Lorinda Davidson

Tbx20 dose-dependently regulates transcription factor networks required for mouse heart and motoneuron development

To elucidate the function of the T-box transcription factor Tbx20 in mammalian development, we generated a graded loss-of-function series by transgenic RNA interference in entirely embryonic stem cell-derived mouse embryos. Complete Tbx20 knockdown resulted in defects in heart formation, including hypoplasia of the outflow tract and right ventricle, which derive from the anterior heart field (AHF), and decreased expression of Nkx2-5 and Mef2c, transcription factors required for AHF formation. A mild knockdown led to persistent truncus arteriosus (unseptated outflow tract) and hypoplastic right ventricle, entities similar to human congenital heart defects, and demonstrated a critical requirement for Tbx20 in valve formation. Finally, an intermediate knockdown revealed a role for Tbx20 in motoneuron development, specifically in the regulation of the transcription factors Isl2 and Hb9, which are important for terminal differentiation of motoneurons. Tbx20 could activate promoters/enhancers of several genes in cultured cells, including the Mef2c AHF enhancer and the Nkx2-5 cardiac enhancer. The Mef2c AHF enhancer relies on Isl1- and Gata-binding sites. We identified a similar Isl1 binding site in the Nkx2-5 AHF enhancer, which in transgenic mouse embryos was essential for activity in a large part of the heart, including the outflow tract. Tbx20 synergized with Isl1 and Gata4 to activate both the Mef2c and Nkx2-5 enhancers, thus providing a unifying mechanism for gene activation by Tbx20 in the AHF. We conclude that Tbx20 is positioned at a critical node in transcription factor networks required for heart and motoneuron development where it dose-dependently regulates gene expression.

Multiple mouse biological loading and monitoring system for MRI

The use of mice to study models of human disease has resulted in a surge of interest in developing mouse MRI. The ability to take 3D, high‐resolution images of live mice allows significant insight into anatomy and function. However, with imaging times on the order of hours, high throughput of specimens has been problematic. To facilitate high throughput, concurrent imaging of multiple mice has been developed; however, this poses further complexities regarding the ease and rapidity of loading several animals. In this study, custom‐built equipment was developed to streamline the preparation process and to safely maintain seven mice during a multiple‐mouse imaging session. Total preparation time for seven mice was ∼24 min. ECG and temperature were monitored throughout the scan and maintained by regulating anesthetic and heating. Proof of principle was demonstrated in a 3‐h imaging session of seven mice. Magn Reson Med 52:709–715, 2004.

Retrospective gating for mouse cardiac MRI.

Cardiac MR imaging in small animals presents some difficulties due to shorter cardiac cycles and smaller dimensions than in human beings, but prospectively gated techniques have been successfully applied. As with human imaging, there may be certain applications in animal imaging for which retrospective gating is preferable to prospective gating. For example, cardiac imaging in multiple mice simultaneously is one such application. In this work we investigate the use of retrospective gating for cardiac imaging in a mouse. Using a three‐dimensional imaging protocol, we show that image quality with retrospective gating is comparable to prospectively gated imaging. We conclude that retrospective gating is applicable for small animal cardiac MRI and show how it can be applied to the problem of cardiac MRI in multiple mice. Magn Reson Med, 2006.

Ultrasound-guided left-ventricular catheterization: a novel method of whole mouse perfusion for microimaging

We describe a novel technique to perform whole-body perfusion fixation in mice with specific relevance to micro-imaging. With the guidance of high-frequency ultrasound imaging, we were able to perfuse fixative and contrast agents via a catheter inserted into the left ventricle, and therefore preserved the integrity of the chest and abdominal cavity. In this preliminary study, our success rate over 15 animals was 73%. We demonstrate applications of this technique for magnetic resonance imaging and micro-CT, but we expect that this method can be generally applied to whole-body perfusions of other small animals in which the intact body is necessary.

Combined magnetic resonance and bioluminescence imaging of live mice

We perform combined magnetic resonance and bioluminescence imaging of live mice for the purpose of improving the accuracy of bioluminescence tomography. The imaging is performed on three live nude mice in which tritium-powered light sources are surgically implanted. High-resolution magnetic resonance images and multispectral, multiview bioluminescence images are acquired in the same session. An anatomical model is constructed by segmenting the magnetic resonance images for all major tissues. The model is subsequently registered with nonlinear transformations to the 3-D light exittance (exiting intensity) surface map generated from the luminescence images. A Monte Carlo algorithm, along with a set of tissue optical properties obtained from in vivo measurements, is used to solve the forward problem. The measured and simulated light exittance images are found to differ by a factor of up to 2. The greatest cause of this moderate discrepancy is traced to the small errors in source positioning, and to a lesser extent to the optical properties used for the tissues. Discarding the anatomy and using a homogeneous model leads to a marginally worse agreement between the simulated and measured data.