RESUMO
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.
Assuntos
Células-Tronco Adultas/metabolismo , Cálcio/metabolismo , Ritmo Circadiano , Espaço Intracelular/metabolismo , Células-Tronco Neurais/metabolismo , Células-Tronco Adultas/citologia , Células-Tronco Adultas/efeitos dos fármacos , Animais , Astrócitos/efeitos dos fármacos , Astrócitos/metabolismo , Comportamento Animal/efeitos dos fármacos , Divisão Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Ritmo Circadiano/efeitos dos fármacos , Citosol/metabolismo , Fator de Crescimento Epidérmico/farmacologia , Receptores de Inositol 1,4,5-Trifosfato/metabolismo , Melatonina/metabolismo , Camundongos , Células-Tronco Neurais/citologia , Células-Tronco Neurais/efeitos dos fármacos , Optogenética , Transdução de Sinais/efeitos dos fármacos , Triptaminas/farmacologiaRESUMO
Neural stem cells (NSCs) are maintained in specific regions of the postnatal brain and contribute to its structural and functional plasticity. However, the long-term renewal potential of NSCs and their mode of division remain elusive. The use of advanced in vivo live imaging approaches may expand our knowledge of NSC physiology and provide new information for cell replacement therapies. In this Review, we discuss the in vivo imaging methods used to study NSC dynamics and recent live-imaging results with respect to specific intracellular pathways that allow NSCs to integrate and decode different micro-environmental signals. Lastly, we discuss future directions that may provide answers to unresolved questions regarding NSC physiology.
Assuntos
Células-Tronco Neurais/fisiologia , Animais , Encéfalo/fisiologia , Terapia Baseada em Transplante de Células e Tecidos/métodos , Humanos , Cuidado Pós-Natal/métodos , Transdução de Sinais/fisiologiaRESUMO
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).