Publication Date

2010

Document Type

Dissertation

Committee Members

Ronny Blust (Committee Member), Allen Burton (Committee Member), Christopher Gillen (Committee Member), Courtney Sulentic (Committee Member), Michele Wheatly (Committee Chair)

Degree Name

Doctor of Philosophy (PhD)

Abstract

Endocrine disruption is a complex phenomenon in the sense that endocrine disrupting chemicals (EDCs) are known to act via multiple modes of action, but the mechanisms of actions are poorly understood. In crustaceans, calcium (Ca2+) apart from its role as a second messenger, is also a major constituent of the calcified exoskeleton which undergoes a periodic mineralization/demineralization process known as the molting cycle. Molt cycle is under the control of steroid hormones, ecdysteroids. EDCs disrupt this molting process via their interference with receptor-mediated ecdysteroid signaling. However, the hormonal regulation of Ca2+ flux in crustacean molting is poorly understood. Cyclic nucleotides -cyclic adenosine monophosphate (cAMP) and/or cyclic guanosine monophosphate (cGMP) -play an important role in ecdysteroidogenesis as well as in the regulation of Ca2+ transporters. These three components -nuclear receptors, cyclic nucleotides and Ca2+ transporters -are seemingly inter-connected, and can be affected by endocrine disruption. I investigated a mechanistic pathway involving these three components in the crustacean endocrine disruption in freshwater crayfish, Procambarus clarkii. I selected 17α-ethinyl-estradiol (EE2, a synthetic estrogen) and 17β-estradiol (E2, a natural estrogen) as model EDCs. We determined the expression profiles of i) nuclear estrogen receptors (EcR: Ecdysone Receptor; RXR: Retinoid X Receptor), ii) cyclic nucleotides (cAMP and cGMP), and iii) Ca2+ transporters (SERCA: Sarco/Endoplasmic Reticulum Ca2+-ATPase; CaM: Calmodulin; and PMCA: Plasma Membrane Ca2+-ATPase) in the hepatopancreatic cells of P. clarkii. Finally, we determined the chitinase (Chi) gene expression to substantiate our findings, as Chi expression represents the terminal events in ecdysteroid signaling. During the molt cycle of P. clarkii, all three components were differentially expressed. Significant upregulation of nuclear estrogen receptors, cyclic nucleotides, and Ca2+ transporters was observed in the presence of environmentally relevant concentrations (100 ng/L and 500 ng/L) of EE2 and E2. Chi expression was significantly higher during postmolt stage, and when cray were fish exposed to E2 and EE2. Understanding the roles of these three components in a natural molt cycle will provide insights about pathways associated with ecdysteroid signaling. Determining the effects of EDCs on these components will aid in understanding the mechanisms behind the disruption of this signaling.

Page Count

131

Department or Program

Department of Earth and Environmental Sciences

Year Degree Awarded

2010

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