Lactate dehydrogenase isozymes in the spermatozoa of the domestic fowl, Gallus domesticus and turkey, Meleagris gallopavo.

1. Electrophoresis of extracts of turkey spermatozoa for lactate dehydrogenase activity revealed the usual five tissue LDHs (LDH-1 to LDH-5). 2. The presence of LDH-X (the spermatozoan-specific isozyme) was not obvious. 3. Only one band was present on electrophoresis of fowl spermatozoan extracts and it coincided with LDH-1 (heart type). 4. Kinetic investigations, the use of inhibitors and the heat-stability test confirmed that the fowl spermatozoan LDH was probably LDH-1 and not LDH-X.     

Effects of corticosteroids on short-circuit current across the cecum of the domestic fowl,Gallus domesticus

Summary Both avian corticosteroid hormones, aldosterone and corticosterone, increased short-circuit current across the wall of the ceca of the domestic fowl (Gallus domesticus) in vitro. About 80% of this short-circuit current was inhibited by the Na-channel blocking drug amiloride. Corticosterone was about ten times less potent than aldosterone in increasing short-circuit current and it exerted a similar maximal effect. Cortisol (an endogenous corticosteroid hormone in mammals but not birds) was about ten times less potent than corticosterone and this difference appeared to reflect the presence of the 17-OH group in cortisol. Carbenoxolene, which inhibits 11-hydroxysteroid dehydrogenase, increased the effect of corticosterone. This effect is consistent with inhibition of the metabolism of corticosterone to 11-dehydrocorticosterone. The latter was found to be about 100 times less potent than corticosterone. The effects of both aldosterone and corticosterone (also dexamethasone) were abolished by the mineralocorticoid receptor antagonist spironolactone. The results suggest that corticosterone has an effect similar to aldosterone but in vivo its action may be depressed by the activity of 11-hydroxysteroid dehydrogenase. The sensitivity of the cecal preparations to corticosterone indicates that this hormone could contribute to the regulation of transcecal Na transport (absorption) in vivo.Abbreviations 11-HSD 11-Hydroxysteroid dehydrogenase - sc short-circuit current - KRB Krebs bicarbonate solution     

Activation of dynein adenosine triphosphatase and flagellar motility of demembranated spermatozoa by monovalent salts at 40 degrees C in the domestic fowl, Gallus domesticus

1. At 40 degrees C, around the normal avian body temperature, demembranated fowl spermatozoa with no addition of monovalent chlorides were immotile. 2. Demembranated spermatozoa become motile at 40 degrees C when 0.1-0.5 M concentrations of NH4Cl, NaCl and KCl were added to the reactivation medium, with maximum motility occurring at 0.2-0.3 M in all cases. 3. The addition of NH4Cl, NaCl and KCl also stimulated the ATPase activity of crude dynein extract. In contrast, LiCl did not appreciably affect motility and ATPase activity. 4. These results showed that the flagellar dynein ATPase activity of fowl spermatozoa could be stimulated by the addition of certain monovalent chlorides, except LiCl, and demembranated spermatozoa might be motile at 40 degrees C.     

Histochemical localization of 3 - and 17 -hydroxysteroid dehydrogenases in the male reprodutive tract of the domestic fowl (Gallus domesticus)

Synopsis 3- and 17-hydroxysteroid dehydrogenase activities were studied histochemically in the male reproductive tract of the domestic fowl. 3-hydroxysteroid dehydrogenase was NAD⁺-linked and was capable of metabolizing the three substrates used, namely, pregnenolone, 17-hydroxypregnenolone and dehydroepiandrosterone. 17-hydroxysteroid dehydrogenase oxidized testosterone in the presence of NAD⁺ or NADP⁺. The pattern of distribution of formazan granules was essentially the same with all the substrates used, and they were located in the Leydig cells, seminiferous tubules and the lining epithelia of the entire excurrent duct system of the testis except the rete testis. The activity of both enzymes appeared to be highest in the ductuli efferentes and decreased distally along the tract. The evidence suggests that steroid synthesis may occur in the epithelial lining of the excurrent ducts as well as in the cells of the testis.     

Effects of genetic selection on growth rate and intestinal structure in the domestic fowl (Gallus domesticus)

1. Pieces of small intestine taken from chickens subjected previously to continuous selection, relaxed selection or no selection for rapid growth were used to estimate villus surface area and microvillus development to determine what effects genetic selection might have on factors controlling intestinal function. 2. Crypt size and the rates at which enterocytes migrated out of crypts were also measured, after injection of tritiated thymidine, to determine the time course of microvillus elongation. 3. Differences in growth rates measured between highly selected, relaxed selected or unselected birds were found to be correlated with parallel changes in villus surface area. Selection for growth did not change the density, dimensions or pattern of development of enterocyte microvilli. Microvilli did, however, produce a maximal 20-fold increase in villus surface area under all conditions. 4. Crypt size and enterocyte migration rates did not vary significantly between tissue taken from unselected and relaxed selected chickens. Tissue taken from highly selected birds had a crypt size and enterocyte migration rate 40% higher than values found for the other two groups of chickens. 5. The possibility that early genetic selection increased growth potential by uncoupling diet-induced changes on crypt hyperplasia from secondary effects on villus structure, and that later selection increased growth potential by increasing appetite, is discussed.     

The excretion of urate and cationic electrolytes by the kidney of the male domestic fowl (Gallus domesticus)

Summary Four groups of roosters were obtained from combinations of high and low protein diets (33% or 11%), and two drinking solutions (tap water or 1% NaCl solution). Ureteral urine was analyzed for urate, Na⁺ and K⁺, in both the liquid and precipitated fractions of the urine.Birds fed a high protein diet-salt water combination excreted unusually large amounts of urate. In all groups, most of the excreted urate was in the form of a precipitate. This precipitate also contained large amounts of Na⁺ and K⁺ (Table 2).The presence of abundant urinary urate, due to high protein diet and large NaCl intake, aids in the excretion of Na⁺ and K⁺, by reducing their contribution to the osmotic pressure of the urine. However, some of these cations, although present in the urine precipitates, do not appear to be in the form of monobasic urate salts. The significance of urate in the excretion of electrolytes is discussed.We wish to thank Dr. Paul B. Siegel, Department of Poultry Science, V.P.I. and S.U., who generously supplied the roosters used in this study. We also wish to thank Mr. Douglas Bartley for his determination of urine pH values, and Dr. Alan G. Heath for the use of his microelectrode pH apparatus. This work was supported by USPHS-NIH grant, number AM 14991.     

Ovarian steroidogenesis during follicular maturation in the domestic fowl (Gallus domesticus)

The steroidogenic potential of various physiological compartments within the ovary of the hen were examined using in vitro systems. Three-hour incubations of individual whole small follicles (less than 1 mm-1 cm) or 100,000 collagenase-dispersed theca cells of the five largest ovarian follicles (F1-F5) were conducted in 1 ml of Medium 199 at 37 degrees C in the presence and absence of luteinizing hormone (LH) (0.39, 0.78, 1.56, 3.13 and 6.25 ng), progesterone (5 ng), and dehydroepiandrosterone (DHEA, 5 ng). Steroid output was measured by radioimmunoassay of incubation media. Progesterone was not produced by small follicles although they are a major source of DHEA and estradiol and a significant source of androstenedione. Output of DHEA, androstenedione and estradiol was highly stimulated by LH. The substrate for androstenedione and estradiol in small follicles is probably DHEA. Output of DHEA and androstenedione in theca cells of F2-F5 was stimulated by LH in a dose-related manner. A dose-response relationship between estradiol output and the concentration of LH in media was not apparent in theca cells from F2-F5. Steroidogenesis in theca tissue of large follicles occurs predominantly via the delta 4 pathway. The ability of these theca cells to metabolize progesterone to androstenedione is lost between 36 and 12 h before ovulation. Their ability to metabolize DHEA to androstenedione is still present 12 h before ovulation. Aromatase activity is significantly reduced between 36 and 12 h before ovulation. These data indicate that both large and small follicles can be stimulated by LH. The small follicles are the major source of estrogen. As the large yolky follicles mature, steroidogenesis shifts from the delta 5 to the delta 4 pathway. By 12 h before ovulation, the F1 follicle has lost the ability to convert progesterone to androstenedione. The inability of the largest ovarian follicle to convert progesterone to androstenedione contributes at least in part to the preovulatory increase in the plasma concentration of progesterone that generates the preovulatory LH surge by positive feedback.     

The passage of spermatozoa through the vitelline membrane in the domestic fowl,Gallus gallus

Summary The developing outer layer of the vitelline membrane of the ovum in the posterior part of the infundibulum of the domestic fowl contains many spermatozoa in nearly parallel orientation with its inner layer. When the acrosomal region of a spermatozoon approaches or contacts the inner layer, promptly undergoes the acrosome reaction. The outer acrosomal membrane and overlying plasma membrane fuse together and the apical region of the acrosome opens, so that the acrosomal contents are released. Meanwhile the spermatozoon remains a time in contact with the surface of the inner layer, and the network of the inner layer just under the tip of the sperm head begins to be dissolved. This dissolution extends downward forming a tunnel, approximately 9 m in diameter. The spermatozoon then passes through the inner layer obliquely via the central region of the tunnel and arrives at the perivitelline space.The authors are greatly indebted to assoc. prof. Dr. Osamu Koga for his valuable advices. The authors also wish to thank Mr. Takayuki Mori for his helpful suggestions and technical advices. This investigation was supported by a grant from the Ministry of Education of Japan (156185)     

Demonstration of W chromosome-specific repetitive DNA sequences in the domestic fowl,Gallus g. domesticus

Evidence is presented to demonstrate the presence of W chromosome-specific repetitive DNA sequences in the female White Leghorn chicken, Gallus g. domesticus, based on two different experimental approaches. First, ³H-labelled, female chicken DNA was hybridized with excess, unlabelled, mercurated, male DNA, and unhybridized single-stranded ³H-DNA (³H-SHU-DNA) was recovered by SH-Sepharose and hydroxyapatite column chromatography. Approximately 24% of the hybridizable ³H-SHU-DNA was female-specific and localized on the W chromosome. The second approach was to examine female-specific DNA fragments among the digests of chicken DNA with various restriction endonucleases. Among them, we found that digestion with XhoI produced two prominent female-specific bands of 0.60 kb (= kilobase pairs) and 1.1 kb. The 0.60 kb fragment was isolated and ³H-labelled by nick-translation. Female-specificity of the ³H-XhoI—0.60 kb DNA was judged to be at least 95% under the conditions of hybridization with membrane filter-bound DNA. Presence of amplified XhoI—0.60 kb DNA on the W chromosome seems to be limited to different lines of G. g. domesticus and no such repeat was detected in three species belonging to other genera in the order Galliformes and in three species belonging to other avian orders.     

Fertilizing competency of multiple ovulated eggs in the domestic fowl (Gallus domesticus)

Fertilizing competency of multiple ovulated eggs in the domestic fowl was examined by fertilization in vitro and early development in culture. Normal laying hens (White Leghorn) were treated with 75 IU of PMSG for 7 days followed by injection of anterior pituitary extracts from chickens (CAPE). Ovulation began to occur 7.5 h after injection of CAPE. These hens ovulated 1-7 ova but some premature ovulation of GV stage ova were observed. In vitro fertilization of the multiple ovulated ova was examined by inseminating 10(6)-10(7) sperm onto the germinal disks in m-Ringer's solution. The gamete or zygote nuclei were detected by DNA specific fluorescence using DAPI (4',6'-diamidino-2-phenylindole) in the histological section prepared from the germinal disk. Process of fertilization was examined in the eggs incubated for 4 h after insemination in DMEM + liquid albumen at 41 degrees C under the atmosphere of 5% CO2 in air. Fertilization rate of the total multiple ovulated eggs was 55% (11/20), in which 90% (9/10) and 10% (1/10) in the eggs recovered 7.5-8.5 h and 9.0-9.5 h after CAPE injection were obtained, respectively. Normal pronuclei were formed in five eggs of those recovered 7.5-8.5 h after CAPE injection. Early development after fertilization in vitro was also examined by incubation for 12 h in DMEM + liquid albumen at 41 degrees C under the atmosphere of 5% CO2 in air. Although development in vitro was delayed compared to that in utero condition, normal development was observed in naturally and multiple ovulated eggs.(ABSTRACT TRUNCATED AT 250 WORDS)     

Cell membrane amino acid transport processes in the domestic fowl (Gallus domesticus)

Intestinal absorption of amino acids in the chicken occurs by way of processes which are concentrative, Na+-dependent and dependent upon metabolic energy in the form of ATP. Intestinal transport is carrier-mediated, subject to exchange transport (trans-membrane effects) and is inhibitable by sugars, reagents which inactivate sulfhydryl groups, potassium ion, and by deoxpyridoxine, an anti-vitamin B6 agent. It is stimulated by phlorizin, a potent inhibitor of sugar transport, and in Na+-leached tissue by modifiers of tissue cyclic AMP levels, e.g. theophylline, histamine, carbachol and secretin. Separate transport sites with broad, overlapping specificities function in the intestinal absorption of the various classes of common amino acids. A simple model for these sites includes one for leucine and other neutral amino acids, one for proline, beta-alanine and related imino and amino acids, one for basic amino acids, and one for acidic amino acids. Absorption of amino acids appears to be widespread in occurrence in the digestive tract of the domestic fowl; transport has been reported to be present in the crop, gizzard, proventriculus, small intestine and in the colon. By the end of the first week of life post-hatch, the caecum loses its ability to transport. Similarly, the yolk sac loses its ability by the second day post-hatch. Intestinal transport was noted before hatch and was found to be maximal immediately post-hatch. A requirement for Ca2+ appears to be lost after the first week of life post-hatch. The cationic amino acids appear to be reabsorbed by a common mechanism in the kidney. Transport rates of leucine measured in the intestine or in the erythrocyte were found to cluster about discrete values when many individual chickens were surveyed; such patterns may be an expression of gene differences between individuals. Two lines of chickens have been developed, one high and the other low uptake, through selective breeding based on the ability of individual birds to absorb leucine in erythrocytes. High leucine absorbing chickens were found to be more effective in absorbing lysine and glycine, were more effectively stimulated by Na+, had greater erythrocyte Na+, K+-ATPase activity, and their erythrocytes contained about 20% less Na+ than low line erythrocytes. The underlying genetic difference between these lines may reside at the level of the Na+, K+-ATPase and (or) with a regulatory gene determining carrier copies. Amino acid transport in erythrocytes was noted to be highest in pre-hatch chicks and to diminish during post-hatch development.(ABSTRACT TRUNCATED AT 400 WORDS)