Name: mesoscopic optical imaging of whole mouse heart
Uploaded: 2021-10-14T14:15:00
Description: Watch this Scientific Journal Video about Mesoscopic Optical Imaging of Whole Mouse Heart at JoVE.com

This project has received fundings from the European Union&#8217;s Horizon 2020 research and innovation programme under grant agreement No 952166 (REPAIR), MUR under the FISR program, project FISR2019_00320 and Regione Toscana, Bando Ricerca Salute 2018, PERCARE project.

All animal handling and procedures were performed in accordance with the guidelines from Directive 2010/63/EU of the European Parliament on the protection of animals used for scientific purposes and conformed to the principles and regulations of the Italian Ministry of Health. The experimental protocol was approved by the Italian Ministry of Health (protocol number 647/2015-PR). All the animals were provided by ENVIGO, Italy. For these experiments, 5 male C57BL/6J mice of 6 months of age were used.
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In this work, a successful approach to clear, stain, and image a whole mouse heart at high resolution was introduced. First, a tissue transformation protocol (CLARITY) was optimized and performed, slightly modified for its application on the cardiac tissue. Indeed, to obtain an efficient reconstruction in 3D of a whole heart, it is essential to prevent the phenomenon of light scattering. The CLARITY methodology allows us to obtain a highly transparent intact heart, but it requires long incubation times when performed pas...

Structural remodeling associated with cardiac diseases can affect electrical conduction and introduce electromechanical dysfunctions of the organ1,2. Current approaches used to predict functional alterations commonly employ MRI and DT-MRI to obtain an overall reconstruction of fibrosis deposition, vascular tree, and fiber distribution of the heart, and they are used to model preferential action potential propagation (APP) paths across the organ3,4. These strategies can provide a beautiful overview of the heart organization. However, their spatial resolution is insufficient to investigate the impact of structural remodeling on cardiac function at the cellular level.
Placing this framework at a different order of magnitude, where single cells can play individual roles on action potential propagation, is challenging. The main limitation is the inefficiency of standard imaging methods to perform high-resolution imaging (micrometric resolution) in massive (centimeter-sized) tissues. In fact, imaging biological tissues in 3D at high resolution is very complicated due to tissue opaqueness. The most common approach to perform 3D reconstructions in entire organs is to prepare thin sections. However, precise sectioning, assembling, and imaging require significant effort and time. An alternative approach that does not demand cutting the sample is to generate a transparent tissue. During the last years, several methodologies for clarifying tissues have been proposed5,6,7,8&#8288;. The challenge to produce massive, transparent, and fluorescently-labeled tissues has been recently achieved by developing true tissue transformation approaches (CLARITY9&#8288;, SHIELD10&#8288;). In particular, the CLARITY method is based on the transformation of an intact tissue into a nanoporous, hydrogel-hybridized, lipid-free form that enables to confer high transparency by the selective removal of membrane lipid bilayers. Notably, this method has been found successful also in cardiac preparation11,12,13,14&#8288;. However, since the heart is too fragile to be suitable for an active clearing, it must be cleared using the passive approach, which requires a long time to confer complete transparency.
In combination with advanced imaging techniques like light-sheet microscopy, CLARITY has the potential to image 3D massive heart tissues at micrometric resolution. In light-sheet microscopy, the illumination of the sample is performed with a thin sheet of light confined in the focal plane of the detection objective. The fluorescence emission is collected along an axis perpendicular to the illumination plane15. The detection architecture is similar to widefield microscopy, making the acquisition much faster than laser scanning microscopes. Moving the sample through the light sheet permits obtaining a complete tomography of big specimens, up to centimeter-sized samples. However, due to the intrinsic properties of the Gaussian beam, it is possible to obtain a very thin (of the order of a few microns) light-sheet only for a limited spatial extension, thus drastically limiting the field of view (FoV). Recently, a novel excitation scheme has been introduced to overcome this limitation and applied for brain imaging, allowing 3d reconstructions with isotropic resolution16.
In this paper, a passive clearing approach is presented, enabling a significant reduction of the clearing timing needed by the CLARITY protocol. The methodological framework described here allows reconstructing a whole mouse heart with micrometric resolution in a single tomographic scan with an acquisition time in the order of minutes.

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mesoscopic optical imaging of whole mouse heart

Both genetic and non-genetic cardiac diseases can cause severe remodeling processes in the heart. Structural remodeling, such as collagen deposition (fibrosis) and cellular misalignment, can affect electrical conduction, introduce electromechanical dysfunctions and, eventually, lead to arrhythmia. Current predictive models of these functional alterations are based on non-integrated and low-resolution structural information. Placing this framework on a different order of magnitude is challenging due to the inefficacy of standard imaging methods in performing high-resolution imaging in massive tissue. In this work, we describe a methodological framework that allows imaging of whole mouse hearts with micrometric resolution. The achievement of this goal has required a technological effort where advances in tissue transformation and imaging methods have been combined. First, we describe an optimized CLARITY protocol capable of transforming an intact heart into a nanoporous, hydrogel-hybridized, lipid-free form that allows high transparency and deep staining. Then, a fluorescence light-sheet microscope able to rapidly acquire images of a mesoscopic field of view (mm-scale) with the micron-scale resolution is described. Following the mesoSPIM project, the conceived microscope allows the reconstruction of the whole mouse heart with micrometric resolution in a single tomographic scan. We believe that this methodological framework will allow clarifying the involvement of the cytoarchitecture disarray in the electrical dysfunctions and pave the way for a comprehensive model that considers both the functional and structural data, thus enabling a unified investigation of the structural causes that lead to electrical and mechanical alterations after the tissue remodeling.

The developed passive clearing setup allows to obtain a cleared adult mouse heart (with a dimension of the order 10 mm x 6 mm x 6 mm) in about 3 months. All the components of the setup are mounted as shown in Figure 1. The negligible temperature gradient between each clearing chamber (of the order of 3&#176;C) allows maintaining the temperature in a proper range across all chambers.
<img alt="Figure 1" class="xfigimg" src="/files/f...

Watch this Scientific Journal Video about Mesoscopic Optical Imaging of Whole Mouse Heart at JoVE.com

Mesoscopic Optical Imaging of Whole Mouse Heart

We report a method for mesoscopic reconstruction of the whole mouse heart by combining new advancements in tissue transformation and staining with the development of an axially scanned light-sheet microscope.

Both genetic and non-genetic cardiac diseases can cause severe remodeling processes in the heart. Structural remodeling, such as collagen deposition ...

This methodology combines improvement of clearing protocol with the eye capability of optical sectioning of the Mesos PIM technology allowing us to perform a meso scopic reconstructions at micrometric resolution. It provides the possibility to image massive cardiac tissues or entire mouse hearts entire solution in a single scan without losing the original three-dimensional tissue organization. After the heart has been cannulated by proximal aorta washed in tyro solution and fixed in 4%Paraform aldehyde in 0.01 molar PBS it is ready to start the clearing protocol.Wash the heart three times in 0.01 molar PBS at four degrees Celsius for 15 minutes. After this step, the heart can be stored in PBS and 0.01%sodium azide at four degrees Celsius for several months. Incubate the heart in 20 milliliters of hydrogel solution in shaking condition at 15 RPM for three days at four degrees Celsius.De-gas the sample at room temperature using a dryer a vacuum pump, and a tube system that connects the dryer to both the pump band and nitrogen pipeline. Place the sample in the dryer and open the vial keeping the cap on it. Close the dryer and remove the oxygen from the tube by opening the nitrogen pipeline.Turn on the vacuum pump to remove the oxygen from the dryer for 10 minutes. Turn off the pump and open the knob of the dryer to the nitrogen pipeline. Then open the nitrogen tap.Once the pressure is equal to the atmospheric pressure carefully open the dryer and quickly close the vial. Keep the heart in the De-gased hydrogel solution at 37 degrees Celsius for about three hours at rest. When the hydrogel is properly polymerized and appears entirely gelatinous, carefully remove the heart from it and place it in a clearing solution in the vial with clearing solution.Change the clearing solution in the vial once every three days to speed up the clarification procedure. Once the heart appears completely clarified remove it from the vial and wash it in 50 milliliters of warmed up PBS for 24 hours in shaking condition. Wash again in 50 milliliters of one X PBS T for 24 hours in shaking condition.Incubate the sample in 0.01 milligrams per milliliters wheat germ aglutinin Alexa floor 633 in three milliliters of one X PBS T in shaking condition at 50 RPM at room temperature for seven days. After the seven day incubation, wash the sample in 50 milliliters of one X PBS T at room temperature in shaking for 24 hours. Incubate the sample in 4%PFA for 15 minutes and then wash it three times in 0.01 molar PBS for five minutes each.Incubate the heart successively in 20 and 47%TDE in 0.01 molar PBS for eight hours each. And then finally at 68%TDE in 0.01 molar PBS to provide the required refractive index of 1.46. Gently fill about 80%of the external quartz quvet with the refractive index medium.Fill the internal quartz quvet with the same refractive index medium. Immerse the sample inside the internal quvet. Gently move the sample to the bottom of the quvet using thin tweezers and arrange the heart with its longitudinal axis parallel to the quvet's main axis to minimize the excitation light path across the tissue during the scanning.Carefully fix the tailored plug above the internal quvet with two screws. Mount the sample to the microscope stage using magnets. Translate the vertical sample stage manually to immerse the internal quvet into the external one.Turn on the excitation light source with a wavelength of 638 nanometers and set it at low power in the order of three milliwatts. Move the sample using the motorized translator to illuminate an inner plate of the tissue. Turn on the HC image live camera software and set the camera trigger on external edge trigger light sheet mode to drive the acquisition trigger of the camera by the custom software controlling the entire setup.Enable auto save in the camera software and set the output folder where the images need to be saved. Manually adjust the sample position in the X Y plane with the linear translators to move the sample to the center of the field of view of the camera sensor. Move the sample along the Z axis using the linear motorized translator to identify heart borders for tomographic reconstruction.Increase the laser power to approximately 20 milliwatts before beginning for the imaging session. Start the tomographic acquisition by clicking the start button in the capture panel of the imaging software, and at the same time start moving the sample along the Z-axis at the constant velocity of six micrometers per second using the motorized translator. The combination of the clarity methodology with TDE as refractive index medium does not significantly change the sample's final volume nor leads to an isotropic deformation of the specimen.The excitation optics produced a light sheet with a minimum waste of about six micrometers that diverged up to 175 micrometers at the edge of the field of view. The synchronization of the camera rolling shutter with the axial scan of the light sheet waste ensured to collect the emission signal only in the sample portion excited with the waste of the light sheet resulting in an average F W H M of about 6.7 micrometers along the entire field of view. The Z point spread function of the microscope was also estimated by a tomographic reconstruction of the fluorescent nanosphere and an F W H M of 6.4 micrometers was estimated by the fit.The potentiality of the system to resolve single cellular membranes in three dimensions with a sufficient signal to noise ratio in the entire organ was also confirmed. The de-gassing procedure is the most critical step of the protocol. If it is not properly performed the tissue could encounter damages indicating during the incubation in a cleaning solution.The presented protocol can be combined with a multi staining protocol to achieve a whole organ reconstruction integrating different biological structures and can be applied to study pathological models.

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Proper Care and Cleaning of the Microscope

Keeping the microscope optics clean is important for high-quality imaging. Dust, fingerprints, excess immersion oil, or mounting medium on or in a microscope causes reduction in contrast and resolution. DIC is especially sensitive to contamination and scratches on the lens surfaces. This protocol details the procedure for keeping the microscope clean.

Keeping the microscope optics clean is important for high-quality imaging. Dust, fingerprints, excess immersion oil, or mounting medium on or in a microscope causes reduction in contrast and resolution. DIC is especially sensitive to contamination and scratches on the lens surfaces. This protocol details the procedure for keeping the microscope clean.

Keeping the microscope optics clean is important for high-quality imaging. Dust, fingerprints, excess immersion oil, or mounting medium on or in a ...

Major Components of the Light Microscope

The light microscope is a basic tool for the cell biologist, who should have a thorough understanding of how it works, how it should be aligned for different applications, and how it should be maintained as required to obtain maximum image-forming capacity and resolution. The components of the microscope are described in detail here.

The light microscope is a basic tool for the cell biologist, who should have a thorough understanding of how it works, how it should be aligned for different applications, and how it should be maintained as required to obtain maximum image-forming capacity and resolution. The components of the microscope are described in detail here.

The light microscope is a basic tool for the cell biologist, who should have a thorough understanding of how it works, how it should be aligned for ...

Phase Contrast and Differential Interference Contrast (DIC) Microscopy

Phase-contrast microscopy is often used to produce contrast for transparent, non light-absorbing, biological specimens. The technique was discovered by Zernike, in 1942, who received the Nobel prize for his achievement. DIC microscopy, introduced in the late 1960s, has been popular in biomedical research because it highlights edges of specimen structural detail, provides high-resolution optical sections of thick specimens including tissue cells, eggs, and embryos and does not suffer from the  phase halos  typical of phase-contrast images. This protocol highlights the principles and practical applications of these microscopy techniques.

Phase Contrast and Differential Interference Contrast DIC Microscopy

This protocol highlights the principles and practical applications of Phase and Differential Interference Contrast (DIC) Microscopy

Phase-contrast microscopy is often used to produce contrast for transparent, non light-absorbing, biological specimens. The technique was discovered by ...

Born Normalization for Fluorescence Optical Projection Tomography for Whole Heart Imaging

Optical projection tomography is a three-dimensional imaging technique that has been recently introduced as an imaging tool primarily in developmental biology and gene expression studies. The technique renders biological sample optically transparent by first dehydrating them and then placing in a mixture of benzyl alcohol and benzyl benzoate in a 2:1 ratio (BABB or Murray s Clear solution). The technique renders biological samples optically transparent by first dehydrating them in graded ethanol solutions then placing them in a mixture of benzyl alcohol and benzyl benzoate in a 2:1 ratio (BABB or Murray s Clear solution) to clear. After the clearing process the scattering contribution in the sample can be greatly reduced and made almost negligible while the absorption contribution cannot be eliminated completely. When trying to reconstruct the fluorescence distribution within the sample under investigation, this contribution affects the reconstructions and leads, inevitably, to image artifacts and quantification errors.. While absorption could be reduced further with a permanence of weeks or months in the clearing media, this will lead to progressive loss of fluorescence and to an unrealistically long sample processing time. This is true when reconstructing both exogenous contrast agents (molecular contrast agents) as well as endogenous contrast (e.g. reconstructions of genetically expressed fluorescent proteins).

We suggest a Born normalized approach for Optical Projection Tomography (BnOPT) that accounts for the absorption properties of imaged samples to obtain accurate and quantitative fluorescence tomographic reconstructions. We use the proposed algorithm to reconstruct the fluorescence molecular probe distribution within small animal organs.

Optical projection tomography is a three-dimensional imaging technique that has been recently introduced as an imaging tool primarily in developmental ...

Semi-automated Optical Heartbeat Analysis of Small Hearts

We have developed a method for analyzing high speed optical recordings from Drosophila, zebrafish and embryonic mouse hearts (Fink, et. al., 2009). Our Semi-automatic Optical Heartbeat Analysis (SOHA) uses a novel movement detection algorithm that is able to detect cardiac movements associated with individual contractile and relaxation events. The program provides a host of physiologically relevant readouts including systolic and diastolic intervals, heart rate, as well as qualitative and quantitative measures of heartbeat arrhythmicity. The program also calculates heart diameter measurements during both diastole and systole from which fractional shortening and fractional area changes are calculated. Output is provided as a digital file compatible with most spreadsheet programs. Measurements are made for every heartbeat in a record increasing the statistical power of the output. We demonstrate each of the steps where user input is required and show the application of our methodology to the analysis of heart function in all three genetically tractable heart models.

We have developed a Semi-automated Optical Heartbeat Analysis method (SOHA) for analyzing high speed optical recordings from Drosophila, zebrafish and embryonic mouse hearts. We demonstrate the application of our methodology to the analysis of heart function in fruit fly and embryonic mouse hearts.

We have developed a method for analyzing high speed optical recordings from Drosophila, zebrafish and embryonic mouse hearts (Fink, et. al., 2009). Our ...

Mesoscopic Fluorescence Tomography for In-vivo Imaging of Developing Drosophila

Visualizing developing organ formation as well as progession and treatment of disease often heavily relies on the ability to optically interrogate molecular and functional changes in intact living organisms. Most existing optical imaging methods are inadequate for imaging at dimensions that lie between the penetration limits of modern optical microscopy (0.5-1mm) and the diffusion-imposed limits of optical macroscopy (&gt;1cm) [1]. Thus, many important model organisms, e.g. insects, animal embryos or small animal extremities, remain inaccessible for in-vivo optical imaging. 
Although there is increasing interest towards the development of nanometer-resolution optical imaging methods, there have not been many successful efforts in improving the imaging penetration depth. The ability to perform in-vivo imaging beyond microscopy limits is in fact met with the difficulties associated with photon scattering present in tissues. Recent efforts to image entire embryos for example [2,3] require special chemical treatment of the specimen, to clear them from scattering, a procedure that makes them suitable only for post-mortem imaging. These methods however evidence the need for imaging larger specimens than the ones usually allowed by two-photon or confocal microscopy, especially in developmental biology and in drug discovery.
We have developed a new optical imaging technique named Mesoscopic Fluorescence Tomography [4], which appropriate for non-invasive in-vivo imaging at dimensions of 1mm-5mm. The method exchanges resolution for penetration depth, but offers unprecedented tomographic imaging performance and it has been developed to add time as a new dimension in developmental biology observations (and possibly other areas of biological research) by imparting the ability to image the evolution of fluorescence-tagged responses over time. As such it can accelerate studies of morphological or functional dependencies on gene mutations or external stimuli, and can importantly, capture the complete picture of development or tissue function by allowing longitudinal time-lapse visualization of the same, developing organism.
The technique utilizes a modified laboratory microscope and multi-projection illumination to collect data at 360-degree projections. It applies the Fermi simplification to Fokker-Plank solution of the photon transport equation, combined with geometrical optics principles in order to build a realistic inversion scheme suitable for mesoscopic range. This allows in-vivo whole-body visualization of non-transparent three-dimensional structures in samples up to several millimeters in size.
We have demonstrated the in-vivo performance of the technique by imaging three-dimensional structures of developing Drosophila tissues in-vivo and by following the morphogenesis of the wings in the opaque Drosophila pupae in real time over six consecutive hours.

Mesoscopic Fluorescence Tomography for In-vivo Imaging of Developing Drosophila

Mesoscopic fluorescence tomography operates beyond the penetration limits of tissue-sectioning fluorescence microscopy. The technique is based on multi-projection illumination and a photon transport description. We demonstrate in-vivo whole-body 3D visualization of the morphogenesis of GFP-expressing wing imaginal discs in Drosophila melanogaster.

Visualizing  developing organ formation as well as progession and treatment of disease often  heavily relies on the ability to optically interrogate ...

In vivo Imaging and Therapeutic Treatments in an Orthotopic Mouse Model of Ovarian Cancer

Human cancer and response to therapy is better represented in orthotopic animal models. This paper describes the development of an orthotopic mouse model of ovarian cancer, treatment of cancer via oral delivery of drugs, and monitoring of tumor cell behavior in response to drug treatment in real time using in vivo imaging system. In this orthotopic model, ovarian tumor cells expressing luciferase are applied topically by injecting them directly into the mouse bursa where each ovary is enclosed. Upon injection of D-luciferin, a substrate of firefly luciferase, luciferase-expressing cells generate bioluminescence signals. This signal is detected by the in vivo imaging system and allows for a non-invasive means of monitoring tumor growth, distribution, and regression in individual animals. Drug administration via oral gavage allows for a maximum dosing volume of 10 mL/kg body weight to be delivered directly to the stomach and closely resembles delivery of drugs in clinical treatments. Therefore, techniques described here, development of an orthotopic mouse model of ovarian cancer, oral delivery of drugs, and in vivo imaging, are useful for better understanding of human ovarian cancer and treatment and will improve targeting this disease.

In vivo Imaging and Therapeutic Treatments in an Orthotopic Mouse Model of Ovarian Cancer

Orthotopic animal models of ovarian cancer replicate better human disease and therefore enhance our understanding of cancer progression and tumor response to therapy. A mouse model receives an intrabursal injection of luciferase-expressing ovarian tumor cells. Treatment is administered via oral gavage. Tumor growth is monitored by in vivo imaging system.

Human cancer and response to therapy is better represented in orthotopic animal models.  This paper describes the development of an orthotopic mouse model ...

A Novel Ex vivo Culture Method for the Embryonic Mouse Heart

Developmental studies in the mouse are hampered by the inaccessibility of the embryo during gestation. Thus, protocols to isolate and culture individual organs of interest are essential to provide a method of both visualizing changes in development and allowing novel treatment strategies. To promote the long-term culture of the embryonic heart at late stages of gestation, we developed a protocol in which the excised heart is cultured in a semi-solid, dilute Matrigel. This substrate provides enough support to maintain the three-dimensional structure but is flexible enough to allow continued contraction. In brief, hearts are excised from the embryo and placed in a mixture of cold Matrigel diluted 1:1 with growth medium. After the diluted Matrigel solidifies, growth medium is added to the culture dish. Hearts excised as late as embryonic day 16.5 were viable for four days post-dissection. Analysis of the coronary plexus shows that this method does not disrupt coronary vascular development. Thus, we present a novel method for long-term culture of embryonic hearts.

A Novel Ex vivo Culture Method for the Embryonic Mouse Heart

Developmental studies in the mouse are hampered by the inaccessibility of the embryo during gestation. To promote the long-term culture of the embryonic heart at late stages of gestation, we developed a protocol in which the excised heart is cultured in a semi-solid, dilute Matrigel.

Developmental studies in the mouse are hampered  by the inaccessibility of the embryo during gestation. Thus, protocols to  isolate and culture individual ...

Optical Mapping of Intra-Sarcoplasmic Reticulum Ca2+ and Transmembrane Potential in the Langendorff-perfused Rabbit Heart

Sarcoplasmic reticulum (SR) Ca2+ handling plays a key role in normal excitation-contraction coupling and aberrant SR Ca2+ handling is known to play a significant role in certain types of arrhythmia. Because arrhythmias are spatially distinct, emergent phenomena, they must be investigated at the tissue level. However, methods for directly probing SR Ca2+ in the intact heart remain limited. This article describes the protocol for dual optical mapping of transmembrane potential (Vm) and free intra-SR [Ca2+] ([Ca2+]SR) in the Langendorff-perfused rabbit heart. This approach takes advantage of the low-affinity Ca2+ indicator Fluo-5N, which has minimal fluorescence in the cytosol where intracellular [Ca2+] ([Ca2+]i) is relatively low but exhibits significant fluorescence in the SR lumen where [Ca2+]SR is in the millimolar range. In addition to revealing SR Ca2+ characteristics spatially across the epicardial surface of the heart, this approach has the distinct advantage of simultaneous monitoring of Vm, allowing for investigations into the bidirectional relationship between Vm and SR Ca2+ and the role of SR Ca2+ in arrhythmogenic phenomena.

Optical Mapping of Intra-Sarcoplasmic Reticulum Ca2+ and Transmembrane Potential in the Langendorff-perfused Rabbit Heart

This article describes the detailed protocol and equipment necessary for dual optical mapping of transmembrane potential (Vm) and free intra-sarcoplasmic reticulum (SR) Ca2+ in the Langendorff-perfused rabbit heart. This method allows for direct observation and quantification of Vm and SR Ca2+ dynamics in the intact heart.

Sarcoplasmic reticulum (SR) Ca2+ handling plays a key role in normal excitation-contraction coupling and aberrant SR Ca2+ handling is known to play a ...

Drosophila Preparation and Longitudinal Imaging of Heart Function In Vivo Using Optical Coherence Microscopy (OCM)

Longitudinal study of the heartbeat in small animals contributes to understanding structural and functional changes during heart development. Optical coherence microscopy (OCM) has been demonstrated to be capable of imaging small animal hearts with high spatial resolution and ultrahigh imaging speed. The high image contrast and noninvasive properties make OCM ideal for performing longitudinal studies without requiring tissue dissections or staining. Drosophila has been widely used as a model organism in cardiac developmental studies due to its high number of orthologous human disease genes, its similarity of molecular mechanisms and genetic pathways with vertebrates, its short life cycle, and its low culture cost. Here, the experimental protocols are described for the preparation of Drosophila and optical imaging of the heartbeat with a custom OCM system throughout the life cycle of the specimen. By following the steps provided in this report, transverse M-mode and 3D OCM images can be acquired to conduct longitudinal studies of the Drosophila cardiac morphology and function. The en face and axial sectional OCM images and the heart rate (HR) and cardiac activity period (CAP) histograms, were also shown to analyze the heart structural changes and to quantify the heart dynamics during Drosophila metamorphosis, combined with the videos constructed with M-mode images to trace cardiac activity intuitively. Due to the genetic similarity between Drosophila and vertebrates, longitudinal study of heart morphology and dynamics in fruit flies could help reveal the origins of human heart diseases. The protocol here would provide an effective method to perform a wide range of studies to understand the mechanisms of cardiac diseases in humans.

Drosophila Preparation and Longitudinal Imaging of Heart Function In Vivo Using Optical Coherence Microscopy OCM

Here, the experimental protocols are described for preparing Drosophila at different developmental stages and performing longitudinal optical imaging of Drosophila heartbeats using a custom optical coherence microscopy (OCM) system. The cardiac morphological and dynamical changes can be quantitatively characterized by analyzing the heart structural and functional parameters from OCM images.

Longitudinal study of the heartbeat in small animals contributes to understanding structural and functional changes during heart development. Optical ...