Mitochondria can be delivered along the axon in association with microtubules, which is important for supplying energy required to maintain neuronal functions. During axonal transport, mitochondria are associated with several motor proteins, such as kinesin for anterograde transport and dynein for retrograde transport. Adaptor proteins, such as Miro and Milton, are connected to mitochondria through kinesin. Although impaired axonal transport of mitochondria has been reported in the presence of Ab, the precise mechanism for this Abinduced impairment remains unclear. In the present study, we attempted to elucidate the mechanism that links the acetylation of GNTI dihydrochloride a-tubulin and Ab-induced impairment of mitochondrial transport in hippocampal neurons cultured in a microfluidic system. To increase a-tubulin acetylation, we used the GSK 4112 Tubastatin A as the HDAC6 inhibitor. Mitochondrial axonal transport was analyzed by measuring the velocity, motility and length of mitochondria. We found that pharmacological inhibition of HDAC6 significantly restored the compromised velocity and motility of the mitochondria of Ab hippocampal neurons to a normal level in both anterograde and retrograde axonal transports. The inhibition of HDAC6 also recovered the length of mitochondria that had been shortened by Ab. These results show that the inhibition of HDAC6 rescued neuronal cells from Ab-induced impairment of mitochondrial axonal transport as well as mitochondrial length, identifying HDAC6 as a potential therapeutic target to modulate AD pathogenesis. It has been reported that Ab alters the level of acetylated atubulin in primary neuronal cultures and cell lines whereas the HDAC6 inhibitor, TBA, promotes the acetylation of a-tubulin. To examine the role of Ab in reducing acetylated atubulin, primary hippocampal neurons were characterized. Impaired axonal transport and mitochondrial dysfunctions occur in the early stages of AD. These changes are induced by Ab which forms amyloid plaques, one of the major hallmarks of AD. Increased GSK3b activity induces hyperphosphorylation of tau, resulting in the formation of neurofibrillary tangles and/or an increase in Ab generation. HDAC6 activity, as a substrate of GSK3b, is increased by GSK3b activation, suggesting a decrease in both the level of atubulin acetylation and axonal transport. To modulate HDAC6 activity, we used TBA as a potent HDAC6 inhibitor, which is more selective than other HDAC6 inhibitors, such as TSA and tubacin. The specificity of TBA to HDAC6 was confirmed in previous studies by homology modeling and enzyme inhibition experiments using 11 HDAC isozymes. TBA affects the acetylation of cytosolic proteins like a-tubulin but not histones. The knockdown of HDAC6 by siRNA resulted in an increased level of acetylated a-tubulin, consistent with the results with the HDAC6 inhibitor.