Histomorphological, Histochemical and Ultrastructural Studies on the Healthy Liver of Yemen Veiled Chameleon (Chamaeleo calyptratus) in Southern Saudi Arabia / Estudios Histomorfológicos, Histoquímicos y Ultraestructurales en el Hígado Sano del Camaleón Velado de Yemen (Chamaeleo calyptratus) en el Sur de Arabia Saudita

SUMMARY: The livers of reptiles are being studied as a model for the link between the environment and hepatic tissue. There have been few investigations on the histology of reptile livers, and very few or no studies have examined the histology of liver of veiled chameleon (Chamaeleo calyptratus). This paper describes the histomorphological, histochemical and ultrastructural characterization of the liver of veiled chameleons in southern Saudi Arabia. Seven Chamaeleo calyptratus were captured in the summer season in Abha City, Aseer region, southern Saudi Arabia. Chamaeleon liver samples were processed for histomorphology, histochemistry and ultrastructure analyses. Morphologically liver of Chamaeleo calyptratus was observed as a large dark brown organ with lighter speckles, which represent melanin deposits. It located at the ventral part of abdominal cavity forward of the stomach. Its dimensions approximately were 3.7 x 2 cm. The liver was a bilobed organ divided into two lobes, right and left lobes. The right one was bigger than the others. The gallbladder was well developed and had an elongated shape, situated between the two lobes and contained the bile for the digestion. Microscopically, the liver was found to be covered by a thick layer of connective tissue, which formed the hepatic capsule. Hepatic parenchyma probably appeared in cross sections as hepatic glandular-like alveoli "acini" or follicular structures with various diameters, each acinus contains approximately four to six hepatocytes, surrounded by sinusoidal capillaries filled with abundant melanomacrophages, which are absent in birds and mammals. Melanomacrophages are common in the hepatic parenchyma's perisinusoidal areas, particularly near portal spaces. Hepatocytes are polyhedral or pyramidal with and mostly contained large, rounded nuclei mostly peripherally located, with prominent dark oval nucleoli. Some of nuclei are eccentric or central position. The cytoplasm appeared spongy or vacuolated and more eosinophilic when stained by hematoxylin-eosin and strongly reactive to PAS staining technique, indicating abundant glycogen content. The reticular fibers that surround hepatocytes, blood arteries, and sinusoids supported the hepatic parenchyma. The blood sinusoids are seen interspersed among hepatocytes of varying sizes. The sinusoidal lumen was bordered by flattened endothelial cells and includes elliptical nucleated erythrocytes and liver macrophages as phagocytes, which are also known as Kupffer cells. Branches of the portal vein, hepatic artery, small bile duct, and lymph vessels were detected in the hepatic portal area "tract" or triad which made up of connective. Hematopoietic tissue was observed in subcapsular region and portal triads. Ultrastructurally, the hepatocyte appeared polyhedric containing a single large rounded basal or eccentric vesicular nucleus with prominent nucleolus. Extensive network of rough endoplasmic reticulum (RER) often arranged in an array parallel to the nuclear membrane with many mitochondria, and Golgi apparatus were described. The cytoplasm contained glycogen granules, vesicles or vacuoles scattered throughout the cytoplasm especially at the apical region were reported. The bile canaliculi and the hepatic "Kupffer" cells were also discussed. This is the first study on the histological characterization of the healthy liver of Yemen veiled chameleon in southern Saudi Arabia. The findings reported here should be used as a reference to compare with the pathological abnormalities of the liver in this animal.

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A morphological, histological and histochemical study of the sexual segment of the kidney of the male Chamaeleo calyptratus (veiled chameleon) / Un estudio morfológico, histológico e histoquímico del segmento sexual del riñón del macho Chamaeleo calyptratus (camaleón velado)

SUMMARY: This study aimed to describe the morphology, histology, and histochemistry of the kidneys and the sexual segment of the kidney (SSK) in Chamaeleo calyptratus (Yemen chameleons or veiled chameleons) collected during the active reproductive season. To achieve this objective, a total of 7 mature male Chamaeleo calyptratus were captured from their natural habitats from different areas in the Aseer region of Saudi Arabia and transferred to the reptilian laboratory where their kidneys were dissected out. Next, a morphological examination was conducted on the kidneys and the SSK was processed for light microscopic examinations with the primary focus placed on the morphology and histology of the SSK. The results of the study showed that Chamaeleo calyptratus possess two, reddish-brown, attenuated kidneys divided into lobes and lobules (lobulated). The histological study showed that the chameleons had no distinct limits between the cortex and medulla of the kidneys. In general, similar with other reptiles, this lizard demonstrated that the kidneys have few nephrons, do not show any nephron loop (loop of Henle), and display few glomeruli. The nephron is generally composed of standard components: renal corpuscle, glomerulus surrounded by a double-walled, the glomerular capsule (Bowman's capsule) and renal tubules (six renal tubular segments differentiated as the ciliated neck segment [between the glomerulus and the proximal segment], proximal segment, ciliated intermediate segment instead of nephron loop that connects proximal and distal, distal segment, terminal segment or sexual segment, and all segments emptying into the collecting ducts. The epithelial tissues lining the lumen of these segments vary from simple cuboidal cells to columnar and tall columnar cells in the SSK. Mature male Chamaeleo calyptratus possess hypertrophied parts of the distal renal segments, which are called the SSK. Histologically, the active SSK consists of simple tall columnar epithelial cells with rounded basal nuclei with clear centralized nucleoli and numerous apical coarse secretory granules which are released from the cells by an apocrine process. Histochemically, the secretory granules stain positively for proteins with mercuric bromophenol blue and react intensely to periodic acid-Schiff (PAS) for neutral carbohydrates. This is the first report about the histology of the SSK of the Chamaeleo calyptratus, and this study adds to the knowledge of the reproductive biology, biodiversity, and reproductive strategies of chameleons. Further studies are required to investigate the ultrastructure of the kidneys and SSK.

RESUMEN: Este estudio tuvo como objetivo describir la morfología, histología e histoquímica de los riñones y el segmento sexual en el riñón (SSK) del Chamaeleo calyptratus (camaleones de Yemen o camaleones con velo) recolectados durante la temporada reproductiva. Un total de 7 machos maduros de Chamaeleo calyptratus fueron capturados en su hábitat natural de diferentes áreas en la región de Aseer de Arabia Saudita y transferidos al laboratorio de reptiles donde fueron disecados los riñones. Posteriormente, se llevó a cabo un examen morfológico de los riñones y se procesó la SSK para exámenes con microscopio óptico con el enfoque principal puesto en la morfología y la histología. Los resultados del estudio mostraron que Chamaeleo calyptratus posee dos riñones atenuados de color marrón rojizo divididos en lóbulos (lobulados). En el estudio histológico se observó que los riñones de los camaleones no tenían límites definidos entre la corteza y la médula. En general, y similar a otros reptiles, este lagarto demostró que los riñones tienen pocas nefronas, no muestran ningún asa de Henle y presentan pocos glomérulos. La nefrona está compuesta por componentes estándar: corpúsculo renal (glomérulo rodeado por una pared de doble pared). Cápsula de Bowman y túbulos renales (seis segmentos tubulares renales diferenciados como el segmento del cuello ciliado [entre el glomérulo y el segmento proximal], segmento proximal, segmento intermedio ciliado en lugar del asa de Henle que conecta proximal y distal, segmento distal, segmento terminal o segmento sexual y todos los segmentos que desembocan en los conductos colectores Los tejidos epiteliales que recubren la luz de estos segmentos varían desde células cuboidales simples hasta células columnares y columnares altas en la SSK. Los machos maduros de Chamaeleo calyptratus poseen partes hipertrofiadas de los segmentos renales distales, denominados SSK. Histológicamente, la SSK activa consiste en células epiteliales cilíndricas altas simples con núcleos basales redondeados, nucléolos centralizados claros y gránulos secretores gruesos apicales que se liberan de las células por un proceso apocrino. Histoquímicamente, los gránulos secretores se tiñen positivamente para las proteínas con azul de bromofenol mercúrico y reaccionan intensamente al ácido periódico-Schiff (PAS) para obtener carbohidratos neutros. Este es el primer informe sobre la histología de la SSK de Chamaeleo calyptratus, y se suma al conocimiento de la biología reproductiva, la biodiversidad y las estrategias reproductivas de los camaleones. Se requieren más estudios para investigar la ultraestructura de los riñones y la SSK.

Fecal adrenal hormone patterns during ovulatory and non-ovulatory reproductive cycles in female veiled chameleons (Chamaeleo calyptratus).

The relationship between the reproductive (hypothalamic-pituitary-gonadal; HPG) and adrenal (hypothalamic-pituitary-adrenal; HPA) hormone axes is complex and can vary depending on the species and environmental factors affecting an individual. In an effort to understand this relationship in female veiled chameleons (Chamaeleo calyptratus), the patterns of fecal metabolites of corticosterone (C), estradiol (E), testosterone (T), and progesterone (P) were analyzed by enzyme immunoassay (EIA) during ovulatory (OC; eggs laid) and non-ovulatory cycles (NOC; no eggs laid). Glucocorticoid (GC) metabolites in the fecal extracts were characterized by HPLC and corticosterone EIA performance was assessed by parallelism, accuracy and precision tests. The results indicated that the assay chosen reliably measured the hormone metabolites present in the fecal extracts. Regular, cyclical hormone metabolite patterns consisting of an E peak followed by peaks of T, P and C in close succession were observed during both ovulatory and non-ovulatory cycles; relative levels of P and C, however, were higher during ovulatory cycles. Corticosterone metabolite levels, in particular, increased throughout vitellogenesis and peaked in late vitellogenesis (in non-ovulatory cycles) or around the time of ovulation, and remained elevated throughout the gravid period, falling just prior to oviposition. The results provide evidence of variation in glucocorticoid production throughout different stages of the reproductive cycle, including a role in the ovulatory process; the physiology, however, remains unclear.

Developmental plasticity in reptiles: Insights into thermal and maternal effects on chameleon phenotypes.

Embryonic environments affect a range of phenotypic traits including sex and reproductive success. I determined (1) how the interaction between incubation temperature and egg size affects sex allocation of Chamaeleo calyptratus and (2) how incubation temperature and maternal parent (clutch) affect water uptake by eggs and body size, growth, and climbing speed of hatchlings and juveniles. Eggs from five clutches were exposed to five temperature treatments with clutches replicated within and among treatments. Temperature affected sex, but only when egg size was included as a factor in analyses. At intermediate (28°C) temperatures, daughters were more likely to be produced from large eggs and sons more likely to be produced from small eggs, while at 25 and 30°C, the pattern of sex allocation was reversed. Temperature and clutch affected water uptake and body size. Nonetheless, the direction of temperature and clutch effects on water uptake by eggs and on the size of hatchlings were not the same and the direction of temperature effects on body sizes of hatchlings and juveniles differed as well. Clutch affected hatchling size but not juvenile size and growth rate. Clutch, but not incubation temperature, affected climbing speed, but the fastest hatchlings were not from the same clutches as the fastest juveniles. The independent effects of incubation temperature and clutch indicate that hatchling phenotypes are influenced largely by conditions experienced during incubation, while juvenile phenotypes are influenced largely by conditions experienced in the rearing environment.

Assessment of the reproductive status of female veiled chameleons (Chamaeleo calyptratus) using hormonal, behavioural and physical traits.

Egg binding is a common reproductive disorder in captive female reptiles leading to premature loss of breeding potential, or in severe cases death. It can result from failure to ovulate (and reabsorb) follicles; follicular stasis, or failure to lay eggs; dystocia. Reproductive status of female veiled chameleons (Chamaeleo calyptratus) in a research colony was assessed using enzyme immunoassay (EIA) of fecal reproductive hormones (estradiol; E2, progesterone; P, and testosterone; T) and their metabolites, ultrasound imaging of the reproductive tract, and receptivity to conspecific males. Periods of follicular growth (vitellogenesis) corresponded with increasing levels of E2, and following ovulation, a distinct change in morphology from round (follicles) to oval (eggs) structures, which was accompanied by a surge in P (>20-fold above baseline). P levels remained elevated throughout the gravid phase until just prior to oviposition. Length/width ratios of follicles and eggs were statistically different, but distinguishing a follicle from an egg based on the ratio was unreliable due to a large overlap in values. In animals that failed to ovulate on their first cycle, follicles began to recede but were not fully reabsorbed and could be distinguished from a second batch of follicles based on their echogenicity. Female receptivity to conspecific males was not related to cycle stage (i.e., previtellogenesis, vitellogenesis, or gravid) or reproductive hormone levels. This study demonstrates the use of ultrasonography and reproductive hormone analysis to assess phase of the reproductive cycle (pre- or post-ovulatory), or confirm ongoing follicular stasis.

Chameleons communicate with complex colour changes during contests: different body regions convey different information.

Many animals display static coloration (e.g. of feathers or fur) that can serve as a reliable sexual or social signal, but the communication function of rapidly changing colours (as in chameleons and cephalopods) is poorly understood. We used recently developed photographic and mathematical modelling tools to examine how rapid colour changes of veiled chameleons Chamaeleo calyptratus predict aggressive behaviour during male-male competitions. Males that achieved brighter stripe coloration were more likely to approach their opponent, and those that attained brighter head coloration were more likely to win fights; speed of head colour change was also an important predictor of contest outcome. This correlative study represents the first quantification of rapid colour change using organism-specific visual models and provides evidence that the rate of colour change, in addition to maximum display coloration, can be an important component of communication. Interestingly, the body and head locations of the relevant colour signals map onto the behavioural displays given during specific contest stages, with lateral displays from a distance followed by directed, head-on approaches prior to combat, suggesting that different colour change signals may evolve to communicate different information (motivation and fighting ability, respectively).