Aβ aggregates are found in the CNS in aging healthy humans and in AD patients. In AD patients these aggregates appear to mainly damage neurons, causing severe brain atrophy in late stages of the disease. However, Aβ aggregates are widespread and are found around neurons, microglia, astrocytes, smooth muscle cells in the vasculature, epithelial cells of the eye, and the choroid plexus. The pathogenesis of these accumulations is not clearly defined. The basis for selective vulnerability of certain tissues and cell types for Aβ aggregates is a largely unknown territory. Drosophila disease models allows for modeling progressive disease and reduction in overall fitness of the fly depending on expressed Aβ variant. We previously conducted a systematic study of 16 different Aβ variants, finding that Aβ1-42 and Aβ3-42 were by far the most neurotoxic variants (). However, the significance of studying Aβ1-42 expression in different cell types of the CNS has not been thoroughly addressed. In the present study, we hence aimed to investigate whether the Aβ1-42 toxicity and aggregation pattern observed in CNS neurons of Drosophila can be observed in other cell types. The transgenic models utilized for this comparative study take advantage of the GAL4-UAS system (). We used the well-studied driver C155-elav-Gal4, directing expression to both neural progenitors and mature neurons of the CNS (); the improved and stronger n-syb-Gal4 driver, directing expression to neurons (); the motor neuron expressing driver D42-Gal4 (); the widely used Drosophila glial driver repo-Gal4 that is expressed in most glial cells in the CNS (; M.R.); and the well-studied GMR-Gal4 driver, directing expression to the fly eye (M.). Our results show that all cell types expressing Aβ1-42 are able to form amyloid aggregates. However, the toxicity is very different: Aβ1-42 in neurons is very toxic, but is less toxic when expressed in glial cells.

Xiong and Montell, 1995 Xiong W.-C.

Montell C. Defective glia induce neuronal apoptosis in the repo visual system of Drosophila.

MacDonald et al., 2006 MacDonald J.M.

Beach M.G.

Porpiglia E.

Sheehan A.E.

Watts R.J.

Freeman M.R. The Drosophila cell corpse engulfment receptor draper mediates glial clearance of severed axons.

Freeman and Doherty, 2006 Freeman M.R.

Doherty J. Glial cell biology in Drosophila and vertebrates.

Wang et al., 2011 Wang L.

Colodner K.J.

Feany M.B. Protein misfolding and oxidative stress promote glial-mediated neurodegeneration in an Alexander disease model.

Colodner and Feany, 2010 Colodner K.J.

Feany M.B. Glial fibrillary tangles and JAK/STAT-mediated glial and neuronal cell death in a Drosophila model of glial tauopathy.

Kretzschmar et al. (2005) Kretzschmar D.

Tschäpe J.

Bettencourt Da Cruz A.

Asan E.

Poeck B.

Strauss R.

Pflugfelder G.O. Glial and neuronal expression of polyglutamine proteins induce behavioral changes and aggregate formation in Drosophila.

Tamura et al. (2009) Tamura T.

Sone M.

Yamashita M.

Wanker E.E.

Okazawa H. Glial cell lineage expression of mutant Ataxin-1 and huntingtin induces developmental and late-onset neuronal pathologies in Drosophila models.

Shiraishi et al., 2014 Shiraishi R.

Tamura T.

Sone M.

Okazawa H. Systematic analysis of fly models with multiple drivers reveals different effects of ataxin-1 and huntingtin in neuron subtype-specific expression.

Caesar et al., 2012 Caesar I.

Jonson M.

Nilsson K.P.

Thor S.

Hammarström P. Curcumin promotes a-beta fibrillation and reduces neurotoxicity in transgenic Drosophila.

The roles of glial cells in modulating neurodegenerative diseases in Drosophila models have begun to be studied in recent years. Glial cells in Drosophila perform similar functions as they do in vertebrates, e.g., providing various support functions to neurons during development, acting on injury to the nervous system, being responsible for the phagocytic clearance of cellular debris and maintaining neuronal survival (; M.R.). To our knowledge, previous studies expressing Aβ1-42 in Drosophila have not targeted glial cells. Our control experiments showed that expression of the cell death gene rpr rendered no transgenic progeny to eclose carrying both Gal4 and UAS-rpr in glia (repo-Gal4). This cross provided an identical outcome as for neuronal expression (n-syb-Gal4) of this cell death protein. Hence, healthy glia cells are essential for Drosophila. This shows that the mild phenotype we observe despite the heavy Aβ1-42 aggregate load must be a result of low sensitivity of glial cells toward Aβ1-42. Studies expressing other proteins causing human neurodegeneration in glial cells of Drosophila reported significant toxicity mediated by protein aggregation from mutant human GFAP associated with Alexander disease (). Expressing human Tau in glial cells resulted in the formation of fibrillary tangles and a reduced lifespan as well as a non-cell-autonomous neuronal cell death (). This study also found that the expression of Tau in both glial cells and neurons caused a striking enhancement of toxicity compared with expression in the respective tissue by itself. Studies byandfound that when expressing the poly-Q proteins Htt and Atxn-1 in neurons and glia, only glia expression resulted in degeneration of the nervous system similarly to findings from (). These studies highlight the different responses that can be seen in neurons and glia. However, and most importantly, the observation of poly-Q and Tau stands in stark contrast to the outcome of our study, where we see a much stronger response in neurons than we do when expressing Aβ1-42 in glial cells. The Aβ1-42 aggregates found in neurons appear intracellular, filling up the soma. The shape strongly suggests that the Aβ1-42 starts to aggregate in the secretory pathway and forms ring-tangles. In contrast, glial expression and eye expression results in extended fibrous aggregates away from the cell soma. Using both the GMR-Gal4 (eye) and the repo-Gal4 (glia) drivers, the expressing cells are spatially separated from the fibrous aggregates and appear mainly extracellular. Although we lack direct evidence it is likely that these aggregates can be detrimental to neighboring cells. The consistent reduction in life span and activity of the glia expressing flies indicates that non-cell-autonomous neurotoxicity may be operative here. One particularly interesting topic in the field of neurodegeneration pertains to the identification of specific structural species associated with toxicity. We have previously showed that immature oligomeric Aβ1-42 aggregates, including protofibrils formed from the Arctic mutation E22G, are severely neurotoxic and showed a pronounced cytotoxic effect in the eye with the GMR-Gal4 driver ( Figure S3 B). However, neurotoxicity could be mitigated by small-molecule therapy that shifted the conformational states toward mature fibrils at the expense of oligomers (). In the current study, we turned to our novel assay for amyloid fibril maturation morphotype staging, combining two LCOs q-FTAA and h-FTAA for fluorescence microspectroscopy. Glial Aβ1-42 aggregates showed a spectral profile indicating significantly more mature fibrils than that observed in aggregates triggered by each of the three neuronal drivers. These data strongly suggest that the toxic species in Drosophila are immature nascent Aβ1-42 fibrils formed intracellularly. To further assess the toxicity differences as a function of aggregate structure we showed that mutant Aβ1-42 A42W with impaired spontaneous fibril formation capacity was more toxic to the fly eye in vivo than mature fibrillar Aβ1-42 WT despite minor accumulation of mutant protein, while less toxic than Aβ1-42 E22G. In glial cells on the other hand this intrinsic property of Aβ1-42 A42W was overridden by some endogenous factor that promoted fibril maturation and hence low cytotoxicity. These results underline that amyloidogenic protein sequence, local concentration, cell-specific protein homeostasis factors, and aggregate conformation are all influencing Aβ1-42 cytotoxicity.