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The olfactory bulb (OB) is one of two regions of the mammalian brain which undergo continuous neuronal replacement during adulthood. A significant fraction of the cells added in adulthood to the bulbar circuitry is constituted by dopaminergic (DA) neurons. We took advantage of a peculiar property of dopaminergic neurons in transgenic mice expressing eGFP under the tyrosine hydroxylase (TH) promoter: while DA neurons located in the glomerular layer (GL) display full electrophysiological maturation, eGFP+ cells in the mitral layer (ML) show characteristics of immature cells. In addition, they also display a lower fluorescence intensity, possibly reflecting different degrees of maturation. To investigate whether this difference in maturation might be confirmed at the gene expression level, we used a fluorescence-activated cell sorting technique on enzymatically dissociated cells of the OB. The cells were divided into two groups based on their level of fluorescence, possibly corresponding to immature ML cells and fully mature DA neurons from the GL. Semiquantitative real-time PCR was performed to detect the level of expression of genes linked to the degree of maturation of DA neurons. We showed that indeed the cells expressing low eGFP fluorescence are immature neurons. Our method can be further used to explore the differences between these two groups of DA neurons.

Microglia are the only immune cell population present in the brain parenchyma. Their vantage position in the central nervous system (CNS) enables these myeloid cells to perform the most disparate of tasks: from the classical immune functions of fighting infections and surveilling the extracellular space for pathogens and damage, to sculpting the neuronal circuitry by pruning unnecessary synapses and assisting neurons in spine formation, aiding in the maintenance of brain homeostasis. The neurotrophin field has always been dominated by the neurocentric view that the primary target of these molecules must be neurons: this holds true even for the Nerve Growth Factor (NGF), which owes its popularity in the neuroscience community to its trophic and tropic activity towards sensory and sympathetic neurons in the peripheral nervous system, and cholinergic neurons in the CNS. The increasing evidence that microglia are an integral part of neuronal computation calls for a closer look as to whether these glial cells are capable of responding directly to NGF. In this review, we will first outline evidence in support of a role for NGF as a molecule mediating neuroimmune communication. Then, we will illustrate some of those non-immune features that have made microglial cells one of the hottest topics of this last decade. In conclusion, we will discuss evidence in support of a microglial function for NGF.

GABA, the main inhibitory neurotransmitter in the adult brain, depolarizes and excites immature neurons because of an initially higher intracellular chloride concentration [Cl−]i due to the delayed expression of the chloride exporter KCC2 at birth. Depolarization-induced calcium rise via NMDA receptors and voltage-dependent calcium channels is instrumental in shaping neuronal circuits and in controlling the excitatory (E)/inhibitory (I) balance in selective brain areas. An E/I imbalance accounts for cognitive impairment observed in several neuropsychiatric disorders. The aim of this review is to summarize recent data on the mechanisms by which alterations of GABAergic signaling alter the E/I balance in cortical and hippocampal neurons in Alzheimer’s disease (AD) and the role of cation-chloride co-transporters in this process. In particular, we discuss the NGF and AD relationship and how mice engineered to express recombinant neutralizing anti-NGF antibodies (AD11 mice), which develop a neurodegenerative pathology reminiscent of that observed in AD patients, exhibit a depolarizing action of GABA due to KCC2 impairment. Treating AD and other forms of dementia with bumetanide, a selective NKCC1 antagonist, contributes to re-establishing a proper E/I balance in selective brain areas, leading to amelioration of AD symptoms and the slowing down of disease progression.

In the brain, the neurotrophin Nerve growth factor (NGF) regulates not only neuronal survival and differentiation, but also glial and microglial functions and neuroinflammation. NGF is known to regulate oligodendrogenesis, reducing myelination in the central nervous system (CNS). In this study, we found that NGF controls oligodendrogenesis by modulating the levels of miR-219a-5p, a well-known positive regulator of oligodendrocyte differentiation. We exploited an NGF-deprivation mouse model, the AD11 mice, in which the postnatal expression of an anti-NGF antibody leads to NGF neutralization and progressive neurodegeneration. Notably, we found that these mice also display increased myelination. A microRNA profiling of AD11 brain samples and qRT-PCR analyses revealed that NGF deprivation leads to an increase of miR-219a-5p levels in hippocampus and cortex and a corresponding down-regulation of its predicted targets. Neurospheres isolated from the hippocampus of AD11 mice give rise to more oligodendrocytes and this process is dependent on miR-219a-5p, as shown by decoy-mediated inhibition of this microRNA. Moreover, treatment of AD11 neurospheres with NGF inhibits miR-219a-5p up-regulation and, consequently, oligodendrocyte differentiation, while anti-NGF treatment of wild type (WT) oligodendrocyte progenitors increases miR-219a-5p expression and the number of mature cells. Overall, this study indicates that NGF inhibits oligodendrogenesis and myelination by down-regulating miR-219a-5p levels, suggesting a novel molecular circuitry that can be exploited for the discovery of new effectors for remyelination in human demyelinating diseases, such as Multiple Sclerosis.

Neurotrophin nerve growth factor (NGF) has been demonstrated to upregulate the gene expression of bradykinin receptor 2 (B2R) on sensory neurons, thus facilitating nociceptive signals. The aim of the present study is to investigate the involvement of B2R in the NGF mechanism of action in nonsensory neurons in vitro by using rat mixed cortical primary cultures (CNs) and mouse hippocampal slices, and in vivo in Alzheimer’s disease (AD) transgenic mice (5xFAD) chronically treated with NGF. A significant NGF-mediated upregulation of B2R was demonstrated by microarray, Western blot, and immunofluorescence analysis in CNs, indicating microglial cells as the target of this modulation. The B2R involvement in the NGF mechanism of action was also demonstrated by using a selective B2R antagonist which was able to reverse the neuroprotective effect of NGF in CNs, as revealed by viability assay, and the NGF-induced long-term potentiation (LTP) in hippocampal slices. To confirm in vitro observations, B2R upregulation was observed in 5xFAD mouse brain following chronic intranasal NGF treatment. This study demonstrates for the first time that B2R is a key element in the neuroprotective activity and synaptic plasticity mediated by NGF in brain cells.