This research project investigates the role of lipid droplets (LDs) in neuronal survival and dysfunction, focussing on two human in vitro models of neurodegenerative diseases (NDs): neuroblastoma SH-SY5Y cell line exposed to 6-OHDA for Parkinson’s, and primary human fibroblasts from Alzheimer’s disease (AD) patients.Valid and reliable experimental tools for LDs detection and quantification were set up in SH-SY5Y. Confocal laser scanning microscopy was combined with image analysis, allowing an automatic and objective quantification of LDs in basal conditions, and in response to LDs modulation and/or mitochondrial stress. Moreover, mitochondrial morphology was evaluated in all experimental conditions.No differences were observed in SH-SY5Y exposed to 6-OHDA compared to untreated cells. However, when subjected to LDs stimulation, the 6-OHDA-treated cells produced higher LDs quantities compared to the controls. This increase could be completely blocked by pharmacological inhibition of LDs synthesis.Human fibroblasts exhibited considerable interindividual diversity in LDs abundance, yet there was no statistically significant difference in LDs content observed in two out of three pairs of AD-healthy patients’ samples. AD fibroblasts showed an increased mitochondrial interconnectivity compared to matching controls. While the importance of lipid metabolism in the mechanisms of NDs is acknowledged, the specific changes in lipid content and metabolism, as well as the role of lipids in mitochondrial stress and neuronal function, remain unclear in normal and pathological states.This work provides tools and first observations for understanding the role of LDs and their possible influence on mitochondria in relevant PD and AD in vitro models.