Microvascular anatomy of the esophagus in larval and adult Xenopus laevis Daudin: a scanning electron microscope study of microvascular corrosion casts and correlative light microscopy

Larval to adult microvascular anatomy of the esophagus was studied in the South African Clawed Toad, Xenopus laevis (Daudin) by scanning electron microscopy of vascular corrosion casts and correlative light microscopy of paraplast embedded stained tissue sections. Analyses of vascular corrosion casts of tadpole esophaguses at premetamorphosis revealed a wide‐meshed, but mature looking capillary bed which during following prometamorphosis increased in density and gained the adult‐like pattern during late metamorphic climax by sprouting and nonsprouting angiogenesis. In adult Xenopus, the esophageal mucosa possessed a dense subepithelial capillary bed fed by one or two esophageal arteries that originated from right and/or left thoracic aorta just distal to the origin of the subclavian arteries. In the adult undistended esophagus, esophageal arteries revealed an undulating course, a pattern that guarantees a continuous blood supply when the esophagus is extremely wide expanded as it is the case when adult Xenopus swallows large prey.     

Microvascularization of the spleen in larval and adult Xenopus laevis: Histomorphology and scanning electron microscopy of vascular corrosion casts

Microvascular anatomy and histomorphology of larval and adult spleens of the Clawed Toad, Xenopus laevis were studied by light microscopy of paraplast embedded serial tissue sections and scanning electron microscopy (SEM) of vascular corrosion casts (VCCs). Histology showed i) that white and red pulp are present at the onset of metamorphic climax (stage 57) and ii) that splenic vessels penetrated deeply into the splenic parenchyma at the height of metamorphic climax (stage 64). Scanning electron microscopy of VCCs demonstrated gross arterial supply and venous drainage, splenic microvascular patterns as well as the structure of the interstitial (extravasal) spaces representing the “open circulation routes.” These spaces identified themselves as interconnected resin masses of two distinct forms, namely “broccoli‐shaped” forms and highly interconnected small resin structures. Arterial and venous trees were clearly identified, as were transitions from capillaries to interstitial spaces and from interstitial spaces to pulp venules. Venous sinuses were not diagnosed (nonsinusal spleen). The splenic circulation in Xenopus laevis is “open.” It is hypothesized that red blood cells circulate via splenic artery, central arteries, penicillar arteries, and red pulp capillaries primarily via “broccoli‐shaped” interstitial spaces, pulp venules and veins into subcapsular veins to splenic veins while lymphocytes circulate also via the interstitial spaces represented by the highly interconnected small resin structures in vascular corrosion casts. In physiological terms, the former most likely represent the fast route for blood circulation, while the latter represent the slow route. J. Morphol. 277:1559–1569, 2016. © 2016 Wiley Periodicals, Inc.     

Microvascularization of the thyroid glands in larval and adult Xenopus laevis‐ histomorphology and scanning electron microscopy of vascular corrosion casts

This study demonstrates gross arterial supply, venous drainage and microvascular patterns of larval and adult thyroid glands in the African Clawed Frog, Xenopus laevis by scanning electron microscopy of vascular corrosion casts and light microscopy of stained serial tissue sections. Results confirm published findings gained by microscopical dissections with respect to gross arterial supply. However, in adult frogs one rather than two thyroid veins are found. This study reveals for the first time that bilaterally located thyroid glands in premetamorphosis have immature capillary networks, lack a clear hierarchy of blood vessels, and show many signs of sprouting angiogenesis. During metamorphic climax, blood vessels gain a clear hierarchy and capillaries form closed networks around thyroid follicles. From climax onwards, non‐sprouting angiogenesis (intussusceptive microvascular growth) becomes the prevailing mode of angiogenesis intensifying follicle capillarization. Due to narrow interfollicular spaces, thyroidal arterioles remain superficial while draining venules are located interfollicularly. In contrast with the mammalian thyroid gland where most thyroid follicles have their own capillary beds, most thyroid follicles in Xenopus share their capillary beds with neighbouring follicles. Consequently, the concept of individual morphological and functional angiofollicular units applicable to adult mammalian and human thyroid glands is not applicable to larval and adult amphibian thyroid glands.     

Microvascular anatomy and histomorphology of extrapulmonary bronchi in adult Xenopus laevis Daudin (Lissamphibia; Anura) point to a role in aerial gas exchange – histomorphology of tissue sections and scanning electron microscopy of vascular corrosion casts

Studies on the amphibian respiratory tract microvascular anatomy are few. Using scanning electron microscopy (SEM) of vascular corrosion casts (VCCs) and light microscopy of perfusion‐fixed tissue sections, we studied the bronchial microvascular anatomy in the adult South African Clawed Toad, Xenopus laevis Daudin. Histomorphology showed that the bronchial wall consists (from luminal to abluminal) of squamous epithelium, subepithelial capillary bed, cartilage rings or cartilage plates, a layer of dense connective tissue, a layer of smooth muscle cells, and squamous epithelium (serosa). SEM of VCCs reveals that bilaterally a ventral, a dorsal (Ø 77.21 ± 7.61 μm), and a caudal bronchial artery supply the bronchial subepithelial capillary bed. The ventral bronchial artery has 3–4 branching orders (interbranching distances: 506.3 ± 392.12 μm; branching angles of first‐ and second‐order bifurcations: 24.60 ± 10.24° and 29.59 ± 14.3°). Casts of bronchial arteries display imprints of flow dividers and sphincters. Cranial and caudal bronchial veins (Ø 154.78 ± 49.68 μm) drain into pulmonary veins. They lack microvenous valves. The location of the dense subepithelial capillary meshwork just beneath the thin squamous bronchial epithelium and its drainage into the pulmonary veins make it likely that in Xenopus, bronchi assist in aerial gas exchange.     

Microvasculature of the mouse eye: Scanning electron microscopy of vascular corrosion casts

Purpose

For drug safety assessment, ophthalmic fundus examination is of diagnostic importance in experimental animals. Interim blood samples are usually collected from the orbital venous sinus in the mouse. This report characterizes the angioarchitecture of the mouse eye.

Methods

In 10 mice, the microvasculature of the eyes of was investigated using scanning electron micrographs of corrosion casts.

Results

The mouse eye was characterized as having a rich vasculature with many thick retinal arteries and a well-developed orbital venous sinus. The eye receives its primary blood supply from the external ophthalmic artery, which is divided into three branches: the central retinal artery, as well as the medial and lateral long posterior ciliary arteries. The central retinal artery is divided into 8-9 radiating retinal arteries. The mouse has an orbital venous sinus around the orbit rather than a well-developed orbital venous plexus in the retrobulbar space as is in the rat. The orbital venous sinus is formed by the episcleral veins, the ophthalmic vein, the superior palpebral vein, inferior palpebral vein and numerous anastomotic veins among these veins.

Conclusions

The mouse ocular vasculature is quite similar to that of rats. It is recommended that the best location for insertion of a capillary tube for collecting blood is in the lateral canthus around the eye where the sinus is larger and is most readily accessible. Functional significance of the vascular patterns of the eye is discussed.     

Renal microvasculature in the adult pipid frog,Xenopus laevis: A scanning electron microscope study of vascular corrosion casts

We studied the opisthonephric (mesonephric) kidneys of adult male and female Xenopus laevis using scanning electron microscopy (SEM) of vascular corrosion casts and light microscopy of paraplast embedded tissue sections. Both techniques displayed glomeruli from ventral to mid-dorsal regions of the kidneys with single glomeruli located dorsally close beneath the renal capsule. Glomeruli in general were fed by a single afferent arteriole and drained via a single thinner efferent arteriole into peritubular vessels. Light microscopy and SEM of vascular corrosion casts revealed sphincters at the origins of afferent arterioles, which arose closely, spaced from their parent renal arteries. The second source of renal blood supply via renal portal veins varied interindividually in branching patterns with vessels showing up to five branching orders before they became peritubular vessels. Main trunks and their first- and second-order branches revealed clear longish endothelial cell nuclei imprint patterns oriented parallel to the vessels longitudinal axis, a pattern characteristic for arteries. Peritubular vessels had irregular contours and were never seen as clear cylindrical structures. They ran rather parallel, anastomosed with neighbors and changed into renal venules and veins, which finally emptied into the ventrally located posterior caval vein. A third source of blood supply of the peritubular vessels by straight terminal portions of renal arteries (vasa recta) was not found.     

Microvascular anatomy of the brain of the adult pipid frog,Xenopus laevis (Daudin): A scanning electron microscopic study of vascular corrosion casts

To demonstrate the 3D microvascular anatomy of the brain of the model organism Xenopus laevis Daudin scanning electron microscopy of vascular corrosion casts was correlated with light microscopy of stained 7 µm thick serial tissues sections. Results showed that supplying arteries descended from the leptomeningeal surface without remarkable branchings straight to the subventricular zone where they branched and capillarized. Capillaries showed few H‐ and/or Y‐shaped anastomoses during their centrifugal course toward the leptomeningeal surface where they drained into cerebral venules and veins. Apart from the accessory olfactory bulb and the vestibule‐cochlear nucleus where capillaries were densely packed, capillaries formed a wide‐meshed 3D network throughout the brain parenchyma and thus contrasted to urodelian brains where hairpin‐shaped capillaries descend from the leptomeningeal vessels into varying depths of the brain parenchyma. In about two‐third of specimens, a closed arterial circle of Willis was found at the base of the brain. If this circle in Xenopus might serve the same two functions as in men is briefly discussed. Choroid plexuses of third and fourth ventricle were found to have a high venous, but a low arterial inflow via one small choroidal artery only. Findings are compared with previous studies on the vascularization of the anuran brain and discrepancies in respect to presence or absence of particular arteries and/or veins in Ranids, Bufonids, and Pipids studied so far are discussed with particular emphasis on the techniques used in the various studies published so far.     

Microvascular anatomy of testes in the adult pipid frog,Xenopus laevis Daudin: A scanning electron microscopic study of vascular corrosion casts

The three-dimensional microvascular anatomy of the testes of adult Xenopus laevis was studied by scanning electron microscopy of vascular corrosion casts. Results showed that testicular arteries branch off from urogenital arteries and approach the testes via the mesorchium. At the dorsal surface of testes, arteries course towards the medial and lateral surfaces of the ovoid testes, pierce the tunica albuginea and run in the subalbugineal space and then penetrate the testicular parenchyma. In the parenchyma, they branch in all directions. At sites where larger branches arise, sphincters are present. Intimal cushions are found at the origin of smaller branches. Terminal arterioles capillarize and form a wide-meshed capillary network around convoluted seminiferous tubules. Inter- and peritubular capillaries cannot be differentiated reliably. Arterio-venous transition distances are short. Arterio-arterial anastomoses and veno-venous anastomoses are present. Testicular venules empty into a venous subalbugineal plexus which shows veno-venous anastomoses of different lengths and calibres. Signs of the ongoing non-sprouting angiogenesis, particularly intussusceptive branch remodelling, reflect the dynamics of the testicular parenchyma. The present study is the first that demonstrates the testicular microvascular anatomy in an anuran species and shows that X. laevis and men share the intricate convoluted seminiferous tubules with associated vascular patterns as a feature in common.     

Vascularization of the brains of the Atlantic and Pacific hagfishes, Myxine glutinosa and Eptatretus stouti: a scanning electron microscope study of vascular corrosion casts

The microvascularization of the brains of the hagfishes, Myxine glutinosa L. and Eptatretus stouti, were studied by scanning electron microscopy (SEM) of microvascular corrosion casts. Sections of these casts were used to determine the vascular territories of defined brain areas. Histological serial sections (10 microm) of the brains served for correlation of findings. Analysis of the microvascular casts of both species revealed that the blood supply to and from these brains arose ventrally and dorsally, respectively. Neither species possesses an arterial circle (Circulus Willisi) and both have similar microvascular patterns. The only difference between Myxine and Eptatretus was that the posterior cerebral artery in Myxine divides into mesencephalic and rhombencephalic branches, and in Eptatretus a third branch, termed telencephalic branch, arises from the posterior cerebral artery. 3D-morphometry revealed that luminal diameters of: 1) intracerebral arteries and arterioles range from 35.11 +/- 5.66 microm (mean +/- SEM) in the hypothalamus to 92.69 +/- 14.48 microm in the thalamus; 2) capillaries range from 17.8 +/- 0.44 microm in the olfactory bulb to 21.70 +/- 0.87 microm in the basal ganglia; and 3) intracerebral venules and veins range from 49.38 +/- 4.17 microm in the hypothalamus to 75.58 +/- 6.59 microm in the rhombencephalon. Interbranching distances of arteries and arterioles range from 179.19 +/- 11.32 microm in the optic tectum to 235.19 +/- 94.64 microm in the hypothalamus. Capillaries range from 91.07 +/- 6.22 microm in the hypothalamus to 116.15 +/- 9.45 microm in the thalamus, and venules and veins range from 137.30 +/- 18.11 microm in the hypothalamus to 189.83 +/- 17.47 microm in the optic tectum. Intervascular distances range from 70.58 +/- 3.58 microm in the olfactory bulb to 89.52 +/- 5.74 microm in the optic tectum. Branching angles of arteries and arterioles range from 38.39 +/- 10.9 degrees in the olfactory bulb to 100.73 +/- 9.4 degrees in the optic tectum, and the branching angles of capillaries range from 74.40 +/- 5.42 degrees in the optic tectum to 90.24 +/- 4.66 degrees in the olfactory bulb. Finally, the branching angles of the venules and veins range from 67.84 +/- 6.83 degrees in the tegmentum of the mesencephalon to 92.30 +/- 6.35 degrees in the optic tectum.     

Scanning electron microscopy of human ocular vascular casts: the submacular choriocapillaris

Vascular casts were prepared from ten human orbits removed at autopsy from patients with no previous history of vascular disease. The ages ranged from 7 months to 68 years. The time interval between death and injection of the orbit was always less than 12 h. The vascular casts were examined under scanning electron microscopy with particular attention to the submacular region. We present photomicrographs of common findings seen in each of our specimens, and discuss our results with respect to current models of the submacular angioarchitecture.     

Development of the vascular bed in the rabbit ear: scanning electron microscopy of vascular corrosion casts

The development of the vascular bed in the rabbit ear was investigated using vascular corrosion casts from animals of various ages. Examination of the casts revealed that the arrangement of the major auricular arteries and veins was determined before birth and was maintained during postnatal growth of the ear. Furthermore, the number of arteries branching off the central ear artery and the lateral arteries did not increase with increasing ear length. Scanning electron microscopic examination of lateral segments of adult ear casts revealed many anastomoses between marginal arteries and veins. These arteriovenous anastomoses (AVAs) occurred singly, in pairs, or in clusters of three to six. Their size and shape were variable, even in the same cast. The central segment of many AVA casts showed surface impressions of endothelial cell nuclei which were different from the impressions on adjacent arteries and veins. Arteriovenous anastomoses were also detected in ear casts from animals as young as 8 days. The density of AVAs in lateral ear segments ranged from 95-165 cm-2 in 8- to 11-day-old rabbits to 80-115 cm-2 in adults. However, estimates of the total number of AVAs in the lateral ear margin indicated that AVAs continued to be formed at a steady rate during growth of the ear. During the early neonatal period the cutaneous capillary plexuses developed prominent tufts projecting toward the skin surface, which were apparently associated with developing hair follicles. These capillary tufts were not seen in casts from fetal or adult rabbits.     

The morphology of cultured melanophores from tadpoles of Xenopus laevis: Scanning electron microscopical observations

Summary Tail-fin melanophores of tadpoles of Xenopus laevis (Daudin) in primary culture were examined scanning electron microscopically in the aggregated and in the dispersed state. After isolation, the melanophores are spherical, but within 24 h they develop thin filopodia for attachment to the substratum. Subsequently, cylinder-like as well as flat sheet-like processes are formed, which adhere to the substratum with terminal pseudopodia and filopodia. The processes of adjacent melanophores contact each other, thus forming an interconnecting network between the melanophores.In the aggregated state the central part of the melanophore is spherical and voluminous. Both the central part and the processes bear microvilli. In melanophores with dispersed melanosomes the central part is much flatter; the distal parts have a thickness that equals a monolayer of melanosomes. The surface of the cell bears only scarce microvilli.These features indicate that melanophores do not have a fixed shape and that pigment migration is accompanied by reciprocal volume transformation between the cell body and its processes.     

Sublethal effect of a weak intermittent magnetic field on the development of Xenopus laevis (Daudin) tadpoles

In three repeated experiments with three different litters of Xenopus laevis (Daudin) tadpoles, three cohorts were reared in an aquarium under the "saw-tooth" magnetic field produced by a television set. Their maturation times are compared with those of three corresponding control cohorts grown in an unexposed aquarium. In the exposed aquarium, the magnetic field amplitude was less than 25 T and the frequency in the extremely-low-frequency and very-low-frequency wavebands. Neither the exposed nor the unexposed cohorts suffered significant mortality and malformations. However, the exposed tadpoles took about 5 days more than the unexposed ones to reach metamorphosis. The differences in mean maturation times between the exposed and control cohorts were extremely significant (P < 0.001). The results show that a biological population can suffer a sublethal effect when exposed to the magnetic field of a TV set for a long time in the course of juvenile life stages, and that this effect can consist of a delay in reaching the adult stage.     

Ontogenic emergence and localization of larval skin antigen molecule recognized by adult T cells of Xenopus laevis: Regulation by thyroid hormone during metamorphosis

Results from previous studies using an inbred strain of Xenopus laevis have led to the proposition that metamorphosis includes the events by which the newly differentiating adult immune system, including T lymphocytes, recognizes and eliminates larval skin cells as 'non-self'. More recently, a larval antigen targeted by adult T cells was identified as a 59 kDa protein with a specific peptide sequence. Using antisera directed against the larval antigen and the peptide, immunohistochemistry and western blotting were done to examine expression of the 59 kDa larval antigen in the skin during larval and metamorphic periods. There was no expression before Nieuwkoop and Faber stage 53. Expression was first seen at the beginning of metamorphic stage 54, when hind limbs appear, and increased thereafter, in apical and skein cells of both trunk and tail regions. In the trunk region, expression started to decrease at stage 58, until it completely disappeared at stage 62 (metamorphic climax). In the tail skin, however, expression persisted throughout the metamorphic stages. Treatment of larvae with thyroid hormone (TH) resulted in repression of expression of the 59 kDa molecule in a dose-dependent manner. Downregulation occurred earlier in the trunk than in the tail skin. These results suggest involvement in metamorphic events of an immunological mechanism: differential expression of the larval antigen in the trunk and tail skin cells due to their differing concentration of TH results in the tail, but not the trunk skin, being selectively attacked by the newly differentiating adult-type immune system.     

Nerve-independent DNA synthesis and mitosis in regenerating hindlimbs of larval Xenopus laevis

Summary Xenopus laevis larvae at stage 52–53 (according to Nieuwkoop and Faber 1956) were subjected to amputation of both limbs at the thigh level as well as to repeated denervations of the right limb. Results obtained in larvae sacrificed during wound healing (1 after amputation), blastema formation (3 days) and blastema growth (5 and 7 days) showed that denervated right limbs have undergone the same histological modifications observed in innervated left limbs and have formed a regeneration blastema consisting of mesenchymal cells with a pattern of DNA synthesis and mitosis very similar to that in presence of nerves. Also, the patterns of cellular density in regenerating right and left limbs were very similar. On the whole, the data here reported show a highly remarkable degree of nerve-independence for regeneration in hindlimbs of larval Xenopus laevis at stage 52–53 and lend some substance to the hypothesis that, in early limbs, there would exist trophic factors capable of replacing those released by nerves, promoting DNA synthesis and mitosis in blastemal cells. Offprint requests to: S. Filoni     

The angioarchitecture of estrous, pseudopregnant and pregnant rabbit ovary as seen by scanning electron microscopy of vascular corrosion casts

Ovarian angioarchitecture was studied by scanning electron microscopy of vascular corrosion casts in estrous, pseudopregnant (stimulated with human chorionic gonadotropin) and pregnant rabbits. In all samples, the proper ovarian branch of the ovarian artery (ramus ovaricus) entered the ovarian hilus near the caudal pole of the organ and ran parallel to the major axis of the hilus. The extraovarian venous drainage was formed by several vessels emptying into a distal large vein. The ramus ovaricus exhibited various degrees of coiling and branched in the medulla. The coiling of the ramus ovaricus and its ramifications were maintained in all samples. A venous meshwork and/or flat vein branches closely enveloped the arterial coils found in the hilus and outer medulla. At this level numerous arteriovenous contacts were demonstrated in all samples. The coiled arteries, prior to entering the ovarian cortex, supplied several small peripheral follicles which were drained by the hilar veins. In the cortex the coiled arteries branched in numerous thin, straight or slightly undulated arterioles which supplied developing estrous follicles and pseudopregnant corpora lutea. The arterioles supplying the pregnant corpora lutea were long, large and tightly spiraled. The venous drainage followed the modifications of the arterial supply. These data demonstrate that ovarian cycle and pregnancy induced significant changes in the cortical vessels, which adapted their structure to the temporary functional needs of the recruited follicles or corpora lutea. Hilar and medullary vessels have permanent structures that may represent morphological devices for (a) a continuous control of the blood flow (spiral arteries) and (b) a local recirculation of endocrine products (arteriovenous contacts) comparable to the ”countercurrent mechanism” previously shown to operate in ovaries of other species, but not yet found in rabbits. Received: 19 June 1996 / Accepted: 7 October 1996