inside Projects-P20-bu — Verstegen Lab

Functional Neuroimaging of Urine Storage and Voiding in Mice

These studies, were funded by an NIH P20 grant DK108276, to form a Resource Development Project within the BIDMC Department of Medicine’s Center for Interdisciplinary Research in Benign Urology.

This Project brings together Department of Radiology Magnetic Resonance Imaging (MRI) physicists and Department of Medicine Physiologists within BIDMC to develop, validate and disseminate methods for functional imaging of mouse brains during cystometrograms (Cystometry is a measurement of bladder filling and emptying).

Neuroimaging in humans who were voiding was pioneered by Gert Holstege’s group in the Netherlands during the 1990s and initially used Positron Emission Tomography (PET) measurements. Later studies shifted to the use of BOLD-MRI (Blood Oxygen Level Dependent MRI) which uses changes in blood oxygen levels detected by MRI to locate brain regions that are changing activation states. This is the primary method used in functional MRI (fMRI). A very good review of the findings in humans related to voiding can be found here.

Mice have become the most common model for research into human disease. Being mammals, mice share considerable anatomy, physiology and metabolism to humans. In addition, considerable efforts by thousands of researchers over the last 40 years has built a huge resource in the form of genetically modified mouse lines that can be used in many areas of research.

However, mice are small, very small, with small brains, thus posing a challenge for studies such as these using fMRI. Using the P20 funding method we have been successful in developing methods for performing fMRI on mice undergoing cystometry. We present some findings below and provide detailed methods on how data collected are analyzed that others may use to further their own studies. As always, there is room for further development and we will update our methods as new components become available.

Look for our first publication on these results in 2020.

We are indebted to Dr. Craig Ferris from Northeastern University for assistance in obtaining preliminary data using awake animals. Dr. Ferris’ information can be found at https://cos.northeastern.edu/people/craig-ferris/.

results

— obtained under urethane anesthesia

Regions that show activation during CMG peak include the Pontine Micturition Center (PMC), the Periaqueductal Gray (PAG) and the Hypothalamus. The PMC is a small center in the hindbrain that we now know activates bladder contractions, whereas the PAG receives neural inputs from the bladder and other brain regions, integrates these signals and activates the PMC. The lateral Hypothalamus (LHA) also project to the PMC and our neuro-urology colleagues have shown optogenetic activation of glutamatergic LHA neurons can cause voiding.

See also the work underway in our Neuro-Urology section, where our colleagues are making tremendous progress in defining the neural pathways that control micturition. See too, their recent publication.

Figure 4 Negative activation in the Hippocampus

Figure 5 The same as Figure 4 but showing the data in the form of the echo planar image scans - the actual fMRI data

— awake studies

Another component to this project is to conduct these studies in mice that are awake throughout the procedures. Maintaining an awake mouse within an MRI instrument without head motion for sufficient time to provide 4-6 cystometry cycles is very challenging. We have been experimenting with alterations to the Bruker bed in which the mouse is placed to reduce head motion. We require the animal to be in place for about 45 minutes to obtain both anatomical an fMRI scans. Figure 6 shows motion changes in 6 directions for a mouse in place for a full 40 minutes of an EPI scan (This mouse was not undergoing cystometry at the time).

Figure 6. The six possible axes of motion that can be made while in a MRI machine. Less is best with ideally lower than 1mm translation and 1 degree of rotation.

— more results

Functional MRI Processing for Mouse Micturition Studies

Functional MRI (fMRI) studies in rodents is somewhat uncommon as it requires some cognitive or physiological property that causes measurable changes in brain activity. These studies, however, lend themselves well to fMRI studies as voiding induced by cystometry has a cyclical property and these studies can be done while the animal is anesthetized with urethane.