Histological organization and its relationship to function in the femur of Alligator mississippiensis

Histological analysis of a growth series of alligator femora tests the correlation between strain milieu and microstructure. From mid‐diaphyseal cross‐sections of these femora (n = 7), vascular canal orientation and density as well as collagen fibre organization were recorded. Throughout ontogeny, the proportion of transverse–spiral (TS) collagen in the dorsal cortex is significantly greater than it is in the ventral cortex (P = 0.008). This regional difference in the proportion of TS collagen is correlated with a regional difference in the state of peak principal strain (compressive or tensile). Nevertheless, the predominant orientation of collagen fibres is longitudinal, which is inconsistent with biomechanical hypotheses that involve peak principal or shear strains. Although the density and orientation of vascular canals do not show significant regional differences (P = 0.26 and P = 0.26, respectively), as with collagen orientation, the vascular canal orientation is predominantly longitudinal. The longitudinal organization of both the vascular canals and the collagen fibres is probably a consequence of longitudinal shifting of subperiosteal osteoid during femoral lengthening. When taken together, these data suggest that growth dynamics is the dominant influence on the histological organization of primary bony tissues in alligator femora.     

Penile Anatomy and Hypotheses of Erectile Function in the American Alligator (Alligator mississippiensis): Muscular Eversion and Elastic Retraction

The intromittent organs of most amniotes contain variable‐volume hydrostatic skeletons that are stored in a flexible state and inflate with fluid before or during copulation. However, the penis in male crocodilians is notable because its shaft does not seem to change either its shape or bending stiffness as blood enters its vascular spaces before copulation. Here I report that crocodilians may have evolved a mechanism for penile shaft erection that does not require inflation and detumescence. Dissections of the cloaca in sexually mature male American alligators (Alligator mississippiensis) show that the cross section of the proximal shaft of the alligator penis contains dense collagenous tissues that do not significantly change shape when fluid is added to the central vascular space. The large amount of collagen in the wall and central space of the alligator penis stiffen the structure so it can be simply everted for copulation and rapidly retracted at its completion. Because no muscles insert directly onto the penis, eversion and retraction must be produced indirectly. My results suggest that the contraction of paired levator cloacae muscles around the anterior end of the cloaca rotates the penis out of the cloacal opening and strains the ligamentum rami that connect the base of the penis to the ischia. When the cloacal muscles relax, the elastic recoil of the ligamentum rami can return the penis to its original position inside the cloaca. Anat Rec, 296:488–494, 2013. © 2013 Wiley Periodicals, Inc.     

Changes in the craniofacial complex and alveolar bone height of young adults: Applications to dental medicine

The few studies on craniofacial complex changes in adults have reported contradictory findings. The aim of this study was to radiographically evaluate changes in the craniofacial complex and alveolar bone height of young adults over a 4‐year period. This prospective study included 82 young adults (62 men; mean age, 19.0 ± 0.8 years; 20 women; mean age, 18.8 ± 0.9 years). Three radiographic examinations were performed longitudinally: the first (T0) was conducted at the start of the study, the second (T1) was conducted 2 years later, and the third (T2) was conducted at the end of the study period. As part of these examinations, lateral cephalograms, maxillary and mandibular anterior periapical radiographs, and bilateral posterior bitewing radiographs were obtained. During the 4‐year follow‐up period, all linear cephalometric measurements were significantly greater in men than in women. The amount of increase in the anterior facial height between T1 and T2 was lower in women than in men (P = 0.029). The rate of uprighting of the upper incisors was lower in men than in women over time (P = 0.020). The apex of the nose moved inferiorly between T0 and T2 (P = 0.006). The average overall change in the alveolar bone height was 0.27 mm and the yearly change was 0.07 mm over the 4‐year period (P< 0.001). Significant changes in the skeletal, dental, and soft tissue, as well as, natural changes in alveolar bone, occur in young adults over time. Clin. Anat. 29:1011–1017, 2016. © 2016 Wiley Periodicals, Inc.     

Development of endothermic metabolic response in embryos and hatchlings of the emu (Dromaius novaehollandiae)

During hatching, there is a maturation of the mechanisms controlling the respiratory physiology involved in endotherm in precocial avian species. Here we examined the timing of the development of an endothermic response of oxygen uptake (MO2) to an alteration of ambient temperature (T(a)) in a model precocial species, the preterm and hatching emu (Dromaius novaehollandiae). Late stage pre-pipped and pipped embryos and hatchlings were measured for responses of MO2 and shell or skin temperature (T(s)) to altered T(a) (DeltaT(a)). MO2 remained unchanged in pre-pipped and internally pipped (IP) embryos at the end of 1.5h exposure to DeltaT(a) of +/-10 degrees C. Externally pipped (EP) embryos responded to a cooling and a warming exposure with marked increase and decrease in MO2, as hatchlings responded to DeltaT(a) with an endothermic change in MO2. The demonstration of the endothermic inverse metabolic response first appearing in EP embryos suggests that pre-EP embryos may also possess the ability to produce the endothermic inverse metabolic response, but they are restricted by the eggshell gas conductance. Late pre-pipped and IP embryos were measured again for responses of [Formula: see text] to DeltaT(a) in air and then in a 40% O(2) environment. The metabolic response of pre-pipped embryos at 90% of incubation was partially altered by switching from air to hyperoxia. IP embryos responded to DeltaT(a) in 40% O(2) with apparent inverse changes in MO2. The late stage emu embryo possesses the ability to produce an endothermic metabolic response at an earlier stage of development than in chickens, but this response is limited by the eggshell gas conductance.     

The tongue as a factor in craniofacial growth. 1. Modification of the linear dimensions of the lower jaw

The influence of the tongue on craniofacial growth was studied in 96 miniature pigs MINI-LEWE. The animals were partially glossectomized at different ages and slaughtered at various post-operative intervals. The skulls were macerated for biometric analysis. Mandibular growth in length was significantly reduced in the animals glossectomized at an age of 12 weeks. Both the overall length and that of the tooth-bearing portion were affected.     

The tongue as a factor in craniofacial growth. 3. The influence of the height and angle of the lower jaw

In the miniature pig MINI-LEWE partial glossectomy had no significant effects on vertical growth of the lower jaw, growth of the upper jaw or overall skull growth. No relationship was found between age at glossectomy or post-surgical interval and the size of the gonion angle. The tongue, peri-oral muscles and the muscles of the floor of the mouth are in a state of functional equilibrium which changes in the course of development, the influence of either the tongue or the peri-oral muscles increasing as a result. Partial disruption of this state of equilibrium leads to dysgnathia.     

The tongue as a factor in craniofacial growth. 2. The influence of the width dimension of the lower jaw

The role played by the tongue in orofacial growth is also indicated by the reduced width of the lower jaw. In miniature pigs MINI-LEWE that were partially glossectomized at an age of 12 weeks lateral growth of the whole lower jaw was reduced after eight months. In animals that had been glossectomized at an age of six weeks lateral growth of the lower jaw was reduced in the region of the 1st deciduous molars and the canines right months after glossectomy.     

The effect of a modified blood supply on the craniofacial growth of the rabbit. 4. Skull and lower jaw asymmetry

Using a microscopic measuring method we were able to detect minor regions of asymmetry in the zygomatic arches of rabbits in which the common carotid artery had been ligated. These are considered in connection with the (also slight) asymmetric gains in the dry weight of the muscles of mastication in the ligated animals (see 2nd Bulletin).     

Establishment, maintenance and modifications of the lower jaw dentition of wild Atlantic salmon (Salmo salar L.) throughout its life cycle

In this paper we elucidate the pattern of initiation of the first teeth and the pattern of tooth replacement on the dentary of wild Atlantic salmon (Salmo salar L.), throughout nearly all stages of its life cycle, using serially sectioned heads and jaws, cleared and stained animals, and X-rays. The dentary teeth are set in one row. Tooth germs appear around hatching, first in odd positions, followed by even positions. From position 8 further backwards, teeth are added in adjacent positions. The first replacement teeth appear in animals of about 30 mm fork length. On the dentary of early life stages (alevins and fry), every position in the tooth row holds a functional (i.e. attached and erupted) tooth and a replacement tooth. The alternating pattern set up anteriorly in the dentary by the first-generation teeth changes in juveniles (parr) whereby teeth are in a similar functional (for the erupted teeth) or developmental stage (for the replacement teeth) every three positions. This pattern is also observed in marine animals during their marine life phase and in both sexes of adult animals prior to spawning (grilse and salmon), but every position now holds either a functional tooth or a mineralised replacement tooth. This is likely due to the fact that replacement tooth germs have to grow to a larger size before mineralisation starts. In the following spring, the dentary tooth pattern of animals that have survived spawning (kelts) is highly variable. The abundance of functional teeth in post-spawning animals nevertheless indicates that teeth are not lost over winter. We confirm the earlier reported lack of evidence for the existence of an edentulous life phase, preceding the appearance of so-called breeding teeth during upstream migration to the spawning grounds, and consider breeding teeth to be just another tooth generation in a regularly replacing dentition. This study shows how Atlantic salmon maintains a functional adaptive dentition throughout its complex life cycle.     

The impact of bone and suture material properties on mandibular function in Alligator mississippiensis: testing theoretical phenotypes with finite element analysis

The functional effects of bone and suture stiffness were considered here using finite element models representing three different theoretical phenotypes of an Alligator mississippiensis mandible. The models were loaded using force estimates derived from muscle architecture in dissected specimens, constrained at the 18th and 19th teeth in the upper jaw and 19th tooth of the lower jaw, as well as at the quadrate-articular joint. Stiffness was varied systematically in each theoretical phenotype. The three theoretical phenotypes included: (i) linear elastic isotropic bone of varying stiffness and no sutures; (ii) linear elastic orthotropic bone of varying stiffness with no sutures; and (iii) linear elastic isotropic bone of a constant stiffness with varying suture stiffness. Variation in the isotropic material properties of bone primarily resulted in changes in the magnitude of principal strain. By comparison, variation in the orthotropic material properties of bone and isotropic material properties of sutures resulted in: a greater number of bricks becoming either more compressive or more tensile, changing between being either dominantly compressive or tensile, and having larger changes in the orientation of maximum principal strain. These data indicate that variation in these model properties resulted in changes to the strain regime of the model, highlighting the importance of using biologically verified material properties when modeling vertebrate bones. When bones were compared within each set, the response of each to changing material properties varied. In two of the 12 bones in the mandible, varied material properties within sutures resulted in a decrease in the magnitude of principal strain in bricks adjacent to the bone/suture interface and decreases in stored elastic energy. The varied response of the mandibular bones to changes in suture stiffness highlights the importance of defining the appropriate functional unit when addressing relationships of performance and morphology.     

The presence of a foramen of Luschka in the American alligator (Alligator mississippiensis) and the continuity of the intraventricular and subdural spaces

In humans and most mammals, there is a notch-like portal, the foramen of Luschka (or lateral foramen), which connects the lumen of the fourth ventricle with the subdural space. Gross dissection, light and scanning electron microscopy, and μCT analysis revealed the presence of a foramen of Luschka in the American alligator (Alligator mississippiensis). In this species, the foramen of Luschka is a notch in the dorsolateral wall of the pons immediately caudal to the peduncular base of the cerebellum, near the rostral end of the telovelar membrane over the fourth ventricle. At the foramen of Luschka there was a transition from a superficial pia mater lining to a deep ependymal lining. There was continuity between the lumen of the fourth ventricle and the subdural space, via the foramen of Luschka. This anatomical continuity was further demonstrated by injecting Evans blue into the lateral ventricle which led to extravasation through the foramen of Luschka and pooling of the dye on the lateral surface of the brain. Simultaneous subdural and intraventricular recordings of cerebrospinal fluid (CSF) pressures revealed a stable agreement between the two pressures at rest. Perturbation of the system allowed for static and dynamic differences to develop, which could indicate varying flow patterns of CSF through the foramen of Luschka.     

Simple,noninvasive system for measuring the heart rate of avian embryos and hatchlings by means of a piezoelectric film

Using a flexible piezoelectric film, the authors developed a simple system to determine noninvasively the heart rate of chicken embryos and hatchlings. The film was piezoelectric polyvinylidene fluoride (PVDF), which is sensitive enough to detect cardiogenic ballistic movements of the egg (ballistocardiogram (BCG)) and precordial movements of the hatchling attributable to cardiac contractions (apexcardiogram (ACG)). The BCG could be detected, during the second half of incubation, by placing the egg on the PVDF film on a soft substrate. The detected signal was found to be a measure of movement velocity. The ACG could be measured when the hatchling's chest wall made contact with the PVDF film installed in a box in which the hatchling was confined. The heart rate was counted from the lag time of autocorrelation calculated for a certain time segment (e.g. 2s) of the BCG and ACG recordings.     

Differences between sliding semi-landmark methods in geometric morphometrics, with an application to human craniofacial and dental variation

Over the last decade, geometric morphometric methods have been applied increasingly to the study of human form. When too few landmarks are available, outlines can be digitized as series of discrete points. The individual points must be slid along a tangential direction so as to remove tangential variation, because contours should be homologous from subject to subject whereas their individual points need not. This variation can be removed by minimizing either bending energy (BE) or Procrustes distance (D) with respect to a mean reference form. Because these two criteria make different assumptions, it becomes necessary to study how these differences modify the results obtained. We performed bootstrapped-based Goodall's F-test, Foote's measurement, principal component (PC) and discriminant function analyses on human molars and craniometric data to compare the results obtained by the two criteria. Results show that: (1) F-scores and P-values were similar for both criteria; (2) results of Foote's measurement show that both criteria yield different estimates of within- and between-sample variation; (3) there is low correlation between the first PC axes obtained by D and BE; (4) the percentage of correct classification is similar for BE and D, but the ordination of groups along discriminant scores differs between them. The differences between criteria can alter the results when morphological variation in the sample is small, as in the analysis of modern human populations.     

Development of the pattern of dentition and dental laminae of costa rican plethodontid salamanders (Amphibia: Urodela)

The pattern of development of teeth and dental laminae of three Costa Rican plethodontids (Amphibia, Urodela, Plethodontidae) was investigated using transparent preparations, light microscopy and scanning electron microscopy. The teeth of the jaws are monostichously positioned, those of the posterior vomeral parts are polystichously arranged. The anterior vomeral parts carry monostichously positioned teeth at the caudal margin; yet, the adult Bolitoglossa subpalmata possesses two lines. As a sex dimorphism adult males display long monocuspid premaxillary teeth which protrude to the outside of the mouth cavity. All species studied possess paired dental laminae in the lower jaw. Nototriton abscondens possesses an unpaired dental lamina in the upper jaw, which is constricted between the unpaired premaxillary and the maxillaries. In contrast, the dental laminae in the upper jaw of B. subpalmata and Oedipina uniformis are segmented into a premaxillary and two maxillary laminae. All species possess a pair of anterior vomeral and a pair of posterior vomeral dental laminae in the adults, whereas the vomeral dental laminae of the subadults are unsegmented. The pattern of dentition is compared with that of Gyrinophilus and Eurycea.     

Detection of antibodies to a pathogenic mycoplasma in American alligators (Alligator mississippiensis), broad-nosed Caimans (Caiman latirostris), and Siamese crocodiles (Crocodylus siamensis)

An epidemic of pneumonia with fibrinous polyserositis and multifocal arthritis emerged in captive American alligators (Alligator mississippiensis) in Florida, United States, in 1995. Mycoplasma alligatoris sp. nov. was cultured from multiple organs, peripheral blood, synovial fluid, and cerebrospinal fluid of affected alligators. In a subsequent experimental inoculation study, the Henle-Koch-Evans postulates were fulfilled for M. alligatoris as the etiological agent of fatal mycoplasmosis of alligators. That finding was remarkable because mycoplasmal disease is rarely fatal in animals. An enzyme-linked immunosorbent assay (ELISA) for the detection of antibodies produced by alligators in response to M. alligatoris exposure was developed by using plasma obtained from naturally infected alligators during the original epidemic. The assay was validated by using plasma obtained during an experimental dose-response study and applied to analyze plasma obtained from captive and wild crocodilian species. The ELISA reliably detected alligator seroconversion (P < 0.05) beginning 6 weeks after inoculation. The ELISA also detected seroconversion (P < 0.05) in the relatively closely related broad-nosed caiman Caiman latirostris and the relatively distantly related Siamese crocodile Crocodylus siamensis following experimental inoculation with M. alligatoris. The ELISA may be used to monitor exposure to the lethal pathogen M. alligatoris among captive, repatriated, and wild crocodilian species.     

Proliferation, apoptosis and expression of non-collagenous proteins: differences between the upper and the lower jaw bone in vitro

One of the effects observed during several screening studies for osteocompatibility in vitro was that cells derived from the upper and lower jaw exhibited distinct differences regarding proliferation. Therefore, the aim of this study was to examine systematically whether a single osteoblast possesses abilities which are specific to the upper or lower jaw. Both human maxillary and mandibular bone samples without any clinical or radiographic evidence of pathology were obtained from 4 male donors aged between 40 and 45 years. Cells were cultured for up to 25 days to investigate in vitro development. Total and apoptotic cell numbers were estimated by image analysis. Cells were identified as bone-like cells using immunocytochemical determination of bone sialoprotein (BSP) and osteocalcin expression. The number of healthy cells was significantly higher for cells of the lower jaw compared to those of the upper jaw. The number of apoptotic cells showed an inverse pattern. The expression pattern of osseo-inductive BSP correlated with the proliferation rate of the cells. The pattern of osteocalcin expression was related to the number of apoptotic cells. Our findings are new but were anticipated regarding the well-known differences in the healing process around implants in the lower jaw versus the upper jaw. Additionally, a relationship between our results and some diseases of the lower/upper jaw seems obvious. Future work on cell responses to biomaterials should define the origin of the cells more precisely.     

Axis development in avian embryos: the ability of Hensen's node to self-differentiate,as analyzed with heterochronic grafting experiments

A series of experiments consisting of transplantation of Hensen's nodes has been conducted to examine axis development in avian embryos. In the first group of experiments, Hensen's nodes from quail embryos were transplanted homotopically and either isochronically or heterochronically to chick embryos, and the structures derived form the grafted nodes were assessed. The grafted Hensen's nodes typically self-differentiated structures appropriate for their stages, and the host embryos developed normally; the structures formed from grafted tissue usually merged caudally with the comparable host structures. Thus, even when the stages of the donor and host tissues were significantly mismatched (e.g., stage 3 donors and stage 9 hosts or vice versa), the graft was unable to repattern the host's neuraxis, and the host was unable to respecify the types of structures derived from the graft. In the second group of experiments, Hensen's nodes from quail embryos were transplanted to sites located just lateral to Hensen's nodes of host chick embryos, thereby providing the potential for development of additional axes. A single axis always resulted in each case in which further development occurred, with the graft self-differentiating its typical stage-specific structures, all of which merged caudally with comparable host structures. A final group of experiments served principally as a control and tested the ability of a part of Hensen's node, when it was transplanted to the extraembryonic germ cell crescent, to organize an ectopic embryo. In these experiments, the entire thickness and length of each Hensen's node, but only the central one-third to one-half of its width, was transplanted to host blastoderms, yet ectopic embryos, complete with induced neuraxes, were formed. Therefore, a part of Hensen's node has the ability to function fully as an organizer when placed in a conducive environment. Collectively, these results provide further documentation of the strong ability of Hensen's node to self-differentiate, and they suggest that once morphogenetic movements are under way, neuraxial structures can form, and characteristic rostrocaudal patterning of the neuraxis can occur, without sustained influence from Hensen's node.     

The development of cytogenetically normal, abnormal and mosaic embryos: a theoretical model

Assisted reproduction and preimplantation genetic diagnosis (PGD) involve various complicated techniques, each of them with its own problems. However, the greatest problem with PGD for chromosome abnormalities is not of a technical nature but is a biological phenomenon: chromosomal mosaicism in the cleavage stage embryo. Here, we present a hypothetical, quantitative model for the development of chromosomally normal, abnormal and mosaic embryos. The arising of mosaicism in 2-8-cell embryos was described by a binomial probability model on the occurrence of mitotic events inducing chromosomal changes in the blastomeres. This model converted the 'mean' rate of mosaicism found in cross-sectional studies (60%) into an equal rate of mosaic embryos at arrival at the 8-cell stage (59.8%). The disappearance of > 90% of the mosaic embryos or the mosaicism itself from surviving embryos during the morula stage was explained by mitotic arrest of most of the mitotically changed cells under increasing cell cycle control. In our model, 25.9 and 14.3% of the embryos at the 8-cell stage are normal and abnormal respectively. The remaining 59.8% of the embryo shows mosaicism: 34.6% of abnormal/normal cells and 25.2% of abnormal/abnormal cells. The high proportion of abnormal and mosaic embryos together explains the high rate of abnormal laboratory findings in PGD for chromosomal abnormalities and aneuploidy screening. The poor representation of a 1- or 2-cell biopsy for the 7- or 6-cell post-biopsy embryo in the case of mosaicism explains the high rate of false-negative and false-positive results.     

Axis development in avian embryos: the ability of Hensen''s node to self-differentiate, as analyzed with heterochronic grafting experiments

A series of experiments consisting of transplantation of Hensen's nodes has been conducted to examine axis development in avian embryos. In the first group of experiments, Hensen's nodes from quail embryos were transplanted homotopically and either isochronically or heterochronically to chick embryos, and the structures derived form the grafted nodes were assessed. The grafted Hensen's nodes typically self-differentiated structures appropriate for their stages, and the host embryos developed normally; the structures formed from grafted tissue usually merged caudally with the comparable host structures. Thus, even when the stages of the donor and host tissues were significantly mismatched (e.g., stage 3 donors and stage 9 hosts or vice versa), the graft was unable to repattern the host's neuraxis, and the host was unable to respecify the types of structures derived from the graft. In the second group of experiments, Hensen's nodes from quail embryos were transplanted to sites located just lateral to Hensen's nodes of host chick embryos, thereby providing the potential for development of additional axes. A single axis always resulted in each case in which further development occurred, with the graft self-differentiating its typical stage-specific structures, all of which merged caudally with comparable host structures. A final group of experiments served principally as a control and tested the ability of a part of Hensen's node, when it was transplanted to the extraembryonic germ cell crescent, to organize an ectopic embryo. In these experiments, the entire thickness and length of each Hensen's node, but only the central one-third to one-half of its width, was transplanted to host blastoderms, yet ectopic embryos, complete with induced neuraxes, were formed. Therefore, a part of Hensen's node has the ability to function fully as an organizer when placed in a conducive environment. Collectively, these results provide further documentation of the strong ability of Hensen's node to self-differentiate, and they suggest that once morphogenetic movements are under way, neuraxial structures can form, and characteristic rostrocaudal patterning of the neuraxis can occur, without sustained influence from Hensen's node.