Lysine uptake was studied at pH 7.4 and 10.0 by rapid filtration methods in brush border membrane vesicles from fifth instar larvae of a model insect, the tobacco hornworm, Manduca sexta (Lepidoptera, sphingidae). At both pH values the uptake was mediated by K+ coupled symport. The uptake rate increased between pH 5.5 and 10, especially so in the alkaline range. The total lysine uptake could be divided into two components based upon lysine's ionic form as a function of pH. Lysine uptake at pH 7.4 was strongly cis-inhibited by arginine but at pH 10 was cis-inhibited and trans-stimulated by many neutral amino acids, e.g. leucine, but not by arginine. Lysine uptake by the arginine-inhibitable component paralleled the titration curve of cationic lysine whereas uptake by the leucine-inhibitable component paralleled that of zwitterionic lysine. Evidently, the brush border membrane contains at least two separate, K(+)-dependent amino acid symporters (co-transporters) that mediate lysine uptake. A cationic amino acid:K+ symporter selects cationic lysine and arginine but not histidine and other amino acids. One or more zwitterionic amino acid:K+ symporters select zwitterionic lysine, possibly arginine, histidine and neutral amino acids. Based upon these substrate repertoires, the zwitterionic symporters are B-type systems whereas the cationic symporter is identical with System R+, which partially resembles System y+. Arginine uptake in vivo is likely to be mediated mainly by System R+ whereas lysine uptake is likely to be mediated by System B-type symporters.