Biology subjects

Gorelick, D. A.

Publications and source records attributed to Gorelick, D. A..

5 recordsLinked to original sources

bioRxiv2018

ahr2, but not ahr1a or ahr1b, is required for craniofacial and fin development and TCDD-dependent cardiotoxicity in zebrafish

The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor that binds environmental toxins and regulates gene expression. AHR also regulates developmental processes, like craniofacial development and hematopoiesis, in the absence of environmental exposures. Zebrafish have three paralogues of AHR: ahr1a, ahr1b and ahr2. Adult zebrafish with mutations in ahr2 exhibited craniofacial and fin defects. However, the degree to which ahr1a and ahr1b influence ahr2 signaling and contribute to fin and craniofacial development are not known. We compared morphology of adult ahr2 mutants and ahr1a/ahr1b single and double mutant zebrafish. We found that ahr1a/ahr1b single and double mutants were morphologically normal while ahr2 mutant zebrafish demonstrated fin and craniofacial malformations. At 5 days post fertilization, both ahr1a/ahr1b and ahr2 mutant larvae were normal, suggesting that adult phenotypes are due to defects in maturation or maintenance. AHR was shown to interact with estrogen receptor alpha, yet it is not known whether these interactions are constitutive or dependent on ahr1 genes. To determine whether estrogen receptors are constitutive cofactors for AHR signaling, we used genetic and pharmacologic techniques to analyze TCDD-dependent toxicity in estrogen receptor and ahr mutant embryos. We found that embryos with mutations in ahr1a/ahr1b or estrogen receptor genes are susceptible to TCDD toxicity while ahr2 mutant embryos are TCDD-resistant. Moreover, pharmacologic blockade of nuclear estrogen receptors failed to prevent TCDD toxicity. These findings suggest that ahr1 genes do not have overlapping functions with ahr2 in fin and craniofacial development or TCDD-dependent toxicity, and that estrogen receptors are not constitutive partners of ahr2.

developmental biology↗

bioRxiv2018

G protein-coupled estrogen receptor is not required for sex determination or ovary function in zebrafish

Estrogens regulate vertebrate development and function through binding to nuclear estrogen receptors alpha and beta (ER, ER{beta}) and the G protein-coupled estrogen receptor (GPER). Studies in mutant animal models demonstrated that ER and ER{beta} are required for normal ovary development and function. However, the degree to which GPER signaling contributes to ovary development and function is less well understood. Previous studies using cultured fish oocytes found that estradiol inhibits oocyte maturation in a GPER-dependent manner, but whether GPER regulates oocyte maturation in vivo is not known. To test the hypothesis that GPER regulates oocyte maturation in vivo, we assayed ovary development and function in gper mutant zebrafish. We found that homozygous mutant gper embryos developed into male and female adults with normal sex ratios and fertility. Adult mutant fish exhibited normal secondary sex characteristics and fertility. Additionally, mutant ovaries were histologically normal. We observed no differences in the number of immature versus mature oocytes in mutant versus wild-type ovaries from both young and aged adults. Furthermore, expression of genes associated with sex determination and ovary function were normal in gper mutant ovaries compared to wild type. Our findings suggest that GPER is not required for sex determination, ovary development or fertility in zebrafish.

developmental biology↗

bioRxiv2017

Assaying uptake of endocrine disruptor compounds in zebrafish embryos and larvae

To study the effects of environmental endocrine disruptors (EEDs) on aquatic animals, embryos and larvae are typically incubated in water containing defined concentrations of EEDs. However, the amount of EED uptake into the animal is often difficult to determine. Using radiolabeled estradiol ([3H]E2), we previously developed a rapid, straightforward assay to measure estradiol uptake from water into zebrafish embryos and larvae. Here, we extend this approach to measure the uptake of two additional EEDs, bisphenol A (BPA) and ethinyl estradiol (EE2). As with E2, the uptake of each compound by individual larvae was low (< 6%), and increased with increasing concentration, duration, and developmental stage. We found that E2 and EE2 had similar uptake under equivalent exposure conditions, while BPA had comparatively lower uptake. One application of this assay is to test factors that influence EED uptake or efflux. It has been suggested that persistent organic pollutants (POPs) inhibit ABC transporters that may normally efflux EEDs and their metabolites, inducing toxicity in aquatic organisms. We measured [3H]E2 levels in zebrafish in the presence or absence of the POP PDBE-100, and cyclosporine A, a known inhibitor of ABC transporters. Neither chemical significantly affected [3H]E2 levels in zebrafish, suggesting that zebrafish maintain estradiol efflux in the presence of PDBE-100, independently of cyclosporine A-responsive transporters. These uptake results will be a valuable reference for EED exposure studies in developing zebrafish, and provide a rapid assay to screen for chemicals that influence estrogen-like EED levels in vivo.

pharmacology and toxicology↗

bioRxiv2017

Nuclear androgen receptor regulates testes organization and oocyte maturation in zebrafish

Androgens act through the nuclear androgen receptor (AR) to regulate gonad differentiation and development. In mice, AR is required for spermatogenesis, testis development and formation of external genitalia in males and oocyte maturation in females. However, the extent to which these phenotypes are conserved in nonmammalian vertebrates is not well understood, because mutations in AR have not been generated in any other species. Here, we generate zebrafish with a mutation in the ar gene and examine the role of AR on sexual differentiation and gonad development. We find that zebrafish AR is not required for male sexual differentiation, since a portion of ar mutants develop a testis. However, we show that in zebrafish, as in mice, AR is required for the development of secondary sexual characteristics and for proper organization of the testis in males and for oocyte maturation in females. Additionally, we find that zebrafish ar mutant males have functional, mature sperm present in their testis, but are infertile due to an inability to release sperm. These findings suggest that AR is required for male sexual development and fertility, but not essential for sexual differentiation in zebrafish. The ar mutant we developed will be useful for modeling human endocrine function in zebrafish.

developmental biology↗

bioRxiv2016

G protein-coupled estrogen receptor regulates heart rate in zebrafish embryos

Estrogens act by binding to estrogen receptors alpha and beta (ER, ER{beta}), ligand-dependent transcription factors that play crucial roles in sex differentiation, tumor growth and cardiovascular physiology. Estrogens also activate the G protein-coupled estrogen receptor (GPER), however the function of GPER in vivo is less well understood. Here we find that GPER is required to maintain normal heart rate in zebrafish embryos. Acute exposure to estrogens increased heart rate in wildtype and in ER and ER{beta} mutant embryos but not in GPER mutants. Nuclear estrogen receptor signaling remained normal in GPER mutant embryos, however GPER mutant embryos exhibited reduced basal heart rate while heart rate was normal in ER and ER{beta} mutants. We detected GPER transcript in discrete regions of the brain but not in the heart. In the brain, we observed gper expression in cells lacking nuclear estrogen receptor activity, suggesting that GPER acts in the brain to regulate heart rate independently of nuclear estrogen receptor signaling. Our results demonstrate that estradiol plays a previously unappreciated role in the acute modulation of heart rate during zebrafish embryonic development and that GPER functions as an autonomous estrogen receptor in vivo to regulate basal heart rate.

pharmacology and toxicology↗