The lack of overt pathology in α-syn germ-line knockout animals (Abeliovich et al., 2000) and the commonly proposed toxic gain-of-function of the α-syn mutant versions (A53T, A30P, E46K; Kruger et al., 1998; Giasson et al., 2002; Klein et al., 2002; Zarranz et al., 2004) led many investigators, us among them, to pursue targeted knockdown of α-syn expression as a potential therapeutic avenue for PD. However, using recombinant adeno-associated virus (rAAV) expressing a short hairpin RNA (shRNA) to knock down α-syn in mature rat substantia nigra (SN) DA neurons we encountered a surprising result: DA neuron degeneration (Gorbatyuk et al., 2010). In our rodent studies, neurodegeneration could be rescued by co-expression of rat α-syn (rendered insensitive to the shRNA) demonstrating that neuronal loss was explicitly due to a toxic loss-of function of α-syn and not due to non-specific shRNA-mediated toxicity. Moreover, by utilizing several distinct shRNA sequences displaying varying efficiencies of endogenous rat α-syn mRNA knockdown we showed that the extent of neuronal loss was dependent on the level of remaining α-syn. The ability to rescue neurons by co-expression of rat α-syn and the fact that toxicity was proportional to the efficiencies of shRNAs demonstrate that neuronal loss was not due to “off-targeting” of other endogenous mRNAs or due to non-specific shRNA toxicity.