Adult neural stem cell activation in mice is regulated by the day/night cycle and intracellular calcium dynamics.

Neural stem cells (NSCs) in the adult brain transit from the quiescent state to proliferation to produce new neurons. The mechanisms regulating this transition in freely behaving animals are, however, poorly understood. We customized in vivo imaging protocols to follow NSCs for several days up to months, observing their activation kinetics in freely behaving mice. Strikingly, NSC division is more frequent during daylight and is inhibited by darkness-induced melatonin signaling. The inhibition of melatonin receptors affected intracellular Ca2+ dynamics and promoted NSC activation. We further discovered a Ca2+ signature of quiescent versus activated NSCs and showed that several microenvironmental signals converge on intracellular Ca2+ pathways to regulate NSC quiescence and activation. In vivo NSC-specific optogenetic modulation of Ca2+ fluxes to mimic quiescent-state-like Ca2+ dynamics in freely behaving mice blocked NSC activation and maintained their quiescence, pointing to the regulatory mechanisms mediating NSC activation in freely behaving animals.

Live imaging of adult neural stem cells in freely behaving mice using mini-endoscopes.

During adulthood, the activation of adult neural stem cells (NSCs) has been mostly studied ex vivo in post-mortem tissues or in vivo in anesthetized animals. This protocol presents an approach that allows for the long-term and minimally invasive investigation of adult NSC activation and physiology in freely behaving animals. By combining specific NSC labeling and mini-endoscopic microscopy, live imaging of NSC division and Ca2+ activity can be performed continuously for 2-3 days and even up to several months. For complete details on the use and execution of this protocol, please refer to Gengatharan et al. (2021).

LRIG1-Mediated Inhibition of EGF Receptor Signaling Regulates Neural Precursor Cell Proliferation in the Neocortex.

Here, we ask how neural stem cells (NSCs) transition in the developing neocortex from a rapidly to a slowly proliferating state, a process required to maintain lifelong stem cell pools. We identify LRIG1, known to regulate receptor tyrosine kinase signaling in other cell types, as a negative regulator of cortical NSC proliferation. LRIG1 is expressed in murine cortical NSCs as they start to proliferate more slowly during embryogenesis and then peaks postnatally when they transition to give rise to a portion of adult NSCs. Constitutive or acute loss of Lrig1 in NSCs over this developmental time frame causes stem cell expansion due to increased proliferation. LRIG1 controls NSC proliferation by associating with and negatively regulating the epidermal growth factor receptor (EGFR). These data support a model in which LRIG1 dampens the stem cell response to EGFR ligands within the cortical environment to slow their proliferation as they transition to postnatal adult NSCs.

The Role of Astrocytes in the Generation, Migration, and Integration of New Neurons in the Adult Olfactory Bulb.

In mammals, new neurons in the adult olfactory bulb originate from a pool of neural stem cells in the subventricular zone of the lateral ventricles. Adult-born cells play an important role in odor information processing by adjusting the neuronal network to changing environmental conditions. Olfactory bulb neurogenesis is supported by several non-neuronal cells. In this review, we focus on the role of astroglial cells in the generation, migration, integration, and survival of new neurons in the adult forebrain. In the subventricular zone, neural stem cells with astrocytic properties display regional and temporal specificity when generating different neuronal subtypes. Non-neurogenic astrocytes contribute to the establishment and maintenance of the neurogenic niche. Neuroblast chains migrate through the rostral migratory stream ensheathed by astrocytic processes. Astrocytes play an important regulatory role in neuroblast migration and also assist in the development of a vasculature scaffold in the migratory stream that is essential for neuroblast migration in the postnatal brain. In the olfactory bulb, astrocytes help to modulate the network through a complex release of cytokines, regulate blood flow, and provide metabolic support, which may promote the integration and survival of new neurons. Astrocytes thus play a pivotal role in various processes of adult olfactory bulb neurogenesis, and it is likely that many other functions of these glial cells will emerge in the near future.

Bioluminescence imaging of the brain response to acute inflammation in living C/EBP reporter mice.

The transcription factor CCAAT enhancer binding protein (C/EBP) is a key regulator of inflammation and immune responses, and recent studies suggest it is involved in inflammatory processes in the nervous system. We generated a transgenic reporter mouse model, carrying the luciferase (luc) gene under the transcriptional control of C/EBP, for visualising C/EBP activity in vivo. Real-time bioluminescence imaging reflecting C/EBP activity was performed in an acute inflammation model, after systemic administration of lipopolysaccharides (LPS), in C/EBP-luc mice. A striking activity of C/EBP was imaged predominantly in the brain of living C/EBP-luc mice in response to LPS, showing for the first time in vivo that C/EBP mediates the brain response to inflammation. Furthermore, dexamethasone, a potent anti-inflammatory agent, diminished the LPS-induced C/EBP activity demonstrating the physiological regulation of bioluminescence intensity in the brain of C/EBP-luc mice. Our results implicate that C/EBP reporter mice have the potential to be a valuable tool for studies on the mechanisms of brain inflammation in vivo and for the noninvasive preclinical evaluation of therapeutic agents targeting neuroinflammatory diseases.