Alterations of sphingolipids have been observed in many neurodegenerative disorders. However, the contribution of these changes to disease pathogenesis is not clear 
. We have uncovered two spontaneous mutants with progressive degeneration of cerebellar Purkinje neurons and widespread lipofuscin accumulation. By positional cloning, we identified the mutations as alleles of the Lass1
gene, which encodes ceramide synthase 1 (CerS1), one of the six ceramide synthases in mammals. Our data demonstrate that deficiency of CerS1/LASS1 leads to dramatic alterations in the levels of sphingolipids in the brain, degeneration of cerebellar Purkinje neurons, and widespread lipofuscin accumulation.
CerS1 is specifically expressed in neurons in the brain and has been shown to produce C180
ceramide species in vitro
or in cultured cells 
. In the brain of Lass1
mutant mice, C18
sphingolipid species were significantly reduced, confirming the role of CerS1 in C18
ceramide biosynthesis in vivo
. Loss of CerS1 also led to a decrease in total ceramide, in agreement with C18
ceramide as a major ceramide in the adult brain 
. However these sphingolipid decreases were accompanied by an increase in the steady state levels of C14
sphingolipid species. In addition, sphingoid bases and their phosphorylated metabolites were drastically increased in the mutant brain.
In contrast to previous reports suggesting that ceramide is proapoptotic in vitro
and can mediate both stress-induced intrinsic and death receptor-mediated extrinsic apoptosis 
, our data suggest that reduction of ceramide or complex sphingolipids results in progressive neuron death, particularly Purkinje cell loss. Intracerebroventricular administration of global ceramide inhibitors has been reported to cause acute neurodegeneration 
. Contrary to in vitro
data, this finding, together with our results, demonstrate that decreases in ceramide synthesis can induce neuron death in vivo
. Sphingolipid homeostasis is critically balanced in the cell under normal conditions, and thus either increases or decreases of ceramide could be detrimental to the cell. Alternatively, ceramide species with different fatty acyl chains may play distinct physiological roles. For example, loss of one of the two worm ceramide synthases, which produce different ceramide species, resulted in opposite outcomes in C. elegans
under hypoxic conditions 
. Perhaps C18
ceramide, or some of its derived sphingolipids such as C18
-sphingomyelin or C18
-cerebrosides, act in a prosurvival fashion in neurons, whereas an increase in other ceramide species, such as C16
ceramide, may be apoptotic 
. Lastly, Purkinje neuron loss may be due to an increase in the sphingoid bases that normally serve as substrates for CerS1. Sphingoid bases, particularly their phosphorylated metabolites, are potent signaling molecules and have been proposed to regulate many signal transduction pathways 
. Thus, elevation of these molecules, particularly dhS and dhS1P, which are increased over ten fold in the CerS1-deficient brain, may impair normal functions of sphingolipid signaling, leading to Purkinje cell death.
Consistent with previous reports demonstrating that sphingolipid biosynthesis is crucial for dendritic development in cultured Purkinje or hippocampal neurons 
, we observed shortened dendritic arbors and decreased density of calbindin immunostaining in the molecular layer in CerS1-deficient cerebella suggesting that dendritic development of mutant Purkinje cells was abnormal. The role of sphingolipids in dendritic development is not clear, but may involve sphingolipid-rich lipid rafts, which are implicated in activity-dependent dendritic development and maintenance of dendritic spines in in vitro
. Dendritic dysfunction or morphological anomalies are often associated with neurodegeneration 
. While we cannot rule out a secondary cause for abnormalities in Purkinje cell dendrites in CerS1 mutant mice, it is possible that defects in dendritic development also contribute to death of these neurons.
In addition to Purkinje cell death, our data clearly demonstrate that deficiency of ceramide biosynthesis can cause the accumulation of lipofuscin, which is often observed in aging brains and in some neurodegenerative diseases. Lipofuscin is known to contain both lipids and proteins that may originate from membrane bound organelles such as mitochondria 
. Accumulation of lipofuscin has been linked to reduced lysosomal hydrolytic capacity, based on observations of lipofuscin deposition in mutants with deficiencies in lysosomal proteins 
. Both increases and decreases in ceramide have been shown to induce autophagy, suggesting that the balance of sphingolipid homeostasis is important for autophagy and/or lysosomal functions 
. Alternatively, conditions that increase the load on lysosomes beyond their capacity may also underlie lipofuscin formation. Given the importance of sphingolipids in protein targeting and as components of membranes, reduced sphingolipid levels may increase the demand on, and/or decrease the capacity of lysosomal function by affecting both lysosomal and non-lysosomal membrane dynamics, and targeting of membrane proteins. Increased load of defective membrane organelles and mislocalized membrane proteins could exceed lysosomal hydrolytic capacity of neurons, resulting in lipofuscin formation.
Previous reports suggest ceramide might also be important during aging 
. One of the yeast genes encoding ceramide synthases, LAG1
ssurance gene 1), was identified in a screen for genes whose expression changed over yeast replicative cycles 
. Deletion of LAG1
was associated with an increase of replicative life span in yeast. However, a LASS1
variant exhibiting increased expression, when combined with specific HRAS1
haplotypes, was suggested to contribute to healthy aging and survival in a human population 
. This finding suggests that appropriately increased ceramide levels might be beneficial to some cells in aging animals. Inversely, given the specificity of Purkinje cell loss observed in CerS1-deficient mice, decreased ceramide levels appear more detrimental to neurons, and accelerate aging phenotypes, including lipofuscin accumulation. Thus our results demonstrate that in addition to neurodegeneration, alteration of ceramide levels can accelerate some aspects of aging. More research is needed to elucidate the role of ceramide and other sphingolipids in aging and neurodegeneration.