The relationship between nitrate influx, BnNrt2 nitrate transporter gene expression and amino acid composition of phloem exudate was investigated during N-deprivation (short-term experiment) and over a growth cycle (long-term experiment) in Brassica napus L. The data showed a positive correlation between γ-aminobutyric acid (GABA) in phloem exudate and nitrate uptake in the short- and the long-term experiments. The hypothesis that this non-protein amino acid could up-regulate nitrate uptake via a long-distance signalling pathway was tested by providing an exogenous GABA supply to the roots. The effect of GABA was compared with the effects of Gln, Glu and Asn, each known to be inhibitors of nitrate uptake. The results showed that GABA treatment induced a significant increase of BnNrt2 mRNA expression, but had less effect on nitrate influx. By contrast, Gln, Glu and Asn significantly reduced nitrate influx and BnNrt2 mRNA expression compared with the control plants. This study provides the first evidence that GABA may act as a putative long-distance inter-organ signal molecule in plants in conjunction with negative control exerted by Gln. The up-regulation effect of GABA on nitrate uptake is discussed in the context of its role in N metabolism, nutritional stress and the recent discovery of a putative role of GABA as a signal molecule in plant development.