Release probability increases towards distal dendrites boosting high-frequency signal transfer in the rodent hippocampus

Dendritic integration of synaptic inputs involves their increased electrotonic attenuation at distal dendrites, which can be counterbalanced by the increased synaptic receptor density. However, during network activity the influence of individual synapses depends on their release fidelity, the dendritic distribution of which remains poorly understood. Here, we employed classical optical quantal analyses and a genetically encoded optical glutamate sensor in acute hippocampal slices of rats and mice to monitor release at CA3-CA1 synapses. We find that their release probability increases with greater distances from the soma. Similar-fidelity synapses tend to group together whereas release probability shows no trends regarding the branch ends. Simulations with a realistic CA1 pyramidal cell hosting stochastic synapses suggest that the observed trends boost signal transfer fidelity, particularly at higher input frequencies. Because high-frequency bursting has been associated with learning, the release probability pattern we have found may play a key role in memory trace formation

Time-Resolved Imaging Reveals Heterogeneous Landscapes of Nanomolar Ca(2+) in Neurons and Astroglia

Maintaining low intracellular calcium is essential to the functioning of brain cells, yet the phenomenology and mechanisms involved remain an enigma. We have advanced a two-photon excitation time-resolved imaging technique, which exploits high sensitivity of the OGB-1 fluorescence lifetime to nanomolar Ca(2+) concentration ([Ca(2+)]) and enables a high data acquisition rate in situ. The [Ca(2+)] readout is not affected by dye concentration, light scattering, photobleaching, micro-viscosity, temperature, or the main known concomitants of cellular activity. In quiescent tissue, standard whole-cell configuration has little effect on resting [Ca(2+)] inside neuronal dendrites or inside astroglia dye-filled via gap junctions. Mapping basal [Ca(2+)] in neurons and astrocytes with submicron resolution unveils heterogeneous concentration landscapes that depend on age and preceding activity. The rich information content represented by such landscapes in acute slices and in vivo promises to unveil the hitherto unexplored, potentially fundamental aspects of brain cell physiology. VIDEO ABSTRACT

Glutamate Imaging Reveals Multiple Sites of Stochastic Release in the CA3 Giant Mossy Fiber Boutons

One of the most studied central synapses which have provided fundamental insights into cellular mechanisms of neural connectivity is the “giant” excitatory connection between hippocampal mossy fibers (MFs) and CA3 pyramidal cells. Its large presynaptic bouton features multiple release sites and is densely packed with thousands of synaptic vesicles, to sustain a highly facilitating “detonator” transmission. However, whether glutamate release sites at this synapse act independently, in a stochastic manner, or rather synchronously, remains poorly understood. This knowledge is critical for a better understanding of mechanisms underpinning presynaptic plasticity and postsynaptic signal integration rules. Here, we use the optical glutamate sensor SF-iGluSnFR and the intracellular Ca2+ indicator Cal-590 to monitor spike-evoked glutamate release and presynaptic calcium entry in MF boutons. Multiplexed imaging reveals that distinct sites in individual MF giant boutons release glutamate in a probabilistic fashion, also showing use-dependent short-term facilitation. The present approach provides novel insights into the basic mechanisms of neurotransmitter release at excitatory synapses

Virus–Host Interactions Between Nonsecretors and Human Norovirus

BACKGROUND & AIMS: Human norovirus infection is the leading cause of acute gastroenteritis. Genetic polymorphisms, mediated by the FUT2 gene (secretor enzyme), define strain susceptibility. Secretors express a diverse set of fucosylated histoblood group antigen carbohydrates (HBGA) on mucosal cells; nonsecretors (FUT2-/-) express a limited array of HBGAs. Thus, nonsecretors have less diverse norovirus strain infections, including resistance to the epidemiologically dominant GII.4 strains. Because future human norovirus vaccines will comprise GII.4 antigen and because secretor phenotype impacts GII.4 infection and immunity, nonsecretors may mimic young children immunologically in response to GII.4 vaccination, providing a needed model to study crossprotection in the context of limited pre-exposure. METHODS: By using specimens collected from the first characterized nonsecretor cohort naturally infected with GII.2 human norovirus, we evaluated the breadth of serologic immunity by surrogate neutralization assays, and cellular activation and cytokine production by flow cytometry. RESULTS: GII.2 infection resulted in broad antibody and cellular immunity activation that persisted for at least 30 days for T cells, monocytes, and dendritic cells, and for 180 days for blocking antibody. Multiple cellular lineages expressing interferon-g and tumor necrosis factor-a dominated the response. Both T-cell and B-cell responses were cross-reactive with other GII strains, but not GI strains. To promote entry mechanisms, inclusion of bile acids was essential for GII.2 binding to nonsecretor HBGAs. CONCLUSIONS: These data support development of withingenogroup, cross-reactive antibody and T-cell immunity, key outcomes that may provide the foundation for eliciting broad immune responses after GII.4 vaccination in individuals with limited GII.4 immunity, including young children

Monitoring single-synapse glutamate release and presynaptic calcium concentration in organised brain tissue

Brain function relies in large part on Ca2+-dependent release of the excitatory neurotransmitter glutamate from neuronal axons. Establishing the causal relationship between presynaptic Ca2+ dynamics and probabilistic glutamate release is therefore a fundamental quest across neurosciences. Its progress, however, has hitherto depended primarily on the exploration of either cultured nerve cells or giant central synapses accessible to direct experimental probing in situ. Here we show that combining patch-clamp with time-resolved imaging of Ca2+ −sensitive fluorescence lifetime of Oregon Green BAPTA-1 (Tornado-FLIM) enables readout of single spike-evoked presynaptic Ca2+ concentration dynamics, with nanomolar sensitivity, in individual neuronal axons in acute brain slices. In parallel, intensity Tornado imaging of a locally expressed extracellular optical glutamate sensor iGluSnFr provides direct monitoring of single-quantum, single-synapse glutamate releases in situ. These two methods pave the way for simultaneous registration of presynaptic Ca2+ dynamics and transmitter release in an intact brain at the level of individual synapses

Multiplex imaging relates quantal glutamate release to presynaptic Ca2+ homeostasis at multiple synapses in situ

Information processing by brain circuits depends on Ca2+-dependent, stochastic release of the excitatory neurotransmitter glutamate. Whilst optical glutamate sensors have enabled detection of synaptic discharges, understanding presynaptic machinery requires simultaneous readout of glutamate release and nanomolar presynaptic Ca2+ in situ. Here, we find that the fluorescence lifetime of the red-shifted Ca2+ indicator Cal-590 is Ca2+-sensitive in the nanomolar range, and employ it in combination with green glutamate sensors to relate quantal neurotransmission to presynaptic Ca2+ kinetics. Multiplexed imaging of individual and multiple synapses in identified axonal circuits reveals that glutamate release efficacy, but not its short-term plasticity, varies with time-dependent fluctuations in presynaptic resting Ca2+ or spike-evoked Ca2+ entry. Within individual presynaptic boutons, we find no nanoscopic co-localisation of evoked presynaptic Ca2+ entry with the prevalent glutamate release site, suggesting loose coupling between the two. The approach enables a better understanding of release machinery at central synapses

Neuronal adaptation involves rapid expansion of the action potential initiation site

Action potential (AP) generation is the key to information processing in the brain. Although APs are normally initiated in the axonal initial segment, developmental adaptation or prolonged network activity may alter the initiation site geometry thus affecting cell excitability. Here we find that hippocampal dentate granule cells adapt their spiking threshold to the kinetics of the ongoing dendrosomatic excitatory input by expanding the AP initiation area away from the soma while also decelerating local axonal spikes. Dual-patch soma-axon recordings combined with axonal Na+ and Ca2+ imaging and biophysical modeling show that the underlying mechanism involves distance-dependent inactivation of axonal Na+ channels due to somatic depolarization propagating into the axon. Thus the ensuing changes in the AP initiation zone and local AP propagation could provide activity-dependent control of cell excitability and spiking on a relatively rapid time scale

A Mechanical Mass Sensor with Yoctogram Resolution

Nanoelectromechanical systems (NEMS) have generated considerable interest as inertial mass sensors. NEMS resonators have been used to weigh cells, biomolecules, and gas molecules, creating many new possibilities for biological and chemical analysis [1-4]. Recently, NEMS-based mass sensors have been employed as a new tool in surface science in order to study e.g. the phase transitions or the diffusion of adsorbed atoms on nanoscale objects [5-7]. A key point in all these experiments is the ability to resolve small masses. Here we report on mass sensing experiments with a resolution of 1.7 yg (1 yg = 10^-24 g), which corresponds to the mass of one proton, or one hydrogen atom. The resonator is made of a ~150 nm long carbon nanotube resonator vibrating at nearly 2 GHz. The unprecedented level of sensitivity allows us to detect adsorption events of naphthalene molecules (C10H8) and to measure the binding energy of a Xe atom on the nanotube surface (131 meV). These ultrasensitive nanotube resonators offer new opportunities for mass spectrometry, magnetometry, and adsorption experiments.Comment: submitted version of the manuscrip

Toward molecular trait-based ecology through integration of biogeochemical, geographical and metagenomic data

Using metagenomic ‘parts lists' to study microbial ecology remains a significant challenge. This work proposes a molecular trait-based approach to biogeography by integrating metagenomic data with external metadata and using functional community composition as readout