Abnormal accumulation of undigested macromolecules, often disease-specific, is a 
major feature of lysosomal and neurodegenerative disease and is frequently 
attributed to defective autophagy. The mechanistic underpinnings of the 
autophagy defects are the subject of intense research, which is aided by genetic 
disease models. To gain an improved understanding of the pathways regulating 
defective autophagy specifically in juvenile neuronal ceroid lipofuscinosis 
(JNCL or Batten disease), a neurodegenerative disease of childhood, we developed 
and piloted a GFP-microtubule-associated protein 1 light chain 3 (GFP-LC3) 
screening assay to identify, in an unbiased fashion, genotype-sensitive small 
molecule autophagy modifiers, employing a JNCL neuronal cell model bearing the 
most common disease mutation in CLN3. Thapsigargin, a sarco/endoplasmic 
reticulum Ca(2+)-ATPase (SERCA) Ca(2+) pump inhibitor, reproducibly displayed 
significantly more activity in the mouse JNCL cells, an effect that was also 
observed in human-induced pluripotent stem cell-derived JNCL neural progenitor 
cells. The mechanism of thapsigargin sensitivity was Ca(2+)-mediated, and 
autophagosome accumulation in JNCL cells could be reversed by Ca(2+) chelation. 
Interrogation of intracellular Ca(2+) handling highlighted alterations in 
endoplasmic reticulum, mitochondrial, and lysosomal Ca(2+) pools and in 
store-operated Ca(2+) uptake in JNCL cells. These results further support an 
important role for the CLN3 protein in intracellular Ca(2+) handling and in 
autophagic pathway flux and establish a powerful new platform for therapeutic 
screening.