Graham M. Fraser

Comparison of generated parallel capillary arrays to three-dimensional reconstructed capillary networks in modeling oxygen transport in discrete microvascular volumes.

We compare RMN to PCA under several simulated physiological conditions to determine how the use of different vascular geometry affects oxygen transport solutions.

Mapping 3-D functional capillary geometry in rat skeletal muscle in vivo

We have developed a novel mapping software package to reconstruct microvascular networks in three dimensions (3-D) from in vivo video images for use in blood flow and O2 transport modeling. An intravital optical imaging system was used to collect video sequences of blood flow in microvessels at different depths in the tissue. Functional images of vessels were produced from the video sequences and were processed using automated edge tracking software to yield location and geometry data for construction of the 3-D network. The same video sequences were analyzed for hemodynamic and O2 saturation data from individual capillaries in the network. Simple user-driven commands allowed the connection of vessel segments at bifurcations, and semiautomated registration enabled the tracking of vessels across multiple focal planes and fields of view. The reconstructed networks can be rotated and manipulated in 3-D to verify vessel connections and continuity. Hemodynamic and O2 saturation measurements made in vivo can be indexed to corresponding vessels and visualized using colorized maps of the vascular geometry. Vessels in each reconstruction are saved as text-based files that can be easily imported into flow or O2 transport models with complete geometry, hemodynamic, and O2 transport conditions. The results of digital morphometric analysis of seven microvascular networks showed mean capillary diameters and overall capillary density consistent with previous findings using histology and corrosion cast techniques. The described mapping software is a valuable tool for the quantification of in vivo microvascular geometry, hemodynamics, and oxygenation, thus providing rich data sets for experiment-based computational models.

Toward a Multiscale Description of Microvascular Flow Regulation: O2-Dependent Release of ATP from Human Erythrocytes and the Distribution of ATP in Capillary Networks

Integration of the numerous mechanisms that have been suggested to contribute to optimization of O2 supply to meet O2 need in skeletal muscle requires a systems biology approach which permits quantification of these physiological processes over a wide range of length scales. Here we describe two individual computational models based on in vivo and in vitro studies which, when incorporated into a single robust multiscale model, will provide information on the role of erythrocyte-released ATP in perfusion distribution in skeletal muscle under both physiological and pathophysiological conditions. Healthy human erythrocytes exposed to low O2 tension release ATP via a well characterized signaling pathway requiring activation of the G-protein, Gi, and adenylyl cyclase leading to increases in cAMP. This cAMP then activates PKA and subsequently CFTR culminating in ATP release via pannexin 1. A critical control point in this pathway is the level of cAMP which is regulated by pathway-specific phosphodiesterases. Using time constants (~100 ms) that are consistent with measured erythrocyte ATP release, we have constructed a dynamic model of this pathway. The model predicts levels of ATP release consistent with measurements obtained over a wide range of hemoglobin O2 saturations (sO2). The model further predicts how insulin, at concentrations found in pre-diabetes, enhances the activity of PDE3 and reduces intracellular cAMP levels leading to decreased low O2-induced ATP release from erythrocytes. The second model, which couples O2 and ATP transport in capillary networks, shows how intravascular ATP and the resulting conducted vasodilation are affected by local sO2, convection and ATP degradation. This model also predicts network-level effects of decreased ATP release resulting from elevated insulin levels. Taken together, these models lay the groundwork for investigating the systems biology of the regulation of microvascular perfusion distribution by erythrocyte-derived ATP.

Microvascular Flow Modeling using In Vivo Hemodynamic Measurements in Reconstructed 3D Capillary Networks

Please cite this paper as: Fraser GM, Goldman D, Ellis CG. Microvascular flow modeling using in vivo hemodynamic measurements in reconstructed 3D capillary networks. Microcirculation 19: 510–520, 2012.

Four-Dimensional Microvascular Analysis Reveals That Regenerative Angiogenesis in Ischemic Muscle Produces a Flawed MicrocirculationNovelty and Significance

Rationale: Angiogenesis occurs after ischemic injury to skeletal muscle, and enhancing this response has been a therapeutic goal. However, to appropriately deliver oxygen, a precisely organized and exquisitely responsive microcirculation must form. Whether these network attributes exist in a regenerated microcirculation is unknown, and methodologies for answering this have been lacking. Objective: To develop 4-dimensional methodologies for elucidating microarchitecture and function of the reconstructed microcirculation in skeletal muscle. Methods and Results: We established a model of complete microcirculatory regeneration after ischemia-induced obliteration in the mouse extensor digitorum longus muscle. Dynamic imaging of red blood cells revealed the regeneration of an extensive network of flowing neo-microvessels, which after 14 days structurally resembled that of uninjured muscle. However, the skeletal muscle remained hypoxic. Red blood cell transit analysis revealed slow and stalled flow in the regenerated capillaries and extensive arteriolar-venular shunting. Furthermore, spatial heterogeneity in capillary red cell transit was highly constrained, and red blood cell oxygen saturation was low and inappropriately variable. These abnormalities persisted to 120 days after injury. To determine whether the regenerated microcirculation could regulate flow, the muscle was subjected to local hypoxia using an oxygen-permeable membrane. Hypoxia promptly increased red cell velocity and flux in control capillaries, but in neocapillaries, the response was blunted. Three-dimensional confocal imaging revealed that neoarterioles were aberrantly covered by smooth muscle cells, with increased interprocess spacing and haphazard actin microfilament bundles. Conclusions: Despite robust neovascularization, the microcirculation formed by regenerative angiogenesis in skeletal muscle is profoundly flawed in both structure and function, with no evidence for normalizing over time. This network-level dysfunction must be recognized and overcome to advance regenerative approaches for ischemic disease.

Modeling steady state SO2-dependent changes in capillary ATP concentration using novel O2 micro-delivery methods

Adenosine triphosphate (ATP) is known to be released from the erythrocyte in an oxygen (O2) dependent manner. Since ATP is a potent vasodilator, it is proposed to be a key regulator in the pathway that mediates micro-vascular response to varying tissue O2 demand. We propose that ATP signaling mainly originates in the capillaries due to the relatively long erythrocyte transit times in the capillary and the short ATP diffusion distance to the electrically coupled endothelium. We have developed a computational model to investigate the effect of delivering or removing O2 to limited areas at the surface of a tissue with an idealized parallel capillary array on total ATP concentration. Simulations were conducted when exposing full surface to perturbations in tissue O2 tension (PO2) or locally using a circular micro-outlet (~100 μm in diameter), a square micro-slit (200 × 200 μm), or a rectangular micro-slit (1000 μm wide × 200 μm long). Results indicated the rectangular micro-slit has the optimal dimensions for altering hemoglobin saturations (SO2) in sufficient number capillaries to generate effective changes in total [ATP]. This suggests a threshold for the minimum number of capillaries that need to be stimulated in vivo by imposed tissue hypoxia to induce a conducted micro-vascular response. SO2 and corresponding [ATP] changes were also modeled in a terminal arteriole (9 μm in diameter) that replaces 4 surface capillaries in the idealized network geometry. Based on the results, the contribution of terminal arterioles to the net change in [ATP] in the micro-vascular network is minimal although they would participate as O2 sources thus influencing the O2 distribution. The modeling data presented here provide important insights into designing a novel micro-delivery device for studying micro-vascular O2 regulation in the capillaries in vivo.

The eye of the beholder: Can patterns in eye movement reveal aptitudes for spatial reasoning?

Mental rotation ability (MRA) is linked to academic success in the spatially complex Science, Technology, Engineering, Medicine, and Mathematics (STEMM) disciplines, and anatomical sciences. Mental rotation literature suggests that MRA may manifest in the movement of the eyes. Quantification of eye movement data may serve to distinguish MRA across individuals, and serve as a consideration when designing visualizations for instruction. It is hypothesized that high‐MRA individuals will demonstrate fewer eye fixations, conduct shorter average fixation durations (AFD), and demonstrate shorter response times, than low‐MRA individuals. Additionally, individuals with different levels of MRA will attend to different features of the block‐figures presented in the electronic mental rotations test (EMRT). All participants (n = 23) completed the EMRT while metrics of eye movement were collected. The test required participants view pairs of three‐dimensional (3D) shapes, and identify if the pair is rotated but identical, or two different structures. Temporal analysis revealed no significant correlations between response time, average fixation durations, or number of fixations and mental rotation ability. Further analysis of within‐participant variability yielded a significant correlation for response time variability, but no correlation between AFD variability and variability in the number of fixations. Additional analysis of salience revealed that during problem solving, individuals of differing MRA attended to different features of the block images; suggesting that eye movements directed at salient features may contribute to differences in mental rotations ability, and may ultimately serve to predict success in anatomy. Anat Sci Educ 9: 357–366.

Different perspectives: Spatial ability influences where individuals look on a timed spatial test

Learning in anatomy can be both spatially and visually complex. Pedagogical investigations have begun exploration as to how spatial ability may mitigate learning. Emerging hypotheses suggests individuals with higher spatial reasoning may attend to images differently than those who are lacking. To elucidate attentional patterns associated with different spatial ability, eye movements were measured in individuals completing a timed electronic mental rotation test (EMRT). The EMRT was based on the line drawings of Shepherd and Metzler. Individuals deduced whether image pairs were rotations (same) or mirror images (different). It was hypothesized that individuals with high spatial ability (HSA) would demonstrate shorter average fixation durations during problem solving and attend to different features of the EMRT than low spatial ability (LSA) counterparts. Moreover, question response accuracy would be associated with fewer fixations and shorter average response times, regardless of spatial reasoning ability. Average fixation duration in the HSA group was shorter than LSA (F(1,8) = 7.99; P = 0.022). Importantly, HSA and LSA individuals looked to different regions of the EMRT images (Fisher Exact Test: 12.47; P = 0.018); attending to the same locations only 34% of the time. Correctly answered questions were characterized by fewer fixations per question (F(1, 8) = 18.12; P = 0.003) and shorter average response times (F(1, 8) = 23.89; P = 0.001). The results indicate that spatial ability may influence visual attention to salient areas of images and this may be key to problem solving processes for low spatial individuals. Anat Sci Educ 10: 224–234.

Reduced uterine perfusion pressure decreases functional capillary density in skeletal muscle

The purpose of this study was to examine the functional and structural capillary density in the reduced uterine perfusion pressure (RUPP) model, which when performed during pregnancy is an established animal model of preeclampsia. We hypothesized that the RUPP model would be associated with capillary rarefaction and impaired capillary perfusion, which would be more pronounced in the pregnant state. Female Sprague-Dawley rats (n = 32) were randomized to nonpregnancy (Nonpregnant) or breeding (Pregnant) at 12 wk of age and again to RUPP or SHAM surgeries on gestational day (GD) 14 (or equivalent age in nonpregnant rats). On GD 20 (or equivalent), capillary structure and perfusion of the extensor digitorum longus were imaged using digital intravital video microscopy. Functional videos were analyzed by a blinded observer to measure capillary density, expressed as capillaries per millimeter intersecting three staggered reference lines (200 μm). Flow was scored as the percentage of capillaries having 1) continuous, 2) intermittent, or 3) stopped flow. Total capillary density was not different between groups. There was a main effect of RUPP surgery resulting in decreased continuous flow vessels (P < 0.01) and increased stopped flow (P < 0.01), which was driven by differences between pregnant animals (Continuous flow: pregnant SHAM 80.1 ± 7.8% vs. pregnant RUPP 67.8 ± 11.2%, P < 0.05) (Stopped flow: pregnant SHAM 8.7 ± 3.2% vs. pregnant RUPP 17.9 ± 5.7%, P < 0.01). Our results demonstrate that the RUPP surgery is associated with a decrease in functional capillary density in skeletal muscle that is more pronounced in the pregnant state, which may contribute to the vascular pathophysiology observed in preeclampsia.

Impact of Incremental Perfusion Loss on Oxygen Transport in a Capillary Network Mathematical Model.

To quantify how incremental capillary PL, such as that seen in experimental models of sepsis, affects tissue oxygenation using a computation model of oxygen transport.