Quantitative in vivo MRI study of Dahl and Sprague-Dawley rat brains in response to salt loading
Kenneth W Fishbein1, Mikayla L Hall1, Mustapha Bouhrara1, Yulia Grigorova1, Jeffrey Long1, Christopher A Morrell1, Edward G Lakatta1, Peter Rapp1, Alexei Y Bagrov1, Richard G Spencer1, and Olga V Fedorova1

1National Institute on Aging, National Institutes of Health, Baltimore, MD, United States

Synopsis

Dahl salt-sensitive rats are a common preclinical model for hypertension. We compared brain morphology and MRI contrast parameters (T2, T2*, MTR and diffusion) in Dahl and Sprague-Dawley rats on low-salt and high-salt diets. Two of five Dahl rats on a high-salt diet exhibited stroke lesions on T2 and diffusion-weighted images. Dahl rats had smaller brain and hippocampus volumes and larger percent ventricular volume relative to Sprague-Dawley rats, regardless of diet. Dahl rats on the high-salt diet had thinner cortex, and longer T2 and shorter T2* in whole brain (excluding lesions and ventricles). Dahl rat brains therefore exhibit distinct morphological and contrast features on MRI, some of which are independent of salt loading.

Objectives

Dahl Salt-Sensitive (DSS) rats1 differ from their “ancestors”, Sprague-Dawley (SD) rats, by developing hypertension, cardiovascular remodeling and fibrosis on a high salt (HS) diet. Hypertension in DSS rats may affect cognitive functions and is a risk factor for hemorrhagic stroke. Interestingly, while DSS rats are frequently used as a model to examine brain vascular and neurochemical adaptation to hypertension, very little has been published on the MRI appearance of the DSS rat brain2-4 and none of these works have described its morphology or contrast in vivo. Thus, we studied (i) the effect of pre-disposition to salt-sensitive hypertension on the difference in brain structures and composition, and (ii) the effect of HS diet on these parameters.

Methods

All rat experiments were approved by the NIH/NIA IACUC. Six-week old SD and DSS rats were given a HS (8% NaCl) or low salt (LS; 0.5% NaCl) diet for 8 weeks; six rats were in each group. Body weight (BW) was measured, systolic blood pressure (SBP) was measured by tail cuff plethysmography, and the animals underwent brain MRI. Imaging was performed on a Bruker Biospec 7T scanner with a 72 mm transmit-receive resonator. Anesthesia was induced by inhalation of isoflurane:O2 mixture and maintained at 2-3% v/v during scanning. Vital signs were continuously monitored and temperature maintained at 37-38°C. Multi-slice data were acquired with 1 mm section thickness and 200μm × 250μm in-plane resolution. T2 and T2*-weighted images were acquired using multi-slice multi-echo spin echo (SE; TE=9ms, TR=4s) and multi-gradient echo (TE1=2.4, ΔTE=3.7ms, TR=2s) sequences, respectively, with 20 echoes. Diffusion-weighted (DW) images (see Fig.1) were acquired with a fat-suppressed SE sequence with TE=27.6ms, δ=7ms, Δ=14ms, b=2 or 212s/mm2 and TR=4s with sensitizing gradients along H/F, A/P and L/R directions. Magnetization-transfer (MT)-weighted images were acquired with a fat-suppressed SE sequence (TE=9ms, TR=3s) preceded by a square, 6kHz-off-resonance pulse of length 0 or 250ms, B1=6μT. ROIs were manually traced on DW images for volume calculations for whole brain and brain regions. T2 and T2* in each ROI were calculated by a 3-parameter fit of mean ROI intensity over all pixels in all slices containing the structure of interest versus TE. R2'=(1/T2*)-(1/T2) was calculated as a measure of B0 and susceptibility inhomogeneity. MT ratio was calculated as MTR=1-Msat/M0. ADC's, mean diffusivity (MD) and fractional anisotropy (FA) were estimated from mean ROI intensities in DW images. Effects of genotype (SD vs. DSS) and diet (LS vs. HS) were evaluated by two-way ANOVA with Newman-Keuls post-hoc pairwise tests. Statistical significance was defined as p<0.05.

Results

DSS rats had lower BW (LS: by 20%; HS: by 32%), and higher SBP (LS: by 31 mmHg; HS: by 97 mmHg) than SD rats fed similar diets (Table 1). After 8 weeks on a HS diet, DSS, unlike SD rats, became hypertensive; SBP was higher by 16mmHg in SD rats on HS versus LS diet (ns), and by 82 mmHg in DSS rats on HS versus LS diet. Total brain and hippocampus volume were smaller in DSS versus SD rats on either diet but cerebellar volumes were similar. Mean cortical thickness was smaller in DSS rats, but only on HS diet. Percent ventricular volume was greater in DSS rats relative to SD rats on LS diet. Obvious stroke lesions were detected on T2 and diffusion-weighted MRI in two DSS rats, both on HS diet (Fig. 3; compare Fig.2). Excluding T2-hyperintense regions, whole-brain T2 was longer and T2* shorter in DSS rats on a HS diet versus control SD rats, resulting in greater R2' (Table 2). No significant differences in MTR were found. FA was generally similar in all groups while ADC in the A/P direction and MD were greater in DSS versus SD rats on the same LS diet.

Discussion

Interestingly, regardless of diet, DSS rat brains were smaller than those of SD rats yet some differences were diet-dependent. As expected, HS diet induced hypertension in DSS rats, resulting in frank stroke lesions in some animals and greater T2 and R2' and shorter T2* in this group in general. This is suggestive of microvascular disease and possibly microbleeds throughout the brain. The latter were evident as hypointensities adjacent to stroke lesions in the T2-weighted images. Behavioral tests are now underway to assess the effects of thinner cortex in DSS rats on HS diet. These results underscore the utility of the DSS rat as a model for hypertension-induced stroke and subclinical, microscopic adaptations while pointing to underlying differences in brain structure and physiology resulting directly from genetic differences between DSS and SD rats.

Acknowledgements

This work was supported by the NIH/National Institute on Aging.

References

1. Dahl LK, Knudsen KD, Heine MA, Leitl GJ. Effects of chronic excess salt ingestion. Modification of experimental hypertension in the rat by variations in the diet. Circ Res 1968;22:11-18.

2. Decano JL, Viereck JC, McKee AC, et al. Early-life sodium exposure unmasks susceptibility to stroke in hyperlipidemic, hypertensive heterozygous Tg25 rats transgenic for human cholesteryl ester transfer protein. Circulation 2009;119:1501-1509.

3. Li Z, Ward BD, Dwinell MR, et al. FMRI and fcMRI phenotypes map the genomic effect of chromosome 13 in Brown Norway and Dahl salt-sensitive rats. NeuroImage 2014;90:403-412.

4. Li C, Li Z, Ward BD et al., Enhancements of resting-state fcMRI networks by prior sensory stimulation. Brain Connectivity 2014;4:760-768.

Figures

Figure 1: Images acquired using a spin-echo diffusion sequence with b = 2 s/mm2 for a 14-week-old Dahl salt-sensitive rat after 8 weeks on a low salt diet, showing typical image quality and resolution achieved in this study.

Figure 2: T2-weighted brain images (TE=72.1ms, TR=4s) of DSS rat on LS diet.

Figure 3: T2-weighted brain images (TE=72.1ms, TR=4s) of DSS rat on HS diet. Numerous stroke lesions are evident as hyperintensities (red arrows) and hypointensities (green arrows). Blue arrows indicate enlarged left lateral ventricle. Note pronounced deviations from midline symmetry.

Table 1: Body weight, systolic blood pressure and morphological data for whole brain and substructures in 14-week-old Dahl salt-sensitive (DSS) and Sprague-Dawley (SD) rats after 8 weeks on a low salt (LS) or high salt (HS) diet.

Table 2: Quantitative MRI parameter data for whole brain and substructures in 14-week-old Dahl salt-sensitive (DSS) and Sprague-Dawley (SD) rats after 8 weeks on a low salt (LS) or high salt (HS) diet. Stroke lesions, evident in the cortex of two DSS rats on HS diet, were excluded from whole-brain parameter data.



Proc. Intl. Soc. Mag. Reson. Med. 24 (2016)
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