Introduction

The Locus Coeruleus (LC) and Substantia Nigra (SN) are neuromelanin (NM) rich nuclei in the brain that are suggested to play a role in the pathology of several neurodegenerative diseases such as Alzheimer’s and Parkinson’s disease^1-4. We have recently shown that Magnetization Transfer (MT) MRI provides good contrast in the LC⁵. Knowledge of the source of contrast is important for the development of a quantitative LC-atrophy estimation method. NM is thought to be the main contributor to the LC/SN-specific MT contrast. In this study, we extend our previous work by comparing the MT properties of an in vivo human LC and SN with measurements on an agar-based phantom containing two commonly used NM models, with the ultimate goal to investigate the MT effect of NM and assess the validity of the phantom.

Methods

MR data from an agar-based phantom containing variable concentrations (0-3 mg/ml) of natural Sepia melanin (M2649, all chemicals from Sigma Chemical Co.) and synthetic Cys-Dopa-Melanin (DAM, synthesized by auto-oxidizing Dopamine (H8502) in the presence of L-Cysteine (168149), and a healthy volunteer (female, 23), were acquired on a 3T Prisma scanner (Siemens Healthineers, Erlangen, Germany). An MT-weighted Turbo Flash (MT-TFL) sequence was acquired, consisting of a multi-shot 3D readout with center-out k-space sampling preceded by a train of off-resonant Gaussian sinc pulses, with adjustable amplitudes and offset frequencies (FO)^5,6. Matching non-MT-TFL images were acquired to obtain MT-ratio images, and monitor the heating of the phantom. A multi-echo gradient echo (ME-GRE) scan and an MP2RAGE were acquired to determine T₁ and T₂* relaxation times. All acquisition parameters are listed in detail in Table 1. MT acquisitions on the phantom were corrected for linear signal changes due to temperature increase, by scaling the data relative to the signal increase in the non-MT scans that were acquired throughout the session. The MTR maps were further corrected by linear scaling for B₁ bias^7,8. Hyperintense voxels in the LC and SN region were selected manually and compared with a similar-sized reference ROI (Pontine Tegmentum). Similarly, melanin and reference ROIs were drawn and compared in the phantom.

Results

The LC showed less MT compared to the reference in the 200-4000Hz FO range, as well as the SN in the 1000-4000Hz FO range. No melanin-specific MT effects were found on either of the melanin models in the phantom (Figure 1). The SN shows less MT relative to its direct environment (though limited contrast compared to the reference ROI). Both NM models show a decreasing T₁ and T₂* with increasing concentration (Figure 3). The LC shows a longer T₁ and no difference in T₂* compared to the reference ROI, while the SN shows no effect on T₁ and a shorter T₂* relative to the reference ROI (Figure 2).

Discussion

In this study, we examined for the first time the MT spectrum of an in vivo LC and compared it to the SN and NM phantoms; this was made feasible by employing a high-resolution MT-TFL sequence. Decreased MT was found in LC and SN, while the NM phantoms did not show such an effect (Figure 1). Furthermore, NM models showed T₁ and T₂* shortening depending on concentration, which was not reproduced in the in vivo LC and SN. This may imply that melanin models are not representative of real NM or that NM is not causing the MT contrast in LC and SN. SN showed more MT compared to LC, especially in the lower range of MT spectrum (200-1000Hz), which may imply a wider direct saturation spectrum; this is in accordance with the shorter T₁ in SN (Figure 3). Finally, the LC showed less MT compared to reference, despite the fact that an increased T_{1 free} was found⁹. This may imply that a fast T_{1 bound} or decreased pool-size-ratio is present in LC, which could result in fast loss of saturation of macromolecular spins¹⁰.

Conclusion

In this study we managed for the first time to acquire the MT spectrum of LC and compare it to SN and NM phantoms. The LC showed a distinct MT spectrum compared to other surrounding structures, including the (also catecholaminergic) SN. Nevertheless, our findings on the phantom may challenge the assumption that NM is the cause of the MT decrease in the LC and SN, or imply that the models are not representative of the in vivo LC and SN. Postmortem and phantom studies with real NM are needed to get conclusive evidence. Characterizing the LC MT spectrum is important to examine its usability as biomarker for neurodegenerative diseases.

Acknowledgements

This work is supported by an NWO VENI [016.158.084] to HILJ and NWO VIDI [016. 178.052] to BAP. The authors thank Harald Moeller and Andre Pampel for helpful discussions regarding MT measurements.

References

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