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Inheritance of the apoE4 allele (ϵ4) increases the risk of developing Alzheimer's disease; however, the mechanisms underlying this association remain elusive. Recent data suggest that inheritance of ϵ4 may lead to reduced apoE protein levels in the CNS. We therefore examined apoE protein levels in the brains, CSF and plasma of ϵ2/2, ϵ3/3, and ϵ4/4 targeted replacement mice. These apoE mice showed a genotype-dependent decrease in apoE levels; ϵ2/2ϵϵ3/3ϵϵ4/4. Next, we sought to examine the relative contributions of apoE4 and apoE3 in the ϵ3/4 mouse brains. ApoE4 represented 30-40% of the total apoE. Moreover, the absolute amount of apoE3 per allele was similar between ϵ3/3 and ϵ3/4 mice, implying that the reduced levels of total apoE in ϵ3/4 mice can be explained by the reduction in apoE4 levels. In culture medium from ϵ3/4 human astrocytoma or ϵ3/3, ϵ4/4 and ϵ3/4 primary astrocytes, apoE4 levels were consistently lower than apoE3. Secreted cholesterol levels were also lower from ϵ4/4 astrocytes. Pulse-chase experiments showed an enhanced degradation and reduced half-life of newly synthesized apoE4 compared with apoE3. Together, these data suggest that astrocytes preferentially degrade apoE4, leading to reduced apoE4 secretion and ultimately to reduced brain apoE levels. Moreover, the genotype-dependent decrease in CNS apoE levels, mirror the relative risk of developing AD, and suggest that low levels of total apoE exhibited by ϵ4 carriers may directly contribute to the disease progression, perhaps by reducing the capacity of apoE to promote synaptic repair and/or Aβ clearance. [ABSTRACT FROM AUTHOR]

Coupling of the group I metabotropic glutamate receptors, mGlu1a and mGlu5a, to the cAMP response element binding protein (CREB) has been studied in Chinese hamster ovary cell lines where receptor expression is under the control of an inducible promoter. Both receptors stimulate CREB phosphorylation with similar time courses, and agonist potency was also comparable between the two receptors. Stimulation of cells in Ca2+-free medium containing EGTA (100 μm), with or without the additional depletion of intracellular stores, caused marked decreases in agonist-mediated responses in both cell lines. Down-regulation of protein kinase C (PKC) activity by phorbol ester treatment, or treatment with the broad spectrum PKC inhibitor Ro 31–8220, partially attenuated both mGlu1a and mGlu5a receptor-mediated responses. Furthermore, stimulation of cells in the absence of extracellular Ca2+ following prior PKC down-regulation resulted in additive inhibitory effects. The involvement of extracellular signal-regulated kinases (ERK1/2), Ca2+/calmodulin or Ca2+/calmodulin-dependent protein kinases was assessed using pharmacological inhibitors. Results indicated that coupling of the group I mGlu receptors to CREB phosphorylation occurs independently of these pathways. Thus, although the[Ca2+]i signatures activated by these mGlu receptors differ, they couple to CREB with comparable potency and recruit similar downstream components to execute CREB phosphorylation. [ABSTRACT FROM AUTHOR]