Parkinson’s disease is characterized by the progressive loss of dopaminergic (DA) neurons in the substantia nigra. The heat-shock protein 70 (Hsp70) reduces protein misfolding and aggregation. It has been shown to protect cells against oxidative stress and apoptotic stimuli in various neurodegenerative disease models. To deliver Hsp70 across cellular membranes and into the brain, we linked it to a cell-penetrating peptide derived from the HIV trans-activator of transcription (Tat) protein. In vitro, Tat-Hsp70 transduced neuroblastoma cells and protected primary mesencephalic DA neurons and their neurites against MPP+-mediated degeneration. In vivo, the systemic application of cell-permeable Hsp70 protected DA neurons of the substantia nigra pars compacta against subacute toxicity of MPTP. Furthermore, Tat-Hsp70 diminished the MPTP induced decrease in DA striatal fiber density. Thus, we demonstrate that systemically applied Tat-Hsp70 effectively prevents neuronal cell death in in vitro and in vivo models of Parkinson’s disease. The use of Tat-fusion proteins might therefore be a valuable tool to deliver molecular chaperones like Hsp70 into the brain and may be the starting point for new protective strategies in neurodegenerative diseases.