CALIFORNIA STATE UNIVERSITY, NORTHRIDGE THE EFFECTS OF SUGAR ALCOHOLS ON SEA URCHIN GASTRULATION IN LOW CALCIUM SEA WATER A Thesis submitted in partial fulfillment of the requirements For the degree of Master of Science In Biology By Edward Holmes May 2015 Copyright 2015, Edward Holmes ii The thesis of Edward Holmes is approved: _____________________________________ ______________________ Lisa Banner, Ph.D. Date __________________________________ ____________________ Stan Metzenberg, Ph.D. Date __________________________________ ____________________ Steven B. Oppenheimer, Ph.D., Chair Date California State University, Northridge iii DEDICATION This research and thesis project has been dedicated to the Holy Trinity. To my heavenly Abba Father who has adopted me as His son To my Lord and Savior Jesus Christ To the Holy Spirit who is my Comforter and Counselor For their perfect love, grace and mercy For their eternal honor and glory iv ACKNOWLEDGEMENTS Thank you: To Dr. Steven Oppenheimer as a mentor and adviser for your patience, encouragement, guidance, and understanding throughout my research and thesis project. To Dr. Stan Metzenberg for your time and constructive criticism as a thesis committee member. To Dr. Lisa Banner for your time and constructive criticism as a thesis committee member. To my parents Roger and Phyllis and my brothers Jeff and Dwayne for your love, support, generousity, and patience which enabled me to complete my research and thesis project. To my brothers and sisters in Jesus Christ from United Campus Ministry (UCM), the Campus Outreach Response Team (CORT), MT28, and Intervarsity Christian Fellowship for your love, encouragement and prayer support. To my laboratory partners Kathy Fernando and Tiffany Smith for their teamwork. To Jouliana Davoudi for her assistance with the statistical analysis of my research and thesis project. v To Samantha Arvizu and Victoria Arvizu for their assistance with the photography of my research and thesis project. This research was supported by NIH, NIGMS SCORE SO648680, MARC, RISE, the Joseph Drown Foundation, and the Sidney Stern Memorial Trust. vi TABLE OF CONTENTS Signature Page ……………………………………………………………………………………………………………………..iii Dedication ……………………………………………………………………………………………………………………………iv Acknowledgements ………………………………………………………………………………………………………………v List of Figures ………………………………………………………………………………………………………………………viii List of Tables ………………………………………………………………………………………………………………………..xii Abstract ……………………………………………………………………………………………………………………………..xvii Introduction ………………………………………………………………………………………………………………………….1 Materials and Methods ……………………………………………………………………………………………………….15 Results ………………………………………………………………………………………………………………………………..26 Discussion …………………………………………………………………………………………………………………………..85 References ………………………………………………………………………………………………………………………….96 vii LIST OF FIGURES Figure 1: Set of photographs showing each L. pictus sea urchin morphological type including: complete archenteron, incomplete archenteron, not invaginated, exogastrulated and dead………………………………………………………………36 Figure 2: Set of photographs showing L. pictus sea urchin embryos at 24-26 hours after fertilization in LCASW controls and in 0.1M, 0.05M, 0.01M, 0.005M and 0.001M adonitol…………………………………………………………………………………………37 Figure 3: Set of photographs showing L. pictus sea urchin embryos at 32-34 hours after fertilization in LCASW controls and in 0.1M, 0.05M, 0.01M, 0.005M and 0.001M adonitol………………………………………………………………………………………..38 Figure 4: Set of photographs showing L. pictus sea urchin embryos at 24-26 hours after fertilization in LCASW controls and in 0.1M, 0.05M, 0.01M, 0.005M and 0.001M L-(-) arabitol………………………………………………………………………………….39 Figure 5: Set of photographs showing L. pictus sea urchin embryos at 32-34 hours after fertilization in LCASW controls and in 0.1M, 0.05M, 0.01M, 0.005M and 0.001M L-(-) arabitol………………………………………………………………………………….40 viii Figure 6: Set of photographs showing L. pictus sea urchin embryos at 24-26 hours after fertilization in LCASW controls and in 0.1M, 0.05M, 0.01M, 0.005M and 0.001M dulcitol………………………………………………………………………………………….41 Figure 7: Set of photographs showing L. pictus sea urchin embryos at 32-34 hours after fertilization in LCASW controls and in 0.1M, 0.05M, 0.01M, 0.005M and 0.001M dulcitol………………………………………………………………………………………….42 Figure 8: Set of photographs showing L. pictus sea urchin embryos at 24-26 hours after fertilization in LCASW controls and in 0.1M, 0.05M, 0.01M, 0.005M and 0.001M D-mannitol……………………………………………………………………………………43 Figure 9: Set of photographs showing L. pictus sea urchin embryos at 32-34 hours after fertilization in LCASW controls and in 0.1M, 0.05M, 0.01M, 0.005M and 0.001M D-mannitol…………………………………………………………………………………..44 Figure 10: Set of photographs showing L. pictus sea urchin embryos at 24-26 hours after fertilization in LCASW controls and in 0.1M, 0.05M, 0.01M. 0.005M and 0.001M D-sorbitol…………………………………………………………………………………….45 Figure 11: Set of photographs showing L. pictus sea urchin embryos at 32-34 hours after fertilization in LCASW controls and 0.1M, 0.05M, 0.01M, 0.005M and 0.001M D-sorbitol…………………………………………………………………………………….46 ix Figure 12: Set of photographs showing L. pictus sea urchin embryos at 24-26 hours after fertilization in LCASW controls and 0.1M, 0.05M, 0.01M, 0.005M and 0.001M xylitol…………………………………………………………………………………………….47 Figure 13: Set of photographs showing L. pictus sea urchin embryos at 32-34 hours after fertilization in LCASW controls and 0.1M, 0.05M, 0.01M, 0.005M and 0.001M xylitol…………………………………………………………………………………………….48 Figure 14: The effects on L. pictus embryos incubated in adonitol and low calcium artificial sea water at the 24-26 hour stage of gastrulation………………………………49 Figure 15: The effects on L. pictus embryos incubated in adonitol and low calcium artificial sea water at the 32-34 hour stage of gastrulation…………………………….50 Figure 16: The effects on L. pictus embryos incubated in L-(-) arabitol and low calcium artificial sea water at the 24-26 hour stage of gastrulation…………………………….51 Figure 17: The effects on L. pictus embryos incubated in L-(-) arabitol and low calcium artificial sea water at the 32-34 hour stage of gastrulation…………………………….52 Figure 18: The effects on L. pictus embryos incubated in dulcitol and low calcium artificial sea water at the 24-26 hour stage of gastrulation…………………………….53 x Figure 19: The effects on L. pictus embryos incubated in dulcitol and low calcium artificial sea water at the 32-34 hour stage of gastrulation…………………………….54 Figure 20: The effects on L. pictus embryos incubated in D-mannitol and low calcium artificial sea water at the 24-26 hour stage of gastrulation…………………………….55 Figure 21: The effects on L. pictus embryos incubated in D-mannitol and low calcium artificial sea water at the 32-34 hour stage of gastrulation…………………………….56 Figure 22: The effects on L. pictus embryos incubated in D-sorbitol and low calcium artificial sea water at the 24-26 hour stage of gastrulation……………………………..57 Figure 23: The effects on L. pictus embryos incubated in D-sorbitol and low calcium artificial sea water at the 32-34 hour stage of gastrulation…………………………….58 Figure 24: The effects on L. pictus embryos incubated in xylitol and low calcium artificial sea water at the 24-26 hour stage of gastrulation……………………………59 Figure 25: The effects on L. pictus embryos incubated in xylitol and low calcium artificial sea water at the 32-34 hour stage of gastrulation……………………………60 xi LIST OF TABLES Table 1: P-value results for each observed morphological characteristic between low calcium artificial sea water (LCASW) control and each of the five adonitol concentrations in LCASW at 24-26 hours after sea urchin fertilization…………………61 Table 2: P-value results for each observed morphological characteristic between low calcium artificial sea water (LCASW) control and each of the five adonitol concentrations in LCASW at 32-34 hours after sea urchin fertilization………………..62 Table 3: P-value results for each observed morphological characteristic between low calcium artificial sea water (LCASW) control and each of the five L-(-) arabitol concentrations in LCASW at 32-34 hours after sea urchin fertilization………………..63 Table 4: P-value results for each observed morphological characteristic between low calcium artificial sea water (LCASW) control and each of the five L-(-) arabitol concentrations in LCASW at 32-34 hours after sea urchin fertilization………………..64 Table 5: P-value results for each observed morphological characteristic between low calcium artificial sea water (LCASW) control and each of the five dulcitol concentrations in LCASW at 24-26 hours after sea urchin fertilization………………..65 xii Table 6: P-value results for each observed morphological characteristic between low calcium artificial sea water (LCASW) control and each of the five dulcitol concentrations in LCASW at 32-34 hours after sea urchin fertilization…………………66 Table 7: P-value results for each observed morphological characteristic between low calcium artificial sea water (LCASW) control and each of the five D-mannitol concentrations in LCASW at 24-26 hours after sea urchin fertilization……………….67 Table 8: P-value results for each observed morphological characteristic between low calcium artificial sea water (LCASW) control and each of the five D-mannitol concentrations in LCASW at 32-34 hours after sea urchin fertilization……………….68 Table 9: P-value results for each observed morphological characteristic between low calcium artificial sea water (LCASW) control and each of the