5ms, 16 echoes, slice thickness 0.3mm, in-plane pixel size 35mm. T2 in CA1 stratum oriens (SO), CA1 pyramidal neuron layer (CA1py), CA1 stratum radiatum (SR), dentate gyrus molecular layer (DG), corpus callosum (CC) and cortex were compared. Results: In T2weighted images, CA1py of WT mice appeared as a dark band at all ages because of lower T2 relative to neighboring SO and SR subfields. Agedependent loss in CA1py contrast was seen only in Tg mice. At 2M, CA1py contrast was similar in Tg and WT mice. But, contrast diminished in 4M old Tg mice, and at 10M, CA1py contrast was lost entirely. Loss of CA1py contrast was resulted from progressive increase in T2 in CA1py: from 35.5ms at 2M to 38.0ms at 10M, along with a decrease in T2 in the adjacent SO and SR subfields, from 38.1ms at 2M to 35.6 at 10M in SO, and from 40.9ms at 2M to 39.2ms at 10M in SR. The cortex, CC and DG of 5xFAD mice did not display any T2 changes with age. Stereological measurements ruled out loss of CA1 pyramidal neuron density in Tg mice. Performance of 5xFAD mice in hippocampally dependent behavioral tasks begin to decline at 4M and worsen at 10M, which parallels the temporal pattern of CA1py T2 increase. Conclusions: Loss of CA1py contrast in the 5xFAD mouse may be a critical step that reflects the onset of AD-like neuropathology. Therefore MR microscopy could be a valuable tool to follow disease progression in preclinical studies that employ 5xFAD mice. (Supported by R01AG027424 and S10RR13880) P3-013
BETA-ALANYL-L-HISTIDINE AMELIORATES MEMORY DECLINE CAUSED BY FEEDING A HIGH FAT DIET IN A TRANSGENIC MOUSE MODEL OF ALZHEIMER’S DISEASE
Tatsuhiro Hisatsune1, Megumi Shibahara2, Bruno Herculano2, Yoshifumi Abe3, J.U.N. Kaneko4, 1The University of Tokyo, Kashiwa, Japan; 2The University of Tokyo, Kashiwa, Japan; 3The University of Tokyo, Kashiwa-shi, Japan; 4The University of Tokyo, Kashiwa, Japan. Contact e-mail: [email protected] Background: Recent epidemiological studies have noted that type 2 diabetes mellitus (T2DM) is a risk factor for Alzheimer’s disease (AD). In a previous study, we utilized a mouse model of T2DM/AD and demonstrated that beta-alanyl-L-histidine, endogenous dipeptide existed in muscle and brain, ameliorated cognitive deficit observed in this T2DM/AD model mouse. Our goal in this study was to elucidate mechanism for the amelioration by the treatment of this dipeptide. For this purpose, we utilized a microarray analysis of expressed gene in the hippocampus of this model mouse either in the absence or the presence of beta-alanyl-L-histidine treatment. Methods: B6C3-Tg (APP swe/PSEN1dE9) 85Dbo/J AD purchased from Jackson Laboratories (Bar Harbor, Maine, USA) was utilized for this study. Animals were fed with high fat diet (HFD-32, containing 32% of fat; CLEA, Japan) from 4 month of age for 8 weeks. For the treatment group, animals were treated with beta-alanyl-L-histidine through drinking bottle at the concentration of 1 g/L (approximately 5 mg/body/day). This treatment was started 2 weeks after the initial feeding with HFD and kept until the end of the experiments. Control animals had access to regular de-ionized auto-
P631
claved drinking water. Animals were tested for contextual fear conditioning task. Twenty-four hours after the behavioral test, mice were sacrificed and perfused transcardially with saline. Then, the right hippocampus was dissected from the brain and was homogenized in TRIzol reagent and stored at -80 C. Gene expression of individual hippocampus was evaluated with SurePrint G3 Mouse GE 8x60K Microarray Kit (Agilent Technologies) under the entrustment of Cell Innovator Co., Fukuoka, Japan. Other sets of animals were utilized for immunohistochemical analysis or animal MRI analysis as described by Chin et al., (2013) J. Neuroinflammation. Results: In this T2DM/AD model, we have observed the decline in spatial memory performance, as detected by the measurement of freezing time in contextual fear conditioning task. We did not detect any change in the amount of beta amyloid plaque between the treatment and the control group, but observed differences in the neuroinflammatry response between the two groups. The beta-alanyl-L-histidine treatment significantly blocked the increase in the neuroinflammatory response as revealed by immnohistochemistry and by diffusion MRI analysis. As a result from microarray analysis of the hippocampal samples (n ¼ 3 in each group), to our surprise, we detected a substantial decrease in the expression of both astrocytic GABA-transpoters in the treatment group in comparison to the control group (GAT2 (Slc6a13): ratio (treat / control) ¼ 0.36 p¼0.0006; BGT1 (Slc6a12): ratio ¼ 0.27, p ¼ 0.0007). In addition, we observed decrease in the expression of BMP7 (ratio 0.52, P ¼ 0.0009) and GFAP (ratio 0.54, P ¼ 0.006) in the treatment group. Conclusions: In conclusion, we can propose a new model for the effect of beta-alanyl-L-histidine on the amelioration of cognitive decline observed in T2DM/AD through the inhibition of the expression of astrocytic GABA transporters. Since, beta-alanine, a cleaved product of this dipeptide, is a substrate of the two astrocytic GABA transporters. We can speculate that beta-alanine, a catabolite of beta-alanyl-Lhistidine by carnosinase, promotes the down-modulation of genes expressing astroytic GABA-transporters after transferred into the central nervous system from the blood flow by taurine / beta-alanine transporter (Slc6A6) at the blood brain barrier. In the affected astrocytes, this cellular event may lead to the inhibition of BMP7 expression then the suppression of astrocytic activation, gliosis. Further study must be definitely required to assist this hypothesis of the suppression of gliosis by the treatment of beta-alanyl-L-histidine. P3-014
LONGITUDINAL CHARACTERIZATION OF CVN MOUSE FOR ALZHEIMER’S DISEASE USING BEHAVIORAL, IMAGING, AND BIOMARKER END-POINTS
Toni Ahtoniemi1, Nina Vartiainen2, Taneli Heikkinen1, Juho Oksman1, Jukka Puoliv€ali1, Marc Cerrada-Gimenez1, Kimmo Lehtim€aki1, Antti Nurmi1, 1Charles River Discovery Research Services, Kuopio, Finland; 2Charles River Discovery Research Services Finland, Kuopio, Finland. Contact e-mail: [email protected] Background: CVN mouse, with APPSweDI mutations and crossed with NOS2 knockout line, exhibits wide range of AD hallmarks including increased insoluble Ab and plaque formation, inflammation, tau phosphorylation, hippocampal neuronal death and behavioral deficits. Validation data from Charles River CVN mouse in-house testing colony shows reproducible AD defects, and positions CVN mouse as one of the most complete rodent AD model available. Methods: Wild-type and CVN mice were studied at 3, 6, 9 and 12 months of age. Behavioral testing battery included various tests including o pen field, Barnes maze, r adial arm water maze and c ontextual fear conditioning. Fine motor deficits were studied by using Motorater kinematic analysis system. At each age point, MRI and MRS evaluation was performed before tissue, CSF and plasma collection for analysis. Immunohistochemical stainings for amyloid plaques and inflammatory cells and biochemical analysis of soluble and insoluble amyloid were performed. Results: Significant behavioral deficits were seen at the age of 6 months in c ontextual fear conditioning and later in Barnes maze and in r adial arm water maze. Robust pathological changes, including increased number of dense amyloid plaques in hippocampus, thalamus and cortex, were evident. Significant inflammatory response was detect ed as microgli osis and CD45-positi ve cells heavily condensed around the plaques in all brain
ALTERATIONS IN THE HIPPOCAMPAL MR CHARACTERISTICS OF A MOUSE MODEL OF AD
Palamadai N. Venkatasubramanian1, Kaushik Govindaraju2, Benjamin Banks2, Frances Lendacki1, Jason Pych1, George Iordanescu1, Alice Wyrwicz1, 1NorthShore University HealthSystem, Evanston, Illinois, United States; 2Northwestern University, Evanston, Illinois, United States. Contact e-mail: [email protected] Background: High resolution MRI has been used previously to examine cortical and hippocampal atrophy in mouse models of AD. We report here a novel alteration in MR contrast specifically in the CA1 pyramidal cell layer (CA1py) in a transgenic mouse that coexpresses five FAD mutations. Methods: 5xFAD mice (Tg) coexpressing 3 APP and 2 PS1 FAD mutations and littermate wildtype (WT) controls were used. Fixed brains from 2, 4 and 10M old mice were imaged at14.1T. T 2 relaxation time maps were obtained using TR 6000ms, TE 7.5ms, 16 echoes, slice thickness 0.3mm, in-plane pixel size 35mm. T2 in CA1 stratum oriens (SO), CA1 pyramidal neuron layer (CA1py), CA1 stratum radiatum (SR), dentate gyrus molecular layer (DG), corpus callosum (CC) and cortex were compared. Results: In T2weighted images, CA1py of WT mice appeared as a dark band at all ages because of lower T2 relative to neighboring SO and SR subfields. Agedependent loss in CA1py contrast was seen only in Tg mice. At 2M, CA1py contrast was similar in Tg and WT mice. But, contrast diminished in 4M old Tg mice, and at 10M, CA1py contrast was lost entirely. Loss of CA1py contrast was resulted from progressive increase in T2 in CA1py: from 35.5ms at 2M to 38.0ms at 10M, along with a decrease in T2 in the adjacent SO and SR subfields, from 38.1ms at 2M to 35.6 at 10M in SO, and from 40.9ms at 2M to 39.2ms at 10M in SR. The cortex, CC and DG of 5xFAD mice did not display any T2 changes with age. Stereological measurements ruled out loss of CA1 pyramidal neuron density in Tg mice. Performance of 5xFAD mice in hippocampally dependent behavioral tasks begin to decline at 4M and worsen at 10M, which parallels the temporal pattern of CA1py T2 increase. Conclusions: Loss of CA1py contrast in the 5xFAD mouse may be a critical step that reflects the onset of AD-like neuropathology. Therefore MR microscopy could be a valuable tool to follow disease progression in preclinical studies that employ 5xFAD mice. (Supported by R01AG027424 and S10RR13880) P3-013
BETA-ALANYL-L-HISTIDINE AMELIORATES MEMORY DECLINE CAUSED BY FEEDING A HIGH FAT DIET IN A TRANSGENIC MOUSE MODEL OF ALZHEIMER’S DISEASE
Tatsuhiro Hisatsune1, Megumi Shibahara2, Bruno Herculano2, Yoshifumi Abe3, J.U.N. Kaneko4, 1The University of Tokyo, Kashiwa, Japan; 2The University of Tokyo, Kashiwa, Japan; 3The University of Tokyo, Kashiwa-shi, Japan; 4The University of Tokyo, Kashiwa, Japan. Contact e-mail: [email protected] Background: Recent epidemiological studies have noted that type 2 diabetes mellitus (T2DM) is a risk factor for Alzheimer’s disease (AD). In a previous study, we utilized a mouse model of T2DM/AD and demonstrated that beta-alanyl-L-histidine, endogenous dipeptide existed in muscle and brain, ameliorated cognitive deficit observed in this T2DM/AD model mouse. Our goal in this study was to elucidate mechanism for the amelioration by the treatment of this dipeptide. For this purpose, we utilized a microarray analysis of expressed gene in the hippocampus of this model mouse either in the absence or the presence of beta-alanyl-L-histidine treatment. Methods: B6C3-Tg (APP swe/PSEN1dE9) 85Dbo/J AD purchased from Jackson Laboratories (Bar Harbor, Maine, USA) was utilized for this study. Animals were fed with high fat diet (HFD-32, containing 32% of fat; CLEA, Japan) from 4 month of age for 8 weeks. For the treatment group, animals were treated with beta-alanyl-L-histidine through drinking bottle at the concentration of 1 g/L (approximately 5 mg/body/day). This treatment was started 2 weeks after the initial feeding with HFD and kept until the end of the experiments. Control animals had access to regular de-ionized auto-
P631
claved drinking water. Animals were tested for contextual fear conditioning task. Twenty-four hours after the behavioral test, mice were sacrificed and perfused transcardially with saline. Then, the right hippocampus was dissected from the brain and was homogenized in TRIzol reagent and stored at -80 C. Gene expression of individual hippocampus was evaluated with SurePrint G3 Mouse GE 8x60K Microarray Kit (Agilent Technologies) under the entrustment of Cell Innovator Co., Fukuoka, Japan. Other sets of animals were utilized for immunohistochemical analysis or animal MRI analysis as described by Chin et al., (2013) J. Neuroinflammation. Results: In this T2DM/AD model, we have observed the decline in spatial memory performance, as detected by the measurement of freezing time in contextual fear conditioning task. We did not detect any change in the amount of beta amyloid plaque between the treatment and the control group, but observed differences in the neuroinflammatry response between the two groups. The beta-alanyl-L-histidine treatment significantly blocked the increase in the neuroinflammatory response as revealed by immnohistochemistry and by diffusion MRI analysis. As a result from microarray analysis of the hippocampal samples (n ¼ 3 in each group), to our surprise, we detected a substantial decrease in the expression of both astrocytic GABA-transpoters in the treatment group in comparison to the control group (GAT2 (Slc6a13): ratio (treat / control) ¼ 0.36 p¼0.0006; BGT1 (Slc6a12): ratio ¼ 0.27, p ¼ 0.0007). In addition, we observed decrease in the expression of BMP7 (ratio 0.52, P ¼ 0.0009) and GFAP (ratio 0.54, P ¼ 0.006) in the treatment group. Conclusions: In conclusion, we can propose a new model for the effect of beta-alanyl-L-histidine on the amelioration of cognitive decline observed in T2DM/AD through the inhibition of the expression of astrocytic GABA transporters. Since, beta-alanine, a cleaved product of this dipeptide, is a substrate of the two astrocytic GABA transporters. We can speculate that beta-alanine, a catabolite of beta-alanyl-Lhistidine by carnosinase, promotes the down-modulation of genes expressing astroytic GABA-transporters after transferred into the central nervous system from the blood flow by taurine / beta-alanine transporter (Slc6A6) at the blood brain barrier. In the affected astrocytes, this cellular event may lead to the inhibition of BMP7 expression then the suppression of astrocytic activation, gliosis. Further study must be definitely required to assist this hypothesis of the suppression of gliosis by the treatment of beta-alanyl-L-histidine. P3-014
LONGITUDINAL CHARACTERIZATION OF CVN MOUSE FOR ALZHEIMER’S DISEASE USING BEHAVIORAL, IMAGING, AND BIOMARKER END-POINTS
Toni Ahtoniemi1, Nina Vartiainen2, Taneli Heikkinen1, Juho Oksman1, Jukka Puoliv€ali1, Marc Cerrada-Gimenez1, Kimmo Lehtim€aki1, Antti Nurmi1, 1Charles River Discovery Research Services, Kuopio, Finland; 2Charles River Discovery Research Services Finland, Kuopio, Finland. Contact e-mail: [email protected] Background: CVN mouse, with APPSweDI mutations and crossed with NOS2 knockout line, exhibits wide range of AD hallmarks including increased insoluble Ab and plaque formation, inflammation, tau phosphorylation, hippocampal neuronal death and behavioral deficits. Validation data from Charles River CVN mouse in-house testing colony shows reproducible AD defects, and positions CVN mouse as one of the most complete rodent AD model available. Methods: Wild-type and CVN mice were studied at 3, 6, 9 and 12 months of age. Behavioral testing battery included various tests including o pen field, Barnes maze, r adial arm water maze and c ontextual fear conditioning. Fine motor deficits were studied by using Motorater kinematic analysis system. At each age point, MRI and MRS evaluation was performed before tissue, CSF and plasma collection for analysis. Immunohistochemical stainings for amyloid plaques and inflammatory cells and biochemical analysis of soluble and insoluble amyloid were performed. Results: Significant behavioral deficits were seen at the age of 6 months in c ontextual fear conditioning and later in Barnes maze and in r adial arm water maze. Robust pathological changes, including increased number of dense amyloid plaques in hippocampus, thalamus and cortex, were evident. Significant inflammatory response was detect ed as microgli osis and CD45-positi ve cells heavily condensed around the plaques in all brain