Mediators and mechanisms responsible for the inhibitory modulation of trout intestinal smooth mus... more Mediators and mechanisms responsible for the inhibitory modulation of trout intestinal smooth muscle were examined using a series of putative mediators and substances known to modulate neurotransmission in mammalian systems. Frequency response relationships to transmural stimulation and concentration response relationships to 5-hydroxytryptamine, carbachol, and substance P were established on paired segments of rainbow trout intestine in vitro in the presence and absence of putative modulatory agents. Modulation of neurally-mediated contractions of trout intestine was achieved with dibutyryl cyclic AMP and forskolin, agents that increase intracellular levels of cyclic AMP. The effect appears to be at the level of the smooth muscle, since the adenylate cyclase activator, forskolin, inhibited muscarinic and serotoninergic contractions as well as transmurally stimulated contractions. Substance P-induced contractions were unaffected by forskolin. The endogenous agonists/neurotransmitters which would increase cyclic AMP levels in rainbow trout intestinal smooth muscle are as yet unknown. The effects do not appear to be modulated by vasoactive intestinal peptide (VIP), calcitonin, calcitonin gene-related peptide (CGRP), or agents that activate 13adrenoceptors. Prostaglandin E 2 (PGE2) and o%-adrenergic agonists are possible agents which will decrease contractility of the smooth muscle. They were only active in the proximal intestine and on transmurally stimulated contractions. The effects of both PGE 2 and c~2-agonists appear to be prejunctional, decreasing release of contractile neurotransmitters in the enteric nervous system.
The purpose of the RECD initiative is to: 1. Develop a framework for research and evaluation in e... more The purpose of the RECD initiative is to: 1. Develop a framework for research and evaluation in early child development; 2. Facilitate indicator development, data collection, and reporting for early child development; with a long term goal to; 3. Build capacity for expertise, resource support, and research in early child development for the government of PEI and the Children's Secretariat. Researchers from the University of Prince Edward Island (UPEI) Centre for Education Research (CER) partnered with the PEI Children's Secretariat to study current issues and challenges facing early child development in PEI. Following an extensive literature review , the Research in Early Child Development (RECD) Initiative was formed in October 2008 to develop a working research framework for early child development, and facilitate indicator development, data collection, and reporting for early child development, including the twelve networks of the PEI Children's Secretariat. The RECD initiative has a long term goal to build capacity for expertise, resource support, and research in Early Child Development, for the PEI Children's Secretariat and early child development community of PEI.
Aquatic toxicology (Amsterdam, Netherlands), Jan 11, 2008
A method to evaluate the expression of three hormone responsive genes, vitellogenin (estrogens), ... more A method to evaluate the expression of three hormone responsive genes, vitellogenin (estrogens), spiggin (androgens), and an androgen receptor (ARbeta) using real-time PCR in threespine stickleback is presented. Primers were designed from previously characterised spiggin and ARbeta sequences, while a homology cloning strategy was used to isolate a partial gene sequence for stickleback vitellogenin (Vtg). Spiggin mRNA was significantly higher in kidneys of field-caught males compared to females by greater than five orders of magnitude while ARbeta levels were only 1.4-fold higher in males. Female fish had four order of magnitude higher liver Vtg expression than wild-captured males. To determine the sensitivity of these genes to induction by hormones, male and female sticklebacks were exposed to 1, 10 and 100 ng/L of methyltestosterone (MT) or estradiol (E2) in a flow-through exposure system for 7 days. Spiggin induction in females, and Vtg induction in males were both detectable at 1...
Fish Physiology and Biochemistry - FISH PHYSIOL BIOCHEM, 1999
The contractility of intestinal smooth muscle of Atlantic salmon (Salmo salar) acclimatized to te... more The contractility of intestinal smooth muscle of Atlantic salmon (Salmo salar) acclimatized to temperatures of 7 and 14 °C was compared over a prolonged period of time (9 months) in freshwater. Initial studies were carried out on intestine isolated from Atlantic salmon to estimate optimal conditions for contractility. Subsequent studies were carried out with the neurotransmitter/agonist 5-HT and TS, which activates neuronal elements in the intestine, in a Krebs-Henseleit solution containing HEPES buffer maintained at pH 7.85 and at the temperature of the experimental group (i.e., 7 or 14 °C). There was a significant interaction with time and temperature for the maximal response of 5-HT (p=0.005), effective concentration producing 50% of the maximal response (EC50) to 5-HT (p=0.026) and maximal response to TS (p=0.002), demonstrating that gastrointestinal contractility of Atlantic salmon is altered by both time (month) and temperature. No significant changes were found with the effec...
The presence of unidentified estrogens and androgens in effluents from pulp and paper mills is we... more The presence of unidentified estrogens and androgens in effluents from pulp and paper mills is well documented. However, their role in effluent effects on fish reproduction remains unclear. The objective of this study was to investigate the hypothesis that reproductive impacts of a modern pulp mill effluent are mediated by androgens and/or estrogens in the effluent. Male and female threespine stickleback were exposed to biologically treated Canadian bleached kraft mill effluent under flow-through conditions in the laboratory at 0, 1, 10 and 100% (v/v) dilutions. After 7 and 21 d of exposure, steroidogenesis was assessed using in vitro incubations of gonadal tissue in both males and females. mRNA expression of the estrogen-regulated gene vitellogenin, and the androgen-responsive gene spiggin were assessed using quantitative RT-PCR in the livers of male and posterior kidneys of female stickleback, respectively. Hepatic 7-ethoxyresorufin-O-deethylase (EROD) activity was assessed in both sexes. Effluent extracts were examined for estrogenic and androgenic bioactivity using receptor binding bioassays, and were screened for pulp and paper related extractives and steroidal androgens using GC-MS. This effluent up-regulated spiggin mRNA in the kidney of female stickleback at 10% and 100% (v/v) effluent at 21 d, but not at 7 d of exposure. This change at the mRNA expression of the gene was associated with an increase in cell height in kidney proximal tubule epithelial cells at 100% effluent after both 7 and 21 d. Liver vitellogenin mRNA in male stickleback was not induced at either 7 or 21 d. EROD was induced at 10 and 100% after 21 d of exposure in both sexes, but not after 7 d of exposure. Despite evidence of exposure to androgens, there was no reduction in steroidogenic capacity at any effluent dilution. Effluent extracts were capable of eliciting the displacement of androgens and estrogens from receptors, but androgenic potency was 4-fold greater. A screen of more than 30 androgenic androstane steroids showed no detections. Hence, the androgenic constituents in this particular effluent remain unknown.
A method to evaluate the expression of three hormone responsive genes, vitellogenin (estrogens), ... more A method to evaluate the expression of three hormone responsive genes, vitellogenin (estrogens), spiggin (androgens), and an androgen receptor (AR) using real-time PCR in threespine stickleback is presented. Primers were designed from previously characterised spiggin and AR sequences, while a homology cloning strategy was used to isolate a partial gene sequence for stickleback vitellogenin (Vtg). Spiggin mRNA was significantly higher in kidneys of field-caught males compared to females by greater than five orders of magnitude while AR levels were only 1.4-fold higher in males. Female fish had four order of magnitude higher liver Vtg expression than wild-captured males. To determine the sensitivity of these genes to induction by hormones, male and female sticklebacks were exposed to 1, 10 and 100 ng/L of methyltestosterone (MT) or estradiol (E2) in a flow-through exposure system for 7 days. Spiggin induction in females, and Vtg induction in males were both detectable at 10 ng/L of MT and E2, respectively. MT exposure did not induce AR expression in the kidneys of female stickleback. In vitro gonadal steroid hormones production was measured in testes and ovaries of exposed stickleback to compare gene expression endpoints to an endpoint of hormonal reproductive alteration. Reduction in testosterone production in ovaries at all three MT exposure concentrations, and ovarian estradiol synthesis at the 100 ng/L exposure were the only effects observed in the in vitro steroidogenesis for either hormone exposure. Application of these methods to assess both androgenic, estrogenic, and anti-steroidogenic properties of environmental contaminants in a single fish species will be a valuable tool for identifying compounds causing reproductive dysfunction in fishes.
Mediators and mechanisms responsible for the inhibitory modulation of trout intestinal smooth mus... more Mediators and mechanisms responsible for the inhibitory modulation of trout intestinal smooth muscle were examined using a series of putative mediators and substances known to modulate neurotransmission in mammalian systems. Frequency response relationships to transmural stimulation and concentration response relationships to 5-hydroxytryptamine, carbachol, and substance P were established on paired segments of rainbow trout intestine in vitro in the presence and absence of putative modulatory agents. Modulation of neurally-mediated contractions of trout intestine was achieved with dibutyryl cyclic AMP and forskolin, agents that increase intracellular levels of cyclic AMP. The effect appears to be at the level of the smooth muscle, since the adenylate cyclase activator, forskolin, inhibited muscarinic and serotoninergic contractions as well as transmurally stimulated contractions. Substance P-induced contractions were unaffected by forskolin. The endogenous agonists/neurotransmitters which would increase cyclic AMP levels in rainbow trout intestinal smooth muscle are as yet unknown. The effects do not appear to be modulated by vasoactive intestinal peptide (VIP), calcitonin, calcitonin gene-related peptide (CGRP), or agents that activate 13adrenoceptors. Prostaglandin E 2 (PGE2) and o%-adrenergic agonists are possible agents which will decrease contractility of the smooth muscle. They were only active in the proximal intestine and on transmurally stimulated contractions. The effects of both PGE 2 and c~2-agonists appear to be prejunctional, decreasing release of contractile neurotransmitters in the enteric nervous system.
Mediators and mechanisms responsible for the inhibitory modulation of trout intestinal smooth mus... more Mediators and mechanisms responsible for the inhibitory modulation of trout intestinal smooth muscle were examined using a series of putative mediators and substances known to modulate neurotransmission in mammalian systems. Frequency response relationships to transmural stimulation and concentration response relationships to 5-hydroxytryptamine, carbachol, and substance P were established on paired segments of rainbow trout intestine in vitro in the presence and absence of putative modulatory agents. Modulation of neurally-mediated contractions of trout intestine was achieved with dibutyryl cyclic AMP and forskolin, agents that increase intracellular levels of cyclic AMP. The effect appears to be at the level of the smooth muscle, since the adenylate cyclase activator, forskolin, inhibited muscarinic and serotoninergic contractions as well as transmurally stimulated contractions. Substance P-induced contractions were unaffected by forskolin. The endogenous agonists/neurotransmitters which would increase cyclic AMP levels in rainbow trout intestinal smooth muscle are as yet unknown. The effects do not appear to be modulated by vasoactive intestinal peptide (VIP), calcitonin, calcitonin gene-related peptide (CGRP), or agents that activate 13adrenoceptors. Prostaglandin E 2 (PGE2) and o%-adrenergic agonists are possible agents which will decrease contractility of the smooth muscle. They were only active in the proximal intestine and on transmurally stimulated contractions. The effects of both PGE 2 and c~2-agonists appear to be prejunctional, decreasing release of contractile neurotransmitters in the enteric nervous system.
The purpose of the RECD initiative is to: 1. Develop a framework for research and evaluation in e... more The purpose of the RECD initiative is to: 1. Develop a framework for research and evaluation in early child development; 2. Facilitate indicator development, data collection, and reporting for early child development; with a long term goal to; 3. Build capacity for expertise, resource support, and research in early child development for the government of PEI and the Children's Secretariat. Researchers from the University of Prince Edward Island (UPEI) Centre for Education Research (CER) partnered with the PEI Children's Secretariat to study current issues and challenges facing early child development in PEI. Following an extensive literature review , the Research in Early Child Development (RECD) Initiative was formed in October 2008 to develop a working research framework for early child development, and facilitate indicator development, data collection, and reporting for early child development, including the twelve networks of the PEI Children's Secretariat. The RECD initiative has a long term goal to build capacity for expertise, resource support, and research in Early Child Development, for the PEI Children's Secretariat and early child development community of PEI.
Aquatic toxicology (Amsterdam, Netherlands), Jan 11, 2008
A method to evaluate the expression of three hormone responsive genes, vitellogenin (estrogens), ... more A method to evaluate the expression of three hormone responsive genes, vitellogenin (estrogens), spiggin (androgens), and an androgen receptor (ARbeta) using real-time PCR in threespine stickleback is presented. Primers were designed from previously characterised spiggin and ARbeta sequences, while a homology cloning strategy was used to isolate a partial gene sequence for stickleback vitellogenin (Vtg). Spiggin mRNA was significantly higher in kidneys of field-caught males compared to females by greater than five orders of magnitude while ARbeta levels were only 1.4-fold higher in males. Female fish had four order of magnitude higher liver Vtg expression than wild-captured males. To determine the sensitivity of these genes to induction by hormones, male and female sticklebacks were exposed to 1, 10 and 100 ng/L of methyltestosterone (MT) or estradiol (E2) in a flow-through exposure system for 7 days. Spiggin induction in females, and Vtg induction in males were both detectable at 1...
Fish Physiology and Biochemistry - FISH PHYSIOL BIOCHEM, 1999
The contractility of intestinal smooth muscle of Atlantic salmon (Salmo salar) acclimatized to te... more The contractility of intestinal smooth muscle of Atlantic salmon (Salmo salar) acclimatized to temperatures of 7 and 14 °C was compared over a prolonged period of time (9 months) in freshwater. Initial studies were carried out on intestine isolated from Atlantic salmon to estimate optimal conditions for contractility. Subsequent studies were carried out with the neurotransmitter/agonist 5-HT and TS, which activates neuronal elements in the intestine, in a Krebs-Henseleit solution containing HEPES buffer maintained at pH 7.85 and at the temperature of the experimental group (i.e., 7 or 14 °C). There was a significant interaction with time and temperature for the maximal response of 5-HT (p=0.005), effective concentration producing 50% of the maximal response (EC50) to 5-HT (p=0.026) and maximal response to TS (p=0.002), demonstrating that gastrointestinal contractility of Atlantic salmon is altered by both time (month) and temperature. No significant changes were found with the effec...
The presence of unidentified estrogens and androgens in effluents from pulp and paper mills is we... more The presence of unidentified estrogens and androgens in effluents from pulp and paper mills is well documented. However, their role in effluent effects on fish reproduction remains unclear. The objective of this study was to investigate the hypothesis that reproductive impacts of a modern pulp mill effluent are mediated by androgens and/or estrogens in the effluent. Male and female threespine stickleback were exposed to biologically treated Canadian bleached kraft mill effluent under flow-through conditions in the laboratory at 0, 1, 10 and 100% (v/v) dilutions. After 7 and 21 d of exposure, steroidogenesis was assessed using in vitro incubations of gonadal tissue in both males and females. mRNA expression of the estrogen-regulated gene vitellogenin, and the androgen-responsive gene spiggin were assessed using quantitative RT-PCR in the livers of male and posterior kidneys of female stickleback, respectively. Hepatic 7-ethoxyresorufin-O-deethylase (EROD) activity was assessed in both sexes. Effluent extracts were examined for estrogenic and androgenic bioactivity using receptor binding bioassays, and were screened for pulp and paper related extractives and steroidal androgens using GC-MS. This effluent up-regulated spiggin mRNA in the kidney of female stickleback at 10% and 100% (v/v) effluent at 21 d, but not at 7 d of exposure. This change at the mRNA expression of the gene was associated with an increase in cell height in kidney proximal tubule epithelial cells at 100% effluent after both 7 and 21 d. Liver vitellogenin mRNA in male stickleback was not induced at either 7 or 21 d. EROD was induced at 10 and 100% after 21 d of exposure in both sexes, but not after 7 d of exposure. Despite evidence of exposure to androgens, there was no reduction in steroidogenic capacity at any effluent dilution. Effluent extracts were capable of eliciting the displacement of androgens and estrogens from receptors, but androgenic potency was 4-fold greater. A screen of more than 30 androgenic androstane steroids showed no detections. Hence, the androgenic constituents in this particular effluent remain unknown.
A method to evaluate the expression of three hormone responsive genes, vitellogenin (estrogens), ... more A method to evaluate the expression of three hormone responsive genes, vitellogenin (estrogens), spiggin (androgens), and an androgen receptor (AR) using real-time PCR in threespine stickleback is presented. Primers were designed from previously characterised spiggin and AR sequences, while a homology cloning strategy was used to isolate a partial gene sequence for stickleback vitellogenin (Vtg). Spiggin mRNA was significantly higher in kidneys of field-caught males compared to females by greater than five orders of magnitude while AR levels were only 1.4-fold higher in males. Female fish had four order of magnitude higher liver Vtg expression than wild-captured males. To determine the sensitivity of these genes to induction by hormones, male and female sticklebacks were exposed to 1, 10 and 100 ng/L of methyltestosterone (MT) or estradiol (E2) in a flow-through exposure system for 7 days. Spiggin induction in females, and Vtg induction in males were both detectable at 10 ng/L of MT and E2, respectively. MT exposure did not induce AR expression in the kidneys of female stickleback. In vitro gonadal steroid hormones production was measured in testes and ovaries of exposed stickleback to compare gene expression endpoints to an endpoint of hormonal reproductive alteration. Reduction in testosterone production in ovaries at all three MT exposure concentrations, and ovarian estradiol synthesis at the 100 ng/L exposure were the only effects observed in the in vitro steroidogenesis for either hormone exposure. Application of these methods to assess both androgenic, estrogenic, and anti-steroidogenic properties of environmental contaminants in a single fish species will be a valuable tool for identifying compounds causing reproductive dysfunction in fishes.
Mediators and mechanisms responsible for the inhibitory modulation of trout intestinal smooth mus... more Mediators and mechanisms responsible for the inhibitory modulation of trout intestinal smooth muscle were examined using a series of putative mediators and substances known to modulate neurotransmission in mammalian systems. Frequency response relationships to transmural stimulation and concentration response relationships to 5-hydroxytryptamine, carbachol, and substance P were established on paired segments of rainbow trout intestine in vitro in the presence and absence of putative modulatory agents. Modulation of neurally-mediated contractions of trout intestine was achieved with dibutyryl cyclic AMP and forskolin, agents that increase intracellular levels of cyclic AMP. The effect appears to be at the level of the smooth muscle, since the adenylate cyclase activator, forskolin, inhibited muscarinic and serotoninergic contractions as well as transmurally stimulated contractions. Substance P-induced contractions were unaffected by forskolin. The endogenous agonists/neurotransmitters which would increase cyclic AMP levels in rainbow trout intestinal smooth muscle are as yet unknown. The effects do not appear to be modulated by vasoactive intestinal peptide (VIP), calcitonin, calcitonin gene-related peptide (CGRP), or agents that activate 13adrenoceptors. Prostaglandin E 2 (PGE2) and o%-adrenergic agonists are possible agents which will decrease contractility of the smooth muscle. They were only active in the proximal intestine and on transmurally stimulated contractions. The effects of both PGE 2 and c~2-agonists appear to be prejunctional, decreasing release of contractile neurotransmitters in the enteric nervous system.
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