Date of Graduation

7-2020

Document Type

Dissertation

Degree Name

Doctor of Philosophy in Biology (PhD)

Degree Level

Graduate

Department

Biological Sciences

Advisor

Christian Tipsmark

Committee Member

Mack Ivey

Second Committee Member

Wayne Kuenzel

Third Committee Member

Steven Beaupre

Keywords

Aquaporins, Claudins, Euryhaline, Medaka, Osmoregulation

Abstract

Japanese medaka (Oryzais latipes) are euryhaline fish, meaning they are capable of surviving in a variety of salinities from fresh water to seawater. The ability to maintain an internal osmotic concentration stems from the phenotypic plasticity of the osmoregulatory organs, the gill, kidney, intestine, and integument. The gill is the main site of osmotic and ionic regulation in fish due to the three-dimensional structure, the direct contact with the outside environment, and the composition of the gill cells. Fish gills are multifunctional as they regulate water movement, acid/base exchange, nitrogenous waste excretion, and ion fluctuations. In freshwater environments, fish are challenged with a passive osmotic gain and ion loss. While the reverse is true in seawater, with passive ion gain and osmotic water loss. Salinity changes and the endocrine system drive regulation of cell type composition and activity.

This dissertation focuses on the Japanese medaka, osmotic regulation patterns in response to salinity fluctuations. Investigation into regulatory patterns of gill aquaporins and claudins were measured in response to exposure to freshwater, seawater, and ion poor water. Hormone in vitro experiments were conducted to understand the effects of prolactin and cortisol on the regulation of aquaporin 1 and aquaporin 3 in the gill. Function and regulation of claudin 30c within the gill in response to salinity was investigated and preliminary experiments for a loss of function study was conducted.

Aquaporin 3 was significantly increased in initial ion poor water exposure. Hormone studies demonstrated the stimulatory effect of prolactin on aquaporin 3, both alone and in combination with cortisol. Aquaporin 1 remained constitutively expressed in all salinities and under hormone exposure. Claudin 30C mRNA was significantly increased in response to ion poor water conditions, and protein expression significantly decreased in seawater versus fresh water. Localization studies showed claudin 30C in the tight junctions of hatchling medaka skin. The work performed in this thesis shows the regulatory patterns of osmoregulatory proteins in medaka, which will aid functional studies within other vertebrate system, as well as show that medaka are a suitable knock down model.

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