Combined fluorescence microscopy and MRI reveals neural dynamics

07 Jun 2017

Integrating the two techniques helps to link microscopic activity with larger behavior of nervous system.

Functional magnetic resonance imaging (fMRI) is an established method to show changes in blood-oxygen levels in different parts of the brain, through an imaging parameter termed the blood oxygenation level dependent (BOLD) contrast.

However, the ways in which the BOLD signal relates to the underlying neural activity and physiological changes under different conditions has remained hard to investigate, not least since the modality's response time to neuronal change can be relatively lengthy, taking a few seconds.

A solution could be to integrate the fMRI technique with two-photon fluorescence microscopy (TPM), which has much higher spatial resolution and can respond rapidly to a number of different functional changes.

Combining the two techniques and carrying out simultaneous TPM and fMRI has been an attractive concept for researchers for some time, but has proven difficult due to practical difficulties co-locating the two distinct systems, as well as the potential effects on fluorescence imaging of the magnetic field strengths involved in the fMRI operation.

Now a project at Purdue University and the University of Minnesota has developed a possible solution, a proof-of-concept platform for the first MRI-compatible integrated TPM-MRI multi-modal imaging system, able to operate at the ultra-high magnetic fields found in fMRI and deliver the micron-level spatial resolution of TPM. The findings were published in Scientific Reports.

"This pilot study paves the way for simultaneous ultra-high-field MRI and high-resolution two-photon microscopy," commented Meng Cui of Purdue University Department of Biological Sciences. "The imaging capability will open new opportunities to study and understand the comprehensive relationships between brain structure and connectivity, neuronal activity and dynamics, brain function and cellular energy metabolism in supporting brain function."

Understanding brain function
The design concept combined a remote laser scanning system located in room adjacent to the MRI magnet room, and an optical imaging module positioned within the MRI instrument. The imaging module included an objective lens, filters and optical mirrors designed to focus the TPM emission onto the entrance port of a 9-meter fiber light guide, taking the fluorescence signal back to a remote photomultiplier detector. The entire in-MRI module contained only glass, brass, plastic and aluminium, fully compatible with the University of Minnesota MRI system and its magnetic field of 16.4 tesla.

To test the integrated TPM-MRI imaging capability, the team applied it to the fixed ex vivo brain of a mouse, positioned in the focal area of the system. According to the team's paper, there was no noticeable change in resolution of the TPM images or shift in their position caused by the operation of the MRI and its associated magnetic fields. The MRI images were similarly unaffected by the modified platform.

The team noted some limitations of this proof-of-concept design, including the use of an objective lens of low NA and the lack of any axial scanning capability. Different lenses of higher NA should improve both the spatial resolution and the efficiency of fluorescence emission collection, while ways to combine the current system with three-dimensional volumetric imaging methods will now be investigated.

If it can be successfully applied to in vivo specimens, the optimized integrated imaging system could provide new insights into the electrophysiological basis of the fMRI BOLD signal, helping to uncover the relationships between neuronal activity and dynamics, and the roles of cellular energy metabolism in supporting brain function.

"Understanding human brain function and dysfunction is the key challenge in neuroscience research, and is also essential for the diagnosis and treatment of brain disorders," said Cui. "There is no clear understanding of what cellular processes cause the MRI signal, and we are left only with hypotheses. We know there is a change in oxygen carried by hemoglobin in the brain, but microscopically nobody knows the details."