Targeting of proteins to specific subcellular locations within pre- and postsynaptic neurons is essential for synapse formation. The heparan sulfate proteoglycan agrin orchestrates postsynaptic differentiation of the neuromuscular junction and may be involved in synaptic development and signaling in the central nervous system (CNS). Agrin is expressed as transmembrane and secretory isoforms with distinct N-termini. We examined the distribution of recombinant agrin in cultured motor and hippocampal neurons by transfection with agrin-GFP constructs. Immunostaining revealed a vesicular transport compartment within all neurites. Plasma membrane insertion and secretion of recombinant agrin were targeted to axonal growth cones of motor neurons; transmembrane agrin-GFP was targeted predominantly to axons and axonal growth cones in hippocampal neurons. We used agrin deletion mutants to show that axonal targeting of agrin depends on multiple domains that function in an additive fashion, including the very N-terminal portions and the C-terminal half of the molecule.