This is Part 2 of a two-part story. See Part 1.
16 February 2012. People carrying one copy of the apolipoprotein E4 gene variant are up to four times more likely to develop Alzheimer’s disease, and that risk rises up to 15-fold in E4 homozygotes—but what makes the protein so harmful? Research suggests that ApoE4 contributes to AD in various ways, some via Aβ, others independent of it (Huang, 2010). At an ApoE workshop held 30 January to 1 February at the Gladstone Institute of Neurological Disease in San Francisco, speakers reported new data along both fronts. Conference organizer Yadong Huang of Gladstone spoke about his lab’s characterization of aging ApoE4 knock-in mice. The data squarely blame the animals’ cognitive deficits on a specific set of cells—GABAergic interneurons in the hilus of the dentate gyrus. For his part, Steven Paul of Weill Cornell Medical College in New York City reported that, surprisingly, ApoE4 speeds neurofibrillary tangle formation and triggers possible neurodegeneration in old AD transgenic mice. Paul found that ApoE4 knock-in mice have abnormal electrophysiology, even seizures. Together, the studies help flesh out ApoE4’s harmful effects at the cellular level. Other researchers reported on the consequences of genetic or chemical modulation of ApoE on cells and animal models, including changes in Aβ dynamics (see Part 1 of this series).
In a prior study, Yaisa Andrews-Zwilling of the Huang lab, and colleagues, reported that female ApoE4 knock-in mice (which express human ApoE4 in place of the endogenous mouse gene) lose hilar GABAergic interneurons as they age (Andrews-Zwilling et al., 2010). By 16 months, cell counts dropped 30 percent. The reduction correlated with learning and memory deficits, which were rescued by a drug that activates GABAA receptors. At the time, the researchers did not know if the neuronal loss was the cause of the cognitive decline or an effect of a broader pathology. However, at the Gladstone meeting, Huang noted that old male ApoE4 knock-in mice have normal numbers of hilar GABAergic interneurons and do not show cognitive deficits. This otherwise unexplained sex difference suggests to Huang that retaining these neurons is key to staying cognitively sharp, and may make males resistant to ApoE4-induced problems.
To address whether loss of hilar GABAergic interneurons in fact causes the cognitive decline, Andrews-Zwilling and Anna Gillespie of the Huang lab used optogenetics to deactivate these cells in living wild-type mice (see Fenno et al., 2011 for a review of the technology). The researchers expressed a light-sensitive chloride pump in hilar GABAergic interneurons; the pump responds to yellow laser light, hyperpolarizing the cells and thereby shutting down their activity. A one-minute pulse of light—shone through glass fibers inserted in the dentate gyrus of freely moving animals—was enough to make wild-type mice perform as poorly as aging ApoE4 knock-in mice on spatial learning and memory tests.
What role do these interneurons play in memory and learning? Huang’s team tested whether optogenetic inhibition of the hilar GABAergic interneurons impairs memory retention, or retrieval, or both. They found that mice expressing the light-sensitive chloride pump learn the location of the hidden platform of the Morris water maze just fine if the laser light is kept off during five days of training trials. The mice forget the location if the light is switched on 24 hours after the last learning trial; but, the animals recall the platform location 48 hours later with the laser turned off. “That means there is a problem with memory retrieval but not retention. In Huang’s view, this sort of cognitive malfunction resembles the predicament of early AD patients, “who cannot remember something today, but tomorrow might remember it,” said Huang.
As a first step toward addressing whether the neuronal deficits in aging ApoE4 mice resemble AD, Huang’s team used induced pluripotent stem (iPS) cells to generate human neurons with ApoE3/E3 or ApoE4/E4 genotypes. The procedure is slow, requiring six to eight weeks to culture the iPS cells and differentiate them into neurons. However, immunostaining and Western blotting suggest that ApoE4 iPS cells make fewer GABAergic interneurons and have increased tau phosphorylation, said Huang.
The theme of tau pathology stood out in Paul’s presentation as well. In collaboration with Patrick Sullivan at Duke University School of Medicine, Durham, North Carolina, Paul and colleagues found evidence for hyperphosphorylated tau, neurofibrillary tangles, and even neurodegeneration in the hippocampus of 15-month-old PDAPP mice expressing human ApoE4—but not in PDAPP mice expressing mouse ApoE or human ApoE3 or E2, or in wild-type mice just expressing human ApoE isoforms (including E4). “Thus, it appears that ApoE4 has amyloid-independent effects in facilitating tau pathology, but only in the presence of human Aβ,” Paul noted in an e-mail to ARF.
In a separate set of studies, Paul reported that about 60 percent of female ApoE4 knock-in mice, and 40 percent of males, have seizure behaviors. These behaviors are mostly mild, for example, head jerks. Nearly a quarter of ApoE4 animals—but not E3 or E2 knock-in mice—have severe clonic (i.e., rhythmic muscle contraction/relaxation) seizures, as well as a lower threshold for pentylenetetrazol-induced seizures and abnormal brain electrical activity measured by electroencephalography (EEG). “This may be the best model of AD-associated seizure disorder, and it is an amyloid-independent behavior,” Paul noted. Scientists have reported epileptiform activity and network instability in several strains of AD transgenic mice (see ARF related news story on Palop et al., 2007; ARF related conference story) and in people with AD or mild cognitive impairment (see ARF related news story).—Esther Landhuis.
This is Part 2 of a two-part story. See Part 1.