Microglial calcium signal acts as a rapid sensor of single neuron damage in vivo

AbstractIn the healthy adult brain microglia, the main immune-competent cells of the CNS, have a distinct (so-called resting or surveying) phenotype. Resting microglia can only be studied in vivo since any isolation of brain tissue inevitably triggers microglial activation. Here we used in vivo two-photon imaging to obtain a first insight into Ca2+ signaling in resting cortical microglia. The majority (80%) of microglial cells showed no spontaneous Ca2+ transients at rest and in conditions of strong neuronal activity. However, they reliably responded with large, generalized Ca2+ transients to damage of an individual neuron. These damage-induced responses had a short latency (0.4–4s) and were localized to the immediate vicinity of the damaged neuron (<50μm cell body-to-cell body distance). They were occluded by the application of ATPγS as well as UDP and 2-MeSADP, the agonists of metabotropic P2Y receptors, and they required Ca2+ release from the intracellular Ca2+ stores. Thus, our in vivo data suggest that microglial Ca2+ signals occur mostly under pathological conditions and identify a Ca2+ store-operated signal, which represents a very sensitive, rapid, and highly localized response of microglial cells to brain damage. This article is part of a Special Issue entitled: 11th European Symposium on Calcium

Long-term in vivo single-cell tracking reveals the switch of migration patterns in adult-born juxtaglomerular cells of the mouse olfactory bulb

The behavior of adult-born cells can be easily monitored in cell culture or in lower model organisms, but longitudinal observation of individual mammalian adult-born cells in their native microenvironment still proves to be a challenge. Here we have established an approach named optical cell positioning system for long-term in vivo single-cell tracking, which integrates red-green-blue cell labeling with repeated angiography. By combining this approach with in vivo two-photon imaging technique, we characterized the in vivo migration patterns of adult-born neurons in the olfactory bulb. In contrast to the traditional view of mere radial migration of adult-born cells within the bulb, we found that juxtaglomerular cells switch from radial migration to long distance lateral migration upon arrival in their destination layer. This unique long-distance lateral migration has characteristic temporal (stop-and-go) and spatial (migratory, unidirectional or multidirectional) patterns, with a clear cell age-dependent decrease in the migration speed. The active migration of adult-born cells coincides with the time period of initial fate determination and is likely to impact on the integration sites of adult-born cells, their odor responsiveness, as well as their survival rate

Endogenous but not sensory-driven activity controls migration, morphogenesis and survival of adult-born juxtaglomerular neurons in the mouse olfactory bulb.

The development and survival of adult-born neurons are believed to be driven by sensory signaling. Here, in vivo analyses of motility, morphology and Ca2+ signaling, as well as transcriptome analyses of adult-born juxtaglomerular cells with reduced endogenous excitability (via cell-specific overexpression of either Kv1.2 or Kir2.1 K+ channels), revealed a pronounced impairment of migration, morphogenesis, survival, and functional integration of these cells into the mouse olfactory bulb, accompanied by a reduction in cytosolic Ca2+ fluctuations, phosphorylation of CREB and pCREB-mediated gene expression. Moreover, K+ channel overexpression strongly downregulated genes involved in neuronal migration, differentiation, and morphogenesis and upregulated apoptosis-related genes, thus locking adult-born cells in an immature and vulnerable state. Surprisingly, cells deprived of sensory-driven activity developed normally. Together, the data reveal signaling pathways connecting the endogenous intermittent neuronal activity/Ca2+ fluctuations as well as enhanced Kv1.2/Kir2.1 K+ channel function to migration, maturation, and survival of adult-born neurons

Optimized ratiometric calcium sensors for functional in vivo imaging of neurons and T lymphocytes

Author Posting. © The Author(s), 2013. This is the author's version of the work. It is posted here by permission of Nature Publishing Group for personal use, not for redistribution. The definitive version was published in Nature Methods 11 (2014): 175-182, doi:10.1038/nmeth.2773.The quality of genetically encoded calcium indicators (GECIs) has improved dramatically in recent years, but high-performing ratiometric indicators are still rare. Here we describe a series of fluorescence resonance energy transfer (FRET)-based calcium biosensors with a reduced number of calcium binding sites per sensor. These ‘Twitch’ sensors are based on the C-terminal domain of Opsanus troponin C. Their FRET responses were optimized by a large-scale functional screen in bacterial colonies, refined by a secondary screen in rat hippocampal neuron cultures. We tested the in vivo performance of the most sensitive variants in the brain and lymph nodes of mice. The sensitivity of the “Twitch” sensors matched that of synthetic calcium dyes and allowed visualization of tonic action potential firing in neurons and high resolution functional tracking of T lymphocytes. Given their ratiometric readout, their brightness, large dynamic range and linear response properties, Twitch sensors represent versatile tools for neuroscience and immunology.2014-07-0

Understanding normal brain aging

Background!#!The d evelopment of surgical techniques and specialization and specifically complication management in pancreatic surgery have improved surgical outcomes as well as oncological results in pancreatic surgery in recent decades. Historical morbidity and especially mortality rates of up to 80% have decreased to below 5% today. This review summarizes the current state of the art in pancreatic cancer surgery.!##!Methods!#!The present literature and clinical experience are summarized to give an overview of the present best practice in pancreatic surgery as one of the most advanced surgical disciplines today.!##!Results!#!Based on the available literature, three important aspects contribute to best patient care in pancreatic surgery, namely, surgical progress, interdisciplinary complication management, and multimodal oncological treatment in case of pancreatic cancer. In addition, minimally invasive and robotic procedures are currently fields of development and specific topics of research.!##!Conclusion!#!In experienced hands, pancreatic surgery-despite being one of the most challenging fields of surgery-is a safe domain today. The impact of multimodal, especially adjuvant, therapy for oncological indications is well established and evidence-based. New technologies are evolving and will be evaluated with high-evidence studies in the near future