Magnetic resonance microscopy (MRM) is an imaging modality that allows for noninvasive acquisition of high-resolution images in intact opaque animals. The zebrafish (Danio rerio) is an important model organism for the study of vertebrate biology. However, optical in vivo studies in zebrafish are restricted to very early developmental stages due to the opacity of the juvenile and adult stages. Application of high resolution MRM has not yet been explored in adult zebrafish. In this study we applied and optimized high resolution MRM methods to examine anatomical structures noninvasively in adult zebrafish. Clear morphological proton images were obtained by T(2)-weighted spin echo and rapid acquisition with rapid acquisition with relaxation enhancement (RARE) sequences which revealed many anatomical details in the entire intact zebrafish at a magnetic field strength of 9.4 T. In addition, in vivo imaging of adult zebrafish revealed sufficient anatomical details. To our knowledge this is the first report of the application of high resolution MRM to study detailed anatomical structures in adult zebrafish.

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Purpose: To optimize high-resolution MR spectroscopy (MRS) for obtaining neurochemical composition of adult zebrafish brain in vivo. Materials and Methods:A flow-through setup for supporting MRS of living zebrafish has been designed. In vivo MR microscopy (MRM) images were obtained using a rapid acquisition with relaxation enhancement (RARE) sequence to select a volume of interest. In vivo MR spectra from zebrafish brain were obtained using an optimized point-resolved spectroscopy (PRESS) sequence preceded by a variable pulse power and optimized relaxation delays (VAPOR) sequence for global water suppression interleaved with outer volume suppression (OVS). In vitro MR spectra in the brain extract were obtained by using correlated spectroscopy (COSY) sequences. Results:Optimized high-resolution localized MRS at 9.4T in conjunction with a strong gradient system, efficient shimming, and the water suppression scheme resulted in a reasonable separation of resonances from various metabolites in vivo from a voxel as small as 3.3 L placed in the zebrafish brain. In addition, more than 14 metabolites were identified in adult zebrafish brain extracts. Conclusion:We have successfully optimized a high-resolution localized in vivo MRS technique to get access to the zebrafish brain, and obtained for the first time the neurochemical composition of the zebrafish brain.

Zebrafish cancer models are fast gaining ground in cancer research. Most tumors in zebrafish develop late in life, when fish are no longer transparent, limiting in vivo optical imaging methods. Thus, noninvasive imaging to track tumor in adult zebrafish remains challenging. In this study, we applied magnetic resonance microimaging (microMRI) to track spontaneous melanomas in stable transgenic zebrafish models expressing an RAS oncoprotein and lacking P53 (mitf:Ras::mitf:GFP X p53(-/-)). Tumors in live adult zebrafish were observed at various locations using a T(2)-weighted fast spin echo sequence at 9.4 T. Further, live imaging of tumors at ultrahigh field (17.6 T) revealed significant tumor heterogeneity. This heterogeneity was also confirmed by the significant differences in transverse relaxation time, T(2) measured in various regions of tumor. To our knowledge, this is the first report demonstrating the application of microMRI to detect the locations, invasion status, and characteristics of internal melanomas in zebrafish and suggesting that noninvasive microMRI can be applied for longitudinal studies to track tumor development and real-time assessment of therapeutic effects in zebrafish tumor models.