Search

The kidneys in two viviparous species of Neotropical lizards, Notomabuya frenata and Aspronema dorsivittatum (Mabuyidae), were investigated by light and scanning electron microscopy in order to determine the presence of the sexual segment of the kidney (SSK) and to study its morphology. The individuals used in this study belong to the Herpetological Collection of the Herpetology Laboratory - Reptiles of the Federal University of Juiz de Fora (CHUFJF-Reptiles) and they were collected between the years 2008 and 2012 from the Cerrado region in the state of Minas Gerais, Brazil. The SSK was present only in sexually mature males (with sperm in the testes / epididymis), whereas it was absent in sexually immature males. The nephron in both species consists of renal corpuscle, proximal convoluted tubule, distal convoluted tubule, collecting duct and sexual segment of the kidney. The SSK of the analyzed species were coated with a simple columnar epithelium, with high cells, basal nucleus and in the apical portion innumerable secretory granules. This study adds to the knowledge on reproductive biology and structures related to reproductive strategies of both lizard species and viviparous Neotropical lizards.

The ultrastructure of renal tubule cells was studied in the European lesser spotted dogfish by the evaluation of thin sections and freeze fracture replicas. Computer-assisted three-dimensional reconstruction of entire nephrons was performed. The distinction of nephron segments and collecting tubule was made using results of previous histological work. The first proximal tubule segment (PI) consists of two subsequent portions, PIa and PIb. PIa is a component of the lateral countercurrent bundle, and PIb, which displays an apical tubulovesicular apparatus and an extended lysosomal compartment, is located in the vicinity of the glomeruli. Rod-shaped intramembrane particles were detected in PIa. The second proximal tubule segment (PII) is a special segment in elasmobranch and teleost fish. PII differs largely from PI in cell morphology and function. The apical cytoplasm was filled with small clear vesicles, and an apical endocytic apparatus was lacking. In the apical cell membrane, rod-shaped particles were revealed by freeze fracture. The apical tight junctions of PI and PII consisted of seven to ten meandering strands. The distal nephron was subdivided into two major segments: early distal tubule (EDT) in the lateral countercurrent bundles and late distal tubule (LDT) in the mesial tissue. The EDT showed marked amplification of basolateral cell membranes. The tight junctions displayed a low number of continuous parallel strands, which is also characteristically found in the diluting segments of other vertebrates. LDT cells showed cytoplasmic studs and rod-shaped intramembraneous particles at the apical cell membrane, thereby resembling type A intercalated cells of collecting duct. The collecting tubule (CT) emerged from the LDT and was part of the countercurrent arrangement inside the lateral bundles. Tight junctions of LDT and CT consisted of many meandering strands in a honeycomb pattern. With immunohistochemistry, binding sites of a polyclonal antibody against an extraplasmic portion of rat gastric H(+)-K(+)-adenosine triphosphatase (ATPase) were observed at the apical cell membrane of PIa, PII, and LDT. From the colocalization of binding sites for the antibody against the transport enzyme with rod-shaped intramembrane particles, we assume that these might be the morphological correlate of gastric H(+)-K(+)-ATPase-like enzyme in the renal tubule.

We modified and improved enzyme digestion and density gradient separation procedures to obtain fractions of proximal and distal renal tubules with high yield and viability. Kidneys from two anesthetized adult Wistar rats were flushed with Krebs-Henseleit buffer (KHB) and then perfused in situ with recirculated KHB containing collagenase and hyaluronidase at 125 mmHg. Cortices were excised, minced, and incubated in KHB containing enzymes for 35 min at 37 degrees C. Dissociated tubules were removed at 10-min intervals, rinsed, and placed in KHB containing 10% calf serum, vitamins, and amino acids at 4 degrees C. Separation was achieved by suspending the tissue in 45% isosmotic Percoll layered over an undiluted Percoll cushion and centrifuging. Proximal tubules sedimented near the cushion. Distal segments were isolated in the uppermost bands of a second 35% Percoll separation. Viability was greater than 95% as measured by lactate dehydrogenase leakage and quantitated by oxygen consumption and ATP content. Basal oxygen consumption was greater than 33 nmol O2 X min-1 X mg protein-1 in all fractions and was stimulated by succinate and inhibited by amiloride and ouabain. Basal ATP content averaged 9.7 nmol/mg ATP. An average 3.3-fold separation for the proximal fraction and 24.5-fold separation for the distal fraction was assessed by the enrichment of six specific enzyme markers, with several of the markers indicating separations up to 32-fold. Isolated tubules also displayed functional responses to parathyroid hormone and vasopressin. Distal, but not proximal, segments demonstrated significantly increased adenosine 3',5'-cyclic monophosphate formation with vasopressin.