The structural units of the vertebral column are the vertebrae, composed of neural and haemal arches and the vertebral body itself, the centrum. Vertebral bodies are joined by intervertebral tissue primarily derived from the notochord [1–3].
The vertebral column results from a strictly controlled segmentation process that occurs in all vertebrate species and is associated to two main structures, the notochord and the somites [4, 5]. The somites are epithelialized spheres of mesoderm that develop on either side of the neural tube, give rise to dermis, skeletal musculature (dermomyotome) and vertebrae (sclerotome) [6, 7]. While the somite contribution to vertebra formation has been extensively studied, particularly in birds [6, 8], the role of the notochord has received less attention. In teleosts, the majority of extant vertebrates, the notochord is composed of a core of large, vacuolated chordocytes, and an epithelial layer of chordoblasts that secrete the notochord sheath . The notochord sheath is a stratified structure, composed of a thin external membrane, with high elastin content, covering a thicker collagenous layer [10, 11].
In chondrichthyans and most osteichthyans, including tetrapods, but not in teleosts, vertebral bodies have a cartilaginous anlage that subsequently either mineralizes or is replaced by bone. In teleosts such as zebrafish, vertebral centra are formed in the absence of cartilage [12, 13]. Indeed, teleost vertebral centra form through the mineralization of the notochord sheath (chordacentrum), which is then surrounded by somite-derived intramembranous bone (autocentrum) [2, 12, 14–16].
In teleosts, such as Atlantic salmon (Salmo salar) or zebrafish (Danio rerio), the notochord plays an important role in early life stages, as its mechanical function is only replaced by the vertebral column in the postembryonic life [2, 17, 18]. In Atlantic salmon, the initial mineralization of the chordacentrum has been described to be associated with cells of the notochord epithelium (chordoblasts), while bone formation, by sclerotome-derived cells (autocentrum), is a second step . In contrast, Inohaya and co-workers suggest that in medaka (Oryzias latipes), only sclerotome-derived cells are involved in chordacentrum and autocentrum mineralization, with no role of chordoblasts . Yet, a recent study  shows that, also in medaka, with conditional ablation of osterix-positive osteoblasts, notochord sheath mineralization is maintained. Therefore, the main cellular and molecular determinants involved in early notochord mineralization are still under debate. Furthermore, the role of extracellular matrix proteins such as osteocalcin, generally expressed by mature and resting osteoblasts  and by hypertrophic chondrocytes [17, 22–24], in that process, remains unclear. In several teleosts, including zebrafish, two Osteocalcin genes (Oc1 and Oc2) have been identified .
In amniotes the vertebral column is divided into five main regions whereas the vertebral column of teleosts is often only subdivided into two main regions, abdominal and caudal (e.g., [26–28]). However, also in zebrafish, several regions can be recognized within the vertebral column. In particular, regions that contain the most anterior and the most posterior vertebrae are highly specialized [15, 29]. Regional differences are not only apparent at the morphological level but also regarding the tendency of vertebrae to fuse. While zebrafish vertebral bodies usually display no pathological fusion , caudal fin vertebrae undergo several fusions as part of regular development [15, 30–32]. Yet, other teleosts, such as Atlantic salmon and other farmed species, are known to suffer frequent pathological vertebral fusions [33, 34]. Whether regional differences in vertebrae morphology and mineralization relate to the susceptibility to fuse remains an open question.
This study aims to characterize mineralization patterns in different regions of the vertebral column (abdominal, caudal, caudal fin region) using various methods to reveal mineral deposition. Subsequently, these patterns are compared with the histogenesis of the arches, revealed through Collagen type II immunostaining, and to the timing of centrum formation. We characterize the proliferation of notochord cells and we also localize proteins related to mineralization, such as Alkaline phosphatase and Osteocalcin. We here provide the first evidence for the early presence of Osteocalcin 1 in mineralizing chordacentra. Finally, we discuss a possible association between timing of centra formation, the mineralization pattern and occurrence of vertebral fusion.