**5.3 IR, IGF-1R signalling and Alzheimer's disease**

The IR and IGF-1R signalling pathway is disturbed in the central nervous system (CNS) of AD patients (Frolich et al. 1998; Frolich et al. 1999; Moloney et al. 2010). Analysis of the mRNA level of insulin and the IR showed a decrease of about 80% in AD patients. Additionally, the expression of the IGF-1R was reduced in AD brains compared to controls (Moloney et al. 2010; Rivera et al. 2005). In contrast, the IGF-1 serum levels of AD patients are increased indicating IGF-1 resistance in AD (Rivera et al. 2005; Vardy et al. 2007). Furthermore, IRS-1 and -2 expression is reduced in AD brains and phosphorylation of IRS-1 at Ser312 and Ser616 is increased, which decreases IRS-1 action characterising AD as "brain type" diabetes (Pilcher 2006)**.** Thus, brains of AD patients are insulin and IGF-1 resistant. Whether these changes are cause or consequence of neurodegeneration is a matter of debate. IGF-1 knockout mice display increase of tau phosphorylation at Ser396 and Ser202 while the tau protein level was not influenced (Cheng et al. 2005). In NIRKO mice, the brain-specific IR knockout mice, tau was hyperphosphorylated at Thr231 (Schubert et al. 2004), whereas IRS-2 knockout mice showed hyperphosphorylation at Ser202 (Schubert et al. 2003). The different phosphorylation patterns of tau in different insulin and IGF-1 resistant mouse models indicate that additional factors may play a role for tau phosphorylation in these models (Freude et al. 2009).

Tg2576 mice express the Swedish mutation of APP (APPsw) and are an established mouse model for analysing amyloid pathology (Vassar et al. 1999; De Strooper 2003; Harada et al. 2006). IRS-2 (IRS-2-/-) or neuron specific IGF-1R knockout (nIGF-1R-/-) in Tg2576 mice protects these mice from premature death and decreases Aβ-accumulation (Freude et al. 2009).

BACE-1 and Presenilin-1/-2, which cleave APP and generate neurotoxic Aβ42, are possible targets for AD treatment since β-secretase cleavage is the rate limiting step of Aβ generation. During ageing, the expression of the neurotrophin receptor tyrosine kinase receptor A (TrkA) and the p75 neurotrophin receptor (p75NTR) changes considerably. Whereas TrkA receptor expression decreases, the p75 neurotrophin receptor increases with age. Human neuroblastoma cells SHSY5Y and primary cultured neurons showed a switch from TrkA to p75NTR expression after treatment with IGF-1 (Costantini, Scrable, and Puglielli 2006). This increases BACE-1 activity via hydrolysis of sphingomyelin and release of ceramide stabilising BACE-1 (Puglielli 2008; Puglielli et al. 2003). It has been shown that embryonic hippocampal neurons treated with Aβ42 as ligand of p75NTR cause cell death. Neurons, which are deficient in p75NTR and also treated with Aβ42, show less cell death. This may represent the molecular mechanism linking IR and IGF-1R signalling pathway to ageing and neurodegeneration (Sotthibundhu et al. 2008).

In *Caenorhabditis elegans* the knockdown of DAF-2, the orthologue of mammalian IR and IGF-1R, reduces Aβ42 toxicity (Cohen et al. 2006). This reduced Aβ42 toxicity results from the activity of the downstream transcription factors DAF-16, the orthologue of mammalian FoxO1 and 3a as well as heat shock transcription factor-1 (HSF-1) (Hsu, Murphy, and Kenyon 2003; Birkenkamp and Coffer 2003; Cohen et al. 2006). The

Role of Central Insulin-Like Growth Factor-1

1998), have been performed.

McNeill 2006; Chrysis et al. 2001).

**6.1 IGF-1 signalling in** *C. elegans*

mammalian FoxO1 and FoxO3a.

**6.2 IGF-1 signalling in** *Drosophila melanogaster*

Ruvkun 1998).

(Murakami 2007).

2004).

Receptor Signalling in Ageing and Endocrine Regulation 227

activation sequence) in *Drosophila*, as well as RNAi (RNA interference) incorparation via feeding of bacteria to *C. elegans* and Cre/loxP system in mice (Roman et al. 2001; Sauer

The CNS is responsible for endocrine release of insulin-like peptides which activate the IR/IGF-1R signalling cascade and subsequently shorten lifespan (Ikeya et al. 2002; Broughton et al. 2005). Confusingly, acute increase of IR/IGF1-R signalling is neuroprotective but reduction of the signalling causes lifespan extension (Bateman and

The impact of IR/IGF-1R-like signalling (IIS) pathway on lifespan was first discovered in *C. elegans*. IIS in *C. elegans* is similar to the pathway in mammals (Taguchi and White 2008). It is activated through binding of insulin-like peptides (INS) to DAF-2 (abnormal Dauer formation-2), the IR/IGF-1R in worms (Kenyon et al. 1993). Upon stimulation of DAF-2 an insulin receptor substrate 1 orthologue (IST-1) is recruited to DAF-2 and AGE-1 (AGEing alteration-1), orthologue to human p110. P110, the catalytic subunit of PI3K, promotes the generation of phosphatidylinositide-triphosphate (PI3,4,5P) which then activates AKT kinase family members (Morris, Tissenbaum, and Ruvkun 1996; Kops et al. 1999; Paradis and

The mutations of DAF-2 and AGE-1 cause lifespan extension in a DAF-16 dependent manner which is a forkhead transcription factor in worms (Kenyon et al. 1993; Lin et al. 1997; Ogg et al. 1997; Morris, Tissenbaum, and Ruvkun 1996) and homologue to the

AGE-1 and DAF-2 mutant worms show enhanced thermotaxis learning behaviour during ageing. This might be due to resistance to neuronal diseases and oxidative stress

The insulin-like peptides (INS) are found throughout the whole body of *C. elegans* (Pierce et al. 2001; Li, Kennedy, and Ruvkun 2003). Specific sensory neurons regulate lifespan through DAF-16 (Alcedo and Kenyon 2004). The ablation of olfactory neurons causes lifespan extension being only partially dependent on DAF-16 indicating an involvement of other factors. The regulation of INS is not well analysed, but the sensory neurons seem to be the major source of INS for peripheral tissues (Alcedo and Kenyon

A study using tissue-specific expression of AGE-1 and DAF-16 revealed that wild-type AGE-1 recovers the extended lifespan of AGE-1 mutants. This was shown for wild-type AGE-1 expressed in neurons and the intestine. In addition, wild-type DAF-16 rescues reduced lifespan of DAF-16/AGE-1 double mutants when expressed in neurons or intestine. Moreover, wild-type DAF-16 exerts its full effect when expressed in neurons and the intestine exclusively. However, DAF-16 expression showed only minor effects when expressed in other tissues (Broughton and Partridge 2009; Iser, Gami, and Wolkow 2007).

*D. melanogaster* posseses endocrine tissues, which are similar to mammals (Toivonen and Partridge 2009). The median neurosecretory cells (mNSCs) of *Drosophila* are localised in the pars intercerebralis. mNSCs generate three of the seven *Drosophila* insulin-like peptides (DILPs). These mNSCs are functionally similar to β-cells of the pancreas which produce

detoxification of Aβ42 by decreased DAF-2 signalling involves two possible mechanisms. First, HSF-1 regulates disaggregation of toxic oligomers followed by degradation of the resulting fragments. Second, DAF-16 regulates the formation of aggregates with high molecular weight and low toxicity, which are built from aggregates with low molecular weight but high toxicity (Aβ hyperaggregation) (Cohen et al. 2006). Recently, Aβ hyperaggregation has been identified as a mechanism of Aβ detoxification in an IGF-1 resistant mouse model of AD (Cohen et al. 2009).

Several ways of Aβ clearance from the brain have been discovered contributing to Aβ detoxification. Aβ clearance is achieved via transport over the blood brain barrier, enzymatic degradation and phagocytosis by microglia. Recently, several enzymes have been discovered which degrade Aβ, e.g. insulin degrading enzyme (IDE), endothelin converting enzyme (ECE), neprilysin, and angiotensin converting enzyme (ACE). The expression of IDE is activated by IR and IGF-1R signalling (Zhao et al. 2004). Transport across the blood brain barrier (BBB) is mediated by distinct receptors. This transport is achieved via binding to the low-density lipoprotein receptor related protein (LRP). The binding of Aβ to LRP occurs directly or in complex with APOE (apolipoprotein E) and/or α2-macroglobulin (α2M). After crossing the BBB, Aβ is transported to peripheral tissues for degradation, for example the liver (Tanzi, Moir, and Wagner 2004). It has been proposed that high IGF-1 levels cause degradation or clearance of Aβ. Tg2576 mice present decreased IGF-1 levels compared to wild type mice. The treatment with IGF-1 yield to increased transport of Aβ from the brain, possibly via the choroid plexus (Carro et al. 2002). Accordingly, the inactivation of the IGF-1R signalling in the choroid plexus caused AD-like pathology (Carro et al. 2006). In contrast, studies in rats and Tg2576 mice using acute, subchronic and chronic IGF-1 treatment found no changes in tau phosphorylation and Aβ concentrations (Lanz et al. 2008). Possibly, chronic peripheral treatment with IGF-1 causes downregulation of the IGF-1R signalling pathway as it has been shown for a cohort of individuals with high serum IGF-1 level but low IGF-1R signalling (Suh et al. 2008). This might explain the conflicting results of the different studies. AD mouse models with induced insulin resistance via high fat diet displayed an exacerbation of amyloid pathology (Ho et al. 2004).

The different studies dealing with IR/IGF-1R signalling and AD prove a connection between this signalling pathways and AD pathology. However, the exact molecular mechanisms need to be elucidated.
