Progress 03/01/10 to 02/28/13
Outputs
OUTPUTS: Events in Year 1 1. USDA Principal Investigators meeting in Washington , D.C. in April 2010. One poster was presented at this meeting: a. Ahearn, G. A. (2010) Synergistic absorption of essential amino acids and metals by crustacean digestive tract. 2. Experimental Biology Conference held in Anaheim, CA from 24-28 April 2010. The participating society for this presentation was the American Physiological Society. One poster was presented at this meeting: a. Ahearn, G. A., Wells, A.L. and Obi, I. (2011) Zn2+-dependent 3H-leucine transport across lobster intestine uses a peptide transporter. Events in Year 2 3. Experimental Biology Conference held in Washington, D. C. from 9-13 April 2011. The participating society for this presentation was the American Physiological Society. Two posters were presented at this meeting: a. Obi, I. and Ahearn, G. A. (2011) Transepithelial D-glucose and D-fructose transport across lobster intestine. b. Ahearn, G. A. Simons, T., and Mozo, J. (2011) Zn2+-dependent 3H-leucine transport across lobster intestine uses a peptide transporter. 4. USDA Principal Investigators meeting in Washington, D. C., from 19-21 April 2011. One poster was presented at this meeting: a. Ahearn, G. A., Mozo, J., Simmons, T., and Wilson, J. (2011) Ion-dependent nutrient transport by shrimp hepatopancreatic brush border membrane vesicles. Events in Year 3 5. Society of Integrative and Comparative Biology meeting in Janaury 2012 held in Charleston, South Carolina in January 2012. Two posters were presented at this meeting: a. Obi, I, Sterling, K. M., Simmons, T., and Ahearn, G. A. (2012) K+-dependent 3H-D-glucose transport by hepatopancreatic BBMV of the marine shrimp, Litopenaeus setiferus. b. Duka, A. and Ahearn, G. A. (2012) Comparative sugar transport by crustacean hepatopancreas and intestine. 6. Experimental Biology Conference held in San Diego, CA in April 2012. The participating society for these presentations was the American Physiological Society. Two posters were presented at this meeting: a. Ahearn, G. A., Obi, I., and Sterling, K. M. (2012) 3H-D-glucose transport is both Na+ and K+-dependent in the marine shrimp, Litopenaeus setiferus. b. Duka, A. and Ahearn, G. A. (2012) Comparative sugar transport by crustacean hepatopancreas and intestine. 7. USDA Principal Investigators meeting in Phoenix, AZ from July 19-20 2012. One 20 minute oral talk presented at this meeting: a. Ahearn, G. A. (2012) "Synergistic absorption of essential amino acids and metals by shrimp digestive tract." Events occurring after grant termination resulting from work conducted in Year 3 8. Experimental Biology Conference held in Boston, MA in April 2013. The participating society for this presentation was the American Physiological Society. One poster was presented at this meeting: a. Duka, A. and Ahearn, G. A. (2013) Leucine and methionine share a Na/K-dependent amino acid transporter in shrimp hepatopancreas. 9. Seminar presented at the Aquamarina Biotechnology Institute in Antofagasta, Chile in May 2013 entitled: "Nutritional physiology of shrimp and other crustaceans and its role in successful aquaculture." PARTICIPANTS: The following personnel have been associated with this project during its 3 years of funding: Principal Investigator: Dr. Gregory A. Ahearn; Research collaborators: Dr. Kenneth Sterling (The Whitney Laboratory, University of Florida, Gainesville, Florida), Dr. Flavia Zanotto (Universitdad Presbiteriana Mackenzie, Sao Paulo, Brazil); Dr. Amy Lane (Department of Chemistry, University of North Florida, Jacksonville, Florida); Laboratory Managers: Ijeoma Obi, Ada Duka; Master of Science graduate students: Anna Wells, Depol Patel, Jennifer Wilson, Tamla Simmons, Ijeoma Obi, Ada Duka; Graduate students from Universitdad Presbiteriana Mackenzie, Sao Paulo, Brazil: Priscila Ortega, Livia Farah; Postbacclaureate student: Julie Mozo. During this 3-year funding period all mentioned graduate students received Master of Science Degrees from the University of North Florida and Julie Mozo finished a 2-year nursing program at the University of North Florida and is now an active nurse in St. Augustine, Florida. TARGET AUDIENCES: Target audiences for the efforts undertaken in this 3-year project would be the general Scientific Community interested in the basic biology of invertebrates, and crustaceans in particular,and how dietary ions stimulate the absorption of essential amino acids and sugars by the invertebrate digestive tract. Of special relevance to this group would be the findings that in shrimp, sugar and amino acid absorption is stimulated by dietary sodium, potassium and metallic cations such as zinc, copper, and manganese. Specific aspects of the work conducted in this project would also be highly relevant to aquacultural managers interested in making improvements in artificial feeds provided to penaeid shrimp pond culture. Modifications in dietary metal and potassium content may improve growth rates of pond penaeid shrimp and lead to faster attainment of market size. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
The objectives of this 3-year study were to use brush border membrane vesicle (BBMV) techniques and radioisotopic tracers to characterize membrane transport processes of the shrimp (Litopenaeus setiferus) digestive tract for sugars and amino acids and to identify dietary factors that stimulate their uptake. The rationale for this work is that knowledge of dietary stimulating factors that enhance the transfer of essential nutrients across the digestive tract of aquacultural species of crustaceans may speed growth and reduce the time to harvest in farmed facilities. Three major findings concerning nutrient absorption by shrimp digestive tract were defined in this study: 1) Luminal metallic cations (Zn2+, Cu2+ and Mn2+, Cd2+, and Co2+) each stimulated the uptake of 3H-L-leucine by shrimp hepatopancreatic BBMV, suggesting a potential synergistic role of the cations in the transmembrane transfer of this amino acid. A model was proposed that involved the formation of bis-complexes in the gut lumen between metals and amino acids during digestion where one metal ion binds two amino acids and uses a dipeptide transport protein to enter hepatopancreatic cells prior to transfer to the blood. Enhanced uptake of both essential metals and essential amino acids occurs by this synergistic process. 2) The essential amino acids L-leucine, L-methionine, and L-phenylalanine share a Na+/K+-dependent amino acid transporter in shrimp hepatopancreas, that is enhanced by luminal acidity (pH 5.0), and is able to use either cation to stimulate the uptake of each amino acid. This shared transport system resembles the Potassium Amino Acid Transporter (KAAT1) found in herbivorous insect gut that uses a K+ gradient to enhance amino acid uptake from dietary constituents. Larval shrimp eat marine algae (rich in K+), while adult shrimp consume flesh (rich in Na+), therefore, both co-transported cations may benefit the animals at different times in their lives. 3) 3H-D-glucose is absorbed by shrimp hepatopancreatic BBMV by a Na+/K+ co-transporter. This transporter differs markedly from the Na+-dependent D-glucose uptake process found in most vertebrate tissues (SGLT1), but is similar to the K+-dependent SGLT4 carrier system found solely in mammal kidney that specifically transports D-fructose and D-mannose. The use of both K+ and Na+ by sugar and amino acid transporters in shrimp digestive tract allows these animals to use K+ as a driver cation for nutrient uptake when they are largely herbivorous (larvae) and change co-transporters to Na+ later in life when they become carnivorous. This information, plus the findings that metallic cations stimulate essential amino acid uptake when the absorption environment is acidic, provides information for the potential development of two growth promoting artificial diets for shrimp in aquacultural facilities, one used during early development and one employed later in life. It is suggested that the combination of two such diets, based on an understanding of shrimp nutritional physiology, might provide faster growth than present diets and could therefore reduce the time to market and the cost for the farmer.
Publications
- Journal articles: Obi, I., Wells, A. L., Ortega, P., Patel, D., Farah, L., Zanotto, F. P. and Ahearn, G. A. (2011) 3H-L-leucine transport by the promiscuous crustacean dipeptide-like cotransporter. J. Exp. Zool. 315: 465-475.
- Simmons, T., Mozo, J., Wilson, J., and Ahearn, G. A. (2012) Cation-dependent nutrient transport in shrimp digestive tract. J. Comp. Physiol. B. 182: 209-216.
- Obi, I. E., Sterling, K. M., and Ahearn, G. A. (2013) K+-dependent 3H-D-glucose transport by hepatopancreatic BBMV of the marine shrimp, Litopenaeus setiferus. J. Comp. Physiol. B.183:61-69.
- Duka, A. and Ahearn, G. A. (2013) L-leucine, L-methionine, and L-phenylalanine share a Na+/K+-dependent amino acid transporter in shrimp hepatopancreas. J. Comp. Physiol. B. (in press).
- Abstracts: Obi, I. and Ahearn, G. A. (2011) Transepithelial D-glucose and D-fructose transport across lobster intestine. FASEB J. 25:1065.1.
- Ahearn, G. A., Obi, I., and Wells, A. L. (2011) Zn2+-dependent 3H-leucine transport across lobster intestine uses a peptide transporter. FASEB J. 25:1065.2.
- Duka, A. and Ahearn, G. A. (2012) Comparative sugar transport by crustacean hepatopancreas and intestine. J. Integ. Biol. 52:e238.
- Obi, I., Sterling, K. M., Simmons, T., and Ahearn, G. A. (2012) K-dependent 3H-D-glucose transport by hepatopancreatic BBMV of the marine shrimp, Litopenaeus setiferus. J. Integ. Comp. Biol. 52:e304
- Ahearn, G. A., Obi, I., and Sterling, K. M. (2012) 3H-D-glucose transport is both Na+- and K+-dependent in the marine shrimp, Litopenaeus setiferus. FASEB J. 26:1111.4.
- Duka, A. and Ahearn, G. A. (2012) Sugar transport by crustacean hepatopancreas and intestine. FASEB J. 26:1111.3.
- Duka, A. and Ahearn, G. A. (2013) Leucine and methionine share a Na/K-dependent amino acid transporter in shrimp hepatopancreas. FASEB J. (in press).
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Progress 03/01/12 to 02/27/13
Outputs
OUTPUTS: OUTPUTS: During Year 3 of a 3-year grant from USDA, results of experimental work supported by this grant were presented as posters or as oral communication at 2 different scientific meetings. Meeting #1 was the Experimental Biology conference held in San Diego, California during April 2012. The participating society for this presentation was the American Physiological Society. Two posters were presented at this meeting: 1. Duka, A. and Ahearn, G. A. (2012) Comparative sugar transport by crustacean hepatopancreas and intestine. FASEB J. 26: 1111.3. 2. Ahearn, G. A., Obi, I., and Sterling, K. M. (2012) 3H-D-glucose transport is both Na+ and K+-dependent in the marine shrimp, Litopenaeus setiferus. FASEB J. 26: 1111.4. Meeting #2 was the USDA Principal Investigators meeting in Phoenix, Arizona from 19-20 July, 2012. One twenty-minute summary of the work conducted over 3 years of USDA funding was presented by Dr. G. A. Ahearn which was entitled "Synergistic absorption of essential amino acids and metals by shrimp digestive tract." PARTICIPANTS: The following personnel have been associated with this project during Year 3: Principal Investigator: Dr. Gregory A. Ahearn; Research collaborators: Dr. Kenneth Sterling (The Whitney Laboratory, University of Florida); Dr. Flavia Zanotto (Universitdade Presbiteriana Mackenzie, Sao Paulo, Brazil; Laboratory Manager, Ijeoma Obi; Master of Science graduate students, Tamla Simmons, Ada Duka, Resheda Likely, Maria Peterson; Post baccalaureate student, Julie Mozo; Undergraduate students, Rania Abdel-Malak and Mihra Rasidovic (Honors student). TARGET AUDIENCES: Target audiences for the efforts undertaken in Year 3 would be the general scientific community interested in the basic biology of invertebrates and how dietary constituents stimulate the absorption of essential amino acids and sugars. Specific aspects of the work conducted in Year 3 would also be highly relevant to aquacultural managers interested in making improvements in artificial feeds provided to penaeid shrimp pond culture. Modifications of dietary metals and both sodium and potassium content may improve growth rates of pond penaeid shrimp and lead to faster attainment of market size. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period. PARTICIPANTS: PARTICIPANTS: The following personnel have been associated with this project during Year 3: Principal Investigator: Dr. Gregory A. Ahearn; Research collaborators: Dr. Kenneth Sterling (The Whitney Laboratory, University of Florida); Dr. Flavia Zanotto (Universitdade Presbiteriana Mackenzie, Sao Paulo, Brazil; Laboratory Manager, Ijeoma Obi; Master of Science graduate students, Tamla Simmons, Ada Duka, Resheda Likely, Maria Peterson; Post baccalaureate student, Julie Mozo; Undergraduate students, Rania Abdel-Malak and Mihra Rasidovic (Honors student). TARGET AUDIENCES: TARGET AUDIENCES: Target audiences for the efforts undertaken in Year 3 would be the general scientific community interested in the basic biology of invertebrates and how dietary constituents stimulate the absorption of essential amino acids and sugars. Specific aspects of the work conducted in Year 3 would also be highly relevant to aquacultural managers interested in making improvements in artificial feeds provided to penaeid shrimp pond culture. Modifications of dietary metals and both sodium and potassium content may improve growth rates of pond penaeid shrimp and lead to faster attainment of market size. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
Outcomes: Hepatopancreatic brush border membrane vesicles (BBMV), made from Atlantic White shrimp (Litopenaeus setiferus), were used to characterize the transport properties of 3H-L-leucine influx by these membrane systems and how other essential amino acids and the cations, sodium and potassium, interact with this transport system. 3H-L-leucine uptake by BBMV was pH-sensitive and occurred against transient transmembrane concentration gradients in both Na+- and K+-containing incubation media, suggesting that either cation was capable of providing a driving force for amino acid accumulation. 3H-L-leucine uptake in NaCl or KCl media were each 3 times greater in acidic pH (pH 5.5) than in alkaline pH (pH 8.5). The essential amino acid, L-methionine, at 20 mM significantly (p < 0.0001) inhibited the 2 min uptakes of 1 mM 3H-L-leucine in both Na+- and K+-containing incubation media. The residual 3H-L-leucine uptake in the two media were significantly greater than zero (p < 0.001), but not significantly different from each other (p > 0.05) and may represent an L-methionine- and cation-independent transport system. 3H-L-leucine influxes in both NaCl and KCl incubation media were hyperbolic functions of [L-leucine], following the carrier-mediated Michaelis-Menten equation. In NaCl, 3H-L-leucine influx displayed a low apparent Km (high affinity) and low apparent Jmax, while in KCl the transport exhibited a high apparent Km (low affinity) and high apparent Jmax. L-methionine (7 and 20 mM) was a competitive inhibitor of 3H-L-leucine influxes in both NaCl and KCl media, producing a significant (p < 0.01) increase in 3H-L-leucine influx Km, but no significant response in 3H-L-leucine influx Jmax. Potassium was a competitive inhibitor of sodium co-transport with 3H-L-leucine, significantly (p < 0.01) increasing 3H-L-leucine influx Km in the presence of sodium, but having negligible effect on 3H-L-leucine influx Jmax in the same medium. These results suggest that shrimp BBMV transport 3H-L-leucine by a single L-methionine-shared carrier system that is enhanced by acidic pH and can be stimulated by either Na+ or K+ acting as co-transport drivers binding to shared activator sites.
Publications
- 1.Obi, I. E., Sterling, K. M. and Ahearn, G. A. (2013) K+-dependent 3H-D-glucose transport by hepatopancreatic BBMV of the marine shrimp, Litopenaeus setiferus. J. Comp. Physiol. B. 183:61-69.
- 2.Duka, A. and Ahearn, G. A. (2013) Leucine and methionine share a Na+/K+-dependent amino acid transporter in shrimp hepatopancreas. FASEB J. (in press) [abstract]
- 3.Duka, A. and Ahearn, G. A. (2013) Leucine and methionine share a Na+/K+-dependent amino acid transporter in shrimp hepatopancreas. J. Comp. Physiol. B (in press).
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Progress 03/01/11 to 02/28/12
Outputs
OUTPUTS: Outputs for Year 2 During Year 2 of a 3-year research grant from USDA, results of experimental work supported by this grant were presented as posters at 3 different scientific meetings. 1. Experimental Biology Conference held in Washington, D. C. from 9-13 April 2011. The participating society for this presentation was the American Physiological Society. Two posters were presented at this meeting: a. Obi, I. and Ahearn, G. A. (2011) Transepithelial D-glucose and D-fructose transport across lobster intestine. b. Ahearn, G. A. Simons, T., and Mozo, J. (2011) Zn2+-dependent 3H-leucine transport across lobster intestine uses a peptide transporter. 2. USDA Principal Investigators meeting in Washington, D. C., from 19-21 April 2011. One poster was presented at this meeting: a. Ahearn, G. A., Mozo, J., Simmons, T., and Wilson, J. (2011) Ion-dependent nutrient transport by shrimp hepatopancreatic brush border membrane vesicles. 3. Society of Integrative and Comparative Biology meeting in Janaury 2012 held in Charleston, South Carolina in January 2012. Two posters were presented at this meeting: a. Obi, I, Sterling, K. M., Simmons, T., and Ahearn, G. A. (2011) K+-dependent 3H-D-glucose transport by hepatopancreatic BBMV of the marine shrimp, Litopenaeus setiferus. b. Duka, A. and Ahearn, G. A. (2011) Comparative sugar transport by crustacean hepatopancreas and intestine. PARTICIPANTS: The following personnel have been associated with this project during Year 2: Principal Investigator: Dr. Gregory A. Ahearn; Research collaborators: Dr. Kenneth Sterling (The Whitney Laboratory, University of Florida); Dr. Flavia Zanotto (Universitdade Presbiteriana Mackenzie, Sao Paulo, Brazil); Laboratory Manager: Ijeoma Obi; Master of Science Graduate students: Anna Wells, Depol Patel, Jennifer Wilson, Tamla Simmons; Post baccalaureate student: Julie Mozo; and graduate students from Universitdade Presbiteriana Mackenzie, Sao Paulo, Brazil: Priscila Ortega and Livia Farah. Extensive training of American and Brazilian graduate students by Drs. Ahearn and Sterling and by I. Obi were conducted throughout Year 2. TARGET AUDIENCES: Target audiences for the efforts undertaken in Year 2 would be the general Scientific Community interested in the basic biology of invertebrates and how dietary ions stimulate the absorption of eseential amino acids and sugars. Specific aspects of the work conducted in Year 2 would also be highly relevant to aquacultural managers interested in making improvements in artificial feeds provided to penaeid shrimp pond culture. Modifications in dietary metal and potassium content may improve growth rates of pond penaeid shrimp and lead to faster attainment of market size. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
Outcome #1: Hepatopancreatic epithelial brush border membrane vesicles (BBMV) of the Atlantic White shrimp, Litopeneaus setiferus, were used to study the cation dependency of sugar and amino acid transport from dietary constituents to epithelial cytoplasm. 3H-D-glucose uptake by BBMV occurred when either transmembrane sodium or potassium gradients were the only driving forces for sugar accumulation, suggesting the presence of a possible coupled transport system capable of using either cation. 3H-Lhistidine transport was only stimulated by a transmembrane potassium gradient, while 3H-leucine uptake was enhanced by either a sodium or potassium gradient. Uptake of 3H-L-leucine was significantly stimulated (p < 0.05) by several metallic cations (e.g., Zn2+, Cu2+, Mn2+, Cd2+, or Co2+) at external pH values of 7.0 or 5.0, suggesting a potential synergistic role of the cations in the transmembrane transfer of amino acids. 3H-L-histidine influxes were hyperbolic functions of external [zinc] or [manganese]. The apparent affinity constant (e.g., Km) for manganese was an order of magnitude smaller (Km = 0.22 μM Mn) than that for zinc (Km = 1.80 μM Zn), while no significant difference (p > 0.05) occurred beween their maximal transport velocities (e.g., Jmax). These results suggest that a number of cation-dependent nutrient transport systems occur on the shrimp brush border membrane and aid in the absorption of these important dietary elements. Outcome #2: Effects of Na, K, sugar inhibitors, and membrane PD on 3H-D-glucose uptake by hepatopancreatic BBMV Atlantic marine shrimp, L. sertiferus, were investigated. 3H-D-glucose uptake was stimulated by both Na and K and membrane PD. Kinetics of 3H-D-glucose influx were hyperbolic functions of both external Na or K, and an induced membrane potential increased influx Jmax, while it lowered Km. 3H-D-glucose influx vs [glucose] in both Na or K media displayed Michaelis-Menten properities. Phloridzin inhibited 0.5 mM 3H-D-glucose influx, requiring 5 mM in NaCl and 10 mM in KCl to significantly reduce hexose transport. Several sugars (D-galactose, alpha-methyl-D-gluco-pyanoside, unlabeled D-glucose, D-fructose, and D-mannose) were used at 75 mM as potential inhibitors of 0.1 mM 3H-D-glucose influx. Only unlabeled D-glucose, D-fructose, and D-mannose significantly (P < 0.05) reduced labeled glucose transport. An additional experiment using increasing concentrations of D-mannose (0, 10, 25, 75, and 100 mM) showed this hexose to be an effective inhibitor of 0.1 mM 3H-D-glucose uptake at concentrations of 75 mM and higher. 3H-D-glucose transport by hepatopancreatic BBMV occurs by a carrier system that uses both Na and K, is enhanced by membrane PD, and is only inhibited by itself, D-fructose, and D-mannose. These properties are similar to those exhibited by the mammalian SLC5A9/SGLT4 transporter, suggesting that an invertebrate analogue of this protein may occur in shrimp.
Publications
- Obi, I., Wells, A. L., Ortega, P., Patel, D., Farah, L., Zanotto, F. P. and Ahearn, G. A. (2011) 3H-L-leucine transport by the promiscuous crustacean dipeptide-like cotransporter. J. Exp. Zool. 315: 465-475.
- Simmons, T., Mozo, J., Wilson, J., and Ahearn, G. A. (2012) Cation-dependent nutrient transport in shrimp digestive tract. J. Comp. Physiol. B. 182: 209-216.
- Obi, I. E., Sterling, K. M., and Ahearn, G. A. (2012) K+-dependent 3H-D-glucose transport by hepatopancreatic BBMV of the marine shrimp, Litopenaeus setiferus. J. Comp. Physiol. B. (submitted for publication).
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Progress 03/01/10 to 02/28/11
Outputs
OUTPUTS: BACKGROUND This 3-year project, examining the synergistic interactions between essential nutrients and heavy metals in penaeid shrimp digestive tract, is an outgrowth and application of 25 years of previous nutritional physiology experiments with the American lobster, Homarus americanus by the Principal Investigator (PI). While the North American lobster fishery remains robust and has not led to the development of aquacultural enterprises supporting the market availability of this species, penaeid shrimp fisheries throughout the world are significantly supported by high-density pond aquaculture. The current USDA grant has used during this first year of funding a significant database developed by the Principal Investigator and numerous national and international colleagues over the last quarter century on the nutritional biology of the lobster to formulate experiments and research protocols that are being extended to shrimp species with the goal of providing physiological information that can be used to enhance shrimp growth rate in pond aquaculture facilities. EXPERIMENTAL PROTOCOLS Most dietary nutrient and ion absorption by crustacean digestive tracts occurs by the hepatopancreatic epithelium in these animals. During the 1980s and 1990s the PI developed an experimental method for studying amino acid, sugar, vitamin, peptide, and ion absorption by lobster hepatopancreatic epithelial cell membranes and published over 25 papers on these properties during this time. This experimental protocol is currently being applied to shrimp hepatopancreas to characterize synergistic interactions during nutrient and ion absorption. In these experiments purified shrimp hepatopancreatic epithelial brush border membrane vesicle are produced and incubated in vitro with radiolabelled organic solutes in the presence of heavy metals or other ions so that rates of trans-membrane transfer processes of these dietary components can be characterized and quantified. These experimental protocols will be modified over the 3-year duration of the current research grant to reach the objectives of the grant and provide information that may be ultimately used to enhance growth rates of shrimp in aquacultural environments. EXPERIMENTAL PERSONNEL The following personnel have been associated with this project during Year 1: Principal Investigator: Dr. Gregory A. Ahearn Laboratory technician: Ijeoma Obi Master of Science graduate students: Jennifer Wilson (MS in 2010), Tamla Simmons (MS expected 2013) Undergraduate researcher: Julie Mozo (BS expected May 2011) EVENTS Results of experimentation during Year 1 will be presented at two meetings during 2011: 1. Experimental Biology Conference, 2011, held in Washington, D.C from 9 - 13 April 2011. A poster of results obtained from experiments conducted during 2010-2011 will be presented at this venue. The participating society for this presentation is the American Physiological Society. 2. USDA Principal Investigators meeting 2011, held in Washington, D. C. from 19-21 April 2011. A second poster of results obtained from experiments conducted during 2010-2011 will be presented at this venue. PARTICIPANTS: The following personnel have been associated with this project during Year 1: Principal Investigator: Dr. Gregory A. Ahearn Laboratory technician: Ijeoma Obi Master of Science graduate students: Jennifer Wilson (MS in 2010), Tamla Simmons (MS expected 2013) Undergraduate researcher: Julie Mozo (BS expected May 2011) During this year Tamla Simmons will be pursuing a Master of Science degree under the guidance of the Principal Investigator. Jennifer Wilson had already completed a MS degree when she worked on this project for the summer of 2010. Julie Mozo is an undergraduate who will graduate in May 2011 and will stay on the project through summer 2011 and perhaps into the fall of 2011 as a postbac student or as a new technician on the project. Other students will be incorporated into the project as they become available. TARGET AUDIENCES: Nothing significant to report during this reporting period. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
Change in knowledge During Year 1 of the current USDA research grant, experimentation with both lobster (Homarus americanus) and penaeid white shrimp (Litopenaeus setiferus led to changes in knowledge that have a direct impact on the objectives of this project: 1. In lobster digestive tract, a broad spectrum, dipeptide-like, membrane co-transport protein was characterized that is able to absorb a wide variety of amino acids as complexes with the heavy metals copper, zinc, and manganese. The amino acid/metal chelates appear to compete with the absorption of dipeptides and suggest a novel means of nutrient absorption in this crustacean. 2. Hepatopancreatic brush border membrane vesicles made from white shrimp (L. setiferus) adapted to 100% seawater displayed significantly greater 3H-L-histidine and 3H-L-leucine uptake than did similar vesicles made from shrimp adapted to 60% seawater, suggesting a down-regulation of nutrient transporters by euryhaline shrimp moving from seawater to estuaries. 3. Sodium and potassium in an incubation buffer individually stimulate the uptake of 3H-D-glucose and 3H-L-leucine by L. setiferus hepatopancreatic brush border membrane vesicles (BBMV), suggesting the synergistic cotransport of ion and nutrient across epithelial membranes. 4. Potassium seems to be a superior stimulator of both sugar and amino acid transport than does sodium, which is opposite to the stimulatory effect of the two cations on nutrient absorption in mammalian digestive tracts. 5. The metals Zn, Cu, Mn Cd and Co all stimulate the uptake of 3H-leucine by L. setiferus hepatopancreatic brush border membrane vesicles, implying a broad-spectrum cotransport process involving both metal and amino acid. 6. Influx kinetics of 3H-L-histidine by L. setiferus BBMV were hyperbolic functions of saline zinc or manganese concentrations from 1 to 10 micromolar metal and followed Michaelis-Menten kinetics. Maximal amino acid transport velocities with the two metals were similar, but the apparent binding affinity of the transporter for manganese was 10 times that of zinc. This implies that it takes 10 times less manganese than zinc to reach maximal absorption of the co-transported amino acid. Change in actions Florida laws do not permit the collection of white shrimp (L. setiferus) in the St. Johns River, Jacksonville, Florida between March and June of the year. During these months the brown shrimp (Farfantepenaeus aztecus), will be collected from offshore waters in the Atlantic and Gulf of Mexico, and will be used in this project to accomplish comparative goals between species. Both shrimp species will first be adapted to full strength seawater and the effects of ions on absorption of both sugars and amino acids under these conditions will be assessed. Additional comparisons between species will be made with shrimp adapted to 75, 50 and 25% seawater. These studies should extend our observations made in Year 1 that a drop in seawater strength appears to result in the down-regulation of ion dependent nutrient transport systems in these animals. Important implications for aquaculture of shrimp in ponds of lowered seawater strength may result from these studies.
Publications
- Ahearn, G. A., Wells, A.L. and Obi, I. (2011) Zn2+-dependent 3H-leucine transport across lobster intestine uses a peptide transporter. FASEB Journal. (in press).
- Obi, I., Wells, A.L., Ortega, P., Patel, D., Farah, L., Zanotto, F. P., and Ahearn, G. A. (2011) 3H-L-leucine transport by the promiscuous crustacean dipeptide-like cotransporter. Journal of Experimental Zoology (submitted for publication).
- Ahearn, G. A., Mozo, J., Simmons, T., Wilson, J. (2011) Ion-dependent nutrient transport by shrimp hepatopancreatic brush border membrane vesicles. (in preparation).
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