**3. IGF-1 signalling in the brain**

#### **3.1 IGF-1's transport across the blood brain barrier**

In the brain, present IGFs arise either by de novo synthesis or by transport from the blood into the brain. So far, the exact mechanisms of how IGFs cross the blood-brain barrier (BBB) are not fully understood. Early *in vitro* studies by Duffy and colleagues on isolated human brain capillaries showed that the affinity of IGF-2 to isolated human brain capillaries was approximately twofold higher than the affinity of IGF-1 binding and binding of both IGFs was nonsaturable over the range of 1 to 200 ng/ml IGF suggesting that there is a very efficient endocytosis mechanism (Duffy, Pardridge, and Rosenfeld 1988). Insulin displaced binding of IGF-1 (50% inhibited by 2 µg/ml Insulin) and IGF-2 (50% inhibited by 0,5 µg/ml Insulin). Furthermore, binding was largely inhibited by adding human serum, which was assumed to be due to the presence of IGFBPs (Duffy, Pardridge, and Rosenfeld 1988). Subsequent studies suggested that there might be a high capacity transport system across the BBB, which is influenced by the IGFBPs (Pan and Kastin 2000). Tores-Aleman and coworkers found that via the choroid plexus epithelium, circulating IGF-1 is transported into the CSF through a mechanism involving the multicargo protein tranporter low-density lipoprotein receptor related protein 2 (LRP2), and concluded that increasing levels of IGF-1 in the CSF, as a result of increasing IGF-1 levels in the serum, might be explained via this transport mechanism (Carro et al. 2000; Carro et al. 2005). LRP1, another membrane cargotransporter, was suggested as targeting platform for the circulating IGF-1/IGFBP-3/ALScomplex (Nishijima et al. 2010). LRP1 is abundantly expressed in brain endothelium and seems to be a cellular receptor for IGFBP-3 (Huang et al. 2003). Recently, it has been demonstrated that neuronal activity increases the permeability of the BBB for IGF-1 through neurovascular coupling (Nishijima et al. 2010). Therefore, serum IGF-1 might influence brain processes like synaptic plasticity and cognition. Furthermore, neurovascular coupling was proposed to locally change cerebral blood flow leading to activation of matrix metalloprotease 9 (MMP9) through diverse mediators (arachdonic acid derivates, ATP, etc.). MMP9 is an IGFBP-3 cleaving protease, which is released in response to neuronal activity (Michaluk and Kaczmarek 2007) and might link neuronal activation to the transport of serum IGF-1 via a LRP1 depending mechanism across the BBB (Nishijima et al. 2010). Hence, neurovascular coupling might result in neurotrophic coupling explaining the neuroprotective effects of physical as well as mental exercise and active social life on brain function (Carro et al. 2000; Fratiglioni, Paillard-Borg, and Winblad 2004).

#### **3.2 Brain IGF receptors**

IGF-1 and to less extend IGF-2 bind and therefore activate the type 1 IGF-receptor (IGF-1R) (Rubin and Baserga 1995; White and Kahn 1994; LeRoith 2000). Several IGF-1R subtypes have been reported, for example the hybrid IGF-1Rs, which are hybrid dimers of the insulinreceptor (IR) and the IGF-1R (see 2.3) binding insulin as well as IGFs with similar affinity (Soos and Siddle 1989; Soos et al. 1990; Siddle et al. 1994; Moxham, Duronio, and Jacobs 1989; Pandini et al. 2002). The physiological significance of these receptor subtypes is still unclear (Russo et al. 2005).

Additionally, there is a type 2 IGF-receptor (IGF-2R) with a short cytoplasmic domain lacking a tyrosine kinase harbouring a higher affinity for IGF-2 than the IGF-1R (Sakano et al. 1991; Oh et al. 1991; Dore, Kar, and Quirion 1997; Braulke 1999; Kiess et al. 1994). This IGF-2R is a cation-independent mannose-6 phosphate (M6P) receptor binding not only IGF-

In the brain, present IGFs arise either by de novo synthesis or by transport from the blood into the brain. So far, the exact mechanisms of how IGFs cross the blood-brain barrier (BBB) are not fully understood. Early *in vitro* studies by Duffy and colleagues on isolated human brain capillaries showed that the affinity of IGF-2 to isolated human brain capillaries was approximately twofold higher than the affinity of IGF-1 binding and binding of both IGFs was nonsaturable over the range of 1 to 200 ng/ml IGF suggesting that there is a very efficient endocytosis mechanism (Duffy, Pardridge, and Rosenfeld 1988). Insulin displaced binding of IGF-1 (50% inhibited by 2 µg/ml Insulin) and IGF-2 (50% inhibited by 0,5 µg/ml Insulin). Furthermore, binding was largely inhibited by adding human serum, which was assumed to be due to the presence of IGFBPs (Duffy, Pardridge, and Rosenfeld 1988). Subsequent studies suggested that there might be a high capacity transport system across the BBB, which is influenced by the IGFBPs (Pan and Kastin 2000). Tores-Aleman and coworkers found that via the choroid plexus epithelium, circulating IGF-1 is transported into the CSF through a mechanism involving the multicargo protein tranporter low-density lipoprotein receptor related protein 2 (LRP2), and concluded that increasing levels of IGF-1 in the CSF, as a result of increasing IGF-1 levels in the serum, might be explained via this transport mechanism (Carro et al. 2000; Carro et al. 2005). LRP1, another membrane cargotransporter, was suggested as targeting platform for the circulating IGF-1/IGFBP-3/ALScomplex (Nishijima et al. 2010). LRP1 is abundantly expressed in brain endothelium and seems to be a cellular receptor for IGFBP-3 (Huang et al. 2003). Recently, it has been demonstrated that neuronal activity increases the permeability of the BBB for IGF-1 through neurovascular coupling (Nishijima et al. 2010). Therefore, serum IGF-1 might influence brain processes like synaptic plasticity and cognition. Furthermore, neurovascular coupling was proposed to locally change cerebral blood flow leading to activation of matrix metalloprotease 9 (MMP9) through diverse mediators (arachdonic acid derivates, ATP, etc.). MMP9 is an IGFBP-3 cleaving protease, which is released in response to neuronal activity (Michaluk and Kaczmarek 2007) and might link neuronal activation to the transport of serum IGF-1 via a LRP1 depending mechanism across the BBB (Nishijima et al. 2010). Hence, neurovascular coupling might result in neurotrophic coupling explaining the neuroprotective effects of physical as well as mental exercise and active social life on brain

**3. IGF-1 signalling in the brain** 

**3.1 IGF-1's transport across the blood brain barrier** 

function (Carro et al. 2000; Fratiglioni, Paillard-Borg, and Winblad 2004).

IGF-1 and to less extend IGF-2 bind and therefore activate the type 1 IGF-receptor (IGF-1R) (Rubin and Baserga 1995; White and Kahn 1994; LeRoith 2000). Several IGF-1R subtypes have been reported, for example the hybrid IGF-1Rs, which are hybrid dimers of the insulinreceptor (IR) and the IGF-1R (see 2.3) binding insulin as well as IGFs with similar affinity (Soos and Siddle 1989; Soos et al. 1990; Siddle et al. 1994; Moxham, Duronio, and Jacobs 1989; Pandini et al. 2002). The physiological significance of these receptor subtypes is still

Additionally, there is a type 2 IGF-receptor (IGF-2R) with a short cytoplasmic domain lacking a tyrosine kinase harbouring a higher affinity for IGF-2 than the IGF-1R (Sakano et al. 1991; Oh et al. 1991; Dore, Kar, and Quirion 1997; Braulke 1999; Kiess et al. 1994). This IGF-2R is a cation-independent mannose-6 phosphate (M6P) receptor binding not only IGF-

**3.2 Brain IGF receptors** 

unclear (Russo et al. 2005).

2 but M6P-containing ligands. This receptor functions in the mediation of endocytosis and lysosomal enzyme trafficking and regulation of apoptotic/ mitogenic effects (Morgan et al. 1987; von Figura and Hasilik 1986; Ghahary et al. 2000). Recent studies have demonstrated that IGF-2 influences memory enhancement via the IGF-2R suggesting that the IGF-2R might transmit some intracellular signalling (Chen et al. 2011).
