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Accurate analysis of the three-dimensional (3D) architecture of developing organs is critical to understanding how developmental defects can be linked with structural abnormalities. Here, we describe a 3D reconstruction technique of the developing kidney including the outer k ...

Organ development requires the coordination of proliferation and differentiation of various cell types. This is particularly challenging in the kidney, where up to 26 different cell types with highly specialized functions are present. Moreover, even though the nephron initially d ...

Design-based stereology is considered the gold-standard method for estimating the total number of glomeruli, and thereby nephrons, in the adult kidney. However, until recently, a design-based method for estimating nephron number in the developing kidney was not available. For such a me ...

The zebrafish has emerged as a powerful model to study organ development and regeneration. It has a number of advantages over other vertebrate model systems. The embryo can be kept transparent throughout embryonic development, which allows direct visualization of the developing orga ...

Organotypic culture is an invaluable technique that allows researchers with the tool to analyze a tissue development in an isolated and well-defined environment. This technique also permits one to study the roles of different signaling systems/signaling molecules and to take advan ...

X-gal staining is a rapid and convenient histochemical technique used to detect reporter gene expression. A prerequisite is the creation or acquisition of transgenic reporter mouse lines, in which the bacterial LacZ gene has been knocked into the gene of interest or placed under the control of ...

This chapter provides a basic protocol to perform fluorescent immunolabeling on embryonic kidney samples. The procedure can be summarized in five steps: permeabilization, primary antibody incubation, washes, secondary antibody incubation, and final washes. This protocol can ...

At the onset of mammalian kidney development, the ureteric bud invades the surrounding metanephric mesenchyme, and genetic studies in the mouse have shown that BMP pathway activity has to be antagonized in the vicinity of the epithelium, a task performed by the secreted BMP antagonist Grem1. H ...

Signaling through the ERK/MAPK pathway within the Wolffian duct and ureteric bud epithelium is critical for kidney induction and branching morphogenesis. ERK signaling is activated by receptor tyrosine kinase such as RET and FGFR2. This protocol describes a method to detect the diphos ...

Chromatin immunoprecipitation (ChIP) is a powerful technique for examining transcription factor recruitment to chromatin, or histone modifications, at the level of specific genomic sequences. As such, it provides an invaluable tool for elucidating gene regulation at the molec ...

Studies of the complex responses of the kidney to acute injury have yielded important insights into mechanisms of tissue injury and repair. A variety of injury models have contributed to this impressive body of knowledge, but the ischemia–reperfusion (IR) model has perhaps been the most wide ...

Congenital Anomalies of the Kidney and Urinary Tract (CAKUT) encompass a spectrum of kidney and urinary tract disorders. Here, we describe two assays that can be used to determine if a mouse has vesicoureteric reflux (VUR) or urinary tract obstruction, two urinary tract defects observed in CAK ...

Nephron number has emerged as a useful parameter for assessing the roles of specific genes and feto-maternal environmental factors in kidney development. Nephron number is also of clinical interest due to increasing evidence suggesting that low nephron number is associated with incr ...

Renal gene therapy may offer new strategies to treat diseases of native and transplanted kidneys. Several experimental techniques have been developed using viral, nonviral, and cellular vectors, although the effectiveness of such techniques varies widely depending upon the vect ...

During embryonic kidney development, the ureteric bud (UB) undergoes repetitive branching to generate the entire renal collecting system. Defects in UB branching result in renal malformations, from hypoplastic kidneys to renal agenesis. Mouse genetics has become an invaluable t ...

We examined the role of semaphorin3a in ureteric bud (UB) branching morphogenesis using mouse metanephric organ culture . In vitro UB injection of Sema3a antisense morpholino resulted in increased branching morphogenesis. Cellular and tissue uptake of oligonucleotides was faci ...

Obstructive nephropathy is the most important cause of renal failure in children. Unilateral ureteral obstruction (UUO) in the neonatal mouse provides a useful model to investigate the response of the developing kidney to urine flow obstruction. Creation of reversible variable par ...

Ultrasound (US) is the most common and least invasive modality for clinical imaging of the kidney. One important application of US in nephrology is the detection and monitoring of structural changes in the kidney. Recent advances in US technology have facilitated the application of similar ...

Lower urinary tract obstruction in mice can lead to end-stage renal disease and death. We have developed a surgical technique to create a cutaneous vesicostomy in mice providing an external outlet for drainage of urine, thereby relieving the obstruction and slowing and/or preventing the de ...

Mitochondrial dysfunction and genomic instability are associated with a number of retinal pathologies including age-related macular degeneration, diabetic retinopathy, and glaucoma. Consequences of mitochondrial dysfunction within cells include elevation of the ra ...