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Morphological characterization of the seminiferous cycle in the goat (Capra hircus): a histological and ultrastructural study.

The cycle of spermatogenesis/seminiferous cycle was investigated in the goat testis using both light and electron microscopy techniques. Using the various cell associations and the accompanying changes in spermatid shape and location, the cycle was divided into eight (8) successive stages. The cycle began with the accomplishment of spermiation (stage 1) and ended with apical migration and close attachment of late maturation phase spermatids at the Sertoli cell apex accompanied by adluminal retention of residual bodies with dense staining inclusions (stage 8). The early stages of the cycle (stages 1-4) were therefore characterized by the presence of only one generation of spermatids, the second one appearing only after the division of secondary spermatocytes in stage 4. Consequently, stages 5-8 had two generations of spermatids; Golgi or cap phase as well as maturation phase spermatids. Although stages 5 to 7 appeared as distinct entities, stages 6 and 7 were rather short-lived and considered as continuations of stage 5. Therefore, the 8 stages of the cycle in the goat were further condensed into 6 main divisions. The duration of each stage was estimated by the frequency of occurrence in sections. Among these, stage 1 had the highest frequency (34%) followed by stages 5-7 (27%). Stages 8 and 4 had the shortest frequency (up to 9%) while stages 2 and 3 had 13% and 12% respectively. These results indicate that, like most domestic species, goats have a cycle of 8 stages with 6 main divisions, the longest being stage 1.

Acrosome↗

Morphological characterization of the female prostate (Skene's gland or paraurethral gland) of Lagostomus maximus maximus.

The Skene's (paraurethral) gland is the histologic homologue to the male prostate. Much experimental work has been done on women and on various rodents. In this study we describe for the first time the anatomical and histological characteristics of the paraurethral gland in the plain viscacha (Lagostomus maximus maximus). This gland is formed by tubuloalveolar adenomers surrounded by connective tissue and smooth muscle fibers. The adenomers are lined by cuboidal or columnar epithelium. In the lumen of the adenomers, PAS positive secretions could be detected. The conduits are lined by bi-stratified epithelium. In some aspects this gland is similar to that of other mammals.

Animals↗

The lysosomes of earthworm chloragocytes: biochemical and morphological characterization.

Nine latent and sedimentable acid hydrolases have been detected in the homogenates of earthworm chloragocytes. Their full activity was revealed by treatment with Triton X-100, a Waring blender treatment, freezing and thawing, hypotonic media or incubation at pH 5 and 25 degrees C. Solubilization paralleled the activation of the enzymes. Together with kinetic studies, these results indicate that the acid hydrolases of the chloragocytes are inside typical lysosome-like particles whose membrane is impermeable to their substrates. It could be shown by density equilibration centrifugation that the lysosomes of those cells constitute a heterogeneous population of subcellular particles distinct from the chloragosomes. Moreover, their digestive function has been directly demonstrated by the capture and degradation of serum albumin. The lysosomes of the chloragocytes have been clearly identified as polyvesicular bodies by electron microscopic analysis of the fractions obtained by density equilibration centrifugation and by examination of the whole tissue, as such or after endocytosis of serum albumin or ferritin. Finally, our results do not support a possible relationship between the lysosomes and the chloragosomes of the chloragocytes.

Animals↗

[Hands and feet of Simian primates: an attempt at morphological characterization].

The carpal height, the length of the metacarpus, the metatarsus, and of all the phalanges of 269 simians were measured, then for each specimen, converted to percentage taking the 3rd metacarpus as the reference unit. They were then compared despite differences between specimens. The average values were determined for the different species, subspecies or families. Mathematical ratios were established: height of the carpus divided by the 3rd metacarpus, carpus divided by the 1st metacarpus, 1st ray of the hand divided by the 2nd, 1st ray of the hand divided by the 1st ray of the foot. CONCERNING THE HAND: For little monkeys, the length of the carpus and the medial metacarpus were similar to those observed in humans. Colobinae exhibited a relatively short thumb column. The length of the thumb was slightly shorter for the platyrrhinii and cercopithecinae, especially macacus. Some hapalidae had a relatively longer thumb than humans. All of their medial fingers were longer than those observed in humans. The big monkeys had a smaller carpus and thumb column, sometimes much smaller than in humans, with a similar length for the metacarpi and the medial fingers, except for gibbons, whose medial fingers were longer. CONCERNING THE FOOT: The 1st ray in humans is nearly as long as the 2nd toe, the other toes being very short. The 1st ray of the hallux of all monkeys was always longer than the ray of the thumb, but shorter than the hallux of humans. The other metatarsi of little monkeys were similar to those in humans, except for the platyrrhinii which had a shorter 2nd metatarsus. The lateral toes, which were much longer than in humans, were very similar to fingers though slightly longer, except for platyrrhinii with much longer toes. The big monkeys had very short metatarsi and slightly smaller lateral toes than humans.

Animals↗

Morphological characterization of skeletal cells in Cbfa1-deficient mice.

To clarify the mechanisms by which core-binding factor-alpha1 (Cbfa1), an essential transcription factor in osteogenesis, functions in osteoblast matrix formation, as well as in chondrocyte differentiation and osteoclastic bone resorption, Cbfa1-deficient embryonic mice were investigated ultrastructurally and histocytochemically at 18.5 days postcoitum. In homozygotic mice, both endochondral and intramembranous ossification were arrested, although bone tissue had already formed at this stage in the wild type. The tibiae of homozygotic mice were characterized by calcified cartilage and alkaline phosphatase (ALP)-positive perichondrium, whereas membranous structures indicating the presence of ALP activity in the lateral portion were observed in the calvariae, rather than the bone tissue. Most of the ALP-positive perichondrial cells in homozygotic tibiae possessed a spindle-shaped cell contour and small cytoplasm, the extracellular matrix of which contained neither type I collagen nor calcifying matrix vesicles. In contrast, some perichondrial cells at the very middle part of tibiae became flattened. In the vicinity of these cells, a thin layer of type I collagen-based calcified matrix, containing osteopontin, bone sialoprotein, or osteocalcin, was observed. In the cartilage of mutant mice, we observed a hypoplasic zone of proliferative chondrocytes, the flattening of hypertrophic chondrocyte-like cells, and calcified chondrocytes which, while not degraded, did display a high level of cell function. Mononuclear osteoclastic cells were found in the perichondrium, near calcified chondrocytes, in mutant mice. Multinuclear osteoclasts possessing H+-ATPase and ruffled borders were also present, although only in limited numbers. Neither the development of ruffled borders nor intracellular polarization was complete. Because the majority of osteogenic cells in Cbfa1-deficient mice can neither form nor calcify the bone matrix, Cbfa1 principally plays essential roles in osteoblastic differentiation and bone matrix formation. Cbfa1 also affects both the proliferation and the differentiation of chondrocytes, whereas its absence prevents normal osteoclast formation and related functions.

Animals↗

Morphological characterization of the retinal degeneration in three strains of mice carrying the rd-3 mutation.

Retinal development in 3 strains of rd-3/rd-3 mutant mice, previously shown to have different rates of degeneration, was studied using light, electron, and immunofluorescence microscopy. The time course and phenotype of the degeneration as well as details on the mechanism of massive photoreceptor cell loss are compared with other known retinal degenerations in mice. Up until postnatal day (P) 10, the retinas of all three strains (RBF, 4Bnr, In-30) develop similarly to those of pigmented and nonpigmented controls. TUNEL-positive cells appear in the outer nuclear layer (ONL) by P14, and reach a maximum in all three mutant strains around P21. Scattered rods and cones form a loose, monolayered ONL by 8 weeks in the albino RBF strain, by 10 weeks in the albino 4Bnr strain, and by 16 weeks in the pigmented In-30 strain. Though the initial degeneration begins in the central retina, there is no preferred gradient of cell death between central and peripheral photoreceptors. Rods and cones are present at all ages examined. During development, stacks of outer segments (OS) form in all three strains though they never achieve full adult lengths, and often have disorganized, atypical OS. Rod opsin is expressed in the developing OS but is redistributed into plasma membrane as OS degeneration proceeds. Retinal pigment epithelial (RPE) cells of all mutant strains contain packets of phagocytosed OS, and their apical processes associate with the distal ends of the OS. At their synaptic sites, photoreceptor terminals contain ribbons apposed to apparently normal postsynaptic triads. As photoreceptors are lost, Müller cells fill in space in the ONL but they do not appear to undergo significant hypertrophy or migration, though during the degeneration, glial fibrillary acidic protein (GFAP) expression is gradually upregulated. Macrophage-like cells are found frequently in the subretinal space after the onset of photoreceptor apoptosis. As OS disappear, the RPE apical processes revert to simple microvilli. Late in the degeneration, some RPE cells die and neighboring cells appear to flatten as if to maintain confluence. In regions of RPE cell loss that happen to lie above retina where the ONL is gone, cells of the inner nuclear layer (INL), wrapped by Müller cell processes, may front directly on Bruch's membrane.

Aging↗

Biodegradable cellulose diacetate-graft-poly(L-lactide)s: enzymatic hydrolysis behavior and surface morphological characterization.

Enzymatic hydrolysis of selected copolymers of cellulose diacetate-graft-poly(L-lactide)s (CDA-g-PLLAs) were conducted with proteinase K for film specimens, which were solely quenched from the molten state or, further, annealed at temperatures below or above their glass transition temperatures. The hydrolysis rates depended seriously on the thermal history, as well as on the graft modification. Especially, the heat treatment, followed by physical aging or crystallization of the originally amorphous materials, was a key factor to control subtly their enzymatic degradation behavior. Atomic force microscopy revealed that the enzymatic hydrolysis transformed the surface of the respective films into a more undulated one with a number of fine protuberances, for example, of several hundred nanometers in height and a few micrometers in width. Attenuated total reflection FTIR spectroscopy ensured selective release of lactyl units from the surface region. In visual appearance, some degraded films exhibited even an iridescent color due to an effect of interference of visible light reflected on the surface. These observations suggest a conception of "spatiotemporally controlled degradation", leading to a new method not only for regulation of the overall rate of degradation but also for fine surface abrasion of polymer materials.

Cellulose↗

Encapsulation of drug reservoirs in fibers by emulsion electrospinning: morphology characterization and preliminary release assessment.

In this paper, we prepared composite fibers via electrospinning from either W/O or O/W emulsion. SEM images demonstrate the beads-in-string structures in these fibers and proved this technique to be an effective method for microencapsulation. As a practical application, Ca-alginate microspheres, which serve as reservoirs for hydrophilic drugs, were prepared in a reverse emulsion and then incorporated into poly (l-lactic acid) (PLLA) fibers by electrospinning. With the bovine serum albumin (BSA) loaded into the microspheres, the beads-in-string structure thus entrapped hydrophilic proteins in hydrophobic polymeric matrix. In the in vitro release test, BSA, which was released from composite fibers, achieved prolonged release profiles and lower burst release rates than those from naked Ca-alginate microspheres. In comparison with other well-established techniques to prepare microcapsules, such as solvent evaporation and spray-drying techniques, emulsion electrospinning features partly competing, partly complementary characteristics. Extension to other emulsion systems will be able to fabricate new types of functional structures.

Alginates↗

Morphological characterization of in-vitro human hair keratinolysis, produced by identified wild strains of Chrysosporium species.

Chrysosporium species were isolated from soil and keratinized material. Primary isolation was performed following the general method of hair baiting on modified Czapek-agar media with washed, defated and sterilized human hair fragments added. Strains were maintained in test tubes of potato dextrose agar at 29 degrees C and cultivated on phytone yeast extract agar at 28 degrees C for 14 days for identification. Isolates were characterized using Van Oorschot's key. Keratinolytic activity was expressed following a subjective scale representing degree/severity of attack upon hair surface and presence of fungal structures observed in substrate. Culture results and characterization methods were effective for soil Chrysosporium strain isolation. A new hair attack mode is described. Of 71 keratinolytic fungal isolates, eight (12%) Chrysosporium species were identified. One keratinolytic Chrysosporium sp. isolate is yet to be identified.

Chrysosporium↗

Transgenic rats carrying the mouse renin gene--morphological characterization of a low-renin hypertension model.

Transgenic rats [TGR; strain name TGR(mRen2)27] harboring the mouse Ren-2 renin gene have been recently generated as a model for the study of primary hypertension that offers the advantage of a clearly-defined genetic alteration. Expression of the mouse Ren-2 gene causes severe hypertension (200 to 260 mm Hg) which is responsive to converting enzyme inhibitors. Compared to control transgene-negative littermates, plasma renin and angiotensin II values are lowered in TGR, whereas plasma prorenin values are strongly elevated. The adrenal gland in TGR shows marked overexpression of mouse renin messenger RNA; in situ hybridization using a 35S-labelled mouse-renin RNA probe reveals that enhanced renin expression is mainly localized to cells of the zona glomerulosa and outer zona fasciculata. Immunohistochemically, renin protein in the TGR adrenal gland is stored in larger quantities than in controls. Adrenal transgene expression probably accounts for most of the elevated plasma prorenin level in TGR, since bilateral adrenalectomy (ADX) causes a significant decrease in prorenin level (318 +/- 79 ng angiotensin I/ml/hr before ADX to 70 +/- 43 ng 4 days after ADX, P less than 0.0005). In the kidney, renin synthesis is almost completely suppressed in TGR. In situ hybridization demonstrates that few juxtaglomerular afferent arterioles express renin. Immunohistochemically, the TGR kidney shows significantly reduced renin and angiotensin II immunoreactivity at the afferent arteriole. Ultrastructural analysis of the afferent arteriolar wall frequently shows the complete absence of renin secretory granules since the granular cells are mostly converted into smooth muscle cells. Beginning at an age of approximately four to six months, TGR develop hypertension-related alterations and pathological lesions in various tissues. In the kidney, the wall thickness of arterioles and arteries is strongly increased, and glomerular lesions including different stages of sclerosis are observed. The thoracic aorta displays a considerable increase in tunica media thickness due to both myocyte hypertrophy and interstitial fibrosis. Coronary arteries and arterioles of the heart are thickened and perivascular fibrosis is observed. The data show that TGR(mRen2)27 transgenic rats display all typical characteristics of hypertensive pathology, making them an interesting model for therapeutic interventions. The fact that these changes occur in animals with a single gene difference to normotensive rats makes them a particularly suitable model for studies on gene-related hypertensive processes.

Adrenal Glands↗

Isolation of pig thyroid lysosomes. Biochemical and morphological characterization.

Open thyroid follicles were prepared by mechanical disruption of pig thyroid fragments through a metal sieve. This procedure allowed preparation of thyroid-cell material depleted of colloid thyroglobulin. Open thyroid follicles were used to prepared a crude particulate fraction, which contained lysosomes, mitochondria and endoplasmic reticulum. These organelles were subfractionated by isopycnic centrifugation on iso-osmotic Percoll gradients. A lysosomal peak was identified by its content of acid hydrolases: acid phosphatase, cathepsin D, beta-galactosidase and beta-glucuronidase. The lysosomal peak was well separated from mitochondria and endoplasmic reticulum. The lysosomal peak, from which Percoll was removed by centrifugation, was taken as the purified lysosome fraction (L). Lysosomes of fraction L were purified 45-55-fold (as compared with the homogenate) and contained about 5% of the total thyroid acid hydrolase activities. Electron microscopy showed that fraction L was composed of an approx. 90% pure population of lysosomes, with an average diameter of 220 nm. Acid hydrolase activities were almost completely (80-90%) released by an osmotic-pressure-dependent lysis. Thyroglobulin was identified by polyacrylamide-gel electrophoresis as a soluble component of the lysosome fraction. In conclusion, a 50-fold purification of pig thyroid lysosomes was achieved by using a new tissue-disruption procedure and isopycnic centrifugation on Percoll gradient. The presence of thyroglobulin indicates that the lysosome population is probably composed of primary and secondary lysosomes. Isolated thyroid lysosomes should serve as an interesting model to study the reactions whereby thyroid hormones are generated from thyroglobulin and released into the thyroid cells.

Acid Phosphatase↗

Primary cell culture and morphological characterization of canine dermal papilla cells and dermal fibroblasts.

Skin biopsies were taken from female dogs, the primary hair follicles isolated and the dermal papilla dissected. After incubation in supplemented Amniomax complete C100 medium in 24-well culture plates, the dermal papilla cells (DPC) grew to confluence within 3 weeks. Thereafter, they were subcultivated every 7 days. Dermal fibroblast (DFB) cultures were established by explant culture of interfollicular dermis in serum-free medium, where they reached confluence in 10 days. They were subcultivated every 5 days. For immunohistochemistry, cells were grown on cover slips for 24 h, fixed and stained with antibodies against collagen IV and laminin. DPC showed an aggregative growth pattern and formation of pseudopapillae. Intensive staining for collagen IV and laminin could be observed until the sixth passage. DFB grew as branching, parallel lines and showed only weak staining for collagen IV and laminin.

Animals↗

Biochemical and morphological characterization of spontaneously hypertensive hyperlipidaemic rats.

1. To develop and characterize a new animal model of hypertension and hyperlipidaemia, we cross-bred spontaneously hypertensive rats (SHR) with spontaneously hyperlipidaemic rats (HLR). 2. A new strain of spontaneously hypertensive hyperlipidaemic rats (SHHR) was established at generation 10 through selective mating of brothers and sisters (systolic blood pressure > 150 mmHg, plasma cholesterol > 150 mg/dL). Cross-bred Wistar-Kyoto (WKY) rats and Sprague-Dawley (SD) rats (SDWKY rats) were used as a control. 3. Adrenaline and noradrenaline levels in the plasma and adrenal medulla of male SHHR were significantly higher than those of male SDWKY rats. The tyrosine hydroxylase activity in the adrenal medulla of male SHHR was significantly higher than that of male SDWKY rats. Low-density lipoprotein expression was found in the plasma of male and female SHHR and HLR. Cholesterol 7alpha-hydroxylase mRNA expression in the liver of male SHHR was lower than that of male SDWKY rats. Endothelium lesions and lipid deposition under the endothelium were observed in the aorta of 24-month-old SHHR, especially female SHHR, but not in age-matched HLR and SDWKY rats. 4. The hypertension of this new animal model of hypertension and hyperlipidaemia may be related to increased catecholamine activity and the hyperlipidaemia may be related to changes in the expression of cholesterol 7alpha-hydroxylase mRNA and lipoprotein profiles. The SHHR may be valuable in the study of mechanisms of atherosclerosis and the evaluation of anti-atherosclerosis drugs as a new strain of cardiovascular disease.

Animals↗

[Diagnosis of pulmonary embolism by transthoracic sonography. Sono-morphologic characterization of pulmonary lesions and comparison with spiral computed tomography].

BACKGROUND AND OBJECTIVE: Pulmonary embolism often extends to pleural surfaces, resulting in pleural-based parenchymal lesions. This study was carried out in order to evaluate the clinical value of transthoracic sonography (TS) for the diagnosis of pulmonary embolism. PATIENTS AND METHODS: A prospective study was performed in 45 patients (27 men, 18 women; mean age: 63.4 years; range: 24-88 years) with suspected pulmonary embolism. Transthoracic sonography was performed in all patients. In addition, helical computed tomography (CT) was carried out in 40 patients. Other diagnostic procedures included the estimation of D-dimeres, echocardiography, venous duplex sonography of the legs and ventilation/perfusion scanning. The diagnosis of pulmonary embolism was accepted when an embolus was detected on CT or as a conclusive result of the other investigations. RESULTS: Pulmonary embolism was established in 33 patients. Peripheral parenchymal lesions suspicious of pulmonary embolism were detected by transthoracic sonography in 26 patients (79%). In seven patients with central pulmonary embolism diagnosed by CT, no peripheral lesions could be detected by sonography. One patient with sonographic signs of pulmonary embolism had a diffuse bronchogenic adenocarcinoma which was diagnosed on autopsy. The sensitivity of transthoracic sonography (computed tomography) for pulmonary embolism was 79% (76%) and the specificity 92% (100%). The positive and negative predictive values of transthoracic sonography for the detection of pulmonary embolism were 96% and 61% (100% and 61%), respectively. The accuracy was 82% (82%). CONCLUSIONS: A substantial number of pulmonary emboli involve the peripheral lung areas. Transthoracic sonography is a non-invasive technique for diagnosing such parenchymal alterations and may serve as an alternative method in the diagnosis of pulmonary embolism.

Adenocarcinoma↗

Morphologic characterization of the pathway of transferrin endocytosis and recycling in human KB cells.

The pathway of transferrin uptake and recycling was investigated in KB cells to attempt to identify the organelles involved in the return of transferrin to the cell surface. Comparison was made with the pathway of internalization of epidermal growth factor (EGF), which has been shown to terminate in lysosomes. A horseradish peroxidase conjugate of transferrin (TF-HRP) was incubated with KB and at 4 degrees C and, at various times after warming to 37 degrees C, the location of TF-HRP was examined at the electron microscopic level. Transferrin, like EGF, was found to enter cells via coated pits and to move to the Golgi region in receptosomes (endosomes). Transferrin was located in the tubular elements of the transreticular portion of the Golgi but not in Golgi stacks. Interestingly, transferrin was not concentrated in the coated pits of the Golgi. In contrast, EGF was highly concentrated there. Transferrin was next detected in tubular elements (approximately equal to 600 A in width and up to 5,000 A in length) that were closely associated with microtubules and in dumbbell-shaped structures. In contrast, EGF was not detected in these structures. These results suggest that those organelles containing transferrin, but not EGF, participate in the return of receptor-bound transferrin to the cell surface.

Biological Transport↗

Studies on the mechanism of urotoxic effects of N,N'-dimethylaminopropionitrile in rats and mice. 1. Biochemical and morphologic characterization of the injury and its relationship to metabolism.

N,N'-Dimethylaminopropionitrile (DMAPN), a major component of the NIAX catalyst ESN, is known to cause urinary bladder dysfunction in exposed workers. In order to investigate the mechanism of DMAPN toxicity, we carried out time-course (0-72 h) and dose-response (175-700 mg/kg) studies on the effects of DMAPN in rats and mice. Treated animals exhibited several signs of toxicity including loss of body weight, reduced water consumption, and bladder urine retention, as well as bladder injury. DMAPN-induced bladder injury was characterized by distended bladders with marked diffuse submucosal and subserosal edema, petechial hemorrhage, and multifocal perivascular inflammatory infiltrates. The qualitative and quantitative analysis of urine indicated hypoosmolality, aciduria, hematuria, proteinuria, and oliguria. Elevated levels of creatinine and urea levels in plasma were indicative of renal dysfunction. Within hours following DMAPN administration, the animals exhibited a significant increase in urinary retention that resolved between 60 and 72 h. Rats excreted about 44% of the administered DMAPN dose unchanged in the urine, while mice excreted only about 6% of the dose. Commercially available DMAPN metabolites, administered by gavage, produced toxic effects less adverse than DMAPN. The biochemical effects of DMAPN included depletion of glutathione and increased lipid peroxidation in target organs, including urinary bladder and kidney. These studies indicate that there are species differences in DMAPN toxicity. The differences may be due to differences in the formation of reactive metabolic intermediates of DMAPN.

Aminopropionitrile↗

Transcellular transport of polymeric IgA in the rat hepatocyte: biochemical and morphological characterization of the transport pathway.

Polymeric IgA (pIgA) is transported by liver parenchymal cells (hepatocytes) from blood to bile via a receptor-mediated process. We have studied the intracellular pathway taken by a TEPC15 mouse myeloma pIgA. When from 1 microgram to 1 mg 125I-pIgA was injected into the saphenous vein of a rat, 36% was transported as intact protein into the bile over a 3-h period. The concentration of transported 125I-pIgA was maximal in bile 30-60 min after injection, and approximately 80% of the total 125I-pIgA ultimately transported had been secreted into bile by 90 min. A horseradish peroxidase-pIgA conjugate (125I-pIgA-HRP) was transported to a similar extent and with kinetics similar to that of unconjugated 125I-pIgA and was therefore used to visualize the transport pathway. Peroxidase cytochemistry of livers fixed in situ 2.5 to 10 min after 125I-pIgA-HRP injection demonstrated a progressive redistribution of labeled structures from the sinusoidal area to intermediate and bile canalicular regions of the hepatocyte cytoplasm. Although conjugate-containing structures began accumulating in the bile canalicular region at these early times, no conjugate was present in bile until 20 min. From 7.5 to 45 min after injection approximately 30% of the labeled structures were in regions that contained Golgi complexes and lysosomes; however, we found no evidence that either organelle contained 125I-pIgA-HRP. At least 85% of all positive structures in the hepatocyte were vesicles of 110-160-nm median diameters, with the remaining structures accounted for by tubules and multivesicular bodies. Vesicles in the bile canalicular region tended to be larger than those in the sinusoidal region. Serial sectioning showed that the 125I-pIgA-HRP-containing structures were relatively simple (predominantly vesicular) and that extensive interconnections did not exist between structures in the sinusoidal and bile canalicular regions.

Animals↗