Animal anatomy has traditionally relied on detailed dissections to produce anatomical illustrations, but modern imaging modalities, such as MRI and CT, now represent an enormous resource that allows for fast non-invasive visualizations of animal anatomy in living animals. These modalities also allow for creation of three-dimensional representations that can be of considerable value in the dissemination of anatomical studies. In this methodological review, we present our experiences using MRI, CT and μCT to create advanced representation of animal anatomy, including bones, inner organs and blood vessels in a variety of animals, including fish, amphibians, reptiles, mammals, and spiders. The images have a similar quality to most traditional anatomical drawings and are presented together with interactive movies of the anatomical structures, where the object can be viewed from different angles. Given that clinical scanners found in the majority of larger hospitals are fully suitable for these purposes, we encourage biologists to take advantage of these imaging techniques in creation of three-dimensional graphical representations of internal structures.

Background: The Fontan procedure has revolutionized the treatment of univentricular hearts. However, it is associated with severe complications such as protein-losing enteropathy, plastic bronchitis, and peripheral edema that may involve the lymphatic circulation. We aimed to assess lymphatic function and morphology in patients with a univentricular circulation. Methods: The functional state of lymphatic vessels in the lower extremities of patients with a Fontan circulation (n=10) was investigated using the novel technique near-infrared fluorescence imaging and compared with an age-, sex-, and weight-matched control group of healthy volunteers (n=10). The lymphatic morphology was described using T2-weighted magnetic resonance imaging, and microvascular permeability was estimated by strain gauge plethysmography. Results: The Fontan patients had 17% lower lymphatic pumping pressure (50±3.1 mm Hg) compared with controls (60±2.8 mm Hg; P =0.0341) and a 62% higher contraction frequency (0.8±0.1 min ⁻¹ ) compared with the healthy controls (0.5±0.1 min ⁻¹ ; P =0.0432). Velocity by which the lymph is moved and refill time after manual emptying of the lymphatic vessels showed no differences between the 2 groups. The thoracic duct was elongated 10% ( P =0.0409) and with an abnormal course in the Fontan patients compared with normal. No difference in microvascular permeability was found between the 2 groups. Conclusions: Patients with a Fontan circulation have an impaired lymphatic pumping capacity and morphologically changed thoracic duct. Our results indicate a challenged lymphatic vasculature in the Fontan circulation and may play a role in the pathogenesis of the complications that are seen in Fontan patients. Clinical Trial Registration: URL: https://www.clinicaltrials.gov . Unique identifier: NCT03379805.

Early tetrapods faced an auditory challenge from the impedance mismatch between air and tissue in the transition from aquatic to terrestrial lifestyles during the Early Carboniferous (350 Ma). Consequently, tetrapods may have been deaf to airborne sounds for up to 100 Myr until tympanic middle ears evolved during the Triassic. The middle ear morphology of recent urodeles is similar to that of early ‘lepospondyl’ microsaur tetrapods, and experimental studies on their hearing capabilities are therefore useful to understand the evolutionary and functional drivers behind the shift from aquatic to aerial hearing in early tetrapods. Here, we combine imaging techniques with neurophysiological measurements to resolve how the change from aquatic larvae to terrestrial adult affects the ear morphology and sensory capabilities of salamanders. We show that air-induced pressure detection enhances underwater hearing sensitivity of salamanders at frequencies above 120 Hz, and that both terrestrial adults and fully aquatic juvenile salamanders can detect airborne sound. Collectively, these findings suggest that early atympanic tetrapods may have been pre-equipped to aerial hearing and are able to hear airborne sound better than fish on land. When selected for, this rudimentary hearing could have led to the evolution of tympanic middle ears.

Background: Videographic material of animals can contain inapparent signals, such as color changes or motion that hold information about physiological functions, such as heart and respiration rate, pulse wave velocity, and vocalization. Eulerian video magnification allows the enhancement of such signals to enable their detection. The purpose of this study is to demonstrate how signals relevant to experimental physiology can be extracted from non-contact videographic material of animals.Results: We applied Eulerian video magnification to detect physiological signals in a range of experimental models and in captive and free ranging wildlife. Neotenic Mexican axolotls were studied to demonstrate the extraction of heart rate signal of non-embryonic animals from dedicated videographic material. Heart rate could be acquired both in single and multiple animal setups of leucistic and normally colored animals under different physiological conditions (resting, exercised, or anesthetized) using a wide range of video qualities. Pulse wave velocity could also be measured in the low blood pressure system of the axolotl as well as in the high-pressure system of the human being. Heart rate extraction was also possible from videos of conscious, unconstrained zebrafish and from non-dedicated videographic material of sand lizard and giraffe. This technique also allowed for heart rate detection in embryonic chickens in ovo through the eggshell and in embryonic mice in utero and could be used as a gating signal to acquire two-phase volumetric micro-CT data of the beating embryonic chicken heart. Additionally, Eulerian video magnification was used to demonstrate how vocalization-induced vibrations can be detected in infrasound-producing Asian elephants.Conclusions: Eulerian video magnification provides a technique to extract inapparent temporal signals from videographic material of animals. This can be applied in experimental and comparative physiology where contact-based recordings (e.g., heart rate) cannot be acquired.

Previously, we showed that the evolution of high acuity vision in fishes was directly associated with their unique pH-sensitive hemoglobins that allow O₂to be delivered to the retina at PO₂s more than ten-fold that of arterial blood (Damsgaard et al., 2019). Here, we show strong evidence that vacuolar-type H⁺-ATPase and plasma-accessible carbonic anhydrase in the vascular structure supplying the retina act together to acidify the red blood cell leading to O₂secretion. In vivo data indicate that this pathway primarily affects the oxygenation of the inner retina involved in signal processing and transduction, and that the evolution of this pathway was tightly associated with the morphological expansion of the inner retina. We conclude that this mechanism for retinal oxygenation played a vital role in the adaptive evolution of vision in teleost fishes.