Neuroanatomy of the equine brain as revealed by high-field (3Tesla) magnetic-resonance-imaging

In this study, the morphology of the horse brain (Equus caballus) is decribed in detail using high field MRI. The study includes sagittal, dorsal, and transverse T2-weighted images at 0.25 mm resolution at 3 Tesla and 3D models of the brain presenting the external morphology of the brain. Representative gallocyanin stained histological slides of the same brain are presented. The images represent a useful tool for MR image interpretation in horses and may serve as a starting point for further research aiming at in vivo analysis in this species.


Introduction
Neuroimaging is increasingly important in veterinary large animal neurology. Magnetic resonance imaging (MRI) is more and more used to evaluate intracranial diseases in horses with neurological signs [1][2][3][4][5][6][7][8][9][10][11][12]. High magnetic field strengths (3 Tesla) are now available in veterinary medicine. Images collected at this field strength provide improvements in image clarity and detail. Despite considerable progress in the technical adaptions of scanners and detection coils to the practical requirements in equine medicine, the description of brain morphology in the horse has been somewhat neglected. Currently, studies describing equine brain anatomy are only available at reduced resolution and from the brain of foals [13,14]. As the number of MRI-investigations of the equine brain grows, so does the need for detailed information about brain structures and special characters of the equine brain, as existing for a number of domestic species [15][16][17][18][19]. The aim of this study was therefore to examine the brain of the horse using high field MRI and to describe the morphology of the equine brain as shown in these images.

Animals
The head of an eight years old warm blood horse was examined post-mortem. The animal was euthanized due to a complicated tarsal fracture. The horse was sedated with 0.4 mg/kg xylazin (Xylazin 2%, CP-Pharma GmbH, Burgdorf, Germany) injected in an intraveneous catheter in a1111111111 a1111111111 a1111111111 a1111111111 a1111111111 the jugular vein. General anesthesia was induced with 0.1 mg/kg diazepam (Diazepam AbZ) and 2.2 mg/kg ketamine (Narketan 10). Euthanasia was performed using 0.12ml/kgkg embutramid (T61). No neurological findings were observed at previous clinical examination. Directly after euthanasia the head was dissected between fourth and fifth cervical vertebrae and was trimmed to fit into a standard human knee coil. MRI was performed 90 minutes after death of the animal. Owner consent was obtained for use of the head in scientific research. All data was anonymized.

MR imaging
MRI scans were performed on a 3-Tesla Magnetom Verio scanner (Siemens Healthcare, Erlangen, Germany) using an eight channel phased array human knee coil. Anatomical images of the entire brain were acquired in transverse, sagittal and dorsal planes, using a T2-weighted spin echo sequence. To achieve a sufficient signal-to-noise ratio (SNR), 32 averages were accumulated obtaining slices of 3 mm thickness with 672x672mm 2 field of view (FOV) at 512 x 512 matrix size resulting in 0.25 mm in-plane resolution and 1mm slice distance. Optimum contrast was obtained with 8500 milliseconds (ms) repetition time (TR) and while the echotime (TE) was adjusted to 12 ms. Image acquisition was anisotropic, which is why sagittal and horizontal planes were not recalculated but obtained in consecutive scan sessions. Total scan time was 4 hours and 55 minutes.

Image processing
Images were reviewed using AMIRA (Mercury Computer Systems) graphical software. This program allows interactive assessment of morphology in all image planes. A 3D model of the outer brain surface was generated based on free hand segmentation of the brain outlines in transverse MR-images. Image segmentation in this context describes the manual tracing of the brain surface. All voxels corresponding to a single anatomical structure in the images are selected and assigned to the same value in the mask. The final mask thus contains information about all selected anatomical structures and, in combination with the original data and polygonal surface reconstruction algorithms, allows the identification of sulci and gyri in 2 D images in association with the produced 3D model. Anatomic structures including the sulci and gyri were identified using a published atlas [20] and by comparison with histological slices produced from the examined brain. Morphological substructures were labelled in transverse images and thereafter located in dorsal and sagittal images using the four-viewer mode of the software. Signal intensities are described in relation to the cortical grey matter as hyperintense (brighter signal compared to cerebral cortex) or hypointense (darker signal as the cerebral cortex).
Vertrieb GmbH Wetzlar, Germany) with section thickness of 350 μm. Celloidin served as a support, guiding the microtome knife through the tissue block and preventing tangentially cut gyri from floating away during subsequent staining procedures. Slicing of the histology slides was performed perpendicular to the brainstem axis to match the same orientation as the MR transverse image slices. The slices were then stained free-floating in gallocyanin-chromalum [21] quenched between two filter papers and two perforated stainless steel plates to prevent distortion of the single slices during dehydration in ethanol, ispopropanol:xylene 1:1, and finally xylene. The sections were coverslipped with Permount.

Results
A 3D rendered model (Fig 1A-1E) demonstrates the morphology of the brain as a whole from a dorsal, lateral, frontal, and ventral as well as from a midsagittal-medial view. T2-weighted high-resolution images with detailed structural identification are provided in transverse, sagittal and dorsal planes shown in . Based on comparison with histological slices, morphological structures of the adult equine brain are labelled in detail in the MR images. A list of anatomical terms and the figures, in which they are displayed, are shown in the Supporting information (S1 Table). The transverse images correspond to a reference line presented along with the 3D model in the right upper corner of the image. Selected examples of histological slides corresponding to the transverse MR-images are presented as Supporting material (S1-15 Figs). Anatomical structures were named according to the Nomina Anatomica Veterinaria [22]. A description of the brain parts, their substructures and connections is given in the following.

Telencephalon
Allocortex. The lateral rhinal fissure (Fissura rhinalis lateralis; Figs 1A, 2-9, 10-12, 18, 20, 21, 27 and 30-34; S1-S8 Figs: Rfi) separates the prominent rhinencephalon from the laterally adjoining telencephalon. The latter can be subdivided into a basal, septal, and a limbic part [23][24][25]. The rostral-most aspect of the basal part constitutes the large olfactory bulb (Bulbus olfactorius; Figs  Caudal to the olfactory tubercle on each side is the rostral perforated substance (Substantia perforata; Fig 19: pfs). It is dorsally continuous with the basal nuclei. The perforated substance is characterized by hyperintense penetrating branches of the striate arteries (ramus striatus of the medial cerebral artery), which supply the internal capsule and the basal nuclei (Fig 6: stra). Together, the olfactory tubercle and the perforated substance build the olfactory area   The amygdaoid complex consists of a cortical-or corticoid-and a subcortical nuclear part with numerous subnuclei, which can be differentiated based on its cytoarchitectonics and histochemistry (S6 Fig: ab) [26]. In our scans, the amygdaloid body can only be seen as a mildly hyperintense ovoid nuclear mass, which is covered ventrally by the paleocortex of the piriform lobe (Lobus piriformis; Figs 7 and 19: pir). Subnuclei are not visible in our MR-images. A slender hypointense fibre bundle runs around the thalamus and the caudate nucleus, which connects the amygdala with the hypothalamus and other basal forebrain regions. This is the terminal stria (Stria terminalis; Figs 6-8; S5-S7 Figs: stt), part of the extended amygdala.
The amygdaloid bodies are associated to the limbic part of the rhinencephalon (Pars limbica rhinencephali [23]. The other components are the cingulate (Gyrus cinguli; Figs 2, 3, 5, 6,  The septal region of the rhinenecphalon (Area septalis, Pars septalis rhinencephali) is seen as a thin sheet of brain tissue below the rostrum of the corpus callosum (Rostrum corporis callosi; Figs 26-28: rccl) extending ventrally to the olfactory tubercle, forming the medial wall of the rostral horn of the lateral ventricle [23]. Based on the topographical relation to the rostral commissure within the lamina terminalis (Fig 26, S5 Fig: lt) [23,27], which extends latero-ventrally in the direction of the external edge of the lateral ventricle. The white matter tract that underlies cingulate cortex   [23,27].
Neocortex. Cortical surface. The cortex of the equine telencephalic hemisphere is intricately folded and the major sulci show a high variability in ramification [23,24]. Even the cerebral vessels can create sulci of considerable depth, which sometimes complicate their clear identification. On the lateral surface of the hemispheres the short almost vertical sylvian fissure  Rostral to the ansate sulcus there is a small furrow on the medial side of the pallium turning to the dorsal surface, which is the cruciate sulcus (Figs 1C-1E, 2, 24, 25, 28 and 29; S1 and S2 Figs: Cru). This area contains the motor cortex of the equine brain [28]. In the rostro-lateral cortical aspect the diagonal sulcus (Figs 1A, 1D, 2, 3, 4, 21-24 and 31-34; S1-S3 Figs: Dia) crosses the lateral surface running towards the sylvian fissure. It is situated between the rostral ectosylvian and presylvian sulcus (Figs 1A, 1C, 1D, 2, 3, 21, 22 and 31-34; S1-S3 Figs: Prs), which turns rostromedially reaching the dorsal surface towards the coronal sulcus. At the base of the frontal lobe, the Subpallial grey matter. The basal ganglia comprise a distributed set of brain structures in the telencephalon and diencephalon. The substantia nigra (Figs 8, 9, 10, 19 and 28-31; S7 and S8 Figs: snr) of the midbrain as well as the subthalamic nucleus (Fig 29: stn), which is located in the caudal diencephalon are both functionally connected to this system. The largest group of these nuclei is the large striate body (Corpus striatum), which is subdivided into two parts (caudate nucleus, lentiform nucleus) by fiber masses of the internal capsule ( , separated from each other by a thin sheet of white matter. In the horse, the head of the caudate nucleus quickly tapers into the tail (cauda) and the body (corpus) is really short compared to other ungulates [17]. Stripes of grey matter connect the head of the caudate nucleus and the putamen. Further caudally, the putamen quickly increases in height and width but never attains the dimensions of the caudate nucleus. The rostral part of the putamen appears on transverse sections as a rather narrow straight band of grey matter stretching from dorso-lateral to ventro-medial. The middle part, the largest, is triangular in shape on the transverse sections. The putamen is laterally bordered by a thin layer of hypointense white matter, the external capsule (Capsula externa; Figs 4-7, 22 and 32; S2-S6 Figs: ec). A hypointense fiber bundle can be seen ventrolateral to the globus pallidus running into the caudal limb of the internal capsule. This is the ansa : cla) appears as a column of hyperintense grey matter located adjacent to the lateral rhinal fissure. It changes its appearance while running caudally, varying from a polymorphic star-shaped to a triangular form. The dorsal part is lying adjacent to the insular cortex (claustrum proper; claustrum insulare) and the ventral part is located next to the prepiriform cortex (also called endopiriform nucleus) [23]. Histological images (S8 Fig) reveal that on the level of the thalamus, the claustrum can be separated into different subnuclei that cannot be seen in the MR-images. Further ventrally the white matter mass does not contain association fibers running from one part of the hemisphere to the other, but only ascending and descending fibers, together referred to as the corona radiata, give rise to the internal capsule. Seen from the dorsal view on the level of the striatum, the curved, fan-shaped internal capsule can be separated into a rostral subcallosal gyrus) and the neurons of the cingulate gyrus with the enthorhinal cortex [23,29]. The ventral (inferior) longitudinal fasciculus runs sagittally next to the subcallosal fascicle along the lateral walls of the ventral and caudal horn of the lateral ventricle [29]. According to differences in density, fiber diameter, and myelination, different layers (strata) can be differentiated (stratum sagittale externum and-internum, (Figs 10 and 11: stse, stsi) [29]. The optic radiation coming from the lateral geniculate body (Corpus genicluatum laterale; Figs 8,9,22  https://doi.org/10.1371/journal.pone.0213814.g020 and 29-32; S7 and S8 Figs: lgb) running to the primary visual cortex along the calcarine fissure can be discerned. It is a thin pathway with moderate hypointensity compared with the adjacent radiation of the corpus callosum within the internal capsule. The pathways of the other sensory (radiation acustica, -sensoria) systems cannot be identified with certainty.  fibers from the olfactory tracts that cross midline in the lamina terminalis in front of the columns of the fornix. In sagittal images the rostral commissure is a round to oval structure within the ventral lamina terminalis. In dorsal images it appears as a characteristic hypointense U-shaped stripe running ventral to the striatum.  Diencephalon. The diencephalon consists of five main components-epithalamus, thalamus metathalamus, subthalamus and hypothalamus (Figs 18 and 19: hyp). The largest, rostralmost part is represented by the nuclear masses of two thalami, which enclose the third ventricle. Along the midline the two large ovoid thalamic complexes are so prominent that the ventricle is restricted here to a largely circular vertical residue bordering the fused parts of the  located between the stria medullaris and the stria terminalis. They protrude the dorsal thalamic surface into the third ventricle (Fig 7; S6 Fig: nad) [30,31].
Hypothalamus. The hypothalamus forms the basal wall of the third ventricle; the hypothalamic sulcus (Sulcus hypothalamicus ; Fig 7, S6 Fig: hs) [33]. The other part is the elongated infundibular stalk (pars infundibularis; Figs 7, 17 and 26: inf), located in the middle between the two lobes of the adenohypophysis in transverse images. The ventral part of the third ventricle builds a large and wide recess extending into the infundibular stalk (infundibular recess ; Fig 17: ir). The pars intermedia of the horse's pituitary gland is large with a hypointense signal intensity. In contrast to other ungulates it completely surrounds the hyperintense infundibular process [33]. It can be clearly seen in sagittal images as a hypointense rim dorsal on the neurohypophysis. A hypophyseal cleft (cavum hypophysis) that characterizes the pituitary gland in many artiodactyls is not present [33]. Rostral S8 Fig: cdc). The epithalamic structures are well-developed in the horse. The pineal body is large and has a lanceolated shape in sagittal images. It houses a deep pineal recess (Fig 26: rpb).

Mesencephalon
The mid-brain of the horse is short, however, has a considerable diameter. In our transverse scans it is square to rectangular in shape. Ventrally it extends from behind the mammillary bodies (Corpus mammillare;  The tectum is dominated by large rostral colliculi (Colliculi rostrales; Figs 10, 22 and 26-28: roc), and the cerebral aqueduct, which is wide and high in the horse. The rostral colliculus is much larger than the caudal one (Colliculus caudalis; Figs 9, 11 and 28; S8 and S10 Figs: cdc). Alternating strata of cells and fibers give the rostral colliculus a faintly layered appearance, best seen in transverse images (Fig 10). The whole colliculus shows eight layers in histological slices (S9 Fig). The superficial zone includes the stratum zonale, stratum griseum superficialis, and stratum opticum [28,34]. The deep zone can be divided into the stratum griseum intermedium and stratum griseum profundum each separated by a stratum album intermedium and -profundum (S9 Fig) [28,34]. Axon bundles arising from the white matter layers of the colliculi link the two rostral colliculi along their whole rostro-caudal extent. This commissure of the rostral colliculi (Commissura colliculi rostrales; Fig 10: ccr) can be seen as a hypointense stripe at the medial basis between the rostral colliculi. At the lateral basis of the colliculus a flat hypointense fibre bundle leave the optic tract medial to the pulvinar to enter the pretectal area and the rostral colliculus. This band is slightly rising over the surface forming the brachium of the rostral colliculus (Brachium colliculi rostralis; Figs 28 and 29: bcr).
The sulcus limitans in the mesencephalic aqueduct seperates the tectum from the ventral tegmentum of the midbrain (Fig 14; S9, S12 and S13 Figs: slm). Long projection fibers enter and exit the prosencephalon mainly via the internal capsule, to continue into the mesencephalon as the crura cerebri. Crura cerebri and nuclei of the mesecenphalic tegmentum constitute the pedunculi cerebri. At the level of the rostral pons, the crural fibers perforate the later and either end in the pontine nuclei or continue into the rostral medulla. The cerebral peduncles accomodate the large substantia nigra (Figs 8, 9, 10, 19 and 28-31; S7 and S8 Figs: snr). A pars reticulata and pars compacta (S8 Fig), which lies medial to the pars reticulata cannot be discerned in our MR-images. The white matter bundles are rather flat and constitute the external parts of the peduncles. The bulk of the white matter bundles divide into a flat ventral part and a rounded (level of habenular nuclei) to triangular dorsolateral part (level of the caudal commissure).
The medial longitudinal fasciculus is caudally continuous with the fasciculus proprius of the spinal cord [23]. It runs close to midline on the ventral aspect of the cerebral aqueduct. Close to the location, where the crus enters the forebrain, the optic tract winds around its ventrolateral surface in an oblique direction on its way to the lateral geniculate body. The area between the crura is termed the interpeduncular fossa, which is pierced by small blood vessels (caudal perforated substance; Fig 26 not labeled). The interpeduncular nucleus is a hyperintense unpaired cell group at the base of the tegmentum between the cerebral crura.
The midbrain is longitudinally traversed by the mesencephalic aqueduct that connects the fourth with the third ventricle. The hyperintense signal around the mesencephalic aqueduct corresponds to the periaqueductal grey matter (Substantia grisea centralis; central grey substance), which is formed by neurons that form a caudal continuation of the diencephalic periventricular nuclei of the hypothalamus. These neuronal masses surround the aqueduct and continue throughout the midbrain. Its shape, as seen in transverse sections, varies at different levels. Scattered throughout the central grey substance are numerous nuclei, which are collectively called the tegmental nuclei. Besides these scattered nuclei the central grey substance contains the nuclei of the oculomotor (Nucleus nervi oculomotorii; Fig 10: nom) and trochlear nerves (Nucleus nervi trochlearis; Fig 11; S10 Fig: nto), and the nucleus of the mesencephalic root of the trigeminal nerve (Nucleus tractus mesencephalicus nervi trigemini, S10 Fig: nmt). Between the fasciculus longitudinalis medialis (Figs 10-15, 19, 20 and 26-28; S9-S14 Figs: flm) and the decussation of the rostral cerebellar peduncles is an area of hyperintense signal corresponding to the oculomotor nucleus. The small oculomotor nerve exits the midbrain ventrally running rostrally towards the orbital fissure.

Metencephalon
Cerebellum. The shape of the large cerebellum is square in sagittal and almost triangular in the transverse planes. Transverse fissures divide the cerebellum into lobes and smaller lobules that are highly irregularly arranged. The equine vermis (Fig 1C: ver) comprises the complete set of lobules found in other mammals [24,25,36,37]. The primary fissure (Fissura prima; Figs 23 and 26-29: Fp) represents the border between the rostral and caudal lobe. The rostral lobe of the vermis is larger than the caudal lobe. This relation is also reflected by the larger rostral trunk of the medullary body (Corpus medullare cerebelli). This is somewhat unique in the horse in contrast to other ungulates that have larger caudal lobes [14,15,[36][37][38]. The medullary branches help to identify the subdivisions of the vermis in sagittal images. From the medullary body, three major branches of white matter take their origin. The rostralmost enters the central lobe (lobule II and [38]. The caudal vermis of the horse shows a characteristic S-shaped deviation of the cerebellar pyramis, tuber and folium (Fig 15). This can also be found in other ungulates and carnivores, but seems to have reached an extreme in the horse [24]. It is interesting to note that in some images the medullary branches within the tuber and/or folium seem to end without a cortical covering (Figs 26 and 28-30; marked by an asterisk), which has also been described in the pig but not in other ungulates. [39,40].
The cerebellar hemispheres are dominated by the massive ansiform (meaning loop-like) lobule (Lobulus ansiformis; Figs 1A, 1C, 13-15, 22-24, 29, 30 and 31-34: ans), which is the hemispheric extension of the vermal folium and tuber [23,38]. Unlike in other mammals, this lobule does not form a horizontal semicircle on its dorsal surface, but rather a straight vertical lobe running more or less parallel to the vermis. Shortly before the caudal end, it turns ven- In dorsal and transverse images, the mildly hyperintense cerebellar nuclei can be seen, embedded in the hypointense white matter. The fastigial nuclei (Nuclei fastigei; Figs 14, 22, 26 and 27: nf) lie directly dorso-lateral to the fastigial recess of the fourth ventricle, whereas the dentate nucleus (Nucleus dentatus; Figs 22, 29 and 30-32: nd) is located more ventro-laterally in the cerebellar hemisphere. Between these two lies the interpositus nucleus (Nucleus interpositus; Figs 14,22,28 and 29: nip), which is composed of two subnuclei (globose and-emboliform nucleus) [38]. Although surrounded by the intensively hypointense medullary body of the cerebellum, the cerebellar nuclei cannot be well visualized.
A large bundle of hypointense fibers originate from the dentate and interpositus nuclei running along the lateral wall of the fourth ventricle in the rostral direction. This bundle is the rostral cerebellar peduncle. Most of these fibers cross in the mesencephalic tegmentum (decussation of the rostral cerebellar peduncles) ending in the contra-lateral red nucleus and the ventral thalamus. The caudal lobe has extensive cortico-pontocerebellar connections via the middle cerebellar peduncles (Figs 11, 12 and 18-21; S11 and S13 Figs: mcp) [23,25].
Medulla. The most characteristic feature of the brainstem is the massive metencephalic pons building the transition from mesencephalon to the medulla oblongata. It is only moderately developed in the horse. It consists of transversely arranged fiber bundles, interspersed with collections of mildly hyperintense pontine nuclei (Nuclei pontis; Figs 11, 12, 28 and 29; S10 and S11 Figs: npo). Cortico-pontine fibers descend from cerebral cortex towards the pontine nuclei, decussate as the transverse fibers of the pons ( Corticospinal tract fibers (Tractus corticospinalis; Figs 12, 17 and 29; S10, S11 and S15 Figs: cst) traverse the pons caudally forming the pyramidal tracts (Figs 13-16, 26, 28 and 29; S12 and S15 Figs: pyr), which are recognizable on each side along the ventral midline. The pyramidal tracts are only small and flat in the horse [41]. At the most caudal pole of the pyramids the cortico-spinal axons cross over the midline and now continue their descent on the contralateral side (Decussatio pyramidalis; Fig 28: dpy).
The mandibular division (Ramus mandibularis nervi trigemini; Figs 31 and 34: man) of the trigeminal nerve contains sensory and motor fibers. The motor nucleus (Fig 29: mtn)  The vestibulocochlear nerve enters the brain stem at the cerebellopontine angle dorsolateral to the emergence of the facial nerve. Its cochlear fibers synapse in dorsal and ventral cochlear nuclei (Figs 13; S12 and S13 Figs: ncv) that are located in the laterally protruding acoustic tubercle (tuberculum acusticum, Fig 14; S13 Fig: tac). This prominence underneath the caudal cerebellar peduncle is flat and rounded in the horse. From here the fibers decussate through the trapezoid body, then ascend in the lateral lemniscus (Lemniscus lateralis; Figs 11, 19 and 29; S10 and S11 Figs: lal) running to the brachium of the caudal colliculus (Brachium colliculi rostralis; Figs 10, 11, 21, 30 and 31; S9 and S10 Figs: bcc). The trapezoid body (Corpus trapezoideum; Figs 1B, 13, 14, 17, 27 and 29; S12 and S13 Figs: tb) is made up of commissural fibers (decussation of the trapezoid body; Figs 14 and 17; S13 Fig: dctb), which originate from either the ventral cochlear nucleus or nucleus of the trapezoid body (Nucleus cochlearis ventralis; Figs 13 and 14; S12 and S13 Figs: ndct), which is also called rostral (superior) olivary nucleus. It can be seen behind the pons as a small transverse ridge.
The caudal continuation of the periaqueductal grey matter is the periventricular grey matter. In its caudal extension through the medulla, numerous nuclear groups can be seen. The topographical relationship between cranial nerve nuclei and the sulcus limitans (Fig 14; S9, S12 and S13 Figs: slm) can serve as a useful landmark in identification of these structures. The lateral sulcus limitans of the fourth ventricle separates the alar and basal plates of the embryonic brain and spinal cord. The derivatives from the alar plate (sensory) lie dorsal or lateral to the sulcus limitans and derivatives from the basal plate (motor) lie ventral or medial to it, which resembles the relationship in the spinal cord.
The medial (Figs 13, 14, 20 and 28; S13 Fig: nvm) and lateral vestibular nuclei (Nuclei vestibulares mediales, -laterales; Figs 13 and 14: S12 and S13 Figs: nvl) are a group of neurons laterally flanking the fourth ventricle. They can be seen as hyperintense areas dorso-lateral to the sulcus limitans and dorsal to the internal fibers of the facial nerve. On the level of these nuclei, a hyperintense band runs vertically from the median sulcus of the fourth ventricle to the trapezoid body. This slender accumulation of cells are the medial raphe nuclei (Fig 14, S13 Fig:  nrm) the tegmental part of the periventricular grey matter.
Further caudally afferent nerve fibers from the viscera unite in the solitary tract (Tractus solitarius; Figs 16; S14 and S15 Figs: sol) carrying information to the nucleus of the solitary tract (nucleus tractus solitarii, Fig 16; S15 Fig: nsol). The tract can be seen as a hypointense spot at the ventrolateral end of the hyperintense nucleus. The cell bands gradually turn medially, meet in midline and close the fourth ventricle. The connection between the two nuclei is the obex, which marks the spino-medullary transition (Fig 26; S15 Fig: obx).
Just rostral to the obex, the nucleus of the solitary tract is bordered medially by the slender motor nucleus of the vagus nerve (Fig 16, S15 Fig: vgn), followed by the conspicuous hypoglossal nucleus (Nucleus nervi hypoglossi; Figs 15 and 16; S14 and S15 Figs: hypn). The nucleus of the soliary tract is dorsally bordered with the nucleus gracilis and nucleus cuneatus.
Ventrally within the reticular formation of the medulla an elongated nuclear column contains motor neurons associated with three cranial nerves, the branchiomotor glossopharyngeal, -vagus and accessory nerve [35]. This is the ambiguus nucleus (Nucleus ambiguus; S13 Fig: amb). The most striking nucleus of the medulla is the inhomogenously hyperintense caudal (inferior) olivary nuclei. It consists of a convoluted band of cells that presents itself as a characteristic serpentine profile. It is located dorsolateral to the pyramid. Axons arising from the nucleus ambiguus pass laterally and slightly ventrally to exit the medulla at the level of the caudal olivary nucleus (Nucleus olivaris; Figs 15-17 and 27-30; S14 and S15 Figs: oli).

Discussion
In contrast to the number of atlases in small animals, to our knowledge there is currently no comparable morphological study in the horse. The present paper is intended to serve as a broad introduction to equine brain morphology. Using a 3 Tesla MR system, images with high spatial resolution and contrast were acquired and described for the use in large animal neurology. However, some factors limit the scope and applicability of the present investigation. In this study, we described one individual horse without taking intraspecies variation due to breed, age, sexual dimorphism, reproductive cycle and other confounding factors into account. It is therefore vital to scan more individuals to reflect the anatomy of the majority of individuals in further studies. To achieve sufficient image quality and to provide morphological detail we have used a 1.5-hour scan time per plane. The resolution of most clinical scanners, which can accommodate horses might be lower and and it is unlikely that MR-images with a comparable resolution and contrast can be produced in a routine in-vivo examination. However, detailed images will improve the diagnosis and understanding of equine central nervous system diseases, especially those affecting distinct structural or functional parts [45] and may also help to establish functional MRI in equine neuroradiology [46].
In general, the equine brain reflects the mammalian blueprint. The characteristics of the ungulate brain incuding the impressive expansion of the neocortex and it's intensive gyrification are obvious in the scans. The pattern of the equine gyri and sulci is by far more complex than in carnivores and in other ungulates. It has been shown that the level of gyrification in mammals is associated with the increase in body mass [47][48][49][50]. However, when species of similar brain weights were compared ungulate brains were significantly more gyrencephalic [51]. It has been proposed that next to the allometric expansion, ungulate-specific sulci have developed in association with special senses of the animals. One important example is the diagonal sulcus, which includes the somatosensory area of the cortex. Sensory information from the lips which are an important explorative organ is processed in the cortex surrounding the diagonal gyrus. Nostrils and tongue are widely represented in the area rostral to the suprasylvian sulcus. [42,44]. The scaling of specialized areas in the cortex also has an indirect impact on the whole brain volume as areas involved in associative processes containing a somatosensory component may also increase.
Due to expansion of the temporal lobe in horses, the insular cortex is is deeply hidden in the depth of the sylvian-and oblique sulcus (opercularisation), and is no longer visible from the outer surface [52]. Little is known about other functional cortical fields in the horse. It is unclear whether the development of the oblique sulcus and gyrus can be seen as a result of a specialized perceptual processing demand or merely based on the evolutionary trend of increased volume of the temporal lobe [53].
The horse has been referred to as being a macrosmatic mammal [23]. Interestingly there is a certain discrepancy between the size of the large rhinencephalon and the rather small olfactory bulbs. In other ungulates as the pig and cow, the linkeage between the two structures is stronger [16][17][18]40]. The same holds for the hippocampus and parahippocampal gyrus that appear small in relation to the massive rhinecephalon. Detailed allometric investigations on the brain and its parts could solve the riddle between volume and function [54,55]. It is uncertain, as to whether this discrepancy is due to a phylogenetic trend for the volume of the allocortex (hippocampus) to decrease in association with volume expansion of the neocortex [56] or if the structures develop independently.
The descending tracts from the motor and premotor region in the cortex running to the internal capsule and continue to form the cerebral peduncles are not well developed. The pyramids are comparatively small [47]. The bulk of the tracts in the peduncles is rather expansive and flattened in the medioventral part. The thickest part runs dorsolaterally on the brainstem. Minimal motor control of the distal limbs in horses probably account for this comparatively underdeveloped efferent system [41].
The shape of the equine cerebellum has been described as beeing very chracteristic amongst ungulates [37,38]. In general, one can see a vertical orientation rather than lateral expansion. A characteristic feature of the equine cerebellum is the deviation of the vermis. There seems to be no rule as to why this shape actually develops. Bolk [38] describes a relationship between the bend of the tuber, folium and declive of the vermis (together forming the median cerebellar lobe) and development and growth of the ansiform lobule (Figs 11-16 and 21-27). He proposes that the sharper the bend of the vermis the smaller is the ansiform lobule. The growth of the tuber results in lateral deviation of the caudal part of the vermis causing an S-shaped bend [38]. A striking feature of the cerebellum are the blind ending medullary fascicles. This was first observed in histological examinations of the pig's cerebellum [57]. It was proposed to be a developmental impediment, thus creating a mechanical barrier due to a lack of space or compression through blood vessels. It was also described to be a physiological cortical aplasia [23,58,59]. Although there are also descriptions of this suspected aplasia in anatomical studies of other species, we propose this finding as to be an artifact that it is probably due to the sharp turns of the foliae out of the sagittal plane, which give the impression of a blind ending of the medullary branches. Other imaging planes do not confirm the presence of this supposed aplasia.