Comparative histological, histochemical and ultrastructural studies of the nephron of selected...

Biologia 68/3: 546—558, 2013Section ZoologyDOI: 10.2478/s11756-013-0181-7

Comparative histological, histochemical and ultrastructural studiesof the nephron of selected snakes from the Egyptian area

Ahmed A. Allam1,2 & Rasha E. Abo-Eleneen2

1King Saud University, College of Science, Zoology Department, P.O. Box 2455, Riyadh 11451, Saudi Arabia;e-mail: [email protected] University, Faculty of Science, Zoology Department, Beni-Suef 65211, Egypt

Abstract: The present study was aimed to compare and contrast the histochemical, histological and ultrastructural vari-ations (microanatomical differences) in the nephrons of selected snake species, Eryx jaculus (Boidae), Psammophis sibilans(Colubridae), Naja haje (Elapidae) and Echis pyramidum (Viperidae) from Egypt. The structural studies were carried outby conventional light and electron microscopy. The nephron, the renal unit of snakes, consists of renal corpuscle, proximaltubule, intermediate segment, distal tubule and collecting tubule. The renal corpuscles have large capillaries with clear anddark fenestrated endothelial cells. The proximal tubule showed long microvilli, cytoplasmic vacuoles, developed endoplasmicreticulum and abundant mitochondria. The intermediate segment was lined by ciliated cells. The lining cells of the distaltubules showed few microvilli, abundant dense mitochondria and clear vesicles of mucous appeared in the terminal por-tion. The collecting tubules consisted of mucous cells. In summary, the ultra-structure studies of nephrons revealed severalinterspecies similarities and also some intra-species differences in species of snakes.

Key words: Eryx jaculus; Psammophis sibilans; Naja haje; Echis pyramidum; kidney

Introduction

The kidney is the main organ of excretion in snakes.The kidney regulates the fluid balance of the body andthe electrolyte in the urine (Gambarian 1994). In recentyears the reptilian kidney has been increasingly stud-ied due its pivotal role in the phylogenetical scale ofthis group of vertebrates (Anderson 1960; Roberts &Schmidt-Nielsen 1966; Davis & Schmidt-Nielsen 1967;Schmidt-Nielsen & Davis 1968; Davis et al. 1976; Gabri1983a, b; Gabri & Butler 1984; Soares & Fava deMoraes 1984; Solomon 1985). The previous reports de-scribed the reptilian nephron as composed of renal cor-puscle, proximal, intermediate and distal tubules andcollecting duct (Bishop 1959; Kahlil et al. 1974a, b;Gabri 1983a). Marked variations have been found inthe ultra structure of the tubular cells which has beenattributed to the enormous variety of species and habi-tats of the animals (Gabri 1983a). The first histologicaldescription of a squamate nephron was made on theEuropean natricine snake Natrix natrix (L., 1758) byGampert (1866).Reptile kidneys have some similarities and also

major differences with mammalian kidneys. Impor-tant anatomical similarities in the tubular system in-clude the renal Bowman’s capsule, proximal convo-luted tubule, distal tubule, collecting duct and ureter.Reptiles lack the loop of the henle and instead havea short intermediate segment. A reptile kidney con-tains a few thousand nephrons whereas a human kid-

ney has about one million nephrons (Peek & McMillian1979a).The anatomy of the renal and urinary system

of both birds and reptiles is markedly different fromthat of mammals (Canny 1998). Whatever type of kid-ney particular vertebrate may have, all kidneys consistof morphologic and functional units called nephrons.Structural modifications of nephrons have enabled ver-tebrates to solve the problems of their environmen-tal conditions whether in fresh or salt water or onland (Gambarian 1978). Carbohydrate content in thenephron plays a significant role in the renal physiologyof mammals (Schimming & Vicentini 2000).The reptilian kidney has been studied by classi-

cal histological and physiological techniques (Gampert1866; Regaud & Policard 1903; Huber 1906; Bishop1959). More recently, several ultrastructural studieson the renal corpuscles have been published (Fernán-dez et al. 1978; Gabri & Butler 1984; Solomon 1985).Reptiles have simple glomeruli whereas others specieslack glomeruli. The glomerular ultrafiltration of rep-tiles does not show the complex capillary network asseen in the mammalian glomeruli (Beyenbach 2004).Many adaptations of reptiles to their environmentalconditions are the production of hypertonic urine andthe ability to secrete uric acid which is the major endmetabolic product of nitrogenous compounds. Theseadaptations indicate that nephrons play an importantrole in coping with the extreme conditions of water de-privation (Dantzler 2005).

c©2013 Institute of Zoology, Slovak Academy of Sciences

Histology and ultrastructure of snake nephron 547

Figs 1A–D. Light micrographs (H&E, ×100) of the kidney illustrating proximal tubule (PT), intermediate segment (arrow head),glomeruli (G), distal tubule (DT) and collecting tubule (CT, arrow). A: E. jaculus; B: P. sibilans; C: N. haje; D: E. pyramidum.

Kidneys play an essential role in fluid ion bal-ance. The mechanisms of renal handling of water varydepending on structural organization of kidneys andthe environment. The excretion of excess water is byforming dilute urine whereas terrestrial tetrapods re-quire water conservation by the kidney for survival(Nishimura & Fan 2003; Dantzler 2003).The present study was aimed to investigate the

histochemical, histological and ultra structural varia-tions of nephron in Eryx jaculus (L., 1758) (Boidae),Psammophis sibilans Boie, 1827 (Colubridae), Najahaje (L., 1758) (Elapidae) and Echis pyramidum (Ge-offroy Saint-Hilaire, 1827) (Viperidae) found in theNile valley, Delta, Faiyum and desert, respectively. Anattempt was made to correlate the nephrons micro-scopic structure and its chemical contents with thebody fluid, water regulation and renal physiology ofthese snakes.

Material and methods

Snake species10 adult specimens of each species were used. The specieswere from different families. The first snake used was thejavelin sand boa E. jaculus (Boidae) found in sandy areasnear cultivated land and collected from Nile Delta, lower val-ley and northern Sinai. The second species was the stripedsand snake (Abu essuyur snake) P. sibilans (Colubridae) asnake of riverine found in agricultural fields usually in gar-dens and cultivated areas. It appears to climb trees occasion-ally and its distribution includes the Nile Valley and Delta.The third is the Egyptian cobra snake N. haje (Elapidae)which inhabits in agricultural fields of the Nile Delta. It ismost frequently encountered on densely vegetated banks ofrivers or irrigation canals and is distributed in the Nile Val-ley, Delta and Faiyum regions. The fourth is Egyptian saw-scaled viper E. pyramidum (Viperidae) is commonly calledthe Haiya ghariba snake that inhabits in sandy desert inthe northern oases of the western desert including western

548 A.A. Allam & R.E. Abo-Eleneen

Figs 1E–H. Higher magnification of the kidney light micrographs (H&E, ×400) show the renal corpuscle formed of parietal epithelium(PE) and visceral epithelium consisting of podocytes (P) and capillaries (CP) lined by endothelial cells (E). E: E. jaculus; F: P. sibilans;G: N. haje; H: E. pyramidum.

area of Faiyum and Cairo. The distribution of the snakesexamined in this study has been earlier described by Saleh(1997).

ChemicalsThe chemicals were purchased from Sigma chemical Com-pany (St Louis, MO, USA). All other chemicals used wereof analytical grade.

Histological and histochemical studiesSnake kidney segments (5 mm in size) were fixed in 10%buffered formalin (pH 7.4) for 24 h. The tissue was dehy-drated in ethyl alcohol followed by two changes of xylene.The tissue was impregnated in paraffin wax and then embed-ded in paraffin wax. Sections (4–5 µm) were cut, de-waxed,hydrated and stained in Mayer’s haemalum solution for 3

min. The sections were stained in eosin for 1 min, washedin tap water and dehydrated in ethanol as described above.Haematoxylin and Eosin stained sections were prepared ac-cording to the method of Mallory (1988). The sections werestained by periodic acid / Schiff’s (PAS) for polysaccha-rides investigations according to McManus (1946). To dif-ferentiate neutral mucopolysaccharides from the acid mu-copolysaccharides, PAS/AB method was employed accord-ing to Mowry (1956). The bromophenol blue was used forproteins (Mazia et al. 1953). The sections were examinedand photographed using Leitz microscope.

Transmission electron microscope (TEM) studiesThe kidneys were cut into small pieces measuring 1 mm3 andimmediately fixed in fresh 3% glutraldehyde-formaldehydeat 4◦C for 18–24 h (pH 7.4) and then post-fixed in isotonic


Figs 2A–D. Light micrograph of the kidney (PAS, ×400). A: Strong amount of PAS positive material in glomerular tuft (G), distal tubule(arrow) and collecting tubule (CT) in E. jaculus. B: Neutral mucosubstances in the glomerulus (G) in P. sibilans. C: Accumulationof PAS positive material in the glomerulus (G), proximal tubule (PT) and collecting tubule (CT) in N. haje. D: Large amounts ofpositive material in the glomerulus (G) and moderate amount in the proximal tubule (arrow head) in E. pyramidum.

1% osmium tetraoxide for one hour at 4◦C. Serial dehydra-tion in alcohol was carried out in the following order: 30min in 50% alcohol, 2 times in 70% alcohol each for 15 min,in 80% alcohol for 15 min, in 90% alcohol for 15 min, then2 times in absolute alcohol each for 30 min. The specimenswere then passed through propylene oxide solutions 2 timeseach for 10 min.

The specimens were embedded in spur resin started bypassing the specimens in propylene oxide then in propyleneoxide-resin mixture at the ratio of 1:1 for 1 hour then inpropylene oxide-resin mixture at the ratio of 1:3 overnight.The samples were left in fresh pure resin at room tempera-ture overnight. In the next day, the specimens were trans-ferred to capsules containing fresh resin and placed in anoven at 60◦C for one day so that polymerization would

harden the blocks. Semithin sections were cut from theseblocks at 1 µm thickness by ultracut Reichert-Jung ultra-microtone with the aid of glass knives, stained with toluidineblue and examined by light microscope. To detect the areaof interest, ultrathin sections were then prepared using theultramicrotome glass knives, stained with uranylacetate andlead citrate and examined with a Joel CX 100 transmissionelectron microscope operated at an accelerating voltage of60 KV.

Results

Histological and histochemical studiesThe nephron of E. jaculus, P. sibilans, N. haje andE. pyramidum consisted of renal corpuscle, proximal


Figs 2E–H. Light micrograph of the kidney (PAS-Alcian, ×400). E: Both acid and neutral mucosubstances in the intermediate segment(IS) and collecting tubule (CT) in E. jaculus. F: Strong amounts of PAS and Alcian blue in the collecting tubule (CT) in P. sibilans.G: Strong amounts of PAS and Alcian blue stained material (mixed polysaccharides) in the collecting tubule (CT) in N. haje. H:Accumulation of mixture of acid and neutral polysacchaides (magenta colour) in the intermediate segment (IS) and collecting tubule(CT) in E. pyramidum.

tubule, intermediate segment, distal and collectingtubules (Figs 1A–H). The renal corpuscle of the presentspecies is varied in its size. In P. sibilans and N. haje,glomerulus is large and urinary space is wider than inE. jaculus and E. pyramidum. The renal corpuscle inthe species appears as dense round tuft of capillariesand formed of parietal epithelium and visceral epithe-

lium consisting of podocytes and capillaries lined byendothelial cells (Figs 1E–H).In the four snake species, the proximal tubule is

lined by simple columnar epithelium. This epitheliumhas abundant clear vesicles and basophilic granules inthe apical cytoplasm. The cell apex exposed to the lu-men of the tubule exhibits a brush border. The upper


Figs 2I–L. Light micrograph of the kidney (Bromophenol blue, ×400). I: Moderate reaction for total protein in E. jaculus. J: Highlevels of protein containing materials in the glomerulus (G), the intermediate segment (IS) and the proximal tubule (PT) in P. sibilans.K: Weak reaction for total protein in N. haje. L: Moderate reaction for total protein in E. pyramidum.

portion of intermediate tubules is lined by columnarcells larger than those observed in the proximal tubulewhile the lower portion is lined by cubodial cells. Allcells of intermediate tubules have a brush border intheir apical poles. The distal tubule is a short segmentwhich shows a wide lumen and is lined by cuboidalepithelium. No brush border is observed in the distaltubule epithelium. The collecting tubule is composedof large columnar cells with basally located nuclei. Theapical pole of the cells is very irregular showing smallprocesses towards the lumen (Figs 1A–D). The large

diameter of the collecting tubule in N. haje (Fig. 1C) isnoted.Histochemically, the renal corpuscle of the four

species shows a similar positive PAS reaction in thebasal lamina and the visceral epithelia (Figs 2A–D).The collecting tubule of E. jaculus (Fig. 2A) and N.haje (Fig. 2C) are strongly PAS-positive while the col-lecting tubule of P. sibilans (Fig 2B) and E. pyramidum(Fig. 2D) are moderately PAS-positive.The application of PAS-Alcian blue revealed a

strong magenta color indicating the presence of neutral


Figs 3A–D. Glomerulus electron micrograph. A: Podacytes (P), urinary space (US), rounded nucleus (N), mitochondria (M), mesangialcells (MS) and lysosomes (L) in E. jaculus (Scale 10 µm). B: Podacytes (P) resting on the basal lamina (arrow). The podocyte give riseto primary processes (PP) which in turn gives numerous secondary foot processes (FS) that rest on the basal lamina, irregular nucleus(N), mitochondria (M), mesangial cells (MS) and lysosomes (L) in P. sibilans (Scale 2 µm). C: Podacytes (P) give rise to primaryprocesses (PP) which give a rise to numerous secondary foot processes (FS) resting on the basal lamina, urinary space (US), irregularnucleus (N), mitochondria (M), mesangial cells (MS) and lysosomes (L) in N. haje (Scale 2 µm). D: Podacytes (P) resting on thebasal lamina (arrow). The podocyte give rise to primary processes (PP) which in turn gives numerous secondary foot processes (FS),urinary space (US), irregular nucleus (N), mitochondria (M), mesangial cells (MS) and lysosomes (L) E. pyramidum (Scale 2 µm).

and acidic mucin in the intermediate segment and in thecollecting tubules of the four species (Figs 2E–H). Bro-mophenol blue staining was used to reveal the proteindistribution in the renal sections of the present snakes.In P. sibilans, kidney revealed intense reaction for to-tal proteins as indicated by the resulting intense bluecolor (Fig. 2J). The kidney of E. jaculus (Fig. 2I) andE. pyramidum (Fig. 2L) showed a moderate amount ofprotein. In N. haje (Fig. 2K), the kidney showed weakprotein stain.

Ultrastructure studiesThe renal corpuscle of the four species was smalland consisted of glomerulus (capillary endothelium,glomerular basement membrane and mesangium) andBowman’s capsule. The glomerulus extended from thepodacyte cell body and the finest branches of one cell.The pedicles were interdigitated with those of an ad-jacent cell. Capillary endothelium was composed of en-dothelial cells with a central nucleus protruding into thecapillary lumen (Figs 3A–D). The cytoplasm contained

the typical organelles and some small rod-shaped bod-ies. In E. jaculus (Fig. 3A), the capillary endotheliumusually presented a reticulated appearance althoughtypical fenestrated region was found in P. sibilans, N.haje and E. pyramidum. The mesangium or central in-tercapillary region of the four species were embeddedin a collagenous matrix and composed of a fine fila-mentous material similar to and continuous with theglomerular basement membrane. The podocytes of P.sibilans, N. haje and E. pyramidum showed irregularnuclei with peripheral aggregations of heterochromatin(Figs 3B–D) while nucleus of podocytes of E. jaculuswas round in shape and heterochromatin appeared frag-mented (Fig. 3A).In all the species, the proximal tubule consisted

of large, clear and dark columnar cells with a well-developed brush border (Figs 4A–D). The cell mem-brane is without basal infoldings. The cell junctionsand some narrow intercellular spaces are also found.The luminal surface of the proximal tubule of E. jac-ulus (Fig. 4A) has long microvilli. The apical aspect


Figs 4A–D. Proximal tubule electron micrograph. A: The cells have tightly packed microvilli (MV), few vacuole (V), lysosomes (L) andmitochondria (M) in E. jaculus (Scale 2 µm). B: Microvilli (MV), numerous mitochondria (M), endoplasmic reticulum (ER), vacuole(V), nucleus (N) and lysosomes (L) in P. sibilans (Scale 2 µm). C: Microvilli (MV), mitochondria (M), vacuole (V), rounded nucleus(N) and lysosomes (L) in N. haje (Scale 10 µm). D: Microvilli (MV), mitochondria (M), vacuole (V), nucleus (N) and lysosomes (L)in E. pyramidum (Scale 10 µm).

of the cytoplasm showed numerous lysosomes and vac-uoles in E. jaculus and P. sibilans (Figs 4A, B) but inN. haje and E. pyramidum (Figs 4C, D) the apical as-pect of the cytoplasm was filled with circular profiles ofendoplasmic reticulum, few vacuoles and lysosomes. Inall species, each cell had a large, oval, centrally locatednucleus. Mitochondria with an electrodense matrix andfew cristae were scattered throughout the cytoplasm.In all species, intermediate segment had a transi-

tional region between the proximal tubule and distaltubule. Three regions of clear and dark ciliated cellswere found. The upper and lower transitional regions ofthe intermediate segment had cells with similar featuresto those of the proximal and distal tubules, respectively.The ciliated cells had an irregular or indented nucleusand few organelles (Figs 5A–D).The apical pole of the lining cells of the distal

tubule showed few microvilli. In E. jaculus and E. pyra-midum (Figs 6A, D), the mitochondria were small,

dense and concentrated at the apical part. Also, therewere few mitochondria scattered in the cytoplasm whilein P. sibilans and N. haje, the cytoplasm showed abun-dant dense mitochondria and occasionally some lyso-somes (Figs 6B, C).The collecting tubules of the four species were con-

sisted of mucous cells. These cells showed a basal nu-clei and interdigitations between the lateral margins ofadjacent cells (Figs 7A–D). The cytoplasm was filledby mucous granules and few mitochondria. In N. haje(Fig. 7C), the mucous cells were more abundant than inother species. Mucous cells were seen in E. pyramidum(Fig. 7D) having a clear granular content while in theothers species (Figs 7A–C) these cells contained densegranular.

Discussion

The results of this study demonstrate that the nephrons


Figs 5A–D. Electron micrograph of the intermediate segment. A: Ciliated cells (C) in E. jaculus (Scale 2 µm). B: Mitochondria (M),lipid droplet (LD) and cilia (C) in P. sibilans (Scale 2 µm). C: Cilia (C), mitochondria (M), endoplasmic reticulum (ER) and nucleus(N) in N. haje (Scale 2 µm). D: Cilia (C), vacuoles (V) and nucleus (N) in E. pyramidum (Scale 2 µm).

of E. jaculus, P. sibilans, N. haje and E. pyramidumhave simple glomeruli, longer distal tubules, wide innermedullary insulation and high medullary cortical ratio.These structural modifications might help in maximiz-ing re-absorption of the glomerular filtrate to preventwater loss in the snake’s dry environment. Collectingtubules are longer in animals adapted to a limited wa-ter supply as compared to animals which have unlimitedaccess to water (Gambarian 1978). These snakes obtainwater from its food plus that produced in its body bycellular respiration and burning of fat. Uric acid ex-cretion allows hyperosmolarity of urine which requirestubular re-absorption for osmoregulation and enhanc-ing concentration ability (Dantzler & Braun 1980).The renal corpuscle varied in size among snake

taxa. In P. sibilans and N. haje, glomerulus is largerand urinary space is wider than in E. jaculus and E.pyramidum. The size of renal corpuscles was directlycorrelated to filtration rate (Davis & Schnerman 1971).Davis et al. (1974) stated that the filtration of the jux-amedullary nephron is significantly higher than the su-perficial nephrons.

The fine structure of the renal corpuscle of thepresent species does not differ greatly from that de-scribed for the lizard Podarcis taurica (Pallas, 1814)(Gabri 1983a). The renal corpuscles are composedof visceral and parietal layers. The epithelial cell(podocytes) of the visceral layer bear trabeculae con-nected to pedicles. The glomerular capsule with its at-tenuated epithelium encloses a tuft of capillaries. Thesefindings are in agreement with Fox (1977) who recordedthat the few and small renal corpuscles of lacertilia arecomposed of a tuft of three or four capillaries (glomeru-lus), Bowman’s capsule and mesangium.In all these species, the glomeruli contain neutral

mucosubstances. The neutral mucosubstances are as-sociated with the glomerular basal membranes to pro-vide structural support and play a major role in surfacetransformation process of the glomeruli (Viotto et al.1988; Bashir 2006).The structural modification and carbohydrate con-

tent of both the glomeruli and renal tubules of the cur-rent species might indicate a role in the nitrogenouswaste excretion and the production of hypertonic urine.


Figs 6A–D. Distal tubule electron micrograph. A: The cells display nuclei located in a basal position, apical microvilli (MV), basalmitochondria (M) and thick basal membrane (BM) in E. jaculus (Scale 2 µm). B: Short microvilli (MV), nucleus (N) and basalmembrane (BM) in P. sibilans (Scale 2 µm). C: Microvilli (MV), nucleus (N) and basal membrane (BM) in N. haje (Scale 2 µm). D:Short microvilli (MV), nucleus (N) and basal membrane (BM) in E. pyramidum (Scale 2 µm).

These might be considered as a mechanism to facili-tate excretion, conserve water and maintain body fluidbalance especially in the extreme conditions of waterdeprivation in the hot habitat of these snakes.Podocytes of E. jaculus, P. sibilans, N. haje and

E. pyramidum are large, have irregular pedicels andfew fenestrae and occasionally having a diaphragm.These characteristics limit the glomerular filtrationrate. These podocytes did not contain the bundlesof microfilaments as described by Peek & McMillan(1979b). The mesangial cells of these species are numer-ous. Similar observations were recorded in other reptil-ian species (Davis et al. 1976; Fernández et al. 1978;Peek & McMillan 1979b; Gabri & Butler 1984; Soares& Fava de Moraes 1984; Galaly 2008).The epithelial cells of the proximal tubule showed a

well developed brush border. Marked protrusions of theapical cytoplasm were observed in some cells (Ander-son 1960; Davis & Schmidt-Nielsen 1967; Gabri 1983a).These protrusions have been related to the secretionof uric acid by the cells and may be important in theexchange of fluids (Gabri 1983a). No segmentation ofthe proximal tubule of the four species has been found

as in the case of other reptiles (Gabri 1983a; Soares& Fava-De Moraes 1984; Solomon 1985). In the basalportion, marked intercellular spaces were observed sim-ilar to those described in other reptiles (Anderson 1960;Roberts & Schmidt-Nielsen 1966; Davis & Schmidt-Nielsen 1967; Schmidt-Nielsen & Davis 1968; Davis etal. 1976; Solomon 1985).In E. jaculus, P. sibilans, N. haje and E. pyra-

midum, the brush border of the proximal tubules hadlong and densely packed microvilli similar to thatfound in Tiliqua scincoides Smith, 1937 (Schmidt-Nielsen & Davis 1968) and Crocodylus acutus Cu-vier, 1807 (Davis & Schmidt-Nielsen 1967). There wasno basal labyrinth as in frogs (Meseguer et al. 1978;Taugner et al. 1982). The mitochondria were scat-tered throughout the cytoplasm as in desert and trop-ical lizards (Roberts & Schmidt-Nielsen 1966; Soares& Fava-De Moraes 1984). The adaptations of cellsengaged in solute-linked water transport. The abun-dant presence of mitochondria was in water andion-transporting cells where mitochondria had a fewcristae and a dense matrix containing dense gran-ules.


Figs 7A–D. Collecting tubule electron micrograph. A: Mucous granules (MU), nucleus (N) with dense scattered patches of heterochro-matin and intercellular canaliculus (IC) in E. jaculus (Scale 2 µm). B: Mucous granules (MU), mitochondria (M), lipid droplets (arrowhead) and nucleus (N) in P. sibilans (Scale 10 µm). C: Mucous granules (MU) fills the apical region of the cells and intercellularcanaliculus (IC) in N. haje (Scale 10 µm). D: Mucous goblet with clear granular content (MU) and nucleus (N) with dense scatteredpatches of heterochromatin in E. pyramidum (Scale 10 µm).

The morphology of the intermediate segment of thecurrent species is similar to Henle’s loop of mammals(Soares & Fava de Moraes 1984) and intermediate seg-ments of other reptiles (Davis et al. 1976; Fernándezet al. 1978). Ciliated cells were frequently observed inthis segment of the nephron of lower vertebrates. Therole of ciliated cells might be related to the low bloodpressure of these animals which produces a low filtra-tion pressure in the kidney. Cilia are known to have apropulsive action of the ultrafiltrate (Davis et al. 1976).In higher vertebrates, where the blood pressure in-creases, no cilia are usually found in the nephron (Mar-shall 1934; Giebrish 1973; Soares & Fava de Moares1984).

The small microvilli observed in the distal tubulein all four species indicated that the absorptive func-tion of this segment is very low (Fernandez et al. 1978).The cytoplasmic features and the apical surface of thedistal tubular cells resembled those seen in other rep-tiles (Davis & Schmidt-Nielsen 1967; Davis et al. 1976).The numerous small apical vesicles are thought to beinvolved in the re-absorption processes (Malnic et al.1966a, b; Giebisch 1971; Danzler & Holmes 1974).The collecting tubule of E. jaculus, P. sibilans, N.

haje and E. pyramidum, is formed by mucous cells.In N. haje mucous cells are as abundant as in somelizards and snakes (Bishop 1959; Gabe 1959; Ander-son 1960; Fernández et al. 1978; Gabri 1983b; Soares


& Fava de Moares 1984) and contain both sialo-andsulpho-mucins. In some turtles, the mucous cells are lo-cated in the distal portion of the intermediate segmentand not in the collecting duct (Solomon 1985).The mucins in the nephron of reptiles have been

implicated in the excretion of non-soluble urates. Theurate crystals are surrounded by mucus which acts as alubricating agent and avoids any injury to the epithe-lium (Minnich 1972; Davis et al. 1976; Peek & McMil-lan 1979b; Gabri 1983b; Solomon 1985). In the fishthe collecting tubules have cells with mucous droplets(Ottosen 1978; Zuasti et al. 1983). The clear and darkcells with mucous goblets observed in all the species ofsnakes. The mucous secretion could produce a luminallayer that either limits water permeability or preventthe precipitation of inorganic salts thus facilitating theexcretion of insoluble urates (Hickman & Trump 1969;Gabri 1983b).In conclusion, the results of this study showed that

the differences in snake nephron microstructure and his-tochemical data may be dependent on their habitat.The observed structural similarities between P. sibilansand N. haje may also be related to their environmentalconditions.

Acknowledgements

This project was supported by King Saud University, Dean-ship of Scientific Research, college of Science Research Cen-ter.

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Received June 13, 2012Accepted November 15, 2012

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