6-Hydroxydopamine induces dopaminergic cell degeneration via a caspase-9-mediated apoptotic pathway that is attenuated by caspase-9dn expression.

This study showed that primary dopaminergic neurons or the dopaminergic cell line MN9D, when exposed to 15 min of the parkinsonian toxin 6-hydroxydopamine (6-OHDA) in the range of 30-100 microM, underwent delayed degeneration and exhibited hallmarks of apoptosis. These results, along with the absence of any increase in lactate dehydrogenase (LDH) release from the degenerated cells, imply that apoptosis was the dominant mode of cell death. Moreover, a distinct elevation in the measured cellular activities of caspase-9 and -3 but not of caspase-8 points to the caspase-9/caspase-3 cascade as the predominant apoptotic pathway in the degeneration of dopaminergic neurons and MN9D cells. In addition, the presence of caspase-9 or -3 peptide inhibitors but not of caspase-8 inhibitor attenuated cell death significantly, supporting the notion that only the intrinsic apoptotic pathway is utilized to achieve cell death. Finally, overexpression of a mutant caspase-9 with dominant negative phenotype (caspase-9dn) in MN9D cells and primary dopaminergic neurons via the adenovirus and adenoassociated virus gene delivery system, respectively, conferred marked increases in tolerance to the toxicity of 6-OHDA. These results point to the intrinsic caspase-9/caspase-3 cascade as the predominant signaling pathway underlying dopaminergic cell death induced by 6-OHDA and suggest that gene delivery of caspase-9dn can attenuate this pathway and its degenerative consequences.