Anteroposterior Levels Research Articles

Pose Analysis in Free-Swimming Adult Zebrafish, Danio rerio: “Fishy” Origins of Movement Design

Introduction: Movement requires maneuvers that generate thrust to either make turns or move the body forward in physical space. The computational space for perpetually controlling the relative position of every point on the body surface can be vast. We hypothesize the evolution of efficient design for movement that minimizes active (neural) control by leveraging the passive (reactive) forces between the body and the surrounding medium at play. To test our hypothesis, we investigate the presence of stereotypical postures during free-swimming in adult zebrafish, Danio rerio. Methods: We perform markerless tracking using DeepLabCut (DLC), a deep learning pose-estimation toolkit, to track geometric relationships between body parts. We identify putative clusters of postural configurations from twelve freely behaving zebrafish, using unsupervised multivariate time-series analysis (B-SOiD machine-learning software) and of distances and angles between body segments extracted from DLC data. Results: When applied to single individuals, DLC-extracted data reveal a best-fit for 36–50 clusters in contrast to 86 clusters for data pooled from all 12 animals. The centroids of each cluster obtained over 14,000 sequential frames represent an a priori classification into relatively stable “target body postures.” We use multidimensional scaling of mean parameter values for each cluster to map cluster centroids within two dimensions of postural space. From a posteriori visual analysis, we condense neighboring postural variants into 15 superclusters or core body configurations. We develop a nomenclature specifying the anteroposterior level/s (upper, mid, and lower) and degree of bending. Conclusion: Our results suggest that constraining bends to mainly three anteroposterior levels in fish paved the way for the evolution of a neck, fore- and hind limb design for maneuverability in land vertebrates.

Evaluation of bone depth, cortical bone, and mucosa thickness of palatal posterior supra-alveolar insertion site for miniscrew placement

BackgroundThe use of palatal miniscrew offers the possibility to improve the effectiveness of orthodontic expansion devices. Palatal expanders supported by miniscrew can be applied with different clinical protocols. Some authors proposed the use of four palatal miniscrews during miniscrew-supported palatal expansion to maximize skeletal effects in young adults’ treatment. However, bone availability decreases in the posterior paramedian palatal regions, making the positioning of the two-posterior paramedian palatal miniscrews challenging, when it is performed avoiding nasal cavities invasion. Some authors proposed miniscrews insertion in a specific region located laterally to the palatal process of the maxillary bone, and apically relatively to the dento-alveolar process. The aim of this study was to evaluate the bone thickness, cortical bone thickness, and mucosae depth of this anatomical site that, in this study, was defined as palatal posterior supra-alveolar insertion site.ResultsThe evaluation of bone availability of palatal posterior supra-alveolar insertion site at different antero-posterior levels showed that the maximum amount of total bone thickness was found between the second premolar and the first molar. At this level total bone, thickness is significantly (p < .05) greater compared to the other sagittal sites and it offers on average around 2 mm of extra bone depth for miniscrew placement. Cortical bone thickness is adequate for primary miniscrew stability. Overall, cortical bone thickness considered at different insertion sites showed significant statistically (p < .05) differences. The findings of this study showed that palatal mucosa is particularly thick with average values ranging from 4 to 7 mm, and its extension ultimately affects miniscrew length selection. Palatal mucosa thickness showed no clinically significant differences comparing different sagittal and vertical insertion sites. Data also showed that palatal mucosal thickness slightly significantly increases (p < .05) with the inclination of the insertion axis relative to the occlusal plane. Finally, study findings showed that vertical growth pattern can significantly affect considered outcomes (p < .05).ConclusionsPalatal posterior supra-alveolar insertion site is an appropriate site for posterior insertion of palatal miniscrews. Considering high anatomical variation preliminary CBCT evaluation is important to achieve optimal miniscrew placement.

Qualitative and quantitative evaluation of the palatal posterior supra alveolar bone region for miniscrews placement

This study aimed to evaluate the total bone thickness, the cortical bone thickness, and the mucosa depth of specific site that we define Palatal Posterior Supra Alveolar Bone (PPSAB), for the miniscrews insertion. The sample included CBCT records and digital models of 23 subjects (17F 6M age mean 32) retrospectively evaluated. Digital dental casts were superimposed with CBTC. Total Bone thickness, cortical bone, and mucosa depth were evaluated at the first molar upper level. The measurements were performed in coronal view, the scans were evaluated mesially, infraradicularlly, and distally to the first molar. Each scan was considered at four levels in the apico-coronal direction (-4 mm, -2 mm, 0 mm, +2 mm) for a total of 36 measurements per subject. The total bone thickness decrease moves from mesial insertion sites to distal ones. Moreover, it increases in the coronal insertion sites compared to the apical at all the anteroposterior levels. However, at -4 mm (the most apical site evaluated) the total bone thickness decreases due to the proximity to the first molar root. The insertion site with the greatest average total bone thickness (8.68 mm) was mesial to first molar 2 mm apical to the corner palatal point (0 mm). The palatal posterior supra alveolar bone (PPSAB), could be a suitable site for the orthodontic miniscrews insertion in the majority of orthodontic adult patients. However, great variability was observed among different considered subjects. Cortical bone thickness is less than 2 mm. Palatal mucosa was on average more than 4 mm.

Evaluation of palatal bone depth, cortical bone, and mucosa thickness for optimal orthodontic miniscrew placement performed according to the third palatal ruga clinical reference.

This retrospective CBCT study aimed to evaluate the palatal anatomical characteristics using the third palatal ruga as a reliable clinical reference for miniscrew placement. Thirty-six subjects (mean age17.1 y.o. ± 4.1) were randomly selected and their records (CBCT volume and maxillary digital models) were included.BlueSkyPlan CBCT software viewer (BluSkyBio, V4.7) was used to measure the following outcomes at the level of third palatal ruga, 2 mm anteriorly and 2 mm posteriorly: total bone depth, cortical bone thickness, and mucosa thickness. The outcomes were evaluated on lines perpendicular to the palatal mucosa laying on different sagittal planes: the mid-palatal plane, 2 and 4 mm paramedian planes. The maximum mean amount of bone depth was registered 2 mm posteriorly to the third ruga and 4 mm paramedian (9.7 mm). No significant difference was observed between the third ruga insertion site and its corresponding 2 mm posterior site. Cortical bone of palatal vault did not change significantly in anteroposterior direction for all the considered sites. Significant differences were found comparing cortical bone at the suture level with cortical bone 2-mm and 4-mm paramedian at all anteroposterior levels. Palatal mucosa increases its thickness in paramedian insertion sites, and it decreases in posterior insertion sites. Both third palatal ruga and 2 mm posteriorly to third ruga (4 mm paramedian) could be the optimal insertion site for palatal miniscrew placement, depending on individual anatomic conditions. The thickness of the cortical palatal bone showed, at 4 mm paramedian, optimal characteristics for miniscrew primary stability. Palatal mucosa thickness values suggest miniscrew neck extension of 2.0-2.5 mm for optimal mucosa adaptation.

138. Toward adaptive radiotherapy: Morphological and setup variability in head and neck TomoTherapy treatments

PurposeIn head and neck (H&N) radiotherapy, an essential requirement is the reproducibility of the relative position of head and neck during the whole treatment. Moreover, adaptive radiotherapy for H&N cancer would benefit from indicators capable of easily identifying morphological alterations. Aim of this study was to test whether the immobilization device guarantees a reproducible head and neck position and to identify, on set-up verification images, some indicators capable of describing morphological alterations. Methods and materialsA retrospective analysis on 15 patients immobilized using s-shaped head-neck-shoulders masks and treated with TomoTherapy (50–70 Gy/25-33fr) was conducted. For each treatment session two MVCT-CT registrations were performed alternatively optimizing matching on the head or the neck. For each patient mean differences between the two registrations parameters (Δx, Δy, Δz, Δpitch, Δroll, Δyaw) over all treatment sessions were evaluated.Morphological alterations were studied (once a week for the whole treatment duration) by measuring on MVCT images the skin-to-skin distance at 3 different antero-posterior levels: at the center of parotid glands (line1) and 2 cm (line 2) and 4 cm (line 3) anteriorly. Mean percentage variations respect to the first week were evaluated for all patients. A Student’s t-test was carried out (p < 0,01). ResultsMean differences between head and neck registration parameters are reported in Table 1. Except for two patients, mean differences were smaller than 1.7 mm and 1.2°.Mean percentage variations of lines’ length are reported in Fig. 1. For all, a decreasing trend was observed. Variations are statistically significant starting from the 4th week for lines 1 and 2, and only from the 6th week for line 3. ConclusionThe immobilization device enables adequate H&N set-up during the whole treatment for almost all patients. Lines 1 and 2 drawn on MVCT images seem to be suitable for describing morphological alterations that might deserve re-planning.

Anatomical integration of the sacral-hindlimb unit coordinated by GDF11 underlies variation in hindlimb positioning in tetrapods.

Elucidating how body parts from different primordia are integrated during development is essential for understanding the nature of morphological evolution. In tetrapod evolution, while the position of the hindlimb has diversified along with the vertebral formula, the mechanism responsible for this coordination has not been well understood. However, this synchronization suggests the presence of an evolutionarily conserved developmental mechanism that coordinates the positioning of the hindlimb skeleton derived from the lateral plate mesoderm with that of the sacral vertebrae derived from the somites. Here we show that GDF11 secreted from the posterior axial mesoderm is a key factor in the integration of sacral vertebrae and hindlimb positioning by inducing Hox gene expression in two different primordia. Manipulating the onset of GDF11 activity altered the position of the hindlimb in chicken embryos, indicating that the onset of Gdf11 expression is responsible for the coordinated positioning of the sacral vertebrae and hindlimbs. Through comparative analysis with other vertebrate embryos, we also show that each tetrapod species has a unique onset timing of Gdf11 expression, which is tightly correlated with the anteroposterior levels of the hindlimb bud. We conclude that the evolutionary diversity of hindlimb positioning resulted from heterochronic shifts in Gdf11 expression, which led to coordinated shifts in the sacral-hindlimb unit along the anteroposterior axis.

Crossed motor innervation of the base of human tongue.

Muscle fibers of the genioglossus (GG) form the bulk of the muscle mass at the base of the tongue. The motor control of the tongue is critical for vocalization, feeding, and breathing. Our goal was to assess the patterns of motor innervation of GG single motor units (SMUs) in humans. Simultaneous monopolar recordings were obtained from four sites in the base of the tongue bilaterally at two antero-posterior levels from 16 resting, awake, healthy adult males, who wore a face mask with airway pressure and airflow sensors. We analyzed 69 data segments in which at least one lead contained large action potentials generated by an SMU. Such potentials served as triggers for spike-triggered averaging (STA) of signals recorded from the other three sites. Spontaneous activity of the SMUs was classified as inspiratory modulated, expiratory modulated, or tonic. Consistent with the antero-posterior orientation of GG fibers, 44 STAs (77%) recorded ipsilateral to the trigger yielded sharp action potentials with a median amplitude of 52 μV [interquartile range (IQR): 25-190] that were time shifted relative to the trigger by about 1 ms. Notably, 48% of recordings on the side opposite to the trigger also yielded sharp action potentials. Of those, 17 (29%) had a median amplitude of 63 μV (IQR: 39-96), and most were generated by tonic SMUs. Thus a considerable proportion of GG muscle fibers receive a crossed motor innervation. Crossed innervation may help ensure symmetry and stability of tongue position and movements under normal conditions and following injury or degenerative changes affecting the tongue.

Lasting Pure-Motor Deficits after Focal Posterior Internal Capsule White-Matter Infarcts in Rats

Small white-matter infarcts of the internal capsule are clinically prevalent but underrepresented among currently available animal models of ischemic stroke. In particular, the assessment of long-term outcome, a primary end point in clinical practice, has been challenging due to mild deficits and the rapid and often complete recovery in most experimental models. We, therefore, sought to develop a focal white-matter infarction model that can mimic the lasting neurologic deficits commonly observed in stroke patients. The potent vasoconstrictor endothelin-1 (n=24) or vehicle (n=9) was stereotactically injected into the internal capsule at one of three antero-posterior levels (1, 2, or 3 mm posterior to bregma) in male Sprague-Dawley rats. Endothelin-injected animals showed highly focal (~1 mm(3)) and reproducible ischemic infarcts, with severe axonal and myelin loss accompanied by cellular infiltration when examined 2 and 4 weeks after injection. Only those rats injected with endothelin-1 at the most posterior location developed robust and pure-motor deficits in adhesive removal, cylinder and foot-fault tests that persisted at 1 month, without detectable sensory impairments. In summary, we present an internal capsule stroke model optimized to produce lasting pure-motor deficits in rats that may be suitable to study neurologic recovery and rehabilitation after white-matter injury.

The Temporal Sequence of the Mammalian Neocortical Neurogenetic Program Drives Mediolateral Pattern in the Chick Pallium

The six-layered neocortex permits complex information processing in all mammalian species. Because its homologous region (the pallium) in nonmammalian amniotes has a different architecture, the ability of neocortical progenitors to generate an orderly sequence of distinct cell types was thought to have arisen in the mammalian lineage. This study, however, shows that layer-specific neuron subtypes do exist in the chick pallium. Deep- and upper-layer neurons are not layered but are segregated in distinct mediolateral domains in vivo. Surprisingly, cultured chick neural progenitors produce multiple layer-specific neuronal subtypes in the same chronological sequence as seen in mammals. These results suggest that the temporal sequence of the neocortical neurogenetic program was already inherent in the last common ancestor of mammals and birds and that mammals use this conserved program to generate a uniformly layered neocortex, whereas birds impose spatial constraints on the sequence to pattern the pallium.

Borders, extent, and topography of human perirhinal cortex as revealed using multiple modern neuroanatomical and pathological markers

Despite rapidly increasing interests in specific contributions of different components of human medial temporal lobe (MTL) to memory and memory impairments in normal aging and in many abnormal conditions such as Alzheimer's disease and Pick's disease, few modern neuroanatomical studies are available about the borders, extent, and topography of human perirhinal areas 35 and 36, which are important components of the MTL memory system. By a combined use of several cellular, neurochemical, and pathological markers, which mainly include neuronal nuclear antigen, calcium-binding proteins (parvalbumin and calbindin-D28k), nonphosphorylated neurofilament protein (SMI-32), Wisteria floribunda agglutinin, and abnormally phosphorylated tau (AT8), this study has revealed that the borders of human perirhinal areas 35 and 36 are significantly different from those defined with conventional Nissl staining. In general, areas 35 and 36 occupy the ventromedial temporopolar and rhinal sulcal regions, the collateral sulcal region, and the anterior two-thirds of fusiform gyrus or occipitotemporal gyrus. Furthermore, the precise borders, extent, and topography of human areas 35 and 36 and adjoining entorhinal cortex were marked at different anteroposterior levels of the MTL with reference to variations of rhinal and collateral sulci and other useful landmarks. These findings would provide reliable neuroanatomical base for the great and yet rapidly increasing number of neuroimaging studies of the human MTL structures in healthy and many abnormal conditions.

Multimodal architectonic subdivision of the rostral part (area F5) of the macaque ventral premotor cortex

We used a cyto-, myelo-, and chemoarchitectonic (distribution of SMI-32 and calbindin immunoreactivity) approach to assess whether the rostral histochemical area F5 of the ventral premotor cortex (PMv) comprises architectonically distinct areas, possibly corresponding to functionally different fields. Three areas were identified, occupying different parts of F5. One area, designated as "convexity" F5 (F5c), extends on the postarcuate convexity cortex adjacent to the inferior arcuate sulcus and is characterized, cytoarchitectonically, by a poorly laminated appearance, resulting from an overall cell population rather homogeneous in size and density. The other two areas, designated as "posterior" and "anterior" F5 (F5p and F5a, respectively), lie within the postarcuate bank at different anteroposterior levels. Major cytoarchitectonic features of F5p are a layer III relatively homogeneous in cell size and density, a cell-dense layer Va, and the presence of relatively large pyramids in layer Vb. Major cytoarchitectonic features of F5a are the presence of relatively large pyramids in lowest layer III and a prominent, homogenous layer V. Furthermore, our results showed that F5c and F5p border caudally with a caudal PMv area corresponding to histochemical area F4, providing additional evidence for a general subdivision of the macaque PMv into a caudal and a rostral part, corresponding to F4 and to the F5 complex, respectively. The present data, together with other functional and connectional data, suggest that the three rostral PMv areas F5p, F5a, and F5c correspond to distinct cortical entities, possibly involved in different aspects of motor control and cognitive motor functions.

Neuroanatomical structure of the spinner dolphin (Stenella longirostris orientalis) brain from magnetic resonance images.

High-resolution magnetic resonance (MR) images of the brain of an adult spinner dolphin (Stenella longirostris orientalis) were acquired in the coronal plane at 55 antero-posterior levels. From these scans a computer-generated set of resectioned virtual images in the two remaining orthogonal planes was constructed with the use of the VoxelView and VoxelMath (Vital Images, Inc.) programs. Neuroanatomical structures were labeled in all three planes, providing the first labeled anatomical description of the spinner dolphin brain.

Thalamic neuropathology in the chronic pilocarpine and picrotoxin model of epilepsy

Adult male Wistar rats were injected with 150/0.5, 75/1.5 and 50/2.0 mg/kg of pilocarpine (Pilo) and picrotoxin (PTX) (Pilo/PTX mg/kg). The vast majority of the animals developed status epilepticus (SE), after which they were observed for a period of 120–131 days for the occurrence of spontaneous recurrent seizures (SRS). After the experiments, animals were deeply anesthetized, perfused with a 10% formaldehyde fixative solution and their brains were processed with cresyl violet, Perls and Von Kossa techniques. Cell counts were performed under a regular microscopic grid in diverse anteroposterior levels of the thalamus. Several thalamic nuclei in the epileptic groups, particularly the central medial, central lateral, paracentral, mediodorsal, laterodorsal and lateroposterior, showed intense cell loss, pathologic calcification and iron tissue deposits. Our results are relevant to support the importance of the thalamus in the pathogenesis of the epilepsies.

Anatomy and three-dimensional reconstructions of the brain of a bottlenose dolphin (Tursiops truncatus) from magnetic resonance images.

Cetacean (dolphin, whale, and porpoise) brains are among the least studied mammalian brains because of the formidable challenge of collecting and histologically preparing such relatively rare and large specimens. Magnetic resonance imaging offers a means of observing the internal structure of the brain when traditional histological procedures are not practical. Furthermore, internal structures can be analyzed in their precise anatomic positions, which is difficult to accomplish after the spatial distortions often accompanying histological processing. In this study, images of the brain of an adult bottlenose dolphin, Tursiops truncatus, were scanned in the coronal plane at 148 antero-posterior levels. From these scans a computer-generated three-dimensional model was constructed using the programs VoxelView and VoxelMath (Vital Images, Inc.). This model, wherein details of internal and external morphology are represented in three-dimensional space, was then resectioned in orthogonal planes to produce corresponding series of virtual sections in the horizontal and sagittal planes. Sections in all three planes display the sizes and positions of major neuroanatomical features such as the arrangement of cortical lobes and subcortical structures such as the inferior and superior colliculi, and demonstrate the utility of MRI for neuroanatomical investigations of dolphin brains.

Estrogens modulate experimentally induced apoptosis of granule cells in the adult hippocampus.

Estrogens are known to have broad effects on neuronal plasticity, but their specific role in neuronal cell death has not been determined. In the present study, we investigated the effects of beta-estradiol on an experimental model of apoptosis of granule cells of the dentate gyrus, i.e., apoptosis induced by intraventricular injection of the microtubule polymerization inhibitor colchicine. Cell death was characterized with multiple methods, including TUNEL and DNA electrophoresis. Nonrandom digestion of DNA was observed within 8-10 hours after colchicine injection, followed by condensation and fragmentation of granule cell nuclei and extensive anterograde degeneration of mossy fibers/terminals in 2 days. We compared the outcomes of the above-described manipulation in ovariectomized or sham-operated rats and animals treated daily with beta-estradiol or vehicle. Animals were lesioned with colchicine or vehicle 2 weeks after ovariectomy or sham operation. Beta-estradiol or vehicle was administered for 1 week prior to lesion and was continued for a further 2 weeks. Total numbers and densities of granule cells in different animal groups were counted by stereology in various anteroposterior levels of the hippocampus. Our results show that ovariectomy intensifies colchicine-induced granule cell apoptosis, which is ameliorated by exogenous beta-estradiol. In doses that ameliorate the adverse effect of ovariectomy, exogenous beta-estradiol appears to have no effect of preventing granule cell death in animals with intact ovaries; i.e., an estrogen excess is not more neuroprotective than physiological levels of these hormones. Taken together, our results indicate that estrogen deprivation increases the vulnerability of hippocampal neurons to injury and may predispose to neurological diseases occurring after menopause.

Axial skeletal patterning in mice lacking all paralogous group 8 Hox genes.

We present a detailed study of the genetic basis of mesodermal axial patterning by paralogous group 8 Hox genes in the mouse. The phenotype of Hoxd8 loss-of-function mutants is presented, and compared with that of Hoxb8- and Hoxc8-null mice. Our analysis of single mutants reveals common features for the Hoxc8 and Hoxd8 genes in patterning lower thoracic and lumbar vertebrae. In the Hoxb8 mutant, more anterior axial regions are affected. The three paralogous Hox genes are expressed up to similar rostral boundaries in the mesoderm, but at levels that strongly vary with the axial position. We find that the axial region affected in each of the single mutants mostly corresponds to the area with the highest level of gene expression. However, analysis of double and triple mutants reveals that lower expression of the other two paralogous genes also plays a patterning role when the mainly expressed gene is defective. We therefore conclude that paralogous group 8 Hox genes are involved in patterning quite an extensive anteroposterior (AP) axial region. Phenotypes of double and triple mutants reveal that Hoxb8, Hoxc8 and Hoxd8 have redundant functions at upper thoracic and sacral levels, including positioning of the hindlimbs. Interestingly, loss of functional Hoxb8 alleles partially rescues the phenotype of Hoxc8- and Hoxc8/Hoxd8-null mutants at lower thoracic and lumbar levels. This suggests that Hoxb8 affects patterning at these axial positions differently from the other paralogous gene products. We conclude that paralogous Hox genes can have a unique role in patterning specific axial regions in addition to their redundant function at other AP levels.

Anatomy and three-dimensional reconstructions of the brain of the white whale (Delphinapterus leucas) from magnetic resonance images.

Magnetic resonance imaging offers a means of observing the internal structure of the brain where traditional procedures of embedding, sectioning, staining, mounting, and microscopic examination of thousands of sections are not practical. Furthermore, internal structures can be analyzed in their precise quantitative spatial interrelationships, which is difficult to accomplish after the spatial distortions often accompanying histological processing. For these reasons, magnetic resonance imaging makes specimens that were traditionally difficult to analyze, more accessible. In the present study, images of the brain of a white whale (Beluga) Delphinapterus leucas were scanned in the coronal plane at 119 antero-posterior levels. From these scans, a computer-generated three-dimensional model was constructed using the programs VoxelView and VoxelMath (Vital Images, Inc.). This model, wherein details of internal and external morphology are represented in three-dimensional space, was then resectioned in orthogonal planes to produce corresponding series of "virtual" sections in the horizontal and sagittal planes. Sections in all three planes display the sizes and positions of such structures as the corpus callosum, internal capsule, cerebral peduncles, cerebral ventricles, certain thalamic nuclear groups, caudate nucleus, ventral striatum, pontine nuclei, cerebellar cortex and white matter, and all cerebral cortical sulci and gyri.

Bmp activity establishes a gradient of positional information throughout the entire neural plate.

Bone morphogenetic proteins (Bmps) are key regulators of dorsoventral (DV) patterning. Within the ectoderm, Bmp activity has been shown to inhibit neural development, promote epidermal differentiation and influence the specification of dorsal neurons and neural crest. In this study, we examine the patterning of neural tissue in mutant zebrafish embryos with compromised Bmp signalling activity. We find that although Bmp activity does not influence anteroposterior (AP) patterning, it does affect DV patterning at all AP levels of the neural plate. Thus, we show that Bmp activity is required for specification of cell fates around the margin of the entire neural plate, including forebrain regions that do not form neural crest. Surprisingly, we find that Bmp activity is also required for patterning neurons at all DV levels of the CNS. In swirl/bmp2b(-) (swr(-)) embryos, laterally positioned sensory neurons are absent whereas more medial interneuron populations are hugely expanded. However, in somitabun(-) (sbn(-)) embryos, which probably retain higher residual Bmp activity, it is the sensory neurons and not the interneurons that are expanded. Conversely, in severely Bmp depleted embryos, both interneurons and sensory neurons are absent and it is the most medial neurons that are expanded. These results are consistent with there being a gradient of Bmp-dependent positional information extending throughout the entire neural and non-neural ectoderm.

Retinotectal Ligands for the Receptor Tyrosine Phosphatase CRYPα

The cell adhesion molecule-like tyrosine phosphatase CRYPα is localized on retinal axons and their growth cones. We present evidence that two isoforms of this type IIa phosphatase, CRYPα1 and CRYPα2, have extracellular ligands along the developing retinotectal pathway. Using alkaline phosphatase fusion proteins containing the CRYPα1 ectodomain, we detect a prominent ligand on basement membranes of the early retina, optic stalk, and chiasm. A second ligand is observed in the endfeet region of radial processes in the developing stratum opticum, the site of initial retinal axon invasion. This latter ligand binds CRYPα2 preferentially. Further ligand interactions are detected for both CRYPα protein isoforms in retinorecipient tectal laminae and on retinal fibers themselves. CRYPα thus has cell- and matrix-associated ligands along the entire retinotectal projection. Moreover, these ligands appear to be heterotypic and interact with CRYPα through both its immunoglobulin and fibronectin type III regions. The anteroposterior levels of the ligands are relatively uniform within the retina and tectum, suggesting that the CRYPα protein within retinal axons does not directly recognise topographically graded guidance cues. We propose that CRYPα may have a permissive role in promoting retinal axon growth across the eye and tectum and that its functions are modulated temporally and spatially by isoform-specific interactions with cell- and matrix-associated ligands.

A photothrombotic ring stroke model in rats with remarkable morphological tissue recovery in the region at risk.

The photothrombotic ring stroke model with sustained underperfusion followed by late spontaneous reperfusion (Gu et al. 1999) was employed to study its morphological consequences. The exposed crania of adult male Wistar rats were subjected to a ring-shaped laser irradiation beam simultaneously with intravenous erythrosin B infusion. The ischemic volume was calculated from serial sections throughout the ischemic lesions at 4, 10, 24, 48, and 72 h and 7 days and 28 days after irradiation. The ischemic volume, expressed as a percentage of the ipsilateral hemispheric volume, increased steadily from 4 to 10 to 24 h to reach its maximum value at 48 h after irradiation; at 3 days, 7 days, and 28 days, the ischemic volume was reduced to 75%, 24%, and 22% of the value at 48 h. Evaluation of ischemic volumes at different anteroposterior levels revealed that the reduced ischemic volume at 72 h and later was mainly due to morphological restoration in the centrally located, nonirradiated region at risk. An initial enlargement and development of cystic coagulation necrosis was observed in the cortical areas corresponding to the ring lesion itself. In the region at risk, a gradually deteriorating neuropil and nerve cell morphology were observed over time, with maximum severity at 48 h postirradiation. At this time, most laminae II and III neurons in the region at risk exhibited eosinophilia and pyknosis but no incrustations, with small islands of less damaged neurons randomly scattered. At 72 h and up to 28 days after irradiation, these cell characteristics were no longer observed and the region at risk was well populated with neurons that had a chiefly unremarkable cytological appearance. Neuronal counts in the central part of the region at risk were performed; no significant difference in neuronal density was observed between sham-operated controls and at 28 days after irradiation. In conclusion, the consistent, late spontaneous reperfusion coincided with remarkable tissue recovery as assessed morphologically in the region at risk. The data suggest that nerve cell repair may occur even after the detection, by conventional morphological methods, of prolonged critical ischemic neuronal damage in the setting of acute ischemic stroke.