Origin and development of the pronephros in the chick embryo

Abstract

The process by which the pronephros develops was morphologically examined in chick embryos from Hamburger-Hamilton stage (ST) 8+ to ST34. The intermediate mesoderm, from which the pronephros arises, was first seen as a faint ridge of undifferentiated mesoderm between the segmental plate and lateral plate at ST8+. It formed a cell cord at the level of the 6th to the presumptive 13th somites at ST9 to ST10. This cell cord then separated into dorsal and ventral parts, the former becoming the nephric duct and the latter the tubules by ST14. The primordia of the external glomeruli (PEGs) appeared at ST15 through some epithelial cells protruding in the nephrostome (the opening of the nephric tubule into the body cavity). PEGs formed gradually in the caudal direction until ST18, while the pronephric tubules and PEGs in cranial locations disappeared. At this stage, only a few PEGs remained at the level of the 13th and 14th somites and these developed from ST23 to ST29 to become ultrastructurally similar to the glomeruli of the functional kidney. From these observations in the avian pronephros, we infer that the pronephric duct and tubules both form from a cell cord in the intermediate mesoderm and at the same time, but later develop differently.

Fig. 1

Number of EGs (or PEGs) appearing in the body cavity on each side in chick embryos from ST15 to ST34. PEGs were considered to possess epithelial cells without foot processes (▪), EGs to possess epithelial cells (podocytes) with foot processes (□). The latter appear at ST21, whereas the former are poorly developed and disappear at an early stage.

Fig. 2

LM images of whole-mount preparations (A,B), SEM images of dorsal surfaces of chick embryos from which the ectoderm had been removed (E–I) and LM images of semithin Epon sections (C,D). (A) ST8+ (five somites). Arrowheads: an initial intermediate mesoderm from the level of the presumptive 6th somite (PS6) to about PS9. (B) ST9 (seven somites). Arrowheads: condensed region extending from the level of the 6th somite (S6) to about PS10. (C) Cross-section through PS7 at ST8+ (five somites). Arrowheads: initial intermediate mesoderm. (D) Cross-section through PS8 at ST9 (seven somites). The intermediate mesoderm is elevated slightly toward the ectoderm (arrowheads). (E) ST9+ (eight somites). The intermediate mesoderm is seen from a level slightly cranial to S6 to about the level of PS12 (enclosed by the broken white line). (F) Enlargement of the rectangle in E. Flat, polygonal cells with many filopodia connect loosely with each other. (G) ST10 (ten somites). A cell cord of intermediate mesoderm is arranged from the S6 level to PS14 (arrows). (H) Enlargement of the rectangle in G. The cell cord consists of cells lengthened in the cranio-caudal direction. (I) ST12+ (18 somites). The cranial part of the cell cord at the level cranial to S14 is covered by mesenchymal cells. The caudal part is lengthened caudally to a distance equivalent to about five somites beyond the last somite (arrow). Scale bars in A–E, G and I = 50 µm; F and H = 10 µm.

Fig. 3

LM images of Epon cross-sections (A–E, H) and longitudinal sections (F,G) of chick embryos. (A) ST9+ (eight somites) through the segmental plate at the PS9 level. The intermediate mesoderm is elevated slightly in the dorsal direction (arrowhead). (B–D) ST11 (13 somites), through S10, S13 and the segmental plate at the PS15 level, respectively. The cell arrangement in the intermediate mesoderm differs between the dorsal and ventral parts. (E) ST12 (17 somites), through S14. In the cell mass of the intermediate mesoderm, the dorsal part is a tubule-like structure (arrow) and the ventral part is arranged in two epithelial-like layers (arrowhead). (F) ST13– (19 somites), at the S9 to S11 level. Small cell cords (asterisks) are seen between the primordium of the nephric duct and the peritoneal epithelium. (G) ST13– (19 somites), at the S11 to S13 level. The cell cord (asterisk) connects obliquely between the peritoneal epithelium and the primordium of the nephric duct (arrowheads). (H) ST13 (20 somites), through S14. The nephric tubule joining the nephric duct opens into the body cavity (arrow). Scale bar = 50 µm.

Fig. 4

SEM images (A,B) and LM image of Epon cross-section (C) of chick embryos. (A) ST13 (21 somites). Nephrostomes are recognized on the dorsal surface of the body cavity as small depressions (arrowheads) or depressions lengthened cranio-caudally (arrows). (B) ST15 (27 somites). A protrusion of cells is seen in the large nephrostome (arrow) at the S14 level but not in the small nephrostome (arrowhead). (C) ST15 (27 somites), through S14. The epithelium of the large nephrostome protrudes into the body cavity (arrow). The small cell cord is the primordium of the nephric tubule. Scale bar in A = 50 µm; B and C = 10 µm.

Fig. 5

SEM image (A) and LM images from Epon serial sections (B–D) of chick embryos at ST17. The longitudinal sections (B–D) are cut through the pronephros at almost the same level as A. (A) Ventral surface at the S13 to S16 level. PEGs and nephrostomes (arrowheads) are seen along the dorsal mesentery. (B) Section through the most caudal PEG, at the S14 level. A nephrostome of the mesonephric tubule opens into the body cavity (arrowhead). (C) Section 13 µm lateral to B. The PEG in B is seen continuously in the pronephric tubule (arrow). Caudal to it, a dilated mesonephric tubule with a cell mass inside it (arrowhead) is seen. (D) Section 56 µm lateral to C. The boundary between a slender pronephric duct and a thick mesonephric duct is present at a level between S14 and S15. Scale bar = 50 µm.

Fig. 6

SEM image (A), LM image in semithin Epon section (B) and TEM image (C,D) of PEGs in chick embryos at ST18. (A) A large PEG at the S12 level consists of hemispherical cells with small microvilli. (B) Cross-section at same level as A. The PEG (enclosed by rectangle) is adjacent to the dorsal aorta. A pronephric tubule extends from the nephrostome to the pronephric duct. (C) Enlargement of the area enclosed by the rectangle in B. The PEG consists of epithelial cells with irregular processes and a primordium of a blood vessel (arrowheads). Apoptotic bodies are seen in both epithelium and endothelium in the PEG. (D) Enlargement of the area enclosed by the rectangle in C. Epithelial cells connect tightly with each other via junctional complexes. Scale bar in A and B = 10 µm; C = 2 µm.

Fig. 7

SEM images (A–C), LM and TEM images (D,E) of a PEG, and an SEM image of a vascular cast (E) in chick embryos at ST21. (A) Two PEGs at the S10 and S11 levels differ in size. (B) Enlargement of the area enclosed by the upper rectangle in A. The small PEG consists of hemispherical cells and is not morphologically different from that seen at ST15 (Fig. 4B). (C) Enlargement of the area enclosed by the lower rectangle in A. The large PEG consists of spindle-shaped cells with long primary (white arrows) and short secondary (black-on-white arrows) processes. (D) Vascular tufts are developing in the large PEG. A branch from the dorsal aorta invades into the PEG. (E) High magnification of the area indicated by an arrow in D. Gap and slit membranes are recognized among the processes of the epithelial cells (arrows). A thin basement membrane is present beneath the epithelium. (F) Right-side view of the dorsal aorta and its branches at the S13 to S15 level. White arrows indicate the vascular tufts of the pronephric PEGs, whereas black-on-white arrows indicate those of the mesonephric PIGs. Scale bar in A–D = 10 µm; F = 50 µm; E = 200 nm.

Fig. 8

SEM images (A,B), LM image (C), SEM images of a vascular cast (D,E) and TEM image (F) of a well-developed EG in chick embryos. (A) A large EG at the S15 level at ST26 is a conjugate of two or three EGs and is partly embedded in the peritoneal wall. (B) Enlargement of part of A. Podocytes with well-development foot processes interdigitate with each other on the EGs. (C) A longitudinal section of an EG similar to that in A. The EG and IG are supplied by a common capillary. (D) Left-side view of the dorsal aorta at the level of the 13–15th segmental arteries at ST24. The capillary loops of EGs in the pronephros are seen at the level of the 13–15th segmental arteries (sa13–sa15), while IGs in the mesonephros arise caudal to them. (E) Enlargement of part of D. The vascular tuft consists of a large EG and a small IG. (F) High magnification of the large EG. The wall of the EG consists of the foot processes of podocytes together with endothelial cells and a basement membrane between the two. A slit membrane between foot processes (arrow) and the thin diaphragm of an endothelial pore (arrowhead) are recognized. Scale bars in A and C–E = 50 µm; B = 5 µm; F = 200 nm.

Fig. 9

LM image (A), TEM image (B) and SEM images (C,D) of a degenerating EG. (A) A relatively large EG in the body cavity at ST28. The blood vessel is filled with blood corpuscles. (B) Enlargement of part of A. The wall of the EG consists of mesenchymal cells and extracellular matrix. (C) An EG in the body cavity at ST34. The surface of the EG is covered with hemispherical cells. (D) Enlargement of part of C. The foot processes of the epithelial cells (podocytes) are comparatively short and rounded (arrows). Scale bars in A = 50 µm; B and C = 10 µm; D = 1 µm.

Fig. 10

Schematic colour diagram showing the process of development of the pronephros in chick embryos from ST8+ to ST34. The cream-coloured area shows the region of the mesonephros (which was omitted except for its most cranial portion).