4 “WW“ A HISTOLOGJC STUEW OF THE BRWARY SYSTEM OF TEE NEWBORN BEAGLE DOG Fhesés for ihe fiegrea a? M. 3.. MEGHSSLAN STATE UAWERSSTY mecx G. S?EERS mm I :1 c1515 A HISTOLOGIC STUDY OF THE URINARY SYSTEM OF THE NEWBORN BEAGLE DOG By Frederick G. Speers A THESIS Submitted to Michigan State University in partial fulfillment of the requirements for the degree of MASTER OF SCIENCE Department of Anatomy 1967 645707 I ..r fl _.‘ a} ’3' 1. ACKNOWLEDGMENTS The author wishes to express his sincere appreciation to his major professor, Dr. Al W. Stinson, for his guidance, patience, and advice throughout the course of this investigation. The author wishes to thank Drs. Esther M. Smith, Daris R. Swindler, and Horace C. Hartsell for serving on the guidance committee and giving construc— tive criticism of the manuscript. Special thanks are extended to Dr. M. L. Calhoun, Chairman of Anatomy, for the use of departmental facili- ties. The author is indebted to Dr. Gabel H. Conner, project director of U.S.P.H. Research Contract Number PH 43-65-100 Leukemia Transmission Studies, and Dr. John A. Moore, director of research animals facilities of the preceding project, who made available the dogs used for this study. Special gratitude is extended to Drs. William Adam, Rexford E. Carrow, and James B. Thomas for giving con- structive criticism of this manuscript. ii TABLE OF CONTENTS INTRODUCTION REVIEW OF THE LITERATURE MATERIALS AND METHODS RESULTS AND DISCUSSION Kidney Capsule Cortex Neogenic Zone Ureterogenic Collecting Tube Metanephric Blastemal Cap Renal Vesicle . "Lipped Body" S—shaped Body Maturation Zone Renal Corpuscle . Proximal Convoluted Tubule Loop of Henle . . Distal Convoluted Tubule Macula Densa Juxtaglomerular Apparatus Medulla . . Collecting Tubules Arched and Straight Collecting Ducts Papillary Duct . . . . . . Renal Pelvis Ureter Tunica Mucosa Epithelium Lamina Propria— —Submucosa Tunica Muscularis Tunica Adventitia Urinary Bladder Tunica Mucosa Epithelium Lamina Propria- -Submucosa Tunica Muscularis Tunica Serosa iii Page [—1 HWKOCDCDCDNN \1 U'I +4FJH romam FJFJHFJFJHFAFJHFJ cnonnxnuwcntxrun» FJHFJFJHFJ arqsqxrq—q HFJFH4FJH \owaamnann Male Urethra Prostatic Portion Tunica Mucosa Epithelium Lamina Propria— —Submucosa Membranous Portion Tunica Mucosa . . Epithelium . . . Lamina Propria- -Submucosa Penile Portion Tunica Mucosa Epithelium Lamina Propria— —Submucosa Female Urethra Tunica Mucosa Epithelium Lamina PrOpria- -Submucosa SUMMARY LITERATURE CITED FIGURES iv Figure 10. 11. 12. 13. 14. 15. LIST OF FIGURES Cortex and medulla of the kidney Brown fat within the kidney capsule . . Kidney capsule Ectopic glomeruli within the kidney capsule . . . . . . Metanephric blastemal cap in the neogenic zone of the kidney . . . . . Early renal vesicle in the neogenic zone of the kidney . . . . . . . Late renal vesicle in the neogenic zone of the kidney . . . . . . . . . . "Lipped body" in the neogenic zone of the kidney . . . . . . . . . "Lipped body" in the neogenic zone of the kidney . . . . . . . . . S—shaped body in the neogenic zone of the kidney . . . . . . . . S-shaped body in the neogenic zone of the kidney. State I in the development of the glomerulus . . . . . . . . . . The sanguineous lacuna. Stage II in the development of the glomerulus . Stage III in the development of the glomerulus . . . . . . . . . Stage IV in the development of the glomerulus . . . . . Stage V in the development of the glomerulus . . . . . Page 32 3A 36 38 NO 42 AA 46 A8 50 52 54 56 58 60 Figure 16. 17. -18. 19. 20. 21. 22. 23. 2A. 25. 26. 27. 28. 29. 30. 31. 32. 33. Page Stage VI in the development of the glomerulus. A mature glomerulus . . . . 62 Mature glomerulus with cuboidal epithelium extending into the renal corpuscle at the urinary pole . . . . . . . . . 6H Ectopic glomerulus near an interlobar artery . . . . . . . . . . . . 66 Afferent arteriole entering a glomerulus located in the maturation zone of the cortex . . . . . . . . . . . . 68 Kidney cortex . . . . . . . . . . . 7O Reticular fibers within the cortex of the kidney . . . . . . . . . . . . 72 Medulla of the kidney . . . . . . . . 7A Reticular fibers within the medulla of the kidney . . . . . . . . . . . . 76 Papillary ducts emptying into the renal pelvis . . . . . . . 78 Papillary ducts within the medulla . . . . 8O PAS positive granules in the papillary ducts and the renal pelvis . . . . . . 82 Cross-section of the ureter . . . . . . 8N Transitional epithelium lining the ureter . 86 Transitional epithelium lining the urinary bladder . . . . . . 88 Three layers of smooth muscle composing the muscularis of the urinary bladder . . . 9O Cross-section of the prostatic urethra . . 92 Developing multilobular mucous acinus in the prostrate gland . . . . . . . . 9U Epithelial lining of the membranous urethra . . . . . . . . . . . . 96 vi Figure Page 3U. Cross-section of the proximal portion of the penile urethra . . . . . . . . 98 35. Cross—section of the distal portion of the penile urethra . . . . . . . . . lOO 36. Epithelial lining of the penile urethra near the external urethral orifice . . . 102 37. Epithelial lining of the female urethra . . 10h vii INTRODUCTION The histology of the urinary systemsof adult dogs has been thoroughly investigated.and compared to that of other domestic animals. However, there is little informa- tion available concerning the urinary system of newborn dogs. Although the development of the nephron of other animals has been studied extensively, the developing nephron of the dog has_received little attention. Informa- tion concerning the remainder of the urinary system of newborn dogs is far from being complete. 'It is the purpose of this study to document the histology of the urinary system of the newborn beagle dog. REVIEW OF LITERATURE A thorough review of the literature revealed limited information concerning particular aSpects of the urinary system of newborn dogs. 'A-description of the developing glomerulus (Trabucco and Marquez, 1952) was the most informative article, and descriptions of the glomerular development in the rabbit (Lewis, 1958), rat (Jokelainen, 1963), and man (MacDonald and Emery, 1959) were available for comparison with that of the dog. MacNider (1945) made a study of the lipoid material in the renal epithelium of animals of various ages. This material was present in the cells of Henlefs loop and absent in the proximal convoluted tubules of A-months- old dogs. Davies (1954) reported on protein absorption in fetal and adult mammalian kidneys. He.found PAS positive granules in the proximal convoluted tubules, the collecting ducts, and.the ureteroffetal, newborn, and immature dogs. Mayer and Ottolenghi.(l9u7) stated that.tubular epithelium protruded into the glomerular capsule in l to 12~day—old beagle dogs. This epithelium was encountered in the mature renal corpuscles of both healthy and sick dogs. Bharadwaj (1960) reported on the penis and prostate gland in puppies. A developing os penis was present in the penis, and the prostatic and penile urethrae were lined with transitional epithelium. Trabucco and Marquez (1952) studied the developing metanephros of dog embryos. They described the formation of the S-shaped body of the early nephron and the sanguineous lacuna which gives rise to the glomerulus. Although the information concerning the development of the glomerulus in the dog is limited to the work of Trabucco and Marquez (1952), extensive work has been done in describing the formation and structure of the glomerulus in the rabbit, rat, and man. Lewis (1958) described the development of the metanephric blood vessels of rabbit embryos during different stages of development. He reported that the cells which form the glomerulus are extruded from the thick, proliferative visceral epithelial cells of the glomerular capsule. Jokelainen (1963) studied the metanephros of the rat and described the development of the entire metanephric unit. He stated that the glomerulus developed in situ. Gruenwald and Popper (19AO) examined the kidneys of embryos, fetuses, and children up to 22 months of age and described the relationship of the structure and function of the glomerulus in early postnatal life. The presence of cuboidal cells in the visceral layer of the developing glomerular capsule impaired filtration because of the cell thickness. The visceral epithelium formed an unextendable sac which reduced blood flow through the glomerulus. MacDonald and Emery (1959) studied the development of human glomeruli and described six develOpmental stages. The arrangement of the visceral epithelium of the glomerular capsule and the presence of a vascular pole were the criteria used to classify the different stages. Aoki (1966) reported on the differentiation of the filtering membrane of the human glomerulus. The visceral epithelium developed from part of the vesicular wall while the capillary endothelium developed from mesenchymal cells and solid buds of interstitial capillaries which penetrated into the cleft of the late renal vesicle. A basement membrane made no appearance until the visceral epithelium and the endothelium were situated close to each other. ngkka (1961) reported that the human kidney reached full functional maturity during fetal life but remained inactive until shortly after birth. This study on 16 human fetuses revealed that epitheloid cells in the walls of afferent glomerular arterioles bulged the intima of the arterioles and reduced the bore of the vessels. However, the obstruction was reported to be incomplete, and when the kidneys of babies began functioning, the epitheloid cells lost their turgidity. MATERIAL AND METHODS Seven male and five female newborn beagle dogs were obtained from Dr. John A. Moore of the Leukemia Transmission Studies at the Michigan State University Veterinary Clinic. The animals were killed by placing them in a gas-tight Jar which contained ethyl ether. Each dog's urinary system was dissected out, and portions of a kidney were removed so that transverse, saggital, and coronal sections of the tissue could be obtained. Portions of a kidney from each dog were placed in the following fixatives: Carnoy's (Gridley, 1960), Bouin's (Gridley, 1960), Rossman's (Lillie, 1965), Baker's Formol Calcium for lipids (Lillie, 1965), and a 10% formalin solution buffered with sodium phosphate (Gridley, 1960). The tissues comprising the remainder of the urinary system were placed in Bouin's fixative (Gridley, 1960). All tissues were left in the fixative for 24 hours. The tissues were then dehydrated and cleared in four changes of tetrahydrofuran. Infiltration with Paraplast was carried out under a vaccum of 25 mm/Hg at 580-600 C for two hours before the tissues were embedded. Those tissues procured for frozen sections were fixed in Baker's Formol Calcium fixative (Lillie, 1965), cut at 10 microns, and stained for lipid material with Oil Red 0 (Gridley, 1960). Sections of all other tissues were out 6-8 microns, and the following stains were used: hematoxylin and eosin, Weigert and Van Gieson's connective tissue stain (Mallory, 1944), periodic acid-Schiff reaction (Gridley, 1960), gold chloride stain (Soule, 1962), and luxol fast blue stain combined with periodic acid-Schiff reaction (Shanklin and Nassar, 1959)- The photomicrographs were taken on a Zeiss photo- microscope using ADOX KB14 film. RESULTS AND DISCUSSION Kidney The unipyramidal kidney of the newborn beagle dog is bean-shaped, and its size varies with the size of the puppy. The parenchyma of the kidney is equally divided into a cortex and a medulla (Figure l). The terminal portions of the collecting tubes of the kidney, the papillary ducts, Open on the renal crest which is enclosed by the renal pelvis. Capsule The capsule of the kidney is composed primarily of collagenous and reticular fibers and some smooth muscle. Yadava and Calhoun (1958) reported smooth muscle in the capsule of adult dogs and Calhoun (1967) has observed smooth muscle in the capsule of 6-months-old beagle dogs. Elastic fibers are absent. Brown fat is present within the capsule at various locations around the kidney (Figure 2) and is particularly prevalent in the region of the hilus. Blood vessels are found within the capsule (Figure 3). A kidney from one dog has glomeruli and uriniferous tubules in the loose connective tissue within the capsule at the polar region (Figure 4). Cortex Neogenic Zone The cortex of the kidney consists of an outer neogenic zone and an inner maturation zone. The maturation zone contains the nephron units that are capable of producing urine while all stages in the development of the metanephric nephron are present in the neogenic zone (Figure l). Mitotic figures are found in all stages of-the developing nephrons. The stroma of the neogenic zone consists of mesen- chymal cells which differentiate into interstitial tissue and blood vessels (Aoki, 1966). Aggregations of red blood cells located in the stroma are similar to the small sinus- oids reported by KaZimierczak (1965) in the neogenic zone of newborn pigs. PAS positive granules are scattered throughout the stroma. Ureterogenic Collecting Tube.--The ureterogenic collecting tube extends through the maturation zone into the neogenic zone and terminates in the subcapsular region (Figure 5). This tube consists of columnar and cuboidal epithelial cells which have indefinite cell boundaries. The Spherical and oval nuclei of these cells possess a dis- tinct nucleolus and nuclear membrane and coarse chromatin material. The dilated end of the collecting tube, the ampulla, is surrounded by a group of elongated cells termed the metanephric blastemal cap (Huber, 1905). Metanephric Blastemal Cap.-—The metanephric blastemal cap consists of oval and elongated cells at the dilated~ end of the collecting tube. While these cells possess little cytoplasm and indefinite cell boundaries, they are easily differentiated from the surrounding mesenchymal cells because the cap as a whole appears as a definite structure (Figure 5). The cells comprising the metanephric blastemal cap possess spherical nuclei which contain a distinct nucleolus and nuclear membrane and coarse chromatin material. There is no basement membrane surrounding the metanephric blastemal cap; however, a basement membrane which reacts positively with PAS and stains positively with a reticular stain is present on the outer surface of the ureterogenic collecting tube separating the tube from the metanephric blastemal cap (Figure 5). Osathanondh and Potter (1963) considered the simul- taneous growth of the-ureterogenic collecting tube and the metanephric blastemal cap in human kidneys necessary for the cap cells to differentiate into the renal vesicle. The cells of the arched portion of the cap eVentually lose connection with the cells of the distal portions of the cap, and the latter continue to develop into the renal vesicle. Renal Vesicle.--The early renal vesicle is located in the subcapsular portion of the neogenic zone and consists of epithelial cells with a centrally located oval cavity (Figure 6). The cell boundaries are indistinct and a base- ment membrane surrounds this structure. The epithelial 10 cells contain Spherical or elongated nuclei which possess a distinct nucleolus and nuclear membrane and coarse chromatin material. The development of the renal vesicle consists of two phases (Jokelainen, 1963). Proliferation of the cells occurs during the first phase, and morphodifferentiation of the proliferating cells occurs during the second phase. The renal vesicle increases in size due to the proliferation of the cells during the growth phase, and as the renal vesicle enlarges the cells composing its wall nearest the ureterogenic collecting tubule increase in length (Figure 6). The change in these cells is considered to be the beginning of morphodifferentiation; however, the cells which are mainly involved in this second phase of development of the renal vesicle are the cells of the outer vesicular wall. During the stages of morphodifferentiation the cells of the outer vesicular wall elongate while the cells of the basal portion of the renal vesicle become cuboidal in shape (Figure 7). The cells of the apical portion of the renal vesicle maintain their original shape. The elongated cells of the outer vesicular wall eventually undergo a change in position and shape. The position of the cell changes as the apical portions of these cells are directed toward the base of the renal vesicle. The nuclei appear to migrate toward the apical part of the cell. Jokelainen (1963) described the fate of certain groups of cells which compose the late renal vesicle in rats. He 11 found that the cells in the middle of the outer vesicular wall which change position and shape are the progenitor cells- of the future visceral epithelium of the glomerular capsule and cells which resemble squamous epithelium in the lower outer portion of the renal vesicle are the progenitor cells of the future parietal epithelium of the glomerular capsule. The cells of the upper portion of the renal vesicle are the progenitor cells of the future proximal and distal convoluted tubules and the loop of Henle. Aoki (1966) reported that the proximal convoluted tubule in human kidneys arises from the group of cells which form the glomerular capsule. "Lipped Body".——The late stage of the renal vesicle differentiates into the "lipped body" (Jokelainen, 1963). A cleft appears in the outer vesicular wall separating the progenitor cells of the future glomerular capsule from part of the progenitor cells of the tubular portion of the nephron. The progenitor cells of the tubule are located above the cleft, and the progenitor cells of the future glomerular capsule are located below the cleft. The latter group of cells constitutes the lip-like structure (Figure 8). The basement membrane appears to be continuous around the entire "lipped body," and it lines the walls of the cleft (Figure 9).There is no evidence that the cavity of the newly formed "lipped body" extends into the lip. As the "lipped body" develops its tubular portion gains cellular 12 contact with the ureterogenic collecting tube which will become the collecting duct. S—Shaped Body.--The "lipped body" differentiates into the S-shaped body which is characterized by the S-shaped appearance of the developing tubule and the presence of mature red blood cells within the cleft. The S-shaped body is the first stage which occurs during glomerulogenesis (Figures 10 and 11). Maturation Zone Renal Corpuscle.--All stages in the development of the renal corpuscle are present within the maturation zone. The parietal layer of the glomerular capsule is lined by a layer of simple squamous epithelium, but the visceral layer of the glomerular capsule consists of various arrangements of columnar, cuboidal, and simple squamous epithelium during the developmental stages. The arrangement of the epithelial cells in the visceral layer of the glomerular capsule and the presence of a vascular pole are the criteria for the classification of the develOp— ing glomeruli into six stages similar to those reported by MacDonald and Emery (1959) in fetal and newborn children (Figures 11, 12, 13, 14, 15, and 16). In some instances the epithelium of the proximal convoluted tubules projects into a mature renal corpuscle (Figures 17 and 21). This epithelium possesses a brush border and is similar to that reported by Mayer and l3 Ottolenghi (1947) in 1 to l2—day-old beagle dogs and Finckh and Joske (1954) in 14 to 16—year-old children. Ectopic glomeruli similar to those described by Moffat and Fourman (1964) in adult dogs are present in the kidney. Their most frequent location is within the connective tissue near the interlobar arteries (Figure 18). The afferent glomerular arterioles lack the turgid epitheloid cells similar to those reported by Jaykk; (1961) in human fetuses. However, normal epitheloid cells arewpresent. Proximal Convoluted Tubule.--The.proximal convoluted tubule of the developing nephron changes from columnar-to pyramidal shaped cells as it matures. The cell boundaries are indistinct-and a lumen seems to be absent in immature tubules. Maturation of the proximal tubule is characterized by a lumen-and a brush border on the apical surface of the epithelial cells (Figures 15, 16, and 19). The brush border gives a positive PAS reaction. No intracytoplasmic lipid droplets are present, and no intraeytoplasmic PAS granules similar to those reported by Davies (1954) are present. The cells of the more mature proximal convoluted tubules contain more granular acidophilic cytoplasm than the progenitor cells of these tubules located in the neogenic zone. .Loqp of Henle.--The loop of Henle is found only in the more mature nephrons. This segment of the metanephric tubule consists of descending and ascending portions connected by a thin segment. The epithelium lining the 14 descending.portion resembles that of the.proximal con- voluted tubule with the exception of a decrease in the height of the cells. vThe:thin segment extends.into the medulla and is lined.by-simple-squamouswepithelium (figures 24 and 25)...The epithelium lining-the ascending portion resembles that of the distal convoluted tubule. Distal Convoluted Tubule.~-The distal.convoluted tubule of the developing.nephron changes from.columnar to cuboidal shaped cells.as it matures. “The.ce11 boundaries are indistinct and a lumen seems to be absent in the immature tubule. As in the proximal convoluted tubule, the distal convoluted tubule acquires a lumen as it matures; however, no brush border is present (Figure 19). The nuclei of the cells of the distal convoluted tubule resemble those of the proximal convoluted tubule. Macula Densa.--The macula.densa is the portion of the- distal convoluted tubule which comes in contact with the afferent and possibly the efferent glomerular arteriole. The columnar shaped cells which form the macula densa lack distinct cell boundaries and contain elongated nuclei (Figure 20). The macula-densa is present only in the more mature nephrons. '"Juxtaglomerular,Apparatus.--The Juxtaglomerular apparatus consists of three different groups of cells which are in close.contact with.each other(Figure 20). The first group of cells are the epitheloid cells located in the tunica media of the afferent arteriole near the 15 glomerulus. The second grouploficells compose the macula~ densa located in the distaliconvoluted.tubule. The third group.of cells are known aslthe cells of Goormaghtigh (Kroon, 1960) or glomerular.hilar cellsl These small elongated cells are located in_the angle between the afferent and efferent glomerular.arterioles and are in contactwwith.these.arterioleswandnthe-macula-densea. Medulla The medulla of the kidney consists-of the arched and straight.collecting ducts and.the.papillary ducts. The arched collecting ducts and portions.of the.straight col- lecting ducts form medullary rays which extend into the cortex. .Also, straight segments.of the proximal and distal convoluted tubules and.Henleis leop.are.1ocated within the mudullary ray. Henle's loOp.extends into the medullary to the region of the papillary.ducts-- Variations of the positions of the above.structures.are.present as many of the nephrens are immature. Collecting Tubules . Arched-and Straighthollecti g Duets.-eSince the arched and straight collectingeducts are difficult to dif- ferentiate histologically they will.be described together. As the.ureterogenic colleeting.tube.advanees toward the outer part of the neogenic zone of the cortex the develop- ing_nephrons surrounding it gain attachment to this tube. This is the earliest stage of.the collecting duct. A 16 single layer of cuboidal.shaped cells with distinct cell boundaries lines the ducts (Figures 22 and 23). These cells.contain spherical or elongated nuclei which possess a distinct nucleolus and nuclear membrane and coarse- chromatin material. PAS positive intracytoplasmic granules are present throughout the length of the straight col- lecting ducts. "Papillary Ducts.e—The epithelial lining of the collecting tubule changes-from cuboidal to columnar in the papillary duct (Figures 24 and 25). PAS positive intracyto- plasmic granules similar to-those reported by Davies (1954) and Liu (1962) are present in the cells lining the ducts and within the lumen of the ducts. .These granules are also present in the cells of the transitional epithelium covering the renal crest.and forming the renal pelvis (Figure 26). Renal Pelvis The renal pelvis encloses the renal crest of the kidney and leads to the ureter. Thin transitional epi— thelium lines the renal pelvis while a.lamina propria and smooth muscle fibers are present external to the epi- thelium (Figure 24). l7 Ureter The wall of the ureter consists of a mucosa, a sub- mucosa, a muscularis, and an adventitia (Figure 27). The following-description pertains to the entire length of the ureter. Tunica Mucosa Epithelium Transitional epithelium forms the lining of the ureter. The thickness of the epithelium varies from two to five cells, and the cell boundaries are distinct (Figure 28). IntracytOplasmic PAS positive granules are present. The nuclei of the epithelial cells are spherical and they possess a distinct nucleolus and nuclear membrane and coarse chromatin material. A thin basement membrane is present beneath the epithelium. Lamina Propria—Submucosa The lamina propria consists of collagenous and reticular fibers (Figures 27 and 28), and the submucosa is continuous with the lamina propria. Plasma cells are present within the lamina prOpria and submucosa of the ureters of one puppy, but they are absent in the other ureters (Figure 28). A capillary plexus extends to the epithelium. Tunica-Muscularis The muscularis consists of smooth muscle fibers arranged in a circular manner (Figure 27). Longitudinal 18 arranged smooth muscle fibers are not discernible; however, Calhoun (1959) reported that the tunica muscularis of adult dogs consists of inner and outer longitudinal layers and a middle circular layer. These three layers are com— prised of smooth muscle. Tunica Adventitia The adventitial layer consists of loosely arranged connective tissue with several blood vessels coursing through it (Figure 27). Brown fat is present in the adventitial coat of the ureters of half of the dogs. Urinary Bladder Tunica Mucosa Epithelium The epithelial lining of the urinary bladder consists of transitional epithelium. It is three to four cells in thickness in the contracted bladder and two to three cells in thickness in the distended bladder. The large, polyhedral and cuboidal cells have distinct cells boundaries (Figure 29). The spherical nuclei of these cells possess a distinct nucleolus and nuclear membrane and coarse chromatin material. Lamina Propria-Submucosa The lamina propria and submucosa consist of collagenous, elastic, and reticular fibers. Blood vessels course through the lamina prOpria and submucosa, and a capillary plexus extends to the epithelium. Although a muscularis mucosa 19 has been reported in adult dogs (Moore and Calhoun, 1957) and in 6-months-old beagle dogs (Calhoun, 1967), no muscularis mucosa is present in newborn beagle dogs. Tunica Muscularis Three thick layers of smooth muscle comprise the tunica muscularis (Figure 30). The inner and outer layers are arranged in a longitudinal manner while the middle layer is arranged in a circular manner. Blood vessels course between the layers of muscle. Tunica Serosa The serosa consists of loose collagenous connective tissue and elastic fibers with blood and lymph vessels coursing through it. The serosa is bounded with mesothelial cells of the peritoneum and contains some brown fat, nerves, and ganglia. The peritoneum does not extend to the bladder neck; therefore, an adventitia covers this portion of the bladder. Male Urethra Prostatic Portion Tunica Mucosa Epithelium.--The prostatic urethra is lined by transitional epithelium which.varies from two to four cell layers in thickness. The cell boundaries are distinct in- some areas and faint in other areas. The surface cells 20 are larger than the basal cells and bulge into the lumen (Figure 32). The nuclei of these cells are spherical or oval shaped and possess a prominent nucleolus, a distinct- nuclear membrane, and coarse chromatin material. Lamina Propria-Submucosa.--The lamina propria and submucosa are composed of collagenous, reticular, and elastic fibers. Within the lamina propria and the sub- mucosa is an extensive capillary plexus. A venous plexus is located in the distal portion of the prostatic urethra. The prostatic urethra is surrounded by the prostrate gland composed of developing multilobular mucous acini, and connective tissue which forms the urethral submucosa (Figures 31 and 32). The ducts of the acini empty into the, urethral lumen. Membranous Portion Tunica Mucosa Epithelium.--The epithelium of the membranous urethra consists of a mixture of transitional and stratified squamous epithelium which varies from three to four cells in thickness. In some instances the epithelial cells appear to be differentiating into stratified columnar epi- thelium. Bharadwaj and Calhoun (1959) reported~that» stratified cuboidal or columnar epithelium lined the mem- branous portion of the urethra in adult male dogs. The cell boundaries of the transitional and stratified squamous' epithelium are fairly distinct. The nuclei of these cells 21 may be oval, elongated, or Spherical in shape-(Figure 33), and all.nuclei possess a distinct.nucleolus and nuclear membrane with coarse chromatin material.- .Lamina Propria-Submucosa.-—The.lamina propria and submucosa consist of collagenous, reticular,.and elastic fibers, and the venous plexus coursing through these fibers is less extensive than that of.the penile urethra~ (Figure 33). A rich capillary plexus lies.beneath the epithelium. Skeletal muscle surrounds.the tunica mucosa of the membranous urethra. Penile Portion The proximal portion.of the penilelurethrazis partially enclosed by developing cartilage.whieh will become the os penis. The cartilage does not.extend.te.the ventral side of the urethra. Within the.distal protion of the penis developing cartilage is located.dorsal to the.urethra. Tunica Mucosa Epithelium.--The proximal.portion of the penile urethra is lined by transitional and stratified squamous epithelium, and near the external urethral.orifice the epithelium consists of stratified squamous epithelium and occasional patches of transitional epithelium (Figures 34, 35, and 36). The thickness of the epithelium varies from three to four cells, and the cell boundaries.are distinct. The nuclei of these cells are similar to those in the epithelium of the prostatic urethra. 22 .Lamina Propria—Submucosa.—-The lamina propria and submucosa consist of collagenous, reticular, and elastic fibers and contain a capillary plexus located immediately beneath the epithelium. .The venous plexus.located in the submucosa of the penile urethra is more.extensive than that in the membranous urethra and the distal portion of the prostatic urethra. Female Urethra Tunica Mucosa Epithelium The lining of the female urethra consists of tran- sitional epithelium-in its.proximal portion and pro-- dominately stratified squamous epithelium with patches-of transitional epithelium near the external urethral orifice‘ (Figure 37). The epithelium is three to four cell layers thick with distinct boundaries. The spherical Shaped nuclei possess a distinct nuclear membrane.and nucleolus and coarse chromatin material. Lamina Propria-Submucosa The lamina propria and submucosa consist of colla- genous, reticular, and elastic fibers, and the submucosa possesses a venous plexus which courses throughout the length of the urethra. A predominate amount of circular smooth muscle surrounds the mucosa of the female urethra. 23 Brown fat is present in.the connective tissue adjacent to the smooth muscle as well as blood vessels, lymphatics, and nerves. SUMMARY A histologic study of the urinary system of seven male and five female newborn beagle dogs is reported. The capsule of the kidney.is composed of fibrous connective tissue which contains some brown fat. Ectopic glomeruli are present in the capsule of one kidney. The cortex of the unipyramidal kidney consists of an outer neogenic zone and an inner maturation zone. The early stages of the developing nephron are present in- the neogenic zone, and.the metanephric blastemal cap, renal vesicle, "lipped-body," and the S—shaped-body are described. Six stages in the development of the glomerulus are found in the maturation zone. The maturation zone contains the nephrons which are capable of producing urine. Ectopic glomeruli are present within the connective tissue of the kidney near the interlobar arteries. A brush border is present on the apical portion of the cells of the proximal convoluted tubules but intracytoplasmic lipid droplets and PAS positive granules are not present in these cells. The macula densa and juxtaglomerular apparatus are present in the more mature nephrons near the cortico-medullary boundary. The epithelium of the collecting ducts, papillary ducts, renal crest, and the ureter contains a large amount 24 25 of PAS positive intracytoplasmic granules. The ureter is lined by transitional epithelium of varying thickness. The lamina.propria.and submucosa con— sist of collagenous and reticular fibers, and a capillary plexus extends through the lamina.propria to the epithelium. The muscularis is composed of a circular layer of smooth muscle. Brown fat is present in the adventitia of half of the ureters. The urinary bladder is lined by transitional epi-. thelium of varying thickness.. A-capillary plexus is pres- ent in the lamina propria and.submucosa. ;No muscularis mucosa is present. The muscularis consists of an outer and inner layer of longitudinal smooth muscle and a middle layer of circular smooth muscle. The urinary bladder is covered by a serosa except at.the neck of the bladder, and the serosa consists of loose connective tissue and elastic fibers and contains blood vessels, lymphatics, nerves, ganglia, and brown fat. The lining of the male urethra.consists of transitional epithelium at its origin and becomes predominately strati- fied squamous epithelium with patches of transitional epithelium near the external urethral orifice. A venous plexus is present in the submucosa throughout the male urethra. The prostatic portion of the male urethra is surrounded by the developing prostrate gland while skeletal muscle surrounds the membranous portion. The proximal portion of the penile urethra is partially enclosed by 26 developing-cartilage.as.the cartilage does not extend to the ventral side of-the urethra. .Developing cartilage is located dorsal to the.urethra in.the distal portion of the penis.- The.developing-cartilage.in the penile.urethra will become the os penis. The lining of the female urethra.consists of transi- tional epithelium in the proximal-portion and stratified. squamous epithelium with patches of transitional epithelium in the distal portion. .A venous plexus is present in the submucosa throughout the female urethra.- Giroularly arranged smooth muscle surrounds the female urethra. LITERATURE C ITED 27 LITERATURE CITED Aoki, A. 1960. DevelOpment of the human renal glomerulus, l. Differentiation of the filtering membrane. Anat. Rec., 155:339—352. Bharadwaj, M. B. 1960. Histology of the urethra and accessory sex glands of the domestic animals and observations on the surgical insertion of the urinary bladder-neck into the descending colon in the dog. Thesis, Department of Anatomy, College of Veterinary Medicine, Michigan State University. Bharadwaj, M. B., and Calhoun, M. L. 1959. Histology of the urethral epithelium of domestic animals. Am. J. Vet. Res., 20:841-851. Calhoun, M. L. 1959. Comparative histology of the ureters of domestic animals. Anat. Rec., 133:365—366. Calhoun, M. L. 1967. Personal Communication. Anatomy Department, Michigan State University, East Lansing. Davies, J. 1954. Cytological evidence of protein absorption in fetal and adult mammalian kidneys. Am. J. Anat., 94:45-71. Finckh, E. S., and Joske, R. A. 1954. The occurrence of columnar epithelium in Bowman's capsule. Jour. Path. and Bact., 68:646—648. Gridley, M. F. 1960. Manual of Histologic and Special Staining Techniques. Armed Forces Institute of Pathology, Washington, D. C. Gruenwald, P., and Popper, H. 1940. The histogenesis and physiology of the renal glomerulus in early postnatal life: Histological examinations. J. Urol., 43:452—458. Huber, G. C. 1905. On the development and shape of uriniferous tubules of certain of higher mammals. Am. J. Anat., 4 (Suppl.): ln98. H H Jaykka, S. 1961. The problem of dormant fetal organs: The kidneys, lungs, and the gut. Biologia Neonatorum, 3:3u3"3560 28 29 Jokelainen, P. 1963. An electron microscope study of the early development of the rat metanephric nephron. Acta Anat., 52 (Suppl. 47):l—7l. KaZimierczak, J. 1965. Development of the glomerulus and the juxtaglomerular apparatus. Acta Path. Microbiol. Scand., 65:318—320. Kroon, D. B. 1960. Origin of the PAS—positive granulated e—cells of the juxta-glomerular apparatus. Acta Anat., 41:138—156. Lewis, 0. J. 1958. The development of the blood vessels of the metanephros. J. Anat., 92:84-97. Lillie, R. D. 1965. Histologic Technique and Practical Histochemistry. McGraw—Hill Book Co., New York. Liu, H. C. 1962. The comparative structure of the ureter. Amer. J. Anat., 111:1—15. MacDonald, M. S., and Emery, J. L. 1959. The late intrauterine and postnatal development of human renal glomeruli. Jour. Anat., 93:331—340. MacNider, W. B. 1945. Occurrence of stainable lipoid material in renal epithelium of animals falling in different age segments. Proc. Soc. Exp. Biol. and Med., 58:326—328. Mallory, F. B. 1944. Pathological Technique. W. B. Saunders Co., Philadelphia. Mayer, E., and Ottolenghi, L. A. 1947. Protrusion of tubular epithelium into the space of Bowman's capsule in kidneys of dogs and cats. Anat. Rec., 99:477-509- Moffat, D. B., and Fourman, J. 1964. Ectopic glomeruli in the human and animal kidney. Anat. Rec., 149: 1.11 0 Moore, R. D., and Calhoun, M. L. 1957. Comparative histology of the bladder and proximal urethra of domestic animals. Anat. Rec., 127:338. Osathanondh, V., and Potter, E. L. 1963. Development of human kidney as shown by microdissection. III. Formation and interrelationship of collecting tubules and nephrons. Arch. Path., 76:290—302. 3O Shanklin, W. M., and Nassar, T. K. 1959. Luxol fast blue combined with the periodic acid—Schiff procedure for cytological staining of kidney. Stain Tech., 34: 257-260. Soule, J. D. 1962. Direct staining of reticular fibers with gold chloride. Stain Tech., 37:31-34. Trabucco, A., and Marquez, F. 1952. Structure of the glomerular tuft. J. Urol.,67:235-255. Yadava, R. P., and Calhoun, M. L. 1958. Comparative histology of the kidney of domestic animals. Am. J. Vet. Res., 19:958-968. FIGURES 31 32 Figure l.--Cortex and medulla of the kidney. 1. Neogenic zone of the cortex 2. Maturation zone of the cortex 3. Medullary ray 4. Medulla Hematoxylin and eosin x 75 33 .(ImW*—.a|.9h 42’; . D . l. Iowa” .‘ov . . Lit .. . .tJr. s\I\: \ \ II ' A . I. I II . . . . .. . o o.. .q .4!fl'fiuf.\ .8; .Wfiw . . . D . . O I . . I p :4... L..- . ‘1 - 34 Figure 2.--Brown fat within the kidney capsule. 1. Brown fat 2. Capsule 3. Neogenic zone of the cortex Hematoxylin and eosin x 384 35 36 Figure 3.--Kidney capsule. l. Fibrous connective tissue comprising the kidney capsule 2. Artery coursing through the capsule 3. Vein coursing through the capsule 4. Neogenic zone of the cortex Hematoxylin and eosin x 960 37 38 Figure 4.--Ectopic glomeruli within the kidney capsule. l. Ectopic glomerulus 2. Uriniferous tubule 3. Blood vessel Hematoxylin and eosin x 307 40 Figure 5.--Metanephric blastemal cap in the neogenic zone of the kidney. 1. 2. 5. Metanephric blastemal cap Ureterogenic collecting tube Mitotic figure Tubule of another developing nephron Basement membrane of the ureterogenic collecting tube Hematoxylin and eosin x 1228 41 42 Figure 6.--Early renal vesicle in the neogenic zone of the kidney 1. Early renal vesicle 2. Ureterogenic collecting tube 3. Kidney capsule Hematoxylin and eosin x 1228 43 44 Figure 7.—-Late renal vesicle in the neogenic zone of the kidney. 1. Late renal vesicle 2. Ureterogenic collecting tube 3. Red blood cells among mesenchymal cells Hematoxylin and eosin x 1228 45 46 Figure 8.-—"Lipped body" in the neogenic zone of the kidney. 1. "Lipped body" 2. Ureterogenic collecting tube 3. Progenitor cells of the visceral and parietal epithelium of the glomerular capsule which form the "lip" at this stage of development 4. Red blood cells within the cleft of the "lipped body" 5. Mitotic figure Hematoxylin and eosin x 1228 47 48 Figure 9.-—"Lipped body" in the neogenic zone of the kidney. 1. "Lipped body" 2. Ureterogenic collecting tube 3. Basement membrane which extends into the cleft of the "lipped body" LFB—PAS x 1228 49 50 Figure lO.--S-shaped body in the neogenic zone of the kidney. 1. S-shaped body 2. Ureterogenic collecting tube 3. "Lip" of S-shaped body 4. Progenitor cells of the metanephric tubule 5. Kidney capsule Hematoxylin and eosin x 1228 51 52 Figure ll.--S-Shaped body in the neogenic zone of the kidney. Stage I in the development of the glomerulus. l. S—shaped body 2. Ureterogenic collecting tube 3. Red blood cells in and around the "lip" 4. Tubule of another develOping nephron Hematoxylin and eosin x 1228 53 54 Figure 12.-~The sanguineous lacuna. Stage II in the development of the glomerulus. l. The mushroom-shaped sanguineous lacuna containing red blood cells 2. Cuboidal and columnar epithelium forming the visceral layer of the glomerular capsule 3. Parietal epithelium of the glomerular capsule 4. Proximal tubule Hematoxylin and eosin x 1536 55 56 Figure 13.--Stage III in the development of the glomerulus. 1. 4. 5. Visceral epithelium of the glomerular capsule composed of a single layer of cuboidal cells Parietal epithelium of the glomerular capsule Red blood cells Proximal tubule lacking a lumen Distal tubule lacking a lumen Hematoxylin and eosin x 1536 57 58 Figure l4.-—Stage IV in the development of the glomerulus. l. Visceral epithelium of the glomerular capsule arranged in clefts 2. Parietal epithelium of the glomerular capsule 3. Red blood cells in a capillary loop Hematoxylin and eosin x 1536 59 60 Figure 15.—-Stage V in the development of the glomerulus. l. Visceral epithelium of the glomerular capsule arranged in irregular groups of cells 2. Parietal epithelium of the glomerular capsule 3. Red blood cells 4. Proximal tubule possessing a lumen Hematoxylin and eosin x 1536 61 62 Figure l6.--Stage VI in the develOpment of the glomerulus. A mature glomerulus. 1. Visceral epithelium of the glomerular capsule 2. Parietal epithelium of the glomerular capsule 3. Red blood cells 4. Distal tubule possessing a large lumen 5. Proximal tubule possessing a large lumen Hematoxylin and eosin x 1536 63 64 Figure l7.--Mature glomerulus with cuboidal epithelium extending into the renal corpuscle at the urinary pole. l. Cuboidal epithelium within the renal corpuscle Hematoxylin and eosin x 1200 66 Figure l8.--Ectopic glomerulus near an interlobar artery. 1. EctOpic glomerulus 2. Proximal convoluted tubule 3. Interlobar artery '4. Medulla 5. Nerve fiber 6. Lumen of renal pelvis Hematoxylin and eosin x 300 67 :J {1 " Wt“. 33 "A -* ' 'l":.r'E 3,}. - ' .Q 1 <, 1“! 42' 3, ’r‘ ‘l 35;“ .35. . . 34%;"; 1" ‘- 5. .~. . 1 ' .- “.I"? l ‘ " . '3 ' '1I \r \ ~I I p k " v ‘ \ é 68 Figure l9.--Afferent arteriole entering a glomerulus located in the maturation zone of the cortex. 1. 2. 3. 4. 5. Afferent arteriole Distal convoluted tubule Proximal convoluted tubule with a prominent brush border Basement membrane of the tubule Basement membrane of the glomerulus PAS x 960 59 70 Figure 20.--Kidney cortex. 1. The developing juxtaglomerular apparatus 2. Macula densa Hematoxylin and eosin x 960 71 72 Figure 21.-—Reticular fibers within the cortex of the kidney. Soule's gold chloride stain x 960 73 74 Figure 22.—~Medulla of the kidney. 1. Straight collecting duct 2. Loop of Henle 3. Capillary Hematoxylin and eosin x 760 75 76 Figure 23.-~Reticular fibers within the medulla of the kidney. Soule's gold chloride stain x 512 78 Figure 24.--Papillary ducts emptying into the renal pelvis. 1. Columnar epithelium lining papillary duct L00p of Henle Vasa recta Lumen of renal pelvis Transitional epithelium forming wall of the renal pelvis Smooth muscles fibers within the wall of the renal pelvis Hematoxylin and eosin x 300 79 .34 o O ... a 0" F...‘ 41v r c (.0 xiv... ' _ _ i .5. :2... .0.0 a o ' U .3.» 9“- s . w... ..\.\.. u}. I. ‘34.?» \u 4 v, 80 Figure 25.——Papillary ducts within the medulla. l. Papillary duct 2. Loop of Henle 3. Capillary Hematoxylin and eosin x 760 81 82 Figure 26.--PAS positive granules in the papillary ducts and the renal pelvis. l. PAS positive granules in the papillary duct 2. PAS positive granules in the pelvic epithelium 3. Lumen of renal pelvis PAS x 307 83 84 Figure 27.--Cross-section of the ureter. 1. Transitional epithelium 2. Plasma cells within the lamina propria and submucosa 3. Smooth muscle fibers arranged in a circular manner 4. Blood vessels coursing through the adventitia 5. Submucosa Hematoxylin and eosin x 300 85 86 Figure 28.—-Transitional epithelium lining the ureter. l. Lumen of the ureter 2. Transitional epithelium 3. Plasma cells located in the lamina propria 4. Capillary beneath the epithelium Hematoxylin and eosin x 960 87 88 Figure 29.--Transitional epithelium lining the urinary bladder. l. Lumen of urinary bladder 2. Transitional epithelium 3. Dense collagenous fibers within the lamina propria Hematoxylin and eosin x 960 89 90 Figure 30.—-Three layers of smooth muscle composing the muscularis of the urinary bladder. 1. 2. 3. u. Outer longitudinal layer Middle circular layer Inner longitudinal layer Blood vessel coursing between muscle layers Hematoxylin and eosin x 300 91 92 Figure 3l.--Cross—section of the prostatic urethra. 1. Lumen of prostatic urethra 2. Multilobular mucous acini of the prostrate gland 3. Ductus deferens 4. Blood vessel coursing through the submucosa Hematoxylin and eosin x 75 93 94 Figure 32.--Developing multilobular mucous acinus in the prostrate gland. l. 2. 3. 4. 5. Lumen of the prostatic urethra Transitional epithelium Multilobular mucous acinus Lamina propria Submucosa Hematoxylin and eosin x 307 95 96 Figure 33.--Epithelial lining of the membranous urethra. l. Urethral lumen 2. Venous plexus within the submucosa Hematoxylin and eosin x 384 97 98 Figure 34.--Cross-section of the proximal portion of the penile urethra. l. Venous plexus within the submucosa 2. Developing cartilage which will become the os penis Hematoxylin and eosin x 307 99 100 Figure 35.--Cross-section of the distal portion of the penile urethra. 1. Capillary plexus within the lamina propria 2. Venous plexus within the submucosa Hematoxylin and eosin x 307 lOl . $5 u‘ixxVési. a. 3.... . . \c: ~§$\w}\\a\‘mx I“ u. x . \sv. .9345... . .SW. \ e“. .‘..\t\:.. ....\. . ax... «REV»? «iv . u . as. t _ B... a.» - i .x . S..- t . 13 v.2. kn»... .\ I\ \."n gui'dfl. \o... r . \‘WU. ‘ ..\. in... Vt} y H x . .\_., 102 Figure 36.-—Epithelial lining of the penile urethra near the external urethral orifice. l. Urethral lumen 2. Capillary plexus within the lamina propria Hematoxylin and eosin x 1228 103 104 Figure 37.--Epithelial lining of the female urethra. l. Urethral lumen 2. Submucosa Hematoxylin and eosin x 768 105 ummumuummuwill”1111,111qu 3 1293 03