Distribution and morphology of calcitonin gene-related peptide (CGRP) innervation in flat mounts of whole rat atria and ventricles

Study Purpose: Calcitonin gene-related peptide (CGRP) is widely used as a marker for nociceptive afferent axons. However, the distribution of CGRP-IR axons has not been fully determined in the whole rat heart. To address this, we prepared whole rat heart tissues as flat-mounts, labeled them for CGRP, and acquired high quality images of the whole tissues. Our data shows the rather ubiquitous distribution of CGRP-IR axons in the whole rat heart at single-cell/axon/varicosity resolution for the first time. This study lays the foundation for future studies to quantify the differences in CGRP-IR axon innervation between sexes, disease models, and species.

Data Collection: Immunohistochemically labeled flat-mounts of the right and left atria and ventricles, and the interventricular septum in rats for CGRP were assessed with a Zeiss imager to generate complete montages of the entire atria, ventricles, and septum, and a confocal microscope was used to acquire detailed images of selected regions.

Primary Conclusion: We found that 1) CGRP-IR axons extensively innervated all regions of the atrial walls and the walls of the great vessels including the sinoatrial node region, auricles, atrioventricular node region, superior/inferior vena cava, left pre-caval vein, and pulmonary veins. 2) CGRP-IR axons formed varicose terminals around individual neurons in some cardiac ganglia but passed through other ganglia without making appositions with cardiac neurons. 3) Varicose CGRP-IR axons innervated the walls of blood vessels. 4) CGRP-IR axons extensively innervated the right/left ventricular walls and interventricular septum.

Curator's Notes

Experimental Design: Rats were initially deeply anesthetized with isoflurane and then perfused. The tissues were subsequently separated to create flat-mounts of a rat heart, including attached lungs, trachea, and esophagus. The heart was isolated from other structures, leading to the separation of the atria and ventricles. The brown adipose tissue and aortic arch were removed to expose the connected right atrium (RA) and left atrium (LA). The LA and pulmonary veins (PV) were then separated from the RA, which included the superior vena cava (SVC), left precaval vein (LPCV), and inferior vena cava (IVC). Additionally, the left ventricle (LV) and right ventricle (RV) were separated by cutting along the interventricular septum (IVS). Images from immunohistochemically labeled flat-mounts, obtained through stacks of optical sections (z-step: 1.5 μm for atria and 2 μm for ventricles), were captured using a Zeiss M2 Imager (20x lens; NA 0.8) and seamlessly stitched together to produce full photo montages of the right and left atria and ventricles. A LED light source with a 488 nm wavelength was utilized to visualize the CGRP-IR axons in the tissues.

Completeness: This dataset is complete.

Subjects & Samples: Male (n=4) adult rats (RRID:RGD_10395233) were used in this study.

Primary vs derivative data: The primary data folder is organized with subfolders sorted by subject ID and then sample ID. Within each sample subfolder, you'll find full Z-stack microscopy imaging files in the open image formats OME TIFF and JPX. The raw imaging data in .lif and .czi format is available in the source folder. In the derivative folder, the .psd files are presented as maximum projections and single-optical section images.