Objective

The present cardiac malformations in the chicken as a result of retinoic acid treatment are part of a continuous spectrum, varying from hearts with an intact ventricular septum and a normal course of the subaortic outflow tract to a double outlet right ventricle with a straddling tricuspid orifice or even a double inlet left ventricle. A remarkable observation in this spectrum concerns the correlation of malformations of the inflow and outflow tracts, which is explained as a cardiac looping disturbance. The disturbance of the looping process seems to lead to malalignment of septal components, although, in the chick, retinoic acid does not in general interfere with the formation of these septal components themselves.

Background: In a previous study retinoic acid treatment of chicken hearts has resulted in a spectrum of looping disturbances. Because of a decrease in contraction force of these hearts, the myocardial volume was hypothesized to be altered. Because retinoic acid has been suggested to influence endocardial cushion volumes, these were estimated as well.Methods: The previously studied hearts were used for estimating the absolute volumes of the atrial and ventricular myocardium and of the endocardial cushions by means of Cavalieri's principle. To measure the surface density of the trabeculations according to the isector method, we used retinoic acid treated hearts, which were perfusion fixed and in which the sections were isotropic uniform random. The volumes and surface densities found in the three morphologically distinguished groups, i.e., intact septum, isolated ventricular septal defect and double outlet right ventricle, were compared with those in shams.Results: A significant volume decrease was found in the right ventricular free wall myocardium of the double outlet right ventricle. No significant differences were found in the surface densities of the trabeculae. The volume of the atrioventricular cushion tissue in the double outlet right ventricle hearts was significantly increased. The morphological spectrum observed previously was also expressed in the right ventricular myocardial volume, which appeared to decrease from the least to the most malformed hearts, and in the volume of the atrioventricular cushion tissue, which appeared to increase.Conclusions: Several studies have shown pathology in myocardial and cushion tissue after retinoic acid treatment. In this study we have found a decreased growth of the right ventricular myocardium and an increased growth of the atrioventricular cushion tissue. We suggest that the previously found looping disturbance causes changed hemodynamics, as reported elsewhere, and that these result in changes in growth. We cannot exclude a direct effect of retinoic acid on the myocardium, which has to explain the looping disturbance. Anat.

Objective: To study the acute effect of epinephrine on hemodynamics of noninnervated normal and retinoic-acid-treated embryos. Design: Prospective interventional study design. Methods: A total of 190 stage 15 (50–55 h of incubation) chick embryos were randomly treated with 1 µg all-trans retinoic acid and reincubated. At stage 20 (day 3) and stage 24 (day 4), dorsal aortic flow velocities were measured with a 20-MHz pulsed Doppler velocity meter, in normal and retinoic-acid-treated embryos. Flow velocity waveforms were assessed both before and after the administration of epinephrine (5 or 10 µg). Results: Epinephrine caused a significant increase (p < 0.05) in heart rate, peak and mean velocities, peak acceleration, peak and mean blood flows, stroke volume and dorsal aortic area of both stage 20 and stage 24 normal and retinoic-acid-treated chick embryos. However, before epinephrine administration, stage 24 retinoic-acid-treated embryos displayed a significantly lesser increase in all outcome variables with the exception of dorsal aortic area. This was even observed after epinephrine administration. The effect of retinoic acid on cardiac output could not be compensated by epinephrine application. Conclusion: Epinephrine affects hemodynamics in both normal embryos and retinoic-acid-treated embryos prior to sympathetic innervation. A significant difference in hemodynamics exists between stage 24 normal and retinoic-acid-treated embryos. The underlying mechanism for the observed hemodynamic changes will need to be investigated.

The distribution of the vagal nerve was studied in whole-mount specimens and serial sections of chick embryos after retinoic acid treatment. White Leghorn chick embryos were treated at stage 15 either with 1 microg all-trans retinoic acid (n=11), or with the solvent dimethylsulphoxide (sham-operated embryos, n=8). Eight embryos served as normal controls. At stage 34 all 27 embryos were examined with a dissecting microscope. In order to reveal the vagal patterning, the hearts were removed and whole-mount stained with the HNK-1 antibody. In three hearts of the retinoic acid-treated group a morphologic intracardiac anomaly -- a double outlet right ventricle -- was found. To explore in depth the vagal nerve distribution in the heart, a separate set of hearts of retinoic acid embryos (n=5), sham-operated (n=4) and control embryos (n=5), was devised solely for serial sectioning and staining with the HNK-1 antibody. All hearts of retinoic acid-treated embryos showed a disturbed vagal nerve distribution both over the surface of the heart and within the heart wall. The vagal patterning was not altered in the sham-operated embryos compared to controls. It is concluded that retinoic acid disturbs the development of vagal nerve patterning regardless of the concurrent presence of intracardiac malformations. The mechanism and functional implications remain to be investigated.