Generation of new progenitor cells and their differentiation into neurons in the dentate gyrus takes place throughout the life of rodents and humans 
. The new neurons have shown to extend their axonal projections into the hippocampal CA3 area 
, indicating that they play a functional role in the neuronal network of the hippocampus.
There is an age-dependent decline in the number of immunoreactive BrdU cells, a marker of proliferating cells, in the SGZ of the dentate gyrus in both wild type and APP/PS1 animals. This decrease in neurogenesis with age has been reported previously on rodents and humans 
. Significantly, APP/PS1 mice showed consistently lower numbers of newly generated cells in their DG compared to aged matched wild type littermates when compared at ages 3 months and 6 months, while at 12 and 15 months no difference was seen. This may be due to AD pathology impairing the proliferation of NSC at earlier stages of the life in the AD mouse model, which has been described previously in several studies with various mouse models of AD 
. Also, as the numbers of new neurons were already very low in the older groups, a significant reduction in numbers would be hard to achieve.
The important roles of various factors such as exercise, enriched environment, trauma and growth factors on the regulation of neurogenesis have been studied in detail 
. Growth factors such as epidermal growth factor (EGF), fibroblast growth factor-2 (FGF-2) and glia cell line-derived neurotrophic factor (GDNF) when administered intracerebroventricular (i.c.v.) in rats with ischemic brain injury, increased neurogenesis was observed, and the new neurons were shown to integrate functionally into the network 
. The roles of NGF and BDNF in neurogenesis also has been researched in some detail, and their potential use as a treatment for neurodegenerative disease has been investigated 
. Thus, compounds that can selectively promote proliferation of NSC and prompt their differentiation to neuronal phenotype may be of use as a treatment of neurodegenerative diseases such as AD 
We observed that a 1-week treatment of wild type animals with liraglutide had no effect on BrdU positive cell numbers, but showed increased endogenous cell proliferation as shown by Ki67 immunoreactive positive cells at 3, 6, 12 or 15 months of age. In contrast, APP/PS1 animals treated acutely for 7 days with liraglutide showed a significant increase in BrdU and Ki67 positive cells at ages 3, 6, 12 and 15 months. Amyloid deposition is related to decreased neurogenesis in AD mice. APP/PS1 double transgenic mice develop early-onset amyloidosis, as young as 3 months, with plaque deposits by 5–6 months, progressing with age up to 15 months 
. Beta-amyloid oligomers have been shown to interfere with insulin signaling 
, and since insulin is an important growth factor that also activates stem cells, this impairment in insulin signaling may be a reason for the impairment of stem cell proliferation found in the APP/PS1 mice 
. A second reason for the normalisation of neurogenesis by liraglutide may be the fact that the chronic inflammation response was reduced. Cytokines released during a chronic inflammation response reduced stem cell proliferation 
. We have previously shown that liraglutide reduces the chronic inflammation response in this APP/PS1 mouse strain 
. This would explain why there was little effect on neuronal progenitor proliferation in wild type mice as there were no plaques nor the associated chronic inflammation response. The effects of acute treatment by liraglutide on cell proliferation in-vivo
in AD mice has not been studied previously.
The effects of long-term administration of liraglutide on progenitor cell proliferation in the SGZ of dentate gyrus was examined in APP/PS1 and WT littermates. The 37- day treatment with liraglutide significantly increased the number of immunopositive BrdU cells and also Ki67 positive cells in 3, 6, 12 and 15-months in both APP/PS1 and WT animals compared to saline controls. Previous studies showed that GLP-1 and analogues of GLP-1 induce cell proliferation in vivo
and in vitro
. GLP-1 receptors are expressed all throughout the brain especially on neurons in the hippocampus and cortex 
. Furthermore, GLP-1 also functions as a growth factor, has neuroprotective characteristics and enhances cell survival 
. GLP-1R mediated activation of mitogen activated protein kinase (MAPK) is crucial for hippocampal neurogenesis 
, which is associated with hippocampal-dependent learning and memory 
Differentiation capacity of NSC on acute treatment with liraglutide showed no changes in both WT and APP/PS1 mice at any age. There was no difference in the percentage of newly generated cells expressing mature marker (NeuN) of neurons compared to control group. No difference in neuronal phenotype number was seen in 3-month old WT and APP/PS1 group treated chronically with liraglutide. Interestingly, in chronic treatment with liraglutide, both APP/PS1 and WT littermates showed significant increase in number of cells incorporating BrdU and mature neuronal marker NeuN at 6, 12 and 15 months, indicating increased neuroblast differentiation. Interestingly, both acute and chronic treatment increased neuroblast differentiation in WT and APP/PS1 animals. This suggest acute requirement of liraglutide is sufficient for its trophic properties in neuroblast differentiation but chronic administration is essential for full differentiation into mature neurons.
Not all newly generated cells differentiate into neuron, but some transform into glial phenotype. Liraglutide did not appear to affect the differentiation into glial. There was a trend towards a decrease the number of new cells transforming to glia cells, which was not significant and may be explained by an increase in cells differentiating into neurons. Previous studies have shown that liraglutide significantly decreases activated microglia and activated astrocytes in APP/PS1 mouse brains and WT mouse brains 
, but no research has been conducted previously on effects of incretins on differentiation of NSC into glial phenotypes. The results shown here suggest a chronic requirement of liraglutide to activate the differentiation process of NSC into neurons. However, our study does not rule out that an increased survival of NSC that may contribute to net neurogenic potential of liraglutide. Past studies have shown an increase in neuronal markers (MAP2, b-III-tubulin) in cell cultures and an increase in DCX positive cells in SVZ of rat brain after chronic exendin-4 treatment 
. Additionally, GLP-1 promotes beta cell proliferation 
and differentiation from embryonic stem cells 
. The mechanism of action of liraglutide in differentiation could be exerted via upregulation of Mash1 (involved in neuronal differentiation in hippocampus) 
This is the first study that investigated the proliferation and differentiation properties of GLP-1 analogue, liraglutide in wild type and APP/PS1 animals at different ages. We are also first to look at the effects of acute and chronic treatment with liraglutide on progenitor cell proliferation and differentiation at different ages of wild type and animals with AD pathology.
Based on findings of our study and previous work that showed liraglutide improves cell proliferation in SGZ, increases differentiation of progenitor cells to neurons, this drug and other GLP-1 analogues have therapeutic potential to treat neurodegenerative disorders such as Alzheimer's and Parkinson's disease.