Amino Acid Transport in Reproductive and Muscle Tissues of Ascaris suum

Abstract

14C-Cycloleucine (1-aminocyclopentane-l-carboxylic acid-carboxyl-14C) is actively transported by the ovary-oviduct tissue of ascaris. This transport occurred both from saline and from perienteric fluid. The rate of uptake of the amino acid from saline was not affected by preincubation of the tissues in glucose. The atmosphere (air, air 95%-CO2 5%, N2 95%-C02 5%) did not influence the rate of uptake. The uptake was competitively inhibited by leucine. The Kt of transport was 0.73 mM and the Vmax was 0.228 tumoles/min/g. 14C-Leucine was also taken up by the ovary-oviduct tissue and this uptake was competitively inhibited by isoleucine. Leucine was also rapidly incorporated into protein. The movement of amino acids into muscle tissue was by simple diffusion. Neither facilitated diffusion nor active transport could be demonstrated. The findings are discussed in terms of the physiology of the parasite. The cuticle of Ascaris suum is impermeable to most nutrients (Rogers and Lazarus, 1949; Cavier and Savel, 1952; Castro and Fairbairn, 1969). The worm must obtain food via its intestine and, indeed, several workers have demonstrated that the intestine has the capability to transport some of these nutrients (Sanhueza et al., 1968; Castro and Fairbairn, 1969; Beames, 1971). As the metabolites are absorbed, they pass into the perienteric fluid which bathes the muscle of the body wall and the reproductive tissues. Both tissues have high metabolic rates (Kelly and Smith, 1956; Entner and Gonzales, 1959), and one might expect that each tissue would compete with the other for substrates in the perienteric fluid. Thus, the development of an active transport system for a certain metabolite by a tissue would give that tissue a selective advantage in obtaining that metabolite. The transport of amino acids has been studied to some extent in parasitic helminths (Read et al., 1963). The metabolically inert amino acid, cycloleucine, has been found to be useful in these studies (Harris and Read, 1968). Since the perienteric fluid contains most of the naturally occurring amino acids (Read, pers. comm.), it was the intent of this study to test the capability of the muscle and ovary-oviduct tissues to actively accumulate amino acids. Received for publication 22 July 1971. * Supported in part by Faculty Research Grant No. 34640, North Texas State University. MATERIALS AND METHODS Adult female ascarids were obtained from the intestines of infected pigs at a local abattoir and transported to the laboratory in balanced saline (Harpur, 1963) maintained at 37 C. The worms were immediately dissected and the musclecuticle or the muscle and ovary-oviduct tissue placed in a preincubation solution (AIS) (Baldwin and Moyle, 1947). The tissues were preincubated together in the flasks for 30 min in a shaking water bath at 37 C while being gassed with N2CO2 (95%-5% ). After preincubation, the tissues were removed with forceps, and incubated together in AIS to which the labeled amino acid had been added. Gas was bubbled through the mixture for at least 3 min before stoppering. In those experiments in which perienteric fluid was used as the incubation medium, the fluid was collected from large females, filtered through glass wool into a graduated cylinder packed in ice, transferred to the incubation flasks, and equilibrated at 37 C. The tissues were preincubated in AIS and then placed in the warmed perienteric fluid to which had been added a small quantity of labeled amino acid. After a specified time, the tissues were removed from the incubation flasks, rinsed in 4 changes of AIS, blotted to remove excess moisture, and added to 4 ml of 70% ethanol. The tissues were removed from the ethanol after 24 hr, rinsed in fresh ethanol, and dried to a constant weight in an oven at 90 C. An aliquot of the ethanol extract, which contained the free amino acids, was spotted on a 2-cm circle of this filter paper, dried, and placed in scintillation vials (PPO-POPOP-Toluene). It was counted in a Beckman LS-100 Liquid Scintillation Counter. Standard solutions of labeled amino acids were treated in the same manner. The data are expressed as micromoles of amino acid taken up per gram of ethanol-extracted dry weight. In those experiments in which the incorporation of leucine into protein was measured, the tissues were incubated for 1 hr in the radioactive leucine solution, then removed from the