High affinity transport of taurine and beta-alanine and low affinity transport of gamma-aminobutyric acid by a single transport system in cultured glioma cells.
چکیده
Transport of radioactive taurine, fi-alanine, and yaminobutyrate was studied in a rat spinal glioma cell line, LRM55. Kinetic analyses showed that LRM55 cells possess only low affinity y-aminobutyrate transport (K,,, = 709 + 61 pM) and only high affinity taurine and /3-alanine transport (K,,, = 30 f 3 and 79 + 12 pM, respectively). No high affinity y-aminobutyrate or low affinity taurine or p-alanine transport systems were observed. Maximum rates of uptake were 4.2 + 2.1,4.6 f 1.4, and 1.7 + 0.3 nmol/min/mg of protein for yaminobutyrate, /3-alanine, and taurine, respectively. Net transport of y-aminobutyrate and taurine was demonstrated by amino acid analysis of cells. Control cells contained approximately 38 mM taurine but identification of y-aminobutyrate and p-alanine were uncertain and the amounts too small to be quantified. Influxes of 1 mM taurine, 1 mM p-alanine, and 0.025 PM to 2 mM yaminobutyrate were nearly linear for 2 h, whereas influxes of 1 pM taurine and 1 PM /3-alanine had nearly ceased after 1 h. These differences among the time courses were attributable to the differences in the kinetic parameters of the three compounds as indicated by the accurate fit of a simple computer model (consisting of Michaelis-Menten influx and first order efflux components) to the time courses for all three compounds. Graded replacement of NaCl with choline chloride strongly reduced the influxes of taurine and yaminobutyrate in a nearly identical way. Taurine, fialanine and y-aminobutyrate were mutually competitive inhibitors of the transport of each other. Sixteen amino acids and structural analogs of taurine, p-alanine, and y-aminobutyrate were found to have very similar inhibitory effects on the influxes of all three substrates. The mutual competition and similarities in sodium dependency and inhibition by analogs indicate that low affinity y-aminobutyrate and high affinity taurine and p-alanine transports occur by the same system in LRM55 cells. This suggests that low affinity y-aminobutyrate uptake in other preparations such as brain slices or synaptosomes is carried out, at least in part, by high affinity transport systems for taurine and palanine.