Jean Pierre Dufaure

Levels of testosterone in the plasma and testis of the viviparous lizard (Lacerta vivipara Jacquin) during the annual cycle.

Abstract Testosterone concentration in the plasma and testis were measured by radioimmunoassay in 90 males of viviparous lizards. Determinations were performed at twice monthly intervals from March to October during two annual cycles. Plasma level of testosterone reached a peak of 445 ng/ml in the mating season, then fell abruptly to the value of 2 ng/ml in July, rose in September, then remained at a similar level of approximately 30–40 ng/ml before and after hibernation. Testosterone levels in the testis followed the plasma values except in June when the testicular levels were high although the plasma concentrations were low. This period (June) is characterized by atrophy of the epididymis (secondary sexual character) and of the seminiferous tubules and precedes the renewal of spermatogenesis (early testicular recrudescence). The plasma values of testosterone and their range of variations in this species are the highest presently known in vertebrates.

Secretory activity of the lizard epididymis and its control by testosterone

Epithelial cells of the lizard epididymis produce large secretion granules during sexual activity. A protein constituent of these granules has been detected histochemically and confirmed by autoradiographic studies of the incorporation of [3H]leucine. The proteins synthesized after incorporation of radioactive leucine have been separated by polyacrylamide gel electrophoresis. By means of castration and testosterone administration, it has been shown that the elaboration of proteins is controlled by the activity of the testis, probably via testosterone production. There is a correlation between the appearance of the secretion granules in the epididymal cells and of a particular peak of radioactivity in the electrophoretic distribution.

Levels of testosterone, dihydrotesterone, and androstenedione in the plasma and testis of a lizard (Lacerta vivipara Jacquin) during the annual cycle

Abstract Testosterone (T), dihydrotestosterone (DHT), and androstenedione (A) concentrations in the plasma and testis were measured by simultaneous radioimmunoassay in 114 males of the viviparous lizard. Determinations were performed at approximately twice-monthly intervals from March to October during one annual cycle and agreed with data obtained from other cycles. In the plasma, fluctuations of the three steroid concentrations were parallel. They reached a peak in April (the breeding season) then fell until July and subsequently rose again till September (at the retreat). Plasma T concentrations were considerably higher (390 ng/ml in April; 27 ng/ml in September) than those of DHT (14 ng/ml in April, 2 ng/ml in September) and those of A (12 ng/ml in April, 1 ng/ml in September). In testis, T and A levels rose abruptly in June (T = 12,440 pg/testis; A: 10,000 pg/testis) although T concentrations did not rise significantly in plasma. This phenomenon seems to be related to the dramatic changes which occur in the seminiferous tubules between two successive spermatogenic cycles. Moreover, DHT, which was not detectable in the testis at other times, occurred transiently in June (320 pg/testis). The dissociation between plasma and testicular androgen levels observed during the transition period between two spermatogenetic cycles indicates that steroid hormones might play an important part in the control of decline and/or initiation of spermatogenesis. It suggests that the lizard is a convenient model for studying the action of steroids on spermatogenesis.

Action des hormones androgenes sur l'epididyme d'un reptile lacertilien, Lacerta vivipara jacquin. Effets de la testosterone et de ses principaux mètabolites en culture organotypique

Resume Lepididyme de lezard est controle par le testicule comme le montre la castration, retient selectivement la testosterone et il est capable de convertir cette hormone en 5αDHT et en 5α androstanediols. Dautre part, cet organe cible subit au cours du cycle annuel dimportantes modifications structurales, de son epithelium en particulier. Il est caracterise pendant sa periode dactivite par une secretion abondante. Nous avons etudie in vitro la stimulation de ce tissu par la testosterone, la 5αDHT, le 3α et le 3β androstanediol. En culture organotypique, il est possible dobtenir des cellules epitheliales de taille normale et capables de secreter, en traintant par ces hormones, des explants provenant danimaux castres dont les cellules epitheliales sont fortement regressees. On observe differents degres de stimulation: la 5αDHT et le 3β androstanediol sont plus puissants que le 3α androstanediol, et ce dernier plus actif que la testosterone. La microscopie electronique montre que leffet de la 5αDHT peut etre observe a deux niveaux: augmentation de la taille du noyau et plus particulierement du nucleole et mise en place dun systeme remarquable de reticulum endoplasmique granulaire. Cet organe est propose comme modele detude du mecanisme daction des androgenes.

L'ultrastructure du testicule de lézard vivipare (Reptile, Lacertilien)

SummarySertoli cells of the testis of Lacerta vivipara have been studied electron microscopically in animals obtained between spring and autumn during two years and in animals hypophysectomized in autumn.These cells contain numerous small mitochondria with lamellar cristae, free ribosomes, smooth endoplasmic reticulum moderately developed, several small dictyosomes forming the Golgi complex, lipid droplets and microtubules. There are numerous dense bodies of large size with an heterogeneous content which seem to be of lysosomial nature. Glycogen consists of β particles dispersed at random in the hyaloplasm. Seasonal variations in the content of glycogen are noted. In hypophysectomized animals Sertoli cells contain large amounts of that metabolite whose particles are concentrated in small areas often around the lipid droplets.Possible role of the Sertoli cells concerning mechanical support and nutrition of the germinal cells, production of hormones and phagocytosis of residual bodies are discussed. The variations in the glycogen content are also discussed.RésuméLes cellules de Sertoli du testicule de Lacerta vivipara ont été étudiées en microscopie électronique chez des animaux récoltés entre le printemps et lautomne pendant deux années et chez des animaux hypophysectomisés en automne.Ces cellules contiennent de nombreuses mitochondries de petite taille à crêtes lamellaires, des ribosomes libres, un reticulum endoplasmique lisse moyennement développé, plusieurs petits dictyosomes formant lappareil de Golgi, des liposomes et des microtubules. Elles renferment aussi de nombreux corps denses de grande taille qui paraissent être de nature lysosomiale. Le glycogène a été particulièrement étudié. Il est formé de particules β dispersées au hasard dans le hyaloplasme. Des variations saisonnières dans la teneur en glycogène ont été notées. Chez les hypophysectomisés, les cellules de Sertoli contiennent de grandes quantités de ce métabolite dont les particules sont concentrées dans des petites plages, souvent autour des liposomes.Les rôles possibles des cellules de Sertoli sont discutés: soutien et apport de nourriture aux cellules germinales, production dhormones et phagocytose des corps résiduels. Les variations de la teneur en glycogène sont également discutées.

Uptake of [3H]Testosterone in several organs of the male viviparous lizard (Lacerta vivipara Jacquin) and selective retention by the epididymis

Abstract [1, 2, 6, 7- 3 H]Testosterone was injected into castrated male viviparous lizards to establish retention of the hormone by blood, lung, gut, liver, kidney, penis, and epididymis. Radioactivity in these organs was determined in the spring four times during the 12 hr following administration of the isotope. Retention in blood was greater than in mammls, so that significant differences in retention between the epididymis and nontarget tissues appeared only between 6 and 12 hr after treatment. High levels of radioactivity found in the gut indicate that this route is used for clearance of the exogenous steroid. Selective retention of testosterone by the epididymis during its period of activity (spring) was no longer found in regressed or inactive organs (summer and autumn). Autoradiographs of three tissues indicated that testosterone or a metabolite was concentrated in cell nuclei of the epididymis but not in those of liver and gut.

L'ultrastructure du testicule de lzard vivipare (Reptile, Lacertilien)@@@The fine structure of the lizard testis: I. Les cellules interstitielles@@@I. The interstitial cells

SummaryInterstitial cells of the testis of Lacerta vivipara have been studied electronmicroscopically in animals obtained between spring and autumn.Smooth endoplasmic reticulum and mitochondria with tubular cristae are the most prominent organels, lipid droplets and Golgi apparatus being also well developed.The most significant ultrastructural changes occur between spring and the beginning of summer. In spring, during the hypertrophy of secondary sexual characters, a conspicuous system of vesicles and vacuoles originates from the smooth endoplasmic reticulum and probably also from the Golgi apparatus. At the beginning of summer, when secondary sexual characters are atrophied, vacuoles are less prominent and the smooth endoplasmic reticulum consists of a dense network of typical tubules, often closely associated with the lipid droplets; the cristae of the mitochondria are swollen.These ultrastructural findings are discussed in relation to the production of hormones. The hypertrophy of membrane systems in spring corresponds presumably to production or (and) release of androgen hormones. In the beginning of summer the cell does not produce androgens, but probably is not completely inactive: it may store precursors of hormones.RésuméLultrastructure des cellules interstitielles du testicule de Lacerta vivipara a été étudiée entre le printemps et lautomne pendant deux années.Le retioulum endoplasmique lisse, et les mitochondries à crêtes tabulaires sont les organites les plus remarquables comme dans les autres cellules productrices de stéroïdes, mais les liposomes et lappareil de Golgi sont bien représentés aussi.Les variations ultrastructurales les plus significatives apparaissent entre le printemps et le début de lété. Au printemps, alors que les caractères sexuels secondaires sont hypertrophiés, un système remarquable de vésicules et de vacuoles se développe à partir du reticulum et probablement aussi du Golgi. Au début de lété, lorsque les caractères sexuels secondaires sont atrophiés, les vacuoles sont moins nombreuses et le reticulum forme un réseau dense de tubules typiques, souvent étroitement associés aux liposomes; les crêtes mitochondriales sont gonflées.Ces images sont discutées en fonction de lactivité saisonnière délaboration dhormones. Lhypertrophie des systèmes membranaires au printemps correspond probablement à la production ou (et) à lexcrétion des hormones androgènes. Au début de lété, la cellule nélabore pas dandrogènes, mais nest peut-être pas complètement inactive: elle pourrait stocker des précurseurs hormonaux.

Scanning electron-microscopical study of epididymal secretions in the lizard Lacerta vivipara (Reptilia, Squamata) and of their relationships with spermatozoa

SummaryDuring the breeding season (April, May) the epididymis of the lizard Lacerta vivipara produces voluminous secretory granules which are abundantly discharged into the lumen of the duct where they mingle with spermatozoa. The mode of secretion appears quite unusual with respect to the method by which the cells discharge their products, the granules coming out of the cells like bullets out of a gun barrel. Spermatozoa come into close relationships with discharged granules, dipping into their outer layers. This is probably the way in which the heads of spermatozoa become covered with the epididymal soluble protein (protein L). This mode of secretion in Lacerta is discussed with regard to possible artifacts and compared with that encountered in the epididymis of some other species including mammals.

CYTOSKELETON OF THE SERTOLI CELL: EFFECTS OF FSH AND TESTOSTERONE IN THE IMMATURE PIG TESTIS

In a variety of physiological * and experimental (after hypophysectomy) conditions, ultrastructural studies of the Sertoli cells of pig have shown that evolution of the cytoskeleton, (microfilaments, intermediate-sized filaments, and microtubules) which is organized during prepuberty and regresses after hypophysectomy, might be regulated directly or indirectly (via testosterone production) by pituitary hormones. This point was first analyzed by cultivating small samples of immature (six-week old) porcine testis in well defined hormonal conditions.3 In samples cultivated with (4 x 10-8 M) testosterone alone microfilaments were organized, and in the presence of testosterone plus (2.8 pg/ml) porcine FSH intermediatesized filaments and microtubules appeared additionally.3 It is not possible to ascertain what the effect of purified FSH is in the absence of testosteronc (checked by radioimmunoassay) , because when the LH contaminant of FSH preparation was neutralized by incubation with a homologous anti-LH, Sertoli cells underwent necrosis. In order to know if this noxious effect is due to the absence of testosterone several controls must be performed. In a second set of experiments, Sertoli cells (81.5% Sertoli cells, 18% peritubular cells, and 0.5% Leydig cells) were isolated from immature (six weeks old) pig testis5 and maintained in medium 199 plus 5 % calf serum. The organization of microfilaments was visualized by changes in the shape of the cell studied in phase contrast microscopy on living cells and on cells fixed with absolute methanol (at -20 C) . Without FSH or testosterone (checked by radioimmunoassay) ir. the medium, the flattened cells aggregated in a confluent monolayer. With (0.3 pg/ml) FSH the cells rounded but this effect was avoided or delayed by adding (4 x M) testosterone. FSH caused a disorganization of microfilaments previously identified as actin-like material by immunohistochemistry. A microfilament disrupting agent, ( M ) cytochalasin B, mimicked the effect of FSH although (10-6 M) colchicine (the specific microtubule-disrupting agent) did not. Such changes in the shape of these cells and in the organization of microfilaments may be correlated with a redistribution of Ca2+. Indeed (6 x M) EGTA (a chelator of Ca2*) added to flattened cells caused a cytoplasmic retraction. This shape modification was reversible by changing the medium. With a specific ionophore of Ca2+ (lo- M calcimycine = A 23187) Sertoli cells show a demonstrable decrease in surface area.

A Model System to Study the Morphogenic and Steroidogenic Effects of Various Gonadotropins on the Leydig Cell of the Boar

The boar testis was chosen because it contains a great number of large polyhedric Leydig cells providing a good opportunity to perform morphometric analysis. Our model system consists of small pieces of testis from adult animals hypophysectomized since 1 to 3 months of age, which were maintained in static organ cultures with medium 199, 10% calf serum, and 250 I.U./ml of penicillin (Trowells method, modified by Lasnitzski,2 adapted to pig testis material3). This medium was supplemented or not with hormones (hCG, LH, or FSH) or other components (related antihormones, CAMP, and dibutyryl CAMP). Results were analyzed both by microscopic examination and measures at histological and cytological levels, and by measurements (radi~immunoassays~) of the steroids released into the medium for 24 hours (testosterone, dihydrotestosterone, androstenedione, and dehydroepiandrosterone sulfate). We have previously shown that hypophysectomy4p6 was followed by a dramatic reduction in the size of the Leydig cells, of their nucleus, and of their organelles (particularly those involved in steroidogenesis), but by only a slight decrease in their number, although testosterone was not more detectable in the plasma. In vivo, these modifications were reversed by injection of hCG.6,8 From 2 days to 10 days of culture with 10 I.U./ml of hCG, the size of the cells and of their nucleus increased, cytoplasmic structures associated to steroidogenesis developed, and the four steroids were released into the medium. The full activity was reached after 10 days of culture (testosterone: 80 ng/ml of medium/24 hr corresponding to 13.2 ng/mg of tissue/24 hr); then activity decreased regularly and irreversibly (except for one case).9 The system was further used to study the dose-response effects of hCG, of various hypophyseal gonadotropins (porcine, ovine, bovine, and human LH and FSH), given alone or combined with their related antihormone, and of the intracellular mediator CAMP.