Arenysaurus ardevoli , first paleoneuroanatomical description of a European hadrosaurid

The neuroanatomy of hadrosaurid dinosaurs is well known from North America and Asia. In Europe only a few cranial remains have been recovered with the braincase. Arenysaurus is the first European endocast for which the paleoneuroanatomy has been studied. The resulting data have enabled us to draw ontogenetic, phylogenetic and functional inferences. Arenysaurus preserves the endocast and the inner ear. This cranial material was CT-scanned, and a 3D-model was generated. The endocast morphology supports a general pattern for hadrosaurids with some characters that distinguish to a subfamily level, such as a brain cavity anteroposteriorly shorter or the angle of the major axis of the cerebral hemisphere to the horizontal in lambeosaurines. Both characters are present in the endocast of Arenysaurus . Moreover, osteological features indicate an adult ontogenetic stage while some paleoneuroanatomical features are indicative of a subadult ontogenetic stage and even a juvenile ontogenetic stage. Finally, a comparison with other hadrosaurids reveals that the low values for the angle of the dural peak may be an autapomorphy exclusive to the Parasaurolophus genus. It is hypothesized that the presence of puzzling characters that suggest different ontogenetic stages for this specimen, may reflect some degree of dwarfism in Arenysaurus . Regarding the inner ear, its structure shows differences from the ornithopod clade with respect to the height of the semicircular canals. These differences could lead to a decrease in the compensatory movements of eyes and head, with important implications for the paleobiology and behavior of hadrosaurid taxa such as Edmontosaurus, Parasaurolophus and Arenysaurus . These differences in the vestibular system could be used as a phylogenetical signal. The endocranial morphology of European hadrosaurids sheds new light on the evolution of this group and may reflect the conditions in the archipelago where these animals lived during the Late Cretaceous.


Introduction
Hadrosaurids are the most abundant ornithopod dinosaurs from the Late Cretaceous of Laurasia with a very complete record including ontogenetic series, mummies, eggs, ichnites, etc. (see Lull & Wright, 1942;Horner, Weishampel & Forster, 2004 for reviews). This rich record also includes natural cranial endocasts or complete skulls allowing the generation of silicone or latex rubber models of the endocast (Lambe, 1920;Gilmore, 1924;Ostrom, 1961;Serrano-Brañas et al., 2006;Lauters et al., 2013). The endocranial morphology of hadrosaurids has been studied since the first quarter of the 20th century (as in the case of Edmontosaurus regalis (Lambe, 1920) or Lambeosaurus (Gilmore, 1924)). Nowadays, non-invasive techniques such as CT scans shed new light on the paleoneurology of dinosaurs and other extinct taxa (Witmer et al., 2008;Evans et al., 2009;Godefroit, Bolotsky & Lauters, 2012;Lautenschlager & Hubner, 2013). CT scan techniques are currently common in biology and paleontology in a considerable variety of studies as a way of obtaining digital models of inner regions, as in the case of endocranial morphology, where these cavities are surrounded by matrix. Interestingly, the CT scan allows a 3D visualization with a high or very high resolution depending on the type of CT scan used and the goal of the study.

Material and methods
Studied material: MPZ2008/1 (Figure 1), skull remains of the holotype of the taxon Arenysaurus (Pereda-Suberbiola et al., 2009b). The remains are from the Blasi 3 locality in the town of Arén (Huesca province, NE Spain). Postcranial remains of Arenysaurus have also been recovered (see Cruzado-Caballero et al., 2013). (excluding the olfactory bulbs). This volume value is close to the results obtained by Saveliev et al. (2012) for the adult specimen of the lambeosaurine Amurosaurus AENM1/123 (see Table 1).
On the other hand, the Arenysaurus endocast is considerably constricted lateromedially at the cerebellum level with a maximum width of 31.32 mm in this region, and slightly constricted at the medulla oblonga (26.26 mm). Unfortunately, the vallecula system, described in the anterior part of the endocast of other hadrosaurids, cannot be observed in Arenysaurus due to the hard matrix that covers this area.
The angle of the major axis of the cerebral hemisphere to the horizontal is close to 45° in the endocast. According to Evans et al. (2009), this high angle corresponds to a lambeosaurine shape as opposed to that of hadrosaurines and other ornithopods, where the cerebral hemisphere is positioned more horizontally (Hopson, 1979).
According to Giffin (1989), pontine flexures are virtually absent and the possession of a nearly straight endocranial cavity is derived for "iguanodontids" and hadrosaurids. Further, in lateral view the cerebral hemisphere is not very strongly arched, as is the case in adult lambeosaurines and unlike young individuals (e.g. Parasaurolophus sp. RAM 14000). These different angles are possibly a consequence of more strongly arched frontals in young individuals (Farke et al., 2013).
The olfactory bulbs are located anteroventrally to the cerebral hemisphere and only preserve their base. It has not been possible to reconstruct them completely, because the skull is broken in the anterior part of the frontals. The left bulb is the more complete one, while the right bulb only preserves its ventral part. In anterior view, the left olfactory bulb has an inside-out L-shaped morphology. In this view, it is also possible to observe that the left olfactory bulb is almost half the height of the cerebral hemisphere, as also happens in the adult of Amurosaurus (IRSNB R 279, AENM nos. 1/232 and 1/240; Saveliev et al., 2012;Lauters et al., 2013)

Cranial nerves
Almost all the cranial nerves, excluding nerve I and IV, can be seen to be preserved on the left side.
Nerve II, or the optic nerve (CN II), only preserves its base. This nerve is the most anterior nerve preserved. It is very small, tubular-like and runs parallel to the ventral side of the cerebral The ventral side of the endocast preserves the nerves VI, or the abducens nerves (CN VI). This joins the pituitary, which their exits from the posterior to connect ventrally with the cerebellum.
The nerves are flattened lateromedially and are wider than high.
Nerve VII, or the facial nerve (CN VII), is present and positioned anteriorly to the cochlea and near nerve VIII. This nerve is tube-like, very small and thin, with a slight widening dorsomedially on its distal side. This nerve is ventral to nerve VIII and runs lateroposteriorly.
Nerve VIII, or the vestibulocochlear nerve (CN VIII), is dorsal to nerve VII. This nerve is only partially preserved, showing a very small portion of the base dorsoventrally flattened.
Nerve IX, or the glossopharyngeal nerve (CN IX), is posterior to the cochlea and runs laterally, touching the cochlea in its anteriormost part. This nerve is very slight in its basal part and is tubular-like in shape. At its lateral extreme the nerve is extremely expanded dorsomedially (3.08 mm) and lateromedially (3.02 mm).
Nerves X and XI, the vagus and accessory nerves respectively (CN X and XI), are separated at their base, but then they join to form a single nerve. This joined nerve is very broad anteroposteriorly (6.8 mm) and is clearly lateroposteriorly directed. Nerve XII, or the hypoglossal nerve (CN XII), is the most posterior one. It presents an anteroposteriorly narrow base (2.19 mm) and a dorsoventral height (3.94 mm) that is expanded distally (with an anteroposterior width of 4.69 mm and a dorsoventral height of 5.58 mm). Unlike the joint nerves X and XI, nerve XII is only laterally directed.

Inner ear
The digitally reconstructed vestibular apparatus is complete on the left side whereas the right side just conserves part of the cochlea and the anterior and posterior semicircular canals. The general form of the inner ear is similar to that described in other hadrosaurids (Brown, 1914;Langston, 1960;Ostrom, 1961;Evans et al., 2009;Farke et al., 2013), and, as discussed in Evans et al. ampulla is the largest, followed by the lateral ampulla). Moreover, in lateral view, the cochlea is boomerang-like, convex laterally and concave medially. In anterior view, it presents an S-shape with a sharp distal border and it has a length of 10.72 mm from the foramen vestibulea (Table 4).

Discussion
The endocranial morphology among hadrosaurid dinosaurs is similar and characteristic of the family (Hopson, 1979). At a subfamily level (hadrosaurine-lambeosaurine) there are characters that can help to distinguish between them, such as a brain cavity that is anteroposteriorly shorter A previous paper (Pereda-Suberbiola et al., 2009b) considered that this Arenysaurus specimen belongs to a presumably sole adult individual on the basis of several osteological characteristics.
The paleoneuroanatomical evidence supports this ontogenetic assignation, with features referred to adult hadrosaurid animals that are present in this specimen: an angle of flexure between the cerebellum and cerebral hemisphere that is very small as in lambeosaurine adults, as described by Evans et al. (2009), and the cranial sutures that are difficult to discern in the CT scan as usual in adult specimens.
However, some paleoneuroanatomical features herein reported are indicative of a subadult ontogenetic stage for this specimen (see Table 1 and 3) and even a juvenile ontogenetic stage in the case of the total length of the endocast. According to Evans et al. (2009), however, this difference in the length of the endocast may be due to phylogenetic rather than ontogenetic considerations, as in the case of Hypacrosaurus. Moreover, when we compare the femur length of juvenile and adult lambeosaurines with the femur of Arenysaurus, the latter is nearer to the average for adult Asian than for adult North American lambeosaurines (see Table 3). This puzzling mixture characters from adult and subadult stages may reflect a possible first case of a certain degree of dwarfism evidenced by a hadrosaurid endocast. The hypothesis of a reduction in size due to insularism in European hadrosaurids has been proposed by several authors in the last authors suggest that the phylogenetic differences between the lambeosaurini and parasaurolophini tribes could be assessed in the light of the angle of the dural peak. In these terms, the lambeosaurins presented a wider angle (around 120º) while parasaurolophins presented a more acute angle (approximately 90º). We have observed that angles up to 100º are present for several hadrosaurins and lambeosaurins. In the case of Arenysaurus, this angle is approximately 114º (see Table 2). In sum, the angle of the dural peak may indeed be informative, suggesting that the condition with a greater angle could be a basal character and a lesser angle of 100º may be exclusive to the genus Parasaurolophus. Regarding the inner ear, although the general form is similar to the other hadrosaurids, it is possible to observe small differences in the semicircular canals with respect to the ornithopod clade (see Figure 4). The anterior semicircular canal is tallest at the base of the clade (Dysalotosaurus and Iguanodon), by contrast with some hadrosaurines, where the posterior semicircular canal is slightly taller than the others (Edmontosaurus). Later, in the Lambeosaurinae subfamily, Parasaurolophus and Arenysaurus present anterior semicircular canals that are slightly taller, while in the lambeosaurini tribe they are similar in size to Dysalotosaurus or Iguanodon. In addition, Parasaurolophus and Arenysaurus share a lateral ampulla that is larger than the posterior and the anterior ampullae. in Edmontosaurus, Parasaurolophus and Arenysaurus. If true, this could be related with behavioral patterns that require less agility in the head movements (Sereno et al., 2007).
Likewise, we hypothesize that these differences in the vestibular system, i.e. the different ratios between the height of the anterior and posterior semicircular canal and the size of the ampullae, could be used as a phylogenetic signal to differentiate Edmontosaurus, Parasaurolophus and Arenysaurus from the rest of the hadrosaurids. However, more data are necessary to know the possible influences that these differences could have on phylogenetic interpretations.

Conclusion
We provide the first complete 3D reconstruction of the brain cavity and inner ear of a European lambeosaurine, Arenysaurus. This cranial endocast presents the general pattern known for hadrosaurids and add to the record of hadrosaurid brain cavities from Laurasia. The osteological and paleoneuroanatomical data suggest that Arenysaurus was an adult individual that probably presented a certain degree of dwarfism due to insularity. Thus, Arenysaurus could be the first evidence of how dwarfism could affect hadrosaurid paleoneuroanatomy. Moreover, the paleoneuroanatomical data suggest that the presence of the low angle of the dural peak could be an autapomorphy of the Parasaurolophus genus. Furthermore, the structure of the inner ear shows differences from the ornithopod clade with respect to the height of the semicircular canals.
These differences can be explained principally in terms of a probable decrease in the compensatory movements of eyes and head, which would affect the paleobiology and behavior of these animals. We hypothesize that these differences in the vestibular system could be used as are a phylogenetic signal.      Table 1. Measurements of length and volume for complete brain cavity and various brain regions, calculated from the digital endocasts using digital segmentation in the Avizo 7.1 program.     Measurements of length and volume for complete brain cavity and various brain regions Measurements of length and volume for complete brain cavity and various brain regions, calculated from the digital endocasts using digital segmentation in the Avizo 7.1 program.      Average measurements of the length and volume of the brain cavity with and without olfactory bulbs, the maximum width of the cerebral hemisphere and the length of the femur Average measurements of the length and volume of the brain cavity with and without olfactory bulbs, the maximum width of the cerebral hemisphere and the length of the femur from lambeosaurines in relation to the ontogenetic stage. Average length, width and volume of the brain measurements were obtained from Evans et al. (2009)