Remodeling of energy metabolism revealed by 31P magnetization transfer in a transgenic rat model of Huntington’s disease
Brice Tiret1,2, Maria-Angeles Carrillo-de Sauvage1,2, Huu Phuc Nguyen3,4, Nicole El Massioui5,6, Valérie Doyère5,6, Vincent Lebon1,2, Emmanuel Brouillet1,2, and Julien Valette1,2

1CEA/DSV/I2BM/MIRCen, Fontenay-aux-Roses, France, 2CNRS Université Paris-Saclay UMR 9199, Fontenay-aux-Roses, France, 3Institute of Medical Genetics and Applied Genomics, University of Tuebingen, Tuebingen, Germany, 4Centre for Rare Diseases, University of Tuebingen, Tuebingen, Germany, 5Paris-Saclay Institute of Neuroscience, Université Paris-Sud, UMR 9197, Orsay, France, 6Centre National de la Recherche Scientifique, Orsay, France

Synopsis

Localized 31P MRS with progressive magnetization transfer (MT) is performed in the BACHD transgenic rat model of Huntington’s disease to assess energy metabolism. Localized measurements of the ATP formation rate through creatine kinase and oxidative phosphorylation (ATPsynthase) are performed in the rat brain for the first time. Results show that ATPsynthase rate is reduced by a factor 2, which is partly compensated by higher cerebral concentrations of phosphocreatine to generate ATP via creatine kinase.

TARGET AUDIENCE:

Those interested in 31P MRS, energy metabolism and Huntington’s disease.

Purpose:

In this work we use localized 31P MRS, including progressive magnetization transfer (MT) experiments, to assess energy metabolism in a transgenic rat model of Huntington’s disease (BACHD [1]). Localized measurements of the ATP formation rate through creatine kinase and ATPsynthase are performed in the rat brain for the first time, and reveal an altered metabolic status in BACHD rats.

Methods:

Animal preparation and in vivo 31P MRS: Seven 18 to 24 months old transgenic rats (BACHD) and six age-matched wild type (WT) littermates anesthetised with 1.5-2 % isoflurane were used to estimate forward reaction rates of both ATP synthase (kf-ATPase) and creatine kinase (kf-CK) using 31P magnetization transfer MRS [2]. All measurements were performed using a 11.7T Bruker BioSpec using a 1.5-cm double-tuned surface coil (Rapid Biomed GmbH®). Localisation of a large 11×8×12 mm (Fig. 1) voxel was achieved using an adiabatic ISIS module followed by a selective BISTRO progressive saturation of the γ-ATP peak. We decided to quantify peak (PCr and Pi) attenuation relative to an unsaturated spectrum rather than symmetric saturation, because: i) signal loss due to RF bleed-over was minimal (~5%) during selective symmetric saturation, as assessed on preliminary experiments (Fig. 2A); and ii) possible residual bias would be identical in both groups and thus wouldn’t affect intergroup comparison. Three different saturation times (tsat=0.55, 1.1 and 2.2 s, Fig. 2B) were used for γ-ATP saturation, as well as two unsaturated spectra, one acquired at the beginning and one at the end of each acquisition, which also ensured that signal remained stable throughout the experiment. Each spectrum was acquired in 17 min, making it possible to estimate kf-CK at the individual level in less than 2 hours (including animal preparation) given the fact that symmetric-saturation spectra needed not to be acquired. Due to lower signal to noise ratio on the Pi peak, kf-ATPase was estimated only at group level, with Pi being quantified on spectra averaged over all animals of each group.

Data Analysis: Data were analysed with LCModel using a simulated basis set, and metabolite concentrations were estimated in both groups on unsaturated spectra. PCr concentration was estimated relative to α-ATP concentration which was considered constant (and indeed yielded the same absolute signal on spectra in both groups, see Fig. 3). From the chemical shift difference between Pi and PCr, the pH was also derived. Individual concentrations of PCr, and group concentrations of Pi, were fitted as a non-linear function of saturation time according to modified Bloch equations accounting for chemical exchange. The standard deviation for kf-ATPase was derived from Monte Carlo simulations over 500 repetitions.

Results and Discussion:

The concentration of phosphocreatine (PCr/α-ATP) exhibits a moderate but significant 10% increase in the BACHD group compared to the WT group (1.84±0.15 vs. 1.67±0.11, p=0.026) (Fig. 3). Note that such an increase has also been reported during healthy aging [3]. No pH difference was found, contrasting with our previous results in 3NP-treated rats and Human patients [4], and suggesting a milder disease or a specific metabolic adaptation. There was no difference in kf-CK (0.43±0.07 s-1 vs. 0.44±0.10 s-1 BACHD vs. WT) between the two groups (Fig. 4A). However, considering the increased PCr levels, the total flux of ATP generation through creatine kinase ([PCr]*kf-CK) is increasing by ~10%. In parallel, a large decrease was measured for kf-ATPase (0.23±0.02 s-1 vs. 0.48±0.10 s-1 BACHD vs. WT, p=0.05 based on permutation test on the distribution generated by Monte Carlo simulations) (Fig. 4B). Altogether these results suggest an adaptive mechanism in the transgenic rat model involving a larger PCr pool to compensate for the defect in oxidative phosphorylation reflected by kf-ATPase.

Conclusion:

We demonstrate for the first time the use of localised 31P MT-MRS to quantify both kf-CK and kf-ATPase in the rat brain. This study also points towards a mitochondrial deficit in the BACHD rat model that can be quantified by 31P MT-MRS, with ATP synthase rate reduced by a factor 2. This decrease could be partly compensated by higher cerebral concentrations of phosphocreatine (the origin of which remains to be elucidated, but could be related to peripheral PCr formation) as a readily available energy source.

Acknowledgements

This project was funded by the French National Research Agency (ANR-14-CE15-0007, HDeNERGY project).

References

[1] Yu-Taeger L, Petrasch-Parwez E, Osmand AP, Redensek A, Metzger S, Clemens LE, Park L, Howland D, Calaminus C, Gu X, Pichler B, Yang XW, Riess O, and Nguyen HP, “A novel BACHD transgenic rat exhibits characteristic neuropathological features of Huntington disease,” J Neurosci, vol. 32, no. 44, pp. 15426–38, 2012.

[2] H. Lei, K. Ugurbil, and W. Chen, “Measurement of unidirectional Pi to ATP flux in human visual cortex at 7 T by using in vivo 31P magnetic resonance spectroscopy.,” Proc. Natl. Acad. Sci. U. S. A., vol. 100, no. 24, pp. 14409–14, 2003.

[3] R. Longo, C. Ricci, L. Dalla Palma, R. Vidimari, a Giorgini, J. a den Hollander, and C. M. Segebarth, “Quantitative 31P MRS of the normal adult human brain. Assessment of interindividual differences and ageing effects.,” NMR Biomed., vol. 6, no. 1, pp. 53–7, 1993.

[4] M. M. Chaumeil, J. Valette, C. Baligand, E. Brouillet, P. Hantraye, G. Bloch, V. Gaura, A. Rialland, P. Krystkowiak, C. Verny, P. Damier, P. Remy, A.-C. Bachoud-Levi, P. Carlier, and V. Lebon, “pH as a biomarker of neurodegeneration in Huntington’s disease: a translational rodent-human MRS study,” Journal of Cerebral Blood Flow & Metabolism, vol. 32. pp. 771–779, 2012.

Figures

Fig. 1 Position of a 11×8×12 mm voxel in the rat brain (A) and a representative spectrum acquired in 17 mins (B).

Fig. 2 31P Spectra series acquired with progressive saturation time (0, 0.55, 1.1, and 2.2 s) where saturation was applied symmetric to the γ-ATP peak relative to Pi (A) or on γ-ATP frequency (B). It is possible to see minimal signal loss during symmetric saturation (A) and decrease of PCr signal (and Pi) as a function of tsat on γ-ATP saturated spectra (B).

Fig. 3 Difference between metabolites profile between BACHD and WT littermate rats.

Fig. 4 Normalised signal attenuations for PCr (A) and Pi (B) between the two animal groups (WT in blue squares and BACHD in red circles) as a function of the saturation time tsat.



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