Nitric oxide mediated modulation of norepinephrine transport: identification of a potential target for S-nitrosylation

1 Carrier mediated uptake (uptake-l) transport of norepinephrine (NE) plays a key role in the regulation of sympathetic neurotransmission. Recent inve stigations indicate that nitric oxide (NO) may modulate uptake-1 activity, possibly in a cyclic GMP independent manner. 2 Carrier mediated transport of [H-3-NE] and [H-3-dopamine, DA] was examine d in CHO cells transfected with cDNA for the NE and DA transporters (NET, D AT) respectively. 3 While exposure to the NO donor S-nitroso-N-acetylpenicillamine (100 mu M I, SNAP) significantly reduced [3H-NE] uptake (P < 0.001), no effect on [H- 3-DA] transport was apparent. 4 Comparison of the amino acid sequences for NET and DAT identified cystein e residue 351 in NET, which was not present in DAT. Site-directed mutagenes is of Cys 351 to Ser produced a functional NET that was resistant to the in hibitory effects of SNAP. 5 The presence of SNAP mediated nitrosylation of the cysteine residue in an 8-mer model peptide based around Cys 351 in NET was confirmed by both bioc hemical and mass spectroscopic means. 6 These data indicate the potential regulatory role for NO in modulating sy mpathetic neurotransmission, and further confirm the importance of non-cycl ic GMP dependent mechanisms in mediating the actions of NO.