Skull of the grey heron (Ardea cinerea): Detailed investigation of the orbital region

The skull of the grey heron (Ardea cinerea) was examined with an emphasis on describing the orbital region. In the young (circa sixteen to seventeen days old) heron, the frontal bone (os frontale) and nasal bone (os nasale) comprised separate paired bones, connected by sutures (sutura interfrontalis, sutura internasalis and sutura frontonasalis plana). In adult animals, the relationship between these bones was different: the left and right frontal bone and the left and right nasal bone had grown together, and the frontal bone and nasal bone had fused into a common frontonasal bone (os frontonasale). In the ectethmoid bone (os ectethmoidale), the main components comprised of the orbital and antorbital part of the ectethmoid plate (lamina ectethmoidalis orbitalis et antorbitalis), the lateral process (processus lateralis ectethmoidalis) and the tubercle (tuberculum ectethmoidalis); the left and right ectethmoid plates were fused together to form the ectethmoid sinus (sinus ectethmoidalis) between them. In the young heron, the anatomical and functional link between the frontal and lacrimal bones did not exist yet, nor did the osseous frame of the ectethmoid‐lacrimal complex. Further research into the young heron skulls is needed. This article provides novel insights into the grey heron's orbital region.


| INTRODUCTION
Herons are members of the family Ardeidae (order Pelecaniformes), and the majority of extant species are in the subfamily Ardeinae, known as true or typical herons. The grey heron (Ardea cinerea Linnaeus, 1758) is closely related and similar to the great blue heron (Ardea herodias Linnaeus, 1758), as well as to the cocoi heron (Ardea cocoi Linnaeus, 1766). The subfamily Ardeinae also includes the great white heron (Ardea alba Linnaeus, 1758). Four subspecies of the grey heron have been recognized as follows: A. c. cinerea in Europe, Africa and western Asia, A. c. jouyi Clark, 1907 (found in eastern Asia), A. c. firasa Hartert, 1917 (found in Madagascar) andA. c. monicae Jouanin andRoux, 1963 (found on islands off Banc d'Arguin, Mauritania).
This study expands upon our previous work (Golob, Bavdek, Zajc, Janžekovič, & Klenovšek, 2016), in which dimensions of several bones (scapula, coracoid, humerus, ulna, femur and tibiotarsus) of the adult grey herons were measured and their lengths compared to the results in the work performed by Kellner (1986). In the present study, we were particularly concerned with evaluating the relationships between the bones of orbital area and in comparing the skeletons of a young (around sixteen to seventeen days of age) grey heron with adult specimens. Although Zusi and Livezey (2006) studied the palatine bone, the ossa cranii in many neognathous taxa remain undetermined and the osteology of the heron's orbital region has not been published. Our novel and detailed description of the orbital region contributes to the knowledge required in understanding the imaging techniques of these birds. This is especially important in the veterinary clinic and may also have relevance in other species.

| MATERIALS AND METHODS
The skulls of four adult and young grey herons (Ardea cinerea), as well as one from an adult white heron (Ardea alba), were used in this study.
The young heron had a weight of 740 g, which corresponds to the age of 16-17 days (Creutz, 1981). Their sexes were unknown. Among the adult grey herons, two were kept in the Slovenian Museum of Natural History, Ljubljana, Slovenia, and the others were from the Department of Biology, Faculty of Natural Sciences and Mathematics, University of Maribor, Slovenia. The skeletons in the zoological collection of the Department of Biology were prepared from herons found naturally deceased and collected in localities around Maribor.
The soft tissues of the adult specimens were cleaned by dermestes beetle larvae (Dermestidae), whilst the soft tissue from the skull of the young specimen was carefully macerated (McDonald, 2006). The living grey heron (Figure 1a) was found injured, brought to a shelter (Asylum for protected wild animals, Muta, Slovenia; Animal evidence number: 0006050/015), successfully cured and returned to the wild. The study complies with all national and government ethical guidelines.
Photographs of the skull were taken by the author Golob using a digital camera and presented using the Adobe InDesign CS6 for primary and Adobe Photoshop CS6 for secondary step of the procedure.
The anatomical descriptions were made by direct observation.

| RESULTS
The grey heron's upper jaw was surrounded by maxillary ramphotheca (ramphotheca maxillae) from the apex of the beak to the craniofacial hinge (zona flexoria craniofacialis) and then extending to some extent laterocaudally ( Figure 1a). The ramphotheca was interrupted to accommodate for the nostrils (nares). From the rostral corner of the nasal aperture (apertura nasi ossea) towards the apex of the beak, a longitudinal nasolabial groove (sulcus nasolabialis) (Figures 1a,b) was observed.

F I G U R E 3
The orbital area of about three-month-old grey heron after cranium removal altogether with frontal bones; dorsal view. Legend: co = capitulum oticum; cs = capitulum squamosum; ons = os nasale sinistrum; pfn = processus frontalis nasalis; pfpr = processus frontalis premaxillare; pmn = processus maxillaris nasalis; poq = processus oticus quadrati; pprn = processus premaxillaris nasalis; sutin = sutura internasalis In the adult grey heron, the articular surface of the frontal bone with the lacrimal bone was clearly formed (Figure 6, fal), as was the orbitonasal opening (foramen orbitonasalis, Figure 7, fon) on the medial side of the lacrimal process of the frontal bone, near the border with the lateral margin of the ectethmoid bone. At this level, the lacrimal process formed a longitudinal crest (crista processus lacrimalis, Figure 7, cpl).
In Figure 8, the ectethmoid bone (os ectethmoidale) is shown in an artificial cross-section performed close to the left and right lateral processes (Figure 8, plect). At the base of the lateral process, the incisura (incisura processus lateralis, ipl) was pronounced, and beneath the process, a low-lying tubercle (tuberculum ectethmoidalis, tect) was located. The orbital plates (laminae orbitales, loect) and antorbital plates (laminae antorbitales, laect) surrounded the ectethmoid sinus (sinus ectethmoidalis, sinect). The left and right orbital plates were dorsally connected by a thin, transverse bony lamina dorsalis (presumably the mesethmoidalis).

| DISCUSSION
The distribution of the maxillary ramphotheca of the grey heron (Ardea cinerea) was similar to that observed in the striated heron (Butorides striata) (Hieronymus & Witmer, 2010). It covers the beak as a simple ramphotheca, and the longitudinal nasal sulcus (sul- The base of the process is formed by the laterosphenoid bone (os laterosphenoidale), and the frontal bone is located dorsally as a kind of cover or lid. Livezey and Zusi (2006) reported that such a situation is common in most birds.
In the young heron, the nasal surfaces of the frontal bones fit with the nasal bones on their ventral surface (sutura frontonasalis plana).
This fact, as well as the existence of the frontonasal and frontoparietal fontanelles (fonticulus frontonasalis et fonticulus frontoparietalis), and the loose mutual contact between the frontal bones (sutura interfrontalis), potentially enables adjustment of the complex of dorsal orbital bones to brain development and growth, and perhaps even to the feeding mechanism of young herons. In adult herons, both pairs of frontal and nasal bones are fused into a single frontonasal bone (os frontonasale).
In the supraorbital area, that is as part of it, there are two anguli of the frontals, the caudal and rostral (angulus supraorbitalis caudalis et rostralis). The caudal angulus has also been termed the supraorbital process by Payne and Risley (1976), and the angulus postocularis by Livezey and Zusi (2006). In front of the rostral supraorbital angulus, the frontal bone is rotated ventrally and forms the lacrimal process (processus lacrimalis frontalis), which serves for articulation with the lacrimal bone. This process is well developed in the adult subjects, whilst it appeared as a low crest in the sixteen to seventeen days old grey heron. Thus, the functional relation between the frontal and lacrimal bones develops with the grey herons aged more than seventeen days. Later development is also observed in relation to the composed lacrimal process of frontal bone, amongst others a pronounced medial longitudinal crest; the latter rises above the passage for nerves (foramen orbitonasalis). The ectethmoid-lacrimal complex was absent in the young grey heron: neither the lacrimal bones nor the lateral process of the ectethmoid bones was observed.
The left and right ectethmoid bones are fused in adult grey herons and form the intermediate space (sinus ectethmoidalis). Shufeldt (1889) has described the heron's ectethmoid bone as an unusually thick and bulky bone, which spreads out a wide base for the frontals to rest upon, including a cancellous internal structure. However, that research did not highlight a space (sinus) inside.
Our description of the bones in the juvenile grey heron has interesting value because the study of juvenile birds is relatively rare. Zusi and Livezey (2006)  Consequently, the relations amongst the ossa cranii in many neognathous taxa remain undetermined. Zusi and Livezey (2006) cited the skeletons of immature birds, when they studied the palatine bone (os palatinum), including skeletons of the great blue heron (Ardea herodias) and the boat-billed heron (Cochlearis cochlearis). However, there is no report or comment on the osteology of the heron's orbital region.
In the rostrolateral area of the grey heron's orbit, a prominent skeletal base arises enabling articulation between the lacrimal bone and the frontal/nasal bones (facies articularis frontonasalis lacrimalis), and between the lacrimal and ectethmoid bones (ectethmoid-lacrimal complex) during growth from seventeen days of age onwards. Moreover, a clear craniofacial hinge also develops (zona flexoria craniofacialis), thus allowing reliable, movable skull mechanisms. In the case of our young heron, the right ectethmoid and nasal bones had fused; however, the left ones had not. It is possible that ossification and fusion of the left and the right side of the orbit are not coincident; however, more specimens would be needed to confirm this finding as this fusion may also be present due to biological variation. Therefore our results on the young grey heron are discussed as preliminary data and warrants further research.
Understanding the anatomy and morphology of this species helps to further the knowledge required when undertaking imaging, diagnosis and treatment in the clinic. It also provides more information in which to undertake comparative work with other avian species and highlights the need for understanding the developmental anatomy and morphology in this bird. This study provides a detailed description with valuable new contributions, such as the growth of the lacrimal processes, first description of sinus ethmoidalis, bilateral ossification and not synchronous growth of the left and right bones, to the knowledge of the grey heron's orbital region.