Cell Tissue Res DOI 10.1007/s00441-014-1996-4

REVIEW

Wnt signalling in neuronal differentiation and development Nibaldo C. Inestrosa & Lorena Varela-Nallar

Received: 2 May 2014 / Accepted: 25 August 2014 # Springer-Verlag Berlin Heidelberg 2014

Abstract Wnts are secreted glycoproteins that play multiple roles in early development, including the differentiation of precursor cells. During this period, gradients of Wnts and other morphogens are formed and regulate the differentiation and migration of neural progenitor cells. Afterwards, Wnt signalling cascades participate in the formation of neuronal circuits, playing roles in dendrite and axon development, dendritic spine formation and synaptogenesis. Finally, in the adult brain, Wnts control hippocampal plasticity, regulating synaptic transmission and neurogenesis. In this review, we summarize the reported roles of Wnt signalling cascades in these processes with a particular emphasis on the role of Wnts in neuronal differentiation and development. Keywords Wnt signalling pathway . Neural progenitor cells . Neuronal differentiation . Neuronal maturation . Adult neurogenesis

N. C. Inestrosa (*) Centro de Envejecimiento y Regeneración (CARE), Departamento de Biología Celular y Molecular, Facultad de Ciencias Biológicas, Pontificia Universidad Católica de Chile, 340, P. O. Box 114-D, Santiago, Chile e-mail: [email protected] N. C. Inestrosa Center for Healthy Brain Ageing, School of Psychiatry, Faculty of Medicine, University of New South Wales, Sydney, Australia N. C. Inestrosa Centro de Excelencia en Biomedicina de Magallanes (CEBIMA), Universidad de Magallanes, Punta Arenas, Chile L. Varela-Nallar (*) Center for Biomedical Research, Faculty of Biological Sciences and Faculty of Medicine, Universidad Andres Bello, Republica 239, 8370146 Santiago, Chile e-mail: [email protected]

Introduction Wnts are secreted signalling molecules that activate signalling cascades involved in different aspects of embryonic development including cell fate specification, polarity and migration (Clevers and Nusse 2012; Nusse and Varmus 2012). The Wnt signalling pathway is activated by Wnt ligands, which are members of a family of 19 secreted glycoproteins in mammals that bind to seven-pass transmembrane Frizzled (FZD) receptors to activate different signalling cascades: the canonical Wnt/β-catenin signalling pathway and the non-canonical βcatenin-independent signalling cascades (Gordon and Nusse 2006) (Fig. 1). Importantly, the same ligand may activate different Wnt signalling cascades depending on the receptor context, and activation of a specific pathway may antagonize the activation of other pathways (Mikels and Nusse 2006). Canonical and non-canonical Wnt ligands can compete for binding to FZD receptors, thereby causing reciprocal pathway inhibition of both signalling pathways (Grumolato et al. 2010). In addition to FZDs, other membrane proteins have been identified as receptors or co-receptors of Wnts, including the low-density lipoprotein receptor-related protein 5 (LRP5), LRP6, receptor tyrosine kinase-like orphan receptor 1 (ROR1), ROR2 (Cadigan and Liu 2006; Gordon and Nusse 2006; Green et al. 2008), and the receptor-like tyrosine kinase Ryk that has been implicated in axon guidance in the developing nervous system (Bovolenta et al. 2006; Fradkin et al. 2009). Soluble proteins are also important modulators of Wnt’s actions, where some of them, such as secreted FZD-related proteins (sFRPs), Dickkopf (DKK) proteins and Wnt inhibitory factor 1 (WIF1), act as inhibitors of the Wnt pathway (Cruciat and Niehrs 2013; Kawano and Kypta 2003; Niehrs 2006). There are also other physiological and pharmacological mechanisms that can activate the intracellular component of the canonical Wnt/β-catenin signalling cascade. For example, extracellular R-spondin (Rspo) proteins have been shown

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Fig. 1 The Wnt signalling pathway. Canonical and non-canonical signalling cascades. Binding of a Wnt ligand to a Frizzled (FZD) receptor activates different signalling cascades. In the canonical Wnt/β-catenin signalling pathway (left) the Wnt ligand also interacts with the co-receptor low density lipoprotein-related protein 5 or 6 (LRP5/6) to activate the protein Dishevelled (Dvl) and induce the stabilization of β-catenin by preventing its phosphorylation by glycogen synthase kinase-3β (GSK3β) in a multiprotein complex composed also of the scaffold protein Axin and adenomatous polyposis coli (APC). Consequently, β-catenin accumulates in the cytoplasm and enters the nucleus where it interacts with the transcription factors TCF/LEF to induce the transcription of Wnt target

genes. In the non-canonical Wnt/Ca2+ signalling cascade (middle), activation of the pathway triggers the activation of trimeric G proteins and subsequently phospholipase C (PLC), increasing inositol triphosphate (IP3). IP3 induces the release of Ca2+ from the endoplasmic reticulum (ER), which then induces the activation of calcium/calmodulin-dependent protein kinase II (CamKII) and protein kinase C (PKC). In the noncanonical Wnt/PCP signalling cascade (right), activation of the pathway leads to the activation of the small GTPases Rho and Rac leading to the activation of Rho-associated protein kinase (ROCK) and Jun N-terminal kinase (JNK) to regulate cytoskeleton dynamics

to bind LGR4, LGR5 and LGR6 G-protein coupled receptors to physiologically activate this pathway (Carmon et al. 2011; Glinka et al. 2011). In addition, the long-standing pharmacological agent lithium inhibits glycogen synthase kinase-3β (GSK-3β) intracellularly, thereby stabilizing β-catenin to activate the pathway independently of ligand–receptor interactions (Toledo and Inestrosa 2010; Valvezan and Klein 2012). The activation of the canonical Wnt/β-catenin signalling cascade involves the formation of a Wnt–LRP–FZD complex that activates the scaffold protein Dishevelled (Dvl) and causes the dissociation of a multiprotein destruction complex, consisting of enzymes that phosphorylate and target β-catenin

for proteasomal degradation when the pathway has not been activated (Hart et al. 1999; Liu et al. 2002). Consequently, βcatenin accumulates in the cytoplasm and translocates to the nucleus where it interacts with members of the TCF/LEF1 family of transcription factors to regulate the expression of Wnt target genes (Arrazola et al. 2009; Clevers and Nusse 2012; Nusse and Varmus 2012). In the multiprotein destruction complex, β-catenin is phosphorylated by casein kinase 1α (CK1α) and GSK-3β which are associated with the scaffold protein Axin and adenomatous polyposis coli (APC) (Hart et al. 1998; Ikeda et al. 1998; Kishida et al. 1998; Sakanaka et al. 1998).

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The Wnt/FZD interaction can also activate the noncanonical Wnt/PCP or Wnt/Ca2+ pathways (Fig. 1). In the Wnt/PCP pathway, the phosphorylation of Dvl causes the activation of the small GTPases Rho and Rac and, consequently, the downstream Jun N-terminal kinase (JNK), which regulates cytoskeleton dynamics (Gordon and Nusse 2006; Rosso and Inestrosa 2013; Rosso et al. 2005). In contrast, the Wnt/Ca 2+ pathway is almost exclusively a G-proteindependent signalling pathway (Kohn and Moon 2005). The activation of the Wnt/Ca2+ pathway requires binding of Wnt to FZD on the cell surface membrane to trigger stimulation of heterotrimeric G-proteins (Slusarski et al. 1997a; Slusarski et al. 1997b), which activate phospholipase-C (PLC). PLC causes an increment in intracellular Ca2+ release that decreases cyclic guanosine monophosphate (cGMP) and activates the protein kinases Ca2+/Calmodulin-dependent protein kinase II (CaMKII) and protein kinase-C (PKC) (Kohn and Moon 2005; Montcouquiol et al. 2006; Veeman et al. 2003). In addition, it activates the transcription factor NFAT to induce the transcription of target genes. In the nervous system, Wnt signalling cascades are important for the formation of neuronal circuits. During development, Wnt signalling regulates self-renewal, maintenance and differentiation of neural progenitor cells (Hirabayashi et al. 2004; Machon et al. 2007; Munji et al. 2011) and regulates the development of the cortex and hippocampus (Li and Pleasure 2005; Machon et al. 2007). Wnts not only regulate early embryonic development but are also key regulators of late embryonic and postnatal development of the central nervous system (CNS) (Inestrosa and Arenas 2010; Oliva et al. 2013). Various Wnt ligands regulate synaptic development and function, including the formation and maturation of pre- and postsynaptic sites and neurotransmission. Members of the Wnt family of secreted signalling proteins are implicated in every step of neural development. During vertebrate development, a Wnt signalling gradient that is high in the posterior and low in the anterior is critical for the proper specification of the anterior–posterior axis of the neural plate (Kiecker and Niehrs 2001). In fact, several studies have demonstrated that inhibition of Wnt signalling in the anterior, as well as the activation of Wnt signalling in the posterior, is required for proper anterior-posterior patterning of the early CNS (Esteve et al. 2000; Glinka et al. 1998; Houart et al. 2002; Kazanskaya et al. 2000). During early development, after the neural plate is specified, it invaginates to form the neural tube, a process that is complete once the paired neural folds adhere at the dorsal midline, and which is required for the development of the spinal cord and brain. Neural tube defects cause conditions like spina bifida and anencephaly. It has been determined that the Wnt/PCP signalling pathway is relevant for neural tube closure and is involved in neural tube defects (Curtin et al. 2003; Wen et al. 2010). Also, a role for the Wnt/β-catenin pathway in neural tube closure is indicated by the presence of

neural tube defects in mice with mutations in LRP6 (Carter et al. 2005) and Axin 1 (Perry et al. 1995), and in mice carrying ablated TCF–β-catenin interactions (Wu et al. 2012). Neural progenitor cells that make up the neural tube proliferate, differentiate and migrate to form the many neuronal ganglia, nuclei, and layers of the CNS. The Wnt/β-catenin signalling pathway has been associated with both proliferation and specification of neural progenitor cells. It has been determined that Wnt-1 and Wnt-3a ligands are expressed at the dorsal midline of the developing neural tube and are required for the formation of brain structures; mutant analyses in mice show that the midbrain is absent when Wnt-1 is not expressed (McMahon and Bradley 1990; Thomas and Capecchi 1990), and the hippocampus is absent when Wnt-3a is not expressed (Lee et al. 2000). The Wnt-3a mutant shows decreased cell division within the hippocampal neuroepithelium; however, there remain small hippocampal subfields. This implies that the Wnt-3a ligand primarily regulates proliferation of hippocampal neural precursor cells (Lee et al. 2000). During corticogenesis, the levels of Wnt activity have been shown to be important in controlling the switch between proliferation and differentiation of progenitor cells (Bielen and Houart 2014), which depends on β-catenin. Increased stabilization of β-catenin decreases cell cycle exit in neural precursors, while decreased Wnt/β-catenin signalling is associated with differentiation (Chenn and Walsh 2002; Mutch et al. 2010). Later in development, Wnt/β-catenin signalling induces neural differentiation. In cortical neural precursor cells, Wnt/βcatenin signalling inhibits the self-renewal capacity of progenitors and promotes neuronal differentiation (Hirabayashi et al. 2004). This effect on differentiation is stage-specific, since it is not observed in neural precursor cells at earlier developmental stages (Hirabayashi et al. 2004; Munji et al. 2011). Thus, Wnt signalling regulates both self-renewal and subsequent cell fate specification of neuronal progeny, and these effects depend on transcriptional regulation mediated by the Wnt/β-catenin signalling pathway. The effect on proliferation involves the transcriptional control of genes that regulate the cell cycle (reviewed in Niehrs and Acebron 2012) and neuronal differentiation involves the regulation of the expression of proneural genes such as the neurogenin 1 gene (Hirabayashi et al. 2004). The relevance of the Wnt signalling pathway in normal brain function is supported by the fact that its deregulation has been linked to different pathologies that affect the CNS, including mental disorders, mood disorders and neurodegenerative diseases (De Ferrari and Moon 2006; Inestrosa et al. 2012; Inestrosa and Varela-Nallar 2014; Lovestone et al. 2007; Oliva et al. 2013). Interestingly, the Wnt signalling pathway shows neuroprotective properties in Alzheimer’s disease (AD), a neurodegenerative illness characterized by a progressive loss of cognitive abilities including learning and memory (Ballard et al. 2011) that has been associated with an

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impairment in Wnt signalling (De Ferrari and Inestrosa 2000; Inestrosa and Arenas 2010). In cultured hippocampal neurons and in hippocampal slices, Wnts show neuroprotective properties against the toxicity of the amyloid-β (Aβ) peptide (Alvarez et al. 2004; Cerpa et al. 2010; Chacon et al. 2008; De Ferrari et al. 2003), which is associated with AD pathogenesis. This was supported by in vivo experiments showing that the activation of the Wnt signalling pathway in a double transgenic mouse model of AD reduces spatial memory impairment and the histopathological hallmarks of the disease (Toledo and Inestrosa 2010), indicating that in vivo modulation of the Wnt signalling pathway may have therapeutic value in some pathological conditions.

Wnt signalling in neuronal maturation Compelling evidence indicates that Wnt signalling cascades are important for the regulation of several aspects of neuronal development. Neuronal polarization, axonal and dendritic development and synaptogenesis are crucial for the formation of neuronal circuits. Studies have shown that the Wnt signalling pathway is involved in all these steps of neuronal development. In cerebellar neurons, it has been shown that Wnt-7a induced axon and growth cone remodelling (Hall et al. 2000; Lucas and Salinas 1997). Wnt-mediated changes in growth cone remodelling involve changes in microtubule organization through the regulation of APC (Purro et al. 2008). Wnt-5a also regulates axonal behavior in sympathetic and cortical neurons (Bodmer et al. 2009; Li et al. 2009). Interestingly, in cortical neurons, Wnt-5a stimulates axonal outgrowth and repulsive axon guidance through different receptors. Axonal outgrowth is mediated by the Ryk receptor, whereas axonal repulsion requires both Ryk and FZD receptors (Li et al. 2009). Recently, we determined that FZD5 regulates axon development (Slater et al. 2013). Moreover, by gain- and loss-of-function experiments, we determined that FZD5 regulates a very early event in neuronal development: the establishment of neuronal polarity (Fig. 2). In cultured neurons, FZD5 shows a very polarized distribution being mainly present in the peripheral zone of growth cones. Overexpression of FZD5 induces a loss of polarized distribution of the receptor and induces a mislocalization of axonal proteins, while FZD5 knockdown induces a loss of axonal proteins (Slater et al. 2013). These data suggest that FZD5 is important for neuronal polarity. In addition, when the receptor is overexpressed after the acquisition of neuronal polarity, it does not revert polarization but alters neuronal morphogenesis by decreasing axonal length and increasing dendritic length and arborisation via a JNK-dependent mechanism (Slater et al. 2013). Several studies have shown the involvement of Wnt signalling components in dendrite morphogenesis. In hippocampal neurons, β-catenin is a critical mediator of dendritic morphogenesis.

This effect of β-catenin is independent of gene transcription and is required for dendritic growth induced by depolarization (Yu and Malenka 2003). Another study demonstrated that, in hippocampal neurons, activation of NMDA receptors increases the expression of Wnt-2, which then stimulates dendritic arborization (Wayman et al. 2006), indicating the involvement of Wnt in activity-dependent dendritic development. Wnt-7b also regulates dendritic arborization, and this effect is mediated by a non-canonical Wnt signalling cascade that involves JNK activation (Rosso et al. 2005). In addition to the participation in neuronal polarization and morphogenesis, it is known that Wnt signalling regulates synapse formation and neurotransmission. Electrophysiological recordings in rat hippocampal slices showed that a blockade of Wnt signalling impairs long-term potentiation (LTP) (Chen et al. 2006), indicating the importance of this signalling pathway in synaptic plasticity (Vargas et al. 2014). Importantly, the expression and release of Wnts are regulated by neuronal activity (Chen et al. 2006; Wayman et al. 2006), supporting the notion that these ligands may play a role during neuronal transmission. Regarding the synaptic effects of Wnts, several years ago it was shown in cerebellar neurons that Wnt-7a regulates the clustering of the synaptic vesicle protein synapsin I (Hall et al. 2000; Lucas and Salinas 1997). The effect of Wnt signalling on presynaptic assembly has also been observed in hippocampal neurons, in which Wnt-7a, Wnt-3a and Wnt-7b increase the presynaptic puncta and the synaptic vesicle cycle (Ahmad-Annuar et al. 2006; Cerpa et al. 2008; Davis et al. 2008). Moreover, electrophysiological recordings on adult rat hippocampal slices demonstrated that Wnt-7a increases neurotransmitter release in CA3-CA1 synapses (Cerpa et al. 2008). The presynaptic effects of Wnts may be mediated by Wnt receptors located at the synaptic region. In hippocampal neurons, we determined that the Wnt receptor FZD1 is present at the presynaptic site where it regulates the presynaptic assembly (Varela-Nallar et al. 2009). Also, FZD5 was shown to mediate synaptogenesis induced by Wnt-7a (Sahores et al. 2010). These findings have demonstrated a relevant role for the Wnt signalling pathway in presynaptic differentiation and function. In addition, non-canonical Wnt signalling cascades have been associated with the postsynaptic apparatus. Electrophysiological recordings showed that Wnt-5a increases the amplitude of field excitatory postsynaptic potentials (fEPSP) and upregulates synaptic NMDAR currents, facilitating the induction of LTP (Cerpa et al. 2010, 2011), a two-step effect that is independently mediated by protein kinase C (PKC) and JNK (Cerpa et al. 2011). At the structural level, Wnt-5a modulates postsynaptic assembly, increasing clustering of the postsynaptic density protein-95 (PSD-95) (Farias et al. 2009), which is a key scaffold protein of the postsynaptic density. The Wnt signalling pathway also modulates dendritic spine morphogenesis in cultured hippocampal neurons, an early

Cell Tissue Res Fig. 2 FZD5 in neuronal polarity and morphogenesis. FZD5 is located in the peripheral zone of axonal growth cones. Gain- and loss-of-function experiments demonstrate that FZD5 is important for neuronal polarity. Overexpression of FZD5 in neurons that are already polarized decrease the total length of axons, a process partially prevented by inhibition of JNK, suggesting the Wnt/PCP pathway modulates axon behavior

event in synapse formation that is crucial for synaptic plasticity. We observed that Wnt-5a treatment induces an increase in dendritic protrusions that mature into dendritic spines (VarelaNallar et al. 2010a). Interestingly, the effect on dendritic protrusions was not observed by treatment with Wnt-3a, indicating that there is some level of specificity for this effect (Varela-Nallar et al. 2010a). Time-lapse imaging revealed that Wnt-5a induces the formation of new dendritic spines and increases the size of preexisting ones (Varela-Nallar et al. 2010a). Another ligand that regulates dendritic spines is Wnt-7a, which was shown to increase the density and maturity of dendritic spines through a mechanism involving CamKII (Ciani et al. 2011), suggesting that the non-canonical Wnt/Ca2+ signalling cascade is involved in this effect. Both Wnt-5a and Wnt-7a increase intracellular Ca2+ concentration in neurons (Ciani et al. 2011; Varela-Nallar et al. 2010a), suggesting that a common Ca2+-dependent mechanism may be involved in the Wnt-mediated regulation of dendritic spines. A potential receptor for the effect of Wnts on dendritic spines is ROR2, which is located in dendritic spines (Alfaro et al., unpublished results). We have determined that ROR2 is important for the maturation and maintenance of dendritic spines in hippocampal neurons (Fig. 3), and, interestingly, this receptor mediated an inhibitory effect of Wnt-5a in a voltage-gated K+ current

(Parodi et al., unpublished results), indicating that this may be the receptor or co-receptor mediating the described postsynaptic effects of Wnt-5a.

Wnt signalling in adult neurogenesis In the adult brain, the generation of new neurons continues mainly in two regions, the SVZ in the wall of the lateral ventricles and the SGZ in the dentate gyrus of the hippocampus (Alvarez-Buylla and Garcia-Verdugo 2002; Zhao et al. 2008). In these two neurogenic regions, neural stem cells (NSC) proliferate and differentiate into neuroblasts that in the SVZ migrate through the rostral migratory stream to the olfactory bulb where they became interneurons, and in the SGZ mature into granule neurons that become integrated into the granule cell layer (Ming and Song 2011; Zhao et al. 2008). Many signalling cascades are involved in the regulation of the different steps of neurogenesis being important for the proper balance between the maintenance of the NSC pool and for the differentiation and maturation of newborn neurons (Faigle and Song 2013; Schwarz et al. 2012; Suh et al. 2009). During recent years, increasing evidence has supported a role for the Wnt/β-catenin pathways in the regulation of adult

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Figure 3 ROR2 is involved in dendritic spine development. Wnt ligands regulate spinogenesis in hippocampal neurons through activation of noncanonical Wnt pathways. Recently the tyrosine kinase receptor ROR2, which is located in dendritic spines, has been implicated in this process (unpublished results). In dendritic spines, ROR2 may act either as a receptor or co-receptor of FZD to activate non-canonical signalling pathways

neurogenesis (Varela-Nallar and Inestrosa 2013). In 2005, the group of Fred Gage determined in in vivo experiments that overexpression of Wnt-3 in the dentate gyrus increases cell proliferation and the generation of new neurons, while inhibition of the Wnt signalling pathway reduces proliferation and neurogenesis (Lie et al. 2005). In that study, it was shown that Wnts derived from hippocampal astrocytes induce the differentiation of rat adult hippocampal progenitors (AHPs) and that AHPs express Wnt signalling components. Later on, another study demonstrated in vitro that rat AHPs express several Wnt ligands and receptors, and also that there is an autocrine Wnt stimulation that supports the proliferation and multipotency of AHPs, and therefore it may be relevant for NSC maintenance (Wexler et al. 2009). More recently, it was shown that Wnt-7a is important for multiple steps of adult neurogenesis including NSC self-renewal, neural progenitor cell proliferation, neuronal differentiation and maturation (Qu et al. 2013). These effects of Wnt-7a involve the regulation of cyclin D1 and neurogenin 2 genes, involved in cell cycle control and neuronal differentiation, respectively (Qu et al. 2013).

In addition to Wnts, multiple components and regulators of the Wnt signalling pathway have been associated with adult neurogenesis. GSK-3β is involved in the regulation of neurogenesis mediated by disrupted in schizophrenia 1 (DISC1) (Inestrosa et al. 2012). This protein inhibits GSK-3β, and the impairment in progenitor cell proliferation caused by DISC1 knock-down can be rescued by overexpression of stabilized β-catenin (Mao et al. 2009) In vivo, the impairments in cell proliferation observed in the adult dentate gyrus by DISC1 loss of function can be rescued by administration of a GSK-3β inhibitor (Mao et al. 2009). In the SVZ, retrovirus-mediated expression of a stabilized β-catenin induces the proliferation of type C cells (Adachi et al. 2007), and expression of Axin, a part of the protein complex that mediates the phosphorylation and degradation of β-catenin, decreases cell proliferation (Qu et al. 2010). Taken together, this evidence supports a role for the canonical Wnt signalling pathway in NSC proliferation. Soluble Wnt inhibitors have also been associated with adult neurogenesis. Dkk1 and sFRP3 are negative regulators of adult neurogenesis. Dkk1 prevents the activation of the canonical Wnt/β-catenin signalling cascade by preventing the formation of the FZD/LRP complex, while sFRP3 binds to Wnts preventing their interaction with their receptors (Cruciat and Niehrs 2013). Deletion of Dkk1 and sFRP3 increase neurogenesis and dendrite complexity of newborn neurons (Jang et al. 2013; Seib et al. 2013). Interestingly, both Wnt inhibitors are regulated by physiological stimuli that regulate neurogenesis, suggesting a physiological role for both proteins and for the Wnt signalling pathway in the regulation of neurogenesis in the adult brain. A reduction in sFRP3 is observed during exercise and electroconvulsive stimulation, suggesting that this reduction may be important for the increase in neurogenesis observed in both conditions (Jang et al. 2013). On the other hand, the expression of Dkk1 is increased during aging (Seib et al. 2013), suggesting that it may be involved in the decline of neurogenesis observed in different species during lifespan (Knoth et al. 2010; Kuhn et al. 1996; Leuner et al. 2007; Varela-Nallar et al. 2010b). During aging, a decline in the levels of Wnt-3 and Wnt-3a in the dentate gyrus was also reported (Okamoto et al. 2011), which may also underlie the decline in neurogenesis. In addition, canonical Wnt signalling is increased in vivo by exposure to chronic hypoxia, concomitantly with an increase in cell proliferation and neurogenesis in the SGZ of adult mice (Varela-Nallar et al. 2014). This suggests that the Wnt signalling pathway may also be involved in the effect of mild hypoxia on neurogenesis. The mechanisms involved in the regulation of adult neurogenesis by the Wnt signalling pathway may involve the transcription of proneural Wnt target genes. It has been determined that the expression of the transcription factors NeuroD1 and Prox1 is regulated by the canonical Wnt signalling pathway (Karalay et al. 2011; Kuwabara et al. 2009). NeuroD1 is important for neurogenesis in the SGZ and SVZ

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in the adult brain (Gao et al. 2009), and Prox1 is important for the differentiation and survival of newborn granule cells (Karalay et al. 2011). Both transcription factors identified as Wnt target genes might be important for the positive effect of the Wnt signalling pathway in adult neurogenesis.

Concluding remarks The Wnt signalling pathway plays pivotal roles during the formation, maintenance and function of the CNS, having a wide range of functions that include neuronal differentiation, development and maturation. There are many Wnt proteins and several Wnt receptors and co-receptors that in conjunction paint a complex picture that might be relevant for the fine regulation of neuronal circuit formation and functioning. More recent evidence has shown that the Wnt signalling pathway also regulates the formation of new neurons in the adult brain, where it regulates the proliferation and differentiation of neural progenitor cells. Considering all the neuronal processes that are regulated by the Wnt signalling pathway in brain neurons, it can be expected that additional roles for this pathway in the development, maturation and integration of adult-born neurons might be unveiled in the near future. We thank Felipe G. Serrano (CARE, Department of Cell and Molecular Biology, P. Catholic University of Chile) for artwork. This work was supported by Grants from FONDECYT (No. 1120156) and the Basal Center of Excellence in Science and Technology (CONICYT-PFB12/ 2007) to N.C.I. and by a Grant from FONDECYT (No. 11110012) to L.V.-N.

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