MYOCARDIAL STRUCTURE AND VASCULARIZATION OF THE HEART VENTRICLE IN HOLOCEPHALI: IMPLICATIONS FOR HEART EVOLUTION

El resumen aparece en el Program & Abstracts of the 10th International Congress of Vertebrate Morphology, Barcelona 2013. Anatomical Record, Volume 296, Special Feature — 1: P-075.It has been classically assumed that the ventricle of the primitive vertebrate heart is composed of spongy myocardium, supplied exclusively by oxygen-poor, luminal blood. This idea is on two facts: (1) extant agnathans have a spongy ventricular myocardium, and (2) in avian and mammalian embryos, the formation of trabeculated myocardium precedes the appearance of compact myocardium. Recently, it has been proposed that, like elasmobranchs, the early gnathostomes possess a fully vascularised ventricle composed of mixed myocardium. We tested this idea by studying the structure and vascularisation of the ventricular myocardium in four holocephalan species of the families Chimaeridae and Rhinochimaeridae. Chimaera monstrosa, Hidrolagus affinis and Harriotta raleighana have a spongy myocardium covered by a thin layer of cardiac muscle. In H. raleighana, the compacta is reduced to an extremely fine rim. In all three species there is a well-developed coronary artery system consisting of subepicardial vessels which give off branches that penetrate the myocardial trabeculae. Rhinochimaera atlantica has no compacta and its ventricular coronary artery system is reduced to subepicardial vessels that do not enter the spongy layer. This report is the first to show that in wild living vertebrates, a coronary artery system supplying the whole myocardium exists in the absence of a well-developed compacta, which supports experimental work that shows that myocardial cell proliferation and coronary vascular growth rely on genetically separated programs. We conclude that the mixed ventricular myocardium is primitive for chondrichthyans, and that the lack of compacta in some holocephalans is a derived character. Moreover our results support the hypotheses that the mixed myocardium is the primitive condition in gnathostomes, and that the absence of a compacta in different actinopterygian taxa is the result of its repeated loss during evolution.Proyecto CGL2010-16417/BOS; Fondos FEDE

The bulbus arteriosus of the holocephalan heart

El resumen aparece en el Program & Abstracts of the 10th International Congress of Vertebrate Morphology, Barcelona 2013.Anatomical Record, Volume 296, Special Feature — 1: P-074.Previous work has shown that the outflow tract of the elasmobranch heart, namely the cardiac portion intercalated between the ventricle and the ventral aorta, does not consist of a single component, the conus arteriosus, as has classically been assumed, but two, the myocardial conus arteriosus and the non-myocardial bulbus arteriosus. From the evolutionary perspective, knowledge of the anatomy of the cardiac outflow tract of the holocephali is important, as they are the sister group of elasmobranchs. Our aim is to describe the cardiac outflow tract of four holocephalan species, two of them, Chimaera monstrosa and Hydrolagus affinis of the family Chimaeridae, and the other two, Harriotta raleighana and Rhinochimaera atlantica, of the family Rhinochimaeridae. The cardiac outflow tract of the four species consisted of a myocardial conus arteriosus, furnished with valves, and a bulbus arteriosus devoid of cardiac muscle. Both the bulbus and conus are tubular in shape. The length of the bulbus relative to the total length of the outflow tract is somewhat smaller in the rhinochimaerids (15%-19%) than in the chimaerids (19%-23%). The bulbus is covered by epicardium and is crossed by the main coronary artery trunks. Histologically, the bulbus is mainly composed of elastin and collagen, and, to a lesser extent, by smooth muscle. This suggests that in holocephalans, the bulbus actively helps to protect the gill vasculature from exposure to high-pressure pulses of blood. Our results prove that the bulbus arteriosus is common to chondrichthyans. In addition, they support the hypothesis that the cardiac outflow tract consisted of a conus arteriosus and a bulbus arteriosus from the beginning of the jawed vertebrate radiation, contributing to our understanding of the morphological changes that have occurred at the arterial pole of the heart in both actinopterygians and sarcopterygians.Proyecto CGL2010-16417/BOS; Fondos FEDE

Effect of hyperlipidic diets on normal and abnormal aortic valves in the Syrian hamster: A preliminary study

Effect of hyperlipidic diets on normal and abnormal aortic valves in the Syrian hamster: A preliminary study. MC Fernández 1,2, J Moncayo-Arlandi 1, MT Soto 1, MA López-Unzu 1, B Fernández 1,2 and AC Durán 1,2. 1 Department of Animal Biology, Faculty of Science, University of Málaga, Spain. 2 Biomedical Research Institute of Málaga (IBIMA), University of Málaga, Spain. Bicuspid aortic valve (BAV) is the most frequent human congenital cardiac malformation. It frequently becomes stenotic due to calcification by an atherosclerosis-like process. Hyperlipidic diets have been classically used to induce atherosclerosis in laboratory animals, including Syrian hamsters. The aim here is to evaluate the effect of hyperlipidic diets in hamsters having different incidence of BAVs. We used a unique inbred strain of Syrian hamsters with a high ( 40%) incidence of spontaneous BAV, morphologically similar to that in man, another inbred strain with a low ( 4%) incidence of BAV, and an outbred, second control line, acquired from Charles River Laboratories. Three experimental groups were fed with standard diet supplemented with 2% cholesterol plus 15% butter during five months. In parallel, three control groups were fed with unmodified standard diet. Hyperlipidic diets induced lesions in the aortic valve and ascending aortic wall, i.e. subendothelial lipid deposits, valve sclerosis, and neo-intima in the aorta. We performed a preliminary, qualitative, comparative study of the lesions associated with the different animal populations and valvular phenotypes. Our results indicate that (1) the type and severity of the lesions varied among the three hamster populations, suggesting that genetic factors may be involved; (2) the aortic valve morphology seems not to determine the severity of the valvular lesions. We conclude that our hamster strain with high incidence of BAV is a promising animal model for studies on human aortic stenosis. This work was supported by P10-CTS-6068.Universidad de Málaga. Campus de Excelencia Andalucía Tech. P10-CTS-6068

The Myosin Heavy Chain specific A4.1025 antibody discriminates different cardiac segments in ancient groups of gnathostomes: Morphological and evolutionary implications

El resumen aparece en el Program & Abstracts of the 11th International Congress of Vertebrate Morphology, Washington DC 2016. Anatomical Record, Volume 299, Special Feature: 263.The pan-Myosin Heavy Chain (pan-MyHC) marker MF20 have been reported to show similar, homogeneous signal in the myocardial segments of the heart of teleosts and tetrapods. However, in an ongoing study of the myocardial structure of the dogfish (Scyliorhinus canicula; Chondrichthyes), we observed differential immunostaining of the cardiac segments using another pan-MyHC, the A4.1025 antibody. In order to investigate the relevance of this finding for better understanding of the morphology and evolution of the vertebrate heart, we performed immunohistochemistry, slot blot and western blot in several species of chondrichthyans, actinopterygians and mammals using the above mentioned antibodies. In the dogfish heart, A4.1025 and MF20 specifically recognized MyHC isoforms, although with different degree of affinity. MF20 reactivity was homogeneous and high in all the myocardial segments. However, A4.1025 reactivity was heterogeneous. It was high in the sinus venosus (external layer), atrium and atrioventricular region, low in the ventricle and conus arteriosus, and null in the internal layer of the sinus venosus. A heterogeneous pattern of A4.1025 immunoreactivity was also detected in two other elasmobranchs, a holocephalan, a polypteryform and an acipenseriform. In all of these species, MF20 immunoreactivity was homogeneous. In addition, both markers showed a homogeneous immunoreactivity pattern in teleosts and mammals. Our results indicate that in the hearts of ancient gnathostomes, in all of which a conspicuous conus arteriosus exists, one or more MyHC isoforms with low affinity for A4.1025 show segment-specific distributions. Thus, A4.1025 appears to be an appropriated marker to identify the cardiac segments and their boundaries. We propose that the segmentspecific distribution of MyHC isoforms may generate a particular type of myocardial contractility associated with the presence of a conus arteriosus.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech. CGL2014-52356-P, CEIMAR, BIO 203, FEDE

Anatomical, histochemical and immunohistochemical characterization of the outflow tract of ray hearts (Rajiformes; Chondrichthyes)

El resumen aparece en el Program & Abstracts of the 11th International Congress of Vertebrate Morphology, Washington DC 2016. Anatomical Record, Volume 299, Special Feature: 264.Recent work has shown that the cardiac outflow tract of sharks and chimaeras does not consist of a single myocardial component, the conus arteriosus, as classically accepted, but two, namely, the myocardial conus arteriosus and the non-myocardial bulbus arteriosus. However, the anatomical composition of the outflow tract of the batoid hearts remains unknown. The present study was designed to fill this gap. The material examined consisted of hearts of two species of rays, namely, the Mediterranean starry ray (Raja asterias) and sandy ray (Leucoraja circularis). They were studied using scanning electron microscopy, and histochemical and inmunohistochemical techniques. In both species, the outflow tract consists of two components, proximal and distal with regard to the ventricle. The proximal component is the conus arteriosus; it is characterized by the presence of compact myocardium in its wall and several transverse rows of pocket-shaped valves at its luminal side. Each valve consists of a leaflet and its supporting sinus. Histologically, the leaflet has two fibrosas, inner and outer, and a middle coat, the spongiosa. The distal component lacks myocardium. Its wall consists of smooth muscle cells, elastic fibers and collagen. Thus, it shows an arterial-like structure. However, it differs from the aorta because it is covered by the epicardium and crossed by coronary arteries. These findings indicate that the distal component is morphologically equivalent to the bulbus arteriosus of sharks and chimaeras. In contrast to foregoing descriptions, the valves of the first transverse row are distally anchored to the bulbus arteriosus and not to the ventral aorta. Our findings give added support to the notion that presence of a bulbus arteriosus at the arterial pole of the heart is common to all chondrichtyans, and not an apomorphy of actinopterygians as classically thought.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech. CGL2014-52356-P, CEIMAR, BIO 203, FEDE

Los componentes del tracto de salida cardiaco de los primeros vertebrados mandibulados están conservados en los anfibios

Estudios recientes han modificado las nociones que se habían asumido clásicamente sobre la composición del tracto de salida cardiaco de los condrictios. En este grupo el tracto de salida consta, no sólo de un cono arterioso, de naturaleza miocárdica, sino también de un bulbo arterioso, cuya pared carece de miocardio. El bulbo es un segmento intrapericárdico, interpuesto entre el cono y la aorta ventral, tapizado por epicardio y recorrido por arterias coronarias. Dichos estudios sugieren que este diseño del tracto de salida se corresponde con la condición primitiva del carácter en los vertebrados mandibulados. Además, otra serie de investigaciones han puesto de manifiesto que el bulbo arterioso de los peces es homólogo a las porciones basales intrapericárdicas de la aorta y de la arteria pulmonar de las aves y los mamíferos. Por su parte, aunque la anatomía y fisiología del tracto de salida del corazón de los anfibios constituyen temas frecuentemente tratados en la bibliografía, existe una notable confusión en la terminología empleada para designar esta parte del corazón, motivada, probablemente, por la escasez de estudios microscópicos sobre este componente cardiaco. A tenor de los antecedentes expuestos, se ha estudiado el tracto de salida cardiaco en ejemplares adultos del anuro Xenopus laevis. Dicho estudio ha consistido en el examen anatómico de material fresco, la observación de muestras mediante microscopía electrónica de barrido, la aplicación de técnicas histomorfológicas para microscopía óptica y la realización de reconstrucciones digitales tridimensionales. El tracto de salida cardiaco de X. laevis consta, como el de los vertebrados pisciformes, de dos componentes, uno proximal y otro distal con respecto al ventrículo. El componente proximal es el cono arterioso, que está recorrido internamente por la cresta espiral y presenta, en ambos extremos, sendas hileras de válvulas. La pared del cono es de naturaleza miocárdica y está irrigada por arterias coronarias. El componente distal es asimismo intrapericárdico, está compuesto por musculatura lisa y tejido elástico, está tapizado por epicardio y recorrido por arterias coronarias y muestra una tabicación interna progresivamente más compleja en sentido caudocefálico. Nuestros resultados sugieren que dicho componente distal, que constituye las bases intrapericárdicas de las arterias sistémico-carotídeas y pulmo-cutáneas, es homólogo al bulbo arterioso de los vertebrados pisciformes. Por tanto, el bulbo arterioso se ha conservado desde los primeros vertebrados mandibulados y a partir de él se han formado las bases de los troncos aórticos y pulmonares de los tetrápodos.CGL2014-52356-P (Ministerio de Economía y Competitividad), FPU15/03209 (Ministerio de Educación, Cultura y Deporte), fondos FEDER, Universidad de Málaga.Campus de Excelencia Internacional Andalucía Tech

Hospitalidad y muerte en la Ruta Jacobea navarra. Evidencias arqueológicas e históricas

Una ruta tan transitada a lo largo de más de un milenio como es la ruta jacobea, ha hecho que la hospitalidad y la muerte estén íntimamente ligadas a ella. Además de numerosa documentación referente a este hecho, existen una serie de evidencias arqueológicas en Navarra que conviene tener en cuenta, pues son diversos los enterramientos conocidos y excavados a lo largo del Camino y en otros lugares, que muestran la presencia de peregrinos o bien miembros de alguna de las cofradías santiaguistas de la región, aspecto que se manifiesta, entre otros elementos, por haber sido enterrados con una vieira, símbolo inequívoco de haber realizado la pe- regrinación al sepulcro del Apóstol. Las diversas evidencias arqueológicas se han localizado junto a la ermita de San Salvador de Ibañeta, en Roncesvalles, en Pamplona (entorno de la Catedral, plaza del Castillo, casa del Condestable), en Santa María de Eunate, en el Santo Sepulcro de Estella, en el Santo Sepulcro de Torres del Río, en San Saturnino de Artajona, en el despoblado de Arlás en Peralta y en Santa María de Ujué.Such a well-trodden route as the Camino de Santiago in the last millennium is inevitably bound up with hospitality and death. As well as voluminous documentation referring to these aspects, there is archeological evidence in Navarra that one has to bear in mind, because of the burials that are known and have been excavated along the Camino and in other places, which reveal the presence of pilgrims or members of one of the guilds associated with St James in the region, shown by the fact that a shell, a symbol of having done the pilgrimage to Santiago, is included among other elements of burial. Diverse archeological evidence has been located beside the church of St Salvador of Ibañeta in Roncesvalles, also in Pamplona (by the Cathedral, the Plaza del Castillo and the Casa del Condestable), in Santa Maria de Eunate and in the Holy Sepulchre of Estella, in the Holy Sepulchre of Torres del Río, in San Saturnino in Artajona, and in the abandoned areas of Arlás in Peralta and in Santa María de Ujué

Ventricular myocardial trabeculation in chondrichthyans. Evolutionary implications

The formation of trabeculae (trabeculation) in vertebrates occurs during cardiac development in the three structural types of ventricular myocardium, namely, compact, spongy and mixed. The compact type is mainly composed of compacted muscular fibers and the spongy type of muscular trabeculae. The mixed type, with an inner trabecular and an outer compact layer, has been proposed as the primitive condition in gnathostomes. In vertebrate models, trabeculation initiates following two alternative mechanisms: (1) in chicken and mouse, the endocardial cells evaginate towards the two-layered myocardium; (2) in zebrafish, cardiomyocytes from the mono-layered myocardium invaginate towards the endocardium. Trabeculation in the mixed myocardium has not been described yet. We have studied the mixed myocardium formation in the dogfish (Scyliorhinus canicula, Elasmobranchii) using light, scanning and transmission electron microscopy. At stage 27, the ventricle consists of a two-layered myocardium internally lined by endocardium, both separated by cardiac jelly. Trabeculation starts at stage 28, when small spaces between cardiomyocytes appear, the cardiac jelly become thinner and the endocardium focally contacts the myocardium. At stage 29 the spaces between cardiomyocytes increase in size and get lined by the endocardium, shaping the presumptive trabeculae. At later stages, the trabeculae increase in complexity and the outer cardiomyocytes proliferate and get compacted, delineating the definitive trabeculated and compact myocardia. We conclude that early trabeculation in elasmobranchs matches that described in tetrapods. Thus, the mechanism of trabeculation of the mixed ventricular myocardium has been conserved in the formation of the compact myocardium of tetrapods. Additional studies in actinopterygians with different myocardial types may uncover how trabeculation has evolved during the evolution of gnathostomes.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech. CGL2017-85090-P, FPU15/03209, UMAJI75 and FEDER

La configuración del tracto de salida cardiaco en los vertebrados pisciformes

La noción clásica relativa a la anatomía del corazón de los vertebrados pisciformes ha cambiado notablemente en los últimos años. Anteriormente se asumía que el tracto de salida cardiaco de los condrictios es estructuralmente diferente del de los teleósteos. En los primeros, el tracto de salida es de naturaleza miocárdica y se denomina cono arterioso, mientras que en los segundos se denomina bulbo arterioso y carece de musculatura cardiaca. Según esta noción clásica, en el curso de la evolución de los actinopterigios, el cono arterioso habría disminuido su tamaño de forma concomitante con la aparición y desarrollo del bulbo arterioso. No obstante, algunos trabajos antiguos en los que se describe el corazón de determinadas especies de condrictios y de actinopterigios no teleósteos, tales como los polipteriformes, refieren que el tracto de salida consta de dos porciones, una miocárdica, proximal con respecto al ventrículo, y otra, distal, no miocárdica. Investigaciones recientes han puesto de manifiesto que el tracto de salida cardiaco de los condrictios está constituido en realidad por un componente proximal, el cono arterioso, de naturaleza miocárdica, y un componente distal, no miocárdico, homólogo al bulbo arterioso de los actinopterigios. Estos dos componentes coexisten también en el polo arterioso de los polipterifomes. Asimismo, se ha observado que el cono arterioso está presente en todos los grupos de teleósteos. Este componente cardiaco está muy reducido e incluso llega a ser vestigial tanto en grupos que divergieron tempranamente, como los osteoglosomorfos, como en grupos apicales, tales como los perciformes. En conclusión, en contraposición a la noción clásica, el polo arterioso de todos los vertebrados pisciformes está constituido por dos componentes, uno proximal y el otro distal con respecto al ventrículo, que son el cono arterioso y el bulbo arterioso. Esta configuración se ha conservado en el curso de la evolución de los diversos grupos de vertebrados pisciformes.CGL2014-52356-P (Ministerio de Economía y Competitividad), FPU15/03209 (Ministerio de Educación, Cultura y Deporte), fondos FEDER. Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Coronary artery high take-off in rodents and the possible involvement of Smad2

The coronary arteries (CAs) supply the mammalian heart with oxygenated blood. They arise from the right and left aortic valve sinuses at the aortic root. In humans, the occurrence of a CA arising ectopically from the tubular aorta, a condition called high take-off (HTO), is rare (A (Smad2C>A) allele is associated with HTO in this species. In order to test whether HTO occurs in association with Smad2C>A in other rodents, we examined the anatomical origin of the CAs, by means of stereomicroscopy and a corrosion cast technique, in 3,388 specimens belonging to 17 rodent species. In addition, Smad2 DNA sequence from M. musculus was compared by Blastn analyses with that from six of the species examined in which this sequence is known (Mus spretus, Rattus norvegicus, Apodemus sylvaticus, Myodes glareolus, Mesocricetus auratus, Microtus agrestis). HTO occurred in nine out of 17 species studied. The incidence of HTO ranged from 0.4% to 6.5% (low) in three species and from 15.9% to 25% (high) in six species. The Smad2 sequence showed similarities higher than 75% for the whole gene, and higher than 71% for the intron sequence that includes the rs29725537:C>A Single Nucleotide Polymorphism. The Smad2C>A allele was identified only in M. spretus and A. sylvaticus, with high incidences of HTO, whereas R. norvegicus, M. glareolus, M. auratus and M. agrestis showed low or null incidences. We conclude that HTO is a common trait in rodents, which does not lead to cardiac pathology probably due to the intramyocardial condition of their CAs, as opposed to the human subepicardial CAs. The Smad2C>A allele may be involved in the development of HTO and probably other phenotypes in different rodent species.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech. Funding: CGL2017-85090-P, FPU15/03209, UMAJI75 and FEDER