The effect of temperature and gonadotropin on testicular steroidogenesis in Sarotherodon (tilapia) mossambicus in vitro

Testes of sexually mature Sarotherodon mossambicus were incubated at 15, 22, 30, and 40° with (a) tritiated testosterone and (b) salmon pituitary extract. Formation of 11-keto- and 11β-hydroxytestosterone from the tritiated precursor showed little change in yield between 15 and 30° but yields of glucuronides rose dramatically between 22 and 30° and a significant rise was observed for formation of 5β-androstane-3α, 17β-diol between 15 and 40°. Yields of 3α,17β-dihydroxy-5β-androstan-11-one followed a pattern similar to that of 11-ketotestosterone. With endogenous precursors under the stimulation of salmon pituitary extract, yields of testosterone, 11-ketotestosterone, and 11β-hydroxytestosterone were maximal at 22° after which they declined to very low levels at 40°. Yields of testosterone and 11-ketotestosterone glucuronides while showing a peak at 22° declined much more slowly at higher temperatures than did those of the free steroids. In the absence of pituitary stimulation, levels of all steroids were below the limits of detection. Plasma levels of testosterone (15.3 ± 1.5 ng/ml), 11-ketotestosterone (5.3 ± 2.7 ng/ml), 11β-hydroxytestosterone (5.5 ± 2.6 ng/ml), and their glucuronides (1.5 ± 0.5, 0.14 ± 0.1, and 1.5 ± 0.5 ng/ml, respectively) were measured in fish held at 25°. A rapid nonchromatographic method for the assay of the three free steroids is described and the results are shown to be comparable to those obtained after chromatography.     

Involvement of gonadotrophin and steroid hormones in spermiation in the amago salmon, Oncorhynchus rhodurus, and goldfish, Carassius auratus

Effects of intraperitoneal injections of chum salmon gonadotrophin (SGA) and various steroid hormones (17α,20β-dihydroxy-4-pregnen-3-one; 17α,20β-diOHprog, 17α-hydroxy-progesterone, testosterone, 11-ketotestosterone) on the induction of in vivo spermiation were examined in nonspermiating amago salmon (Oncorhynchus rhodurus) and goldfish (Carassius auratus). A single injection of SGA to amago salmon and two successive injections to goldfish induced strong spermiation responses. Similarly two successive injections of 17α, 20β-diOHprog caused precocious spermiation in both species; however, the response to 17α,20β-diOHprog was of lesser magnitude than that to SGA. The spermiation response of goldfish to 17α-hydroxyprogesterone was similar to that of 17α,20β-diOHprog. Neither testosterone nor 11-ketotestosterone were effective in inducing spermiation in amago salmon, but these steroids were found to be slightly effective in goldfish. Effects of a single injection of SGA on changes in serum levels of 17α,20β-diOHprog and 11-ketotestosterone were also examined in nonspermiating amago salmon. Serum levels of 17α,20β-diOHprog dramatically increased after treatment and peaked on Day 2 and thereafter declined quickly. Similarly, 11-ketotestosterone peaked on Day 2, but the levels remained high throughout the experimental period. Considered together, these findings are consistent with the suggestion that 17α,20β-diOHprog is involved in the process of spermiation in teleosts. It is further suggested that this hormone is a testicular steroidal mediator of gonadotrophin-induced spermiation in amago salmon.     

In vitro metabolism of testosterone in hepatic tissue of a hagfish, Eptatretus burgeri

The metabolism of testosterone in hepatic tissue (combined microsomal and cytosol fractions) of a hagfish, Eptatretus burgeri, was examined in the presence of NADPH and in aerobic or CO-saturated atmosphere. The metabolites formed in CO atmosphere were identified as 4-androstene-3,17-dione, 5α-dihydrotestosterone, 3β-hydroxy-5α-androstan-17-one, and 5α-androstane-3β,17β-diol. From formation of these metabolites of testosterone, the activities of 5α-reductase, 3β- and 17β-hydroxysteroid dehydrogenases were indicated. Under aerobic condition, 7α-hydroxytestosterone, 7α-hydroxy-5α-dihydrotestosterone, 3β,7α-dihydroxy-5α-androstan-17-one and 5α-androstane-3β,7α,17β-triol were formed, in addition to 4-androstene-3,17-dione. These results demonstrate the presence of 5α-reductase, 3β- and 17β-hydroxysteroid dehydrogenases and 7α-hydroxylase in the hagfish liver. Since the hagfish is regarded as a very primitive vertebrate, it may be suggested that these enzymes are phylogenetically old and that the 7α-hydroxylase represents one of the first forms of hepatic cytochrome P-450 appearing during development.     

Further evidence that serotonin may be a physiological melanocyte-stimulating hormone-releasing factor in the lizard Anolis carolinensis

Testes of the hagfish Myxine glutinosa were incubated with [³H]progesterone and [³H]-testosterone and their metabolites identified by chromatography and either chemical reaction followed by crystallisation to constant isotope ratio or by gas chromatography-mass spectrometry. The following metabolites of testosterone were identified: androstenedione (23% yield), 6β-hydroxytestosterone (6.3%), 5α-androstane-3β,7α,17β-triol (4.5%), 5α-androstane-3β,6β,17β-triol (trace), and a solvolysable conjugate of testosterone (0.7%). Testosterone was identified as a metabolite of progesterone (3% yield).     

Cortisol and osmotic adjustment of the euryhaline teleost, Sarotherodon mossambicus

Changes of plasma cortisol concentration in the euryhaline Sarotherodon mossambicus were studied during the first time of adaptation to different external salinity conditions. The fish survived for more than 24 hr after direct transfer to saltwater only when the concentration was not higher than 2.7%. Transfer from fresh water directly to 2.7% saltwater increased the plasma cortisol level only during the first 2 hr. Plasma osmolality and sodium level were raised for more than 72 hr. Fish adapted for some weeks to 3.5% saltwater and transferred back to fresh water showed a similar increase of cortisol concentration which lasted for 6 hr. Plasma osmolality and sodium were lowered for a longer time (more than 6 days). Transfer from fresh water to distilled water was followed by an increase of plasma cortisol for 2 days. The significance of the increase of cortisol can be seen in the induction of metabolic changes necessary for volume regulatory processes. To test this hypothesis, the influence of cortisol injections was studied in a second set of experiments. Pretreatment with cortisol before the transfer from fresh water to 2.7% saltwater reduced the shrinkage of the muscle cell and enlarged especially the extracellular space within the muscle. It is evident that shifts of electrolytes are stimulated by cortisol.     

Immunocytochemical localization of luteinizing hormone-releasing hormone in the brain of the goldfish Carassius auratus

Using an indirect immunofluorescence technique, LH-RH was localized in the brain of the goldfish (Carassius auratus). Immunoreactive cell bodies were inconsistantly found in two hypothalamic nuclei: the anteroventral nucleus preopticus periventricularis (npp) and the posterior nucleus lateralis tuberis (nltp). Numerous immunoreactive fibers were observed in hypothalamic as well as in the extrahypothalamic parts of the brain and in the proximal pars distalis of the pituitary gland. Our results are discussed in relation to the neuroendocrine control of the pituitary gonadotropic function in teleosts.     

Effects of monosodium l-glutamate on pituitary innervation in goldfish, Carassius auratus

Monosodium l-glutamate was injected intraperitoneally into goldfish at a dosage of 2.5 mg/g body wt. The first noticeable effect in the pituitary, 30 min postinjection, was a marked swelling of pituicytes in the neurohypophysis (NH). Eighteen hours postinjection a large number of degenerating type B fibers were found in the NH of the proximal pars distalis (PPD), while all type A fibers, and type B fibers in other parts of the pituitary, remained unaffected. Because glutamate causes a lesion in the nucleus lateral tuberis (NLT), from the anterior margin of the pituitary stalk through to the posterior end of the nucleus, and a minor lesion in the nucleus preopticus periventricularis, it is likely that the NLT is the origin of the degenerative type B fibers in the proximal NH. The significance of these results is discussed in relation to the neuroendocrine regulation of the pituitary functions in goldfish.     

Active vitellogenesis in precocene-treated Locusta migratoria

Vitellogenesis continues unhindered during the first 2 days after precocene II treatment of vitellogenic female locusts bearing terminal oocytes up to 4 mm in length. The growth rate of these oocytes remains normal, but the corpora allata of treated females are inactive insofar as juvenile hormone synthesis is concerned. This response differs fundamentally from that occasioned by extirpation of the corpora allata, wherein oocyte growth ceases shortly there-after.     

High temperature and corticosteroid in the lymphocytes of domestic fowl (Gallus domesticus)

Exposure to an environmental temperature of 42° for 1 hr increased the concentration of endogenously produced corticosteroid bound in the cytoplasm and the quantity of corticosteroid in the nucleus of the lymphoid cells of the bursa of Fabricius, thymus, and spleen of young White Rock chickens. When heat episodes were applied 8–12 hr before the final episode, the increases of corticosteroid in the cells were suppressed relative to the number of preliminary heatings. These results demonstrate an in vivo relationship between environmentally induced corticosteroid response and the potential immunoreactive lymphoid cells.     

Ovarian cholesterol dynamics in teleost Channa punctatus (Bloch): Relationship with reproductive cycle and response to gonadotropins

Thin-layer chromatography was employed to separate free and esterified cholesterol from total ovarian cholesterol in the teleost Channa punctatus. Results clearly showed that free cholesterol was almost double the concentration (5.37 ± 0.32 nmol/mg) of esterified cholesterol (2.67 ± 0.26 nmol/mg). An annual profile of free and esterified cholesterol determined per ovary showed that both altered in different months of the year although fluctuation was more prominent with free cholesterol. In contrast, the study in terms of per milligramovarian tissue, showed fluctuation of free cholesterol only. Among the pituitaries collected in different reproductive stages, prespawning pituitary extract had maximum depletive effect on ovarian total and free cholesterol in comparison to others. Ovine LH (1 μg/100 g body wt) could effectively induce the depletion of ovarian free cholesterol; the most remark-able effect was obtained with a similar dose of SG-G100 but there was insignificant alteration of esterified cholesterol. In hypophysectomized fish, SG-G100 in increasing doses (1 to 10 μg/100 g body wt) evoked a linear response in regard to the ovarian free cholesterol depletion. These findings contrast strikingly with reports on the mammalian ovary.     

Breeding condition, temperature, and the regulation of salt and water by pituitary hormones in the red-spotted newt, Notophthalmus viridescens

In vivo integumental transepithelial potential (TEP) in the Eastern red-spotted newt (Notophthalmus viridescens) increases linearly with external [Na+] from 0.1 to 10 mM and is anion independent. Both integumental TEP and osmotic permeability are higher in laboratory-conditioned (LC, terrestrial) than in breeding-condition (BC, aquatic) newts at temperatures of 5-25 degrees. Prolactin (PRL) treatment of LC newts decreased both TEP and rate of water uptake. Arginine vasotocin (AVT) treatment resulted in a substantial increase in water uptake in LC newts, while little or no AVT response was seen in PRL-treated or BC newts. Hypophysectomy (HX) or ergocryptine treatment increased TEP in BC newts, whereas HX + PRL maintained TEP at control levels. Although ergocryptine and HX were without effect on water uptake in BC newts kept at 5 degrees for 9 days, HX + ACTH increased water uptake. HX produced a substantial fall in serum [Na+] in BC newts, while either PRL or ACTH replacement elevated serum [Na+]. Combined ACTH and PRL treatment returned serum [Na+] to control levels. These data suggest that high endogenous prolactin plays a significant role in maintaining serum [Na+] and integumental permeability and transport characteristics in breeding-condition N. viridescens. Although PRL and ACTH are both sodium retaining in the aquatic breeding stage, these two hormones promote opposite effects on the integument; PRL decreases both water uptake and integumental TEP, whereas ACTH (presumably acting through the adrenals) increases water uptake and possibly TEP.     

Effect of the presence of females on the pituitary-testicular activity in male Japanese quail (Coturnix coturnix japonica)

Five experiments were carried out to study the role of the presence of a female on the reproductive endocrinology of male Japanese quail. In the first three experiments, exposure of an adult male raised in long days to a female for 10 min or 1 week failed to increase plasma testosterone and LH levels; in fact a significant transitory decrease in plasma testosterone was observed, associated with a preceding increase in plasma corticosterone. These changes are interpreted as a result of the stress caused by repeated bleeding or by the continuous presence of a female in a limited space. In the last two experiments, an increase in the maturation rate of immature males could be observed in birds maintained in the continuous presence of females by comparison with birds kept in isolation. The paired males had larger cloacal glands and testes and higher plasma levels of testosterone and LH than the isolated ones. This effect of the female was observed in long days (16L:8D) as well as in marginally stimulating short days (12L:12D).     

Age-related changes in the water and electrolyte metabolism in male rats of the Wistar/TW strain

The changes of water intake, urinary volume, ratio of the water intake to the urinary output (water turnover rate) and urinary concentrations of sodium, potassium and chloride ions were investigated in male rats of the inbred Wistar/Tw strain during aging. Marked increase of the water intake, the urinary output and the water turnover rate as well as marked decrease of the urinary electrolyte concentrations were observed at the age of over 16 months. First signs of increase in the water intake and the water turnover rate were observed at the age of 12-13 months, but the initial reduction of urinary electrolyte concentrations was detected as early as at 7-9 months of age.     

In vivo regulation of steroidogenesis by ovine gonadotropins in male and female mudpuppies,Necturus maculosus Rafinesque

Using steroid radioimmunoassay the in vivo steroidogenic responses of male and female mudpuppies (Necturus maculosus) to single injections of ovine FSH and LH were investigated. In males, the major effect of LH was to stimulate testosterone and 5α-dihydrotestosterone secretion within 2 hr of injection, with a lesser effect on estrogen (estrone and estradiol-17β) secretion. In contrast, FSH primarily stimulated secretion of estrogens. The effects of both FSH and LH on steroid secretion in males were dose related. Castration of males reduced plasma androgen, and estrogen to very low or nondetectable levels and abolished the steroidogenic response to LH. Interrenalectomy prevented a postcastration rise in the progesterone level. In females, FSH did not stimulate steroid secretion but LH increased plasma androgens. Homologous pituitary extracts also stimulated plasma levels of all gonadal steroids measured.     

Endocrine regulation of ovarian development in the autogenous mosquito, Aedes atropalpus

The mosquito Aedes atropalpus is autogenous, i.e., normal ovarian development proceeds in adult females without any exogenous source of protein. Using a radioimmunoassay (RIA) for ecdysone, it was found that the titer of antigen-positive material in whole-body extracts of adult females rose from about 150 pg equiv of β-ecdysone per female at eclosion to an average peak value of 323 pg 32 hr later. The titer then dropped to approximately 100 pg over the next 16 hr. A comparison of the RIA titer with follicle size, vitellin deposition into oocytes, and total extractable soluble proteins from ovaries suggested a causal relationship between the hormone titer and normal ovarian development. Preliminary data also suggest that the ovary is a source of ecdysteroid in these adult animals. Decapitation of adult females from 0 to 4 hr after eclosion completely prevented ovarian development. Animals decapitated 30 min after eclosion and then assayed for ecdysteroid content 32 hr later revealed a hormonal titer 75% less than normal intact controls. An analysis of the 32-hr RIA-positive material by high-pressure liquid chromatography confirmed the presence of β-ecdysone but did not detect any α-ecdysone. Ovarian deficiencies incurred by decapitation were partially restored by injection with β-ecdysone or by topical application of juvenile hormone.     

Age-related changes in pituitary/thyroid function in chickens

Pituitary cyclic AMP-dependent protein kinase (cAMP-PK) activity ratios are not increased in response to thyrotropin-releasing hormone (TRH) in old chickens (retired White Leghorn breeders). This reduced responsiveness may be due to reduced hypothalamic function, reduced thyrotrope function, or to a reduction in TRH membrane receptors. The thyroid cAMP-PK activity ratio of old males does not respond to TRH treatment but the thyroid of old females does have an increased activity ratio ater TRH injection. Both males and females have a much higher basal cAMP-PK activity ratio than young birds. This higher basal level is thought to be due to an increased thyroid-stimulating hormone (TSH) stimulation, and the failure of old males to increase activity after TRH injections may be due to a loss of the thyroid's ability to respond to direct TRH stimulation.     

Corticosteroid receptors in the kidney of chick embryo. III. Nature, properties, and ontogeny of aldosterone receptor

An aldosterone receptor in the cytosol from kidney of chick embryos which had a sedimentation coefficient of 8.2 S and a molecular weight higher than 100,000 was identified. Kinetic analysis at 4 degrees revealed a rapid association of the hormone to the receptor that followed second-order reaction kinetics and a dissociation of pseudo-first-order reaction kinetics. The association (ka) and dissociation (kd) rate constants were, respectively, 4.94 X 10(5) M-1 sec-1 and 8.33 X 10(-6) sec-1. From their ratio a KA value of 5.9 X 10(10) M-1 was calculated. In a series of experiments performed with kidneys of 17-day-old embryos, the KA at equilibrium, obtained from the Scatchard plot, was 3.1 +/- 1.2 X 10(8) M-1, whereas the Nmax was 172 +/- 14 fmol/mg protein. Competition studies with various steroids demonstrated that corticosterone had an affinity for the receptor close to that of aldosterone, thus suggesting a degree of resemblance of the mineralo- and glucocorticoid receptors in the chick embryo. However, the profiles of the binding affinities and capacities during the embryogenesis showed that the aldosterone-binding sites had a pattern completely different from that of the glucocorticoid receptor, indicating that the two receptors are most likely separate entities.     

Concentration of triiodothyronine in the sera of the sea lamprey, Petromyzon marinus, and the brook lamprey, Lampetra lamottenii, at various phases of the life cycle

Triiodothyronine (T3) was measured by double antibody radioimmunoassay in the sera of individuals of all stages of the life cycle in the sea lamprey, Petromyzon marinus, and in larval (ammocoetes) and metamorphosing individuals of the brook lamprey, Lampetra lamottenii. There was no significant difference in results from the two species. Although serum T3 concentration did not vary significantly in ammocoetes maintained at the same temperature throughout the year, concentrations in ammocoetes displayed an inverse response to fluctuations in water temperature. Ammocoetes maintained at temperatures of 7 to 10 degrees and 19 to 21 degrees exhibited significantly different circulating T3 levels of 2400 and 1550 ng dl-1, respectively. Therefore, it is expected that there may be some seasonal variation in the levels of the hormone in ammocoetes in their natural environment. A precipitous collapse of the serum levels of T3 was observed by an early stage of metamorphosis (100 ng dl-1) and no significant difference in levels was observed throughout the remaining period of this phase. Young parasitic adults and adult lampreys collected during their spawning migration (upstream migrants) exhibited circulating T3 concentrations of 40 and 30 ng dl-1, respectively. T3 levels in some upstream migrants were below the limit of detection (less than 5 ng dl-1). The metamorphic decline in serum T3 may simply mark the beginning of the lower T3 levels that characterizes the adult serum.     

Circulating thyroid hormone concentrations in relation to seasonal events in the male Indian garden lizard, Calotes versicolor

Circulating thyroxine (T4) and triiodothyronine (T3) were estimated by RIA in plasma samples of Calotes versicolor collected over a period of 28 months. Monthly observations on seasonal events, viz., molting, whole body O2 consumption, abdominal fat deposition, and testes weight were also made. Plasma T4 ranged from 0.16 to 4.63 ng/ml and T3 from 0 to 1.76 ng/ml during the course of the year. Peaks of T4 and T3 could be related with prehibernating (October/November) and posthibernating (March/April) periods, when scale shedding and O2 consumption were also enhanced. Maximum abdominal fat deposition was found in prehibernatory months (October/November). Thyroid hormone levels declined sharply with entry into hibernation. Abdominal fat deposits gradually decreased, being almost depleted by the end of hibernation. The gradual increase in thyroid hormone concentration in the later half of hibernation may be associated with increased lipolysis and preparation of the oncoming reproductive period. Although T4 and T3 profiles were apparently parallel, discrepancies were observed. Posthibernatory peak of T4 was maintained much longer than that of T3 which declined earlier. Also the rise in the prehibernatory T3 concentration was delayed as compared to that in T4.     

Effects of urotensin I on the isolated dorsal aorta of the chukar, Alectoris graeca.

The effects of purified urotensin I (UI) from the Catostomus urophysis on arterial blood vessels were examined using isolated dorsal aortic strips from a bird, the chukar, Alectoris graeca. UI at concentrations greater than 1 mU/ml produced dose-dependent relaxation. The same effect was more clearly evident in aortic strips contracted by norepinephrine beforehand. The relaxation following UI was not inhibited by propranolol (10⁻⁶M) or atropine (10⁻⁶M), which markedly reduced the relaxation induced by isoproterenol or acetylcholine, respectively. The dose-dependent contractile response to norepinephrine was not significantly affected by UI, whereas it was competitively blocked by phentolamine (10⁻⁷, 10⁻⁶M). Arginine vasotocin (AVT) at low concentrations (0.1 to 1.0 ng/ml) caused relaxation in 50% of the preparations and slight contraction in the other preparations; at higher concentrations AVT caused contraction in all preparations. In comparison with the response to UI, AVT-induced relaxation developed rapidly. UI at concentrations ranging from 10 to 100 mU/ml also elicited relaxation of the strips contracted by AVT. These results suggest that UI-induced aortic relaxation is not mediated through adrenergic or cholinergic receptors, and that it is unlikely that UI and AVT act by similar mechanisms.