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Biomedical subjects

G V Callard

Publications and source records attributed to G V Callard.

At least 73 records · Page 4Linked to original sources
PubMed1983

The differentiation of Leydig cells, steroidogenesis, and the spermatogenetic wave in the testis of Necturus maculosus.

The study of seminiferous tubule--Leydig cell interactions in relation to specific germ cell stages during the cycle of the seminiferous epithelium is extremely difficult in most mammalian species due to the continual presence of different spermatogenetic stages in the testis from the onset of puberty. The problem is also compounded by the uniform distribution of both seminiferous tubules and interstitial tissue throughout the entire testis. This difficulty can be circumvented, however, by studying certain species where there is a topographical distribution of germ cell stages within the testis. The urodele amphibian Necturus maculosus exhibits a breeding cycle during which a longitudinal wave of spermatogenesis occurs along the length of the testis, resulting in a spatial and temporal segregation of differentiating germ cells. Moreover, this topographical pattern of spermatogenesis is also reflected in the degree of development of adjacent Leydig cells. This anatomical arrangement allows distinct testicular regions to be obtained using a dissecting microscope. The isolated zones, containing germ cells and Leydig cells in various stages of development, were analyzed for 17 alpha-hydroxylase, C-17,20-lyase, and aromatase activities (key enzymes for the synthesis of androgens and estrogen), estrogen binding, and cytochrome P-450 content. Functional parameters were then correlated with the morphology of Leydig cells in the various zones observed by both light and electron microscopy. It was found that there existed a distinct correlation between the state of differentiation of the leydig cells, their steroidogenic potential, and the distribution of estrogen receptors. These results in Necturus indicate indicate in this species, at least, the steroidal microenvironment of different germ cell associations may be quite specific.

Androgens↗
PubMed1983

Identification of androgen metabolic pathways in the brain of little brown bats (Myotis lucifugus): sex and seasonal differences.

During the reproductive cycle of the little brown bat (Myotis lucifugus), the display of sexual behavior is out of synchrony with maximal gonadal development, suggesting that changes in plasma hormone levels per se are not the primary determinants of neural responsiveness. In the present study, we investigated pathways of androgen metabolism in the brain of the bat and compared brain aromatase activity in spring and in autumn. Tissue homogenates were incubated with [3H] androstenedione in the presence of an NADH/NADPH-generating system, and radiolabeled products isolated and their authenticity verified by recrystallization to constant specific activity. Aromatase was identified in preoptic/hypothalamic and adjacent limbic areas, and 5 alpha-reductase was detected in all major brain divisions. No gender-based differences in brain aromatase were seen in spring just prior to emergence from hibernation; however, activity was greater in males than in females when measured during maximal breeding activity in autumn. In addition, seasonal changes were evident in the preoptic/hypothalamic regions of both sexes (spring greater than autumn), although estrone yields from limbic tissues remained constant. Thus, in seasonally breeding species, the rate at which circulating androgen is converted to estrogen or other biologically active metabolites in the brain itself may be an important determinant of behavioral or feedback responsiveness.

Androgens↗
PubMed1983

Identification of aromatase activity in rodent pituitary cell strains.

To date, biochemical evidence has been presented for hypophysial aromatization in only one species, a teleost fish, although the pituitary glands of several mammals have been reported to be aromatase negative. To reinvestigate this problem, established clonal strains of rodent pituitary cells (GH3, GH4C1, and AtT20/D16) were incubated at 37 C for 6-48 h in serum-less medium containing [7-3H]androstenedione. Radiolabeled metabolites were isolated by solvent extraction, thin layer chromatography, and phenolic partition. The authenticity of the estrogenic products in both cells and incubation medium was verified by methylation and recrystallization to constant specific activity. Measurement of androgen metabolites was also validated by recrystallization of selected samples. Authentic estrone and 17 beta-estradiol were identified in cultures of the two PRL- and GH-secreting clones, and there were strain differences in the quantity of estrogen produced (GH3 greater than GH4C1). Under the same conditions, aromatization was not detectable in the ACTH-secreting line (AtT20/D16). A time-yield analysis of androgen metabolism in GH4C1 cells showed that aromatization was linear for 12 h after labeling, but that substrate was diverted mainly to 5 alpha-reducing pathways. Large amounts of highly polar metabolites accumulated 24 and 48 h after the addition of [3H]androgen, and subsequent hydrolysis revealed that these were sulfo- and glucuronoconjugates. The metabolic fate of estrogen in GH4C1 cultures was investigated indirectly by adding a radioinert estrone trap together with the radiolabeled androgen substrate and was also tested in separate cultures by adding [3H]estrone and [3H]estradiol directly. Although the two estrogens were interconverted, there was no evidence that formed or added estrogen was extensively metabolized or conjugated. We conclude that the expression of aromatase activity in hypophysial cells is not a property of all transformed lines but may be dictated by the secretory cell type. Although low relative to other metabolites, estrogen yields in cultured GH cells resemble production in primary cultures derived from other tissues known to be estrogen targets, including the hypothalamus.

Androstenedione↗
PubMed1981

Aromatization is cyclic AMP-dependent in cultured brain cells.

When added to primary cultures of adult turtle (Chrysemys) brain, dibutyryl cyclic AMP increases estrogen yields from [3H]testosterone in a dose- and time-dependent manner, although rat FSH, rat LH, and dibutyryl cyclic GMP are relatively ineffective. A phosphodiesterase inhibitor (MIX) by itself increases estrogen yields from the substrate and potentiates the response to dibutyryl cyclic AMP.

1-Methyl-3-isobutylxanthine↗
PubMed1980

Aromatization of androgen to estrogen by cultured turtle brain cells.

Cells obtained from turtle forebrain can be maintained in culture for at least 3 weeks. The cells are capable of aromatizing [3H]androstenedione and [3H]testosterone to estrone and estradiol and several C-19 metabolites. There are marked differences in the quality and quantity of the products formed from the two substrates. Conditions in living cells favor accumulation of 17-hydroxylated steroids. Aromatase activity as measured by estrogen yield increases with time in culture. Estrogen content of 14-day-old cultures may be enhanced or reduced by addition of natural or synthetic steroids. This system may provide a model for studying the regulation of brain aromatization, an essential step in the expression of androgen action on certain behavioral and neuroendocrine responses.

Androstenedione↗
PubMed1978

Phylogenetic distribution of aromatase and other androgen-converting enzymes in the central nervous system.

Metabolism of [3H]androstenedione was studied in brain tissue homogenates of opossum, bird, snake, sea turtle, urodele amphibian, teleost, shark, skate, hagfish, and lobster. Estrone, 17 beta-estradiol, or 17 alpha-estradiol was formed by central neural tissues of all species, with the exception of the opossum, hagfish, and lobster. Aromatase activity was concentrated in the forebrain, although some estrogen was synthesized by mid- or hindbrain homogenates of two lower vertebrates (teleost and skate) and the newly hatched chick. 5 alpha-Androstanedione (5 alpha-androstane-3,17-dione) or 5 alpha-dihydrotestosterone were products of metabolism in several nonmammalian vertebrates and in the invertebrate central nervous system also. 5 alpha-Reductase was found in all major brain divisions. These and previously reported comparative studies indicate that the ability to aromatize and otherwise transform androgen substrates is a primitive characteristic of the brain that has been widely conserved phylogenetically.

Androstenedione↗
PubMed1977

Identification of aromatase in the reptilian brain.

The present study tests the hypothesis that brain aromatase is an "ancient" property of nervous tissue and may be identified in homologues of the limbic system in a non-mammalian vertebrate, the turtle Chrysemys picta. Tissue homogenates (180 mg wet weight/2 ml) were incubated with [7 alpha-3H]androstenedione and cofactors for 60 min at 37 C. Estrone (E1) was isolated and characterized by thin layer chromatography, methylation and recrystallization to constant specific activity. No estradiol-17 beta was detected. Aromatase was found only in the forebrain but was diffusely distributed throughout this major brain division. No other neural or non-neural tissues, including mid- and hindbrain structures, testis, and ovary, synthesized detectable quantities of E1 in our system. The strio-amygdaloid complex of both sexes synthesized more E1 per unit weight than the preoptic-hypothalamic area (POA-HTH) or other forebrain structures. It is possible that the conversion of androgen to estrogen has biological significance in this species since the reaction occurred throughout the physiological temperature range experienced during activity in nature and sex differences in brain aromatase activity during the breeding season were apparent. These experiments in Chrysemys demonstrate that the synthesis of estrogen from androgen by the brain is not limited to mammals, but also occurs at a more primitive level of phylogenetic development. Restriction of aromatase to forebrain structures of the turtle is consistent with the neuroanatomic distribution of enzyme activity in the limbic system of mammals. Estrogen yield from adult turtle brain incubates (3.2-22.6 pmol/g) is more like that reported for fetal (2.7-33 pmol/g) than for adult (0.1-1.9 pmol/g) mammals. We suggest that the in situ synthesis of estrogen by the central nervous system has its orgins early in vertebrate evolution and may be a primitive characteristic of brain-steroid interactions that regulate physiological and behavioral sex in vertebrates.

Amygdala↗