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For decades, "all-or-none" and "kiss-and-run" were thought to be the only major exocytotic release modes in cell-to-cell communication, while the significance of partial release has not yet been widely recognized and accepted owing to the lack of direct evidence for exocytotic partial release. Correlative imaging with transmission electron microscopy and NanoSIMS imaging and a dual stable isotope labeling approach was used to study the cargo status of vesicles before and after exocytosis; demonstrating a measurable loss of transmitter in individual vesicles following stimulation due to partial release. Model secretory cells were incubated with

The delivery of antisense oligonucleotides (ASOs) to specific cell types via targeted endocytosis is challenging due to the low cell surface expression of target receptors and inefficient escape of ASOs from the endosomal pathway. Conjugating ASOs to glucagon-like peptide 1 (GLP1) leads to efficient target knockdown, specifically in pancreatic β-cells. It is presumed that ASOs dissociate from GLP1 intracellularly to enable an ASO interaction with its target RNA. It is unknown where or when this happens following GLP1-ASO binding to GLP1R and endocytosis. Here, we use correlative nanoscale secondary ion mass spectroscopy (NanoSIMS) and transmission electron microscopy to explore GLP1-ASO subcellular trafficking in GLP1R overexpressing HEK293 cells. We isotopically label both eGLP1 and ASO, which do not affect the eGLP1-ASO conjugate function. We found that the eGLP1 peptide and ASO are not detected at the same level in the same endosomes, within 30 min of GLP1R-HEK293 cell exposure to eGLP1-ASO. When we utilized different linker chemistry to stabilize the GLP1-ASO conjugate, we observed more ASO located with GLP1 compared to cell incubation with the less stable conjugate. Overall, our work suggests that the ASO separates from GLP1 relatively early in the endocytic pathway, and that linker chemistry might impact the GLP1-ASO function.

[Image: see text] Mass spectrometry imaging is a field that promises to become a mainstream bioanalysis technology by allowing the combination of single-cell imaging and subcellular quantitative analysis. The frontier of single-cell imaging has advanced to the point where it is now possible to compare the chemical contents of individual organelles in terms of raw or normalized ion signal. However, to realize the full potential of this technology, it is necessary to move beyond this concept of relative quantification. Here we present a nanoSIMS imaging method that directly measures the absolute concentration of an organelle-associated, isotopically labeled, pro-drug directly from a mass spectrometry image. This is validated with a recently developed nanoelectrochemistry method for single organelles. We establish a limit of detection based on the number of isotopic labels used and the volume of the organelle of interest, also offering this calculation as a web application. This approach allows subcellular quantification of drugs and metabolites, an overarching and previously unmet goal in cell science and pharmaceutical development.

The short-lived (26)Al radionuclide is thought to have been admixed into the initially (26)Al-poor protosolar molecular cloud before or contemporaneously with its collapse. Bulk inner Solar System reservoirs record positively correlated variability in mass-independent (54)Cr and (26)Mg*, the decay product of (26)Al. This correlation is interpreted as reflecting progressive thermal processing of in-falling (26)Al-rich molecular cloud material in the inner Solar System. The thermally unprocessed molecular cloud matter reflecting the nucleosynthetic makeup of the molecular cloud before the last addition of stellar-derived (26)Al has not been identified yet but may be preserved in planetesimals that accreted in the outer Solar System. We show that metal-rich carbonaceous chondrites and their components have a unique isotopic signature extending from an inner Solar System composition toward a (26)Mg*-depleted and (54)Cr-enriched component. This composition is consistent with that expected for thermally unprocessed primordial molecular cloud material before its pollution by stellar-derived (26)Al. The (26)Mg* and (54)Cr compositions of bulk metal-rich chondrites require significant amounts (25-50%) of primordial molecular cloud matter in their precursor material. Given that such high fractions of primordial molecular cloud material are expected to survive only in the outer Solar System, we infer that, similarly to cometary bodies, metal-rich carbonaceous chondrites are samples of planetesimals that accreted beyond the orbits of the gas giants. The lack of evidence for this material in other chondrite groups requires isolation from the outer Solar System, possibly by the opening of disk gaps from the early formation of gas giants.

Ichthyosaurs are extinct marine reptiles that display a notable external similarity to modern toothed whales. Here we show that this resemblance is more than skin deep. We apply a multidisciplinary experimental approach to characterize the cellular and molecular composition of integumental tissues in an exceptionally preserved specimen of the Early Jurassic ichthyosaur Stenopterygius. Our analyses recovered still-flexible remnants of the original scaleless skin, which comprises morphologically distinct epidermal and dermal layers. These are underlain by insulating blubber that would have augmented streamlining, buoyancy and homeothermy. Additionally, we identify endogenous proteinaceous and lipid constituents, together with keratinocytes and branched melanophores that contain eumelanin pigment. Distributional variation of melanophores across the body suggests countershading, possibly enhanced by physiological adjustments of colour to enable photoprotection, concealment and/or thermoregulation. Convergence of ichthyosaurs with extant marine amniotes thus extends to the ultrastructural and molecular levels, reflecting the omnipresent constraints of their shared adaptation to pelagic life.

We have measured the abundance and isotopic composition of water in apatites from several lunar rocks representing Potassium (K), Rare Earth Elements (REE), and Phosphorus (P) − KREEP − rich lithologies, including felsites, quartz monzodiorites (QMDs), a troctolite, and an alkali anorthosite. The H-isotope data from apatite provide evidence for multiple reservoirs in the lunar interior. Apatite measurements from some KREEP-rich intrusive rocks display moderately elevated δD signatures, while other samples show δD signatures similar to the range known for the terrestrial upper mantle. Apatite grains in Apollo 15 quartz monzodiorites have the lowest δD values measured from the Moon so far (as low as −749‰), and could potentially represent a D-depleted reservoir in the lunar interior that had not been identified until now. Apatite in all of these intrusive rocks contains 6500 ppm H2O). Complexities in partitioning of volatiles into apatite make this comparison uncertain, but measurements of residual glass in KREEP basalt fragments in breccia 15358 independently show that the KREEP basaltic magmas were low in water. The source of 15358 contained ∼10 ppm H2O, about an order of magnitude lower than the source of the Apollo 17 pyroclastic glass beads, suggesting potential variations in the distribution of water in the lunar interior.

The rate of the transfer of deuterium from D 3 + to the three types (benzylic, aliphatic and aromatic) of bonds of 2-Ethylnaphthalene (and Methylnaphthalene) has been measured in the laboratory. The D/H ratio of each bond was calculated from the mass spectrum of the different isotopomers produced in the source of the GC mass spectrometer. The relations between the D/H ratio of the different isotopomers allow to calculate the internal isotopic fractionation factors (αi−j = (D/H)i/(D/H)j) between the 3 bonds: αBenzylic–Aromatic = 1.99 ± 0.38 and αAliphatic–Aromatic = 1.22 ± 0.39. Isotopic exchange rate constants are ⩾2.2 × 10−7±0.4 s−1, determined at room temperature for 10−3 g of 2-Ethylnaphthalene in contact with 5 × 1012 cm−3 of D 3 + . These results are compatible with those measured in the insoluble organic matter (IOM) isolated from the Orgueil meteorite ( Remusat et al., 2006 ), i.e., αBenzylic–Aromatic = 1.96 ± 0.05 and αAliphatic–Aromatic = 1.35 ± 0.05. Using the calculated dust and H2D+ densities from the models of deuterium chemistry in protostellar disks, the duration needed to enrich the IOM up to its measured D/H ratio is ⩽104±1 s. These laboratory results are in agreement with a late deuteration of the insoluble organic matter taking place in the solar T-Tauri disk by interaction of the organic dust with H2D+.

The insoluble organic matter (IOM) of an unequilibrated enstatite chondrite Sahara (SAH) 97096 has been investigated using a battery of analytical techniques. As the enstatite chondrites are thought to have formed in a reduced environment at higher temperatures than carbonaceous chondrites, they constitute an interesting comparative material to test the heterogeneities of the IOM in the solar system and to constrain the processes that could affect IOM during solar system evolution. The SAH 97096 IOM is found in situ: as submicrometer grains in the network of fine-grained matrix occurring mostly around chondrules and as inclusions in metallic nodules, where the carbonaceous matter appears to be more graphitized. IOM in these two settings has very similar $\delta^{15}N$ and $\delta^{13}C$; this supports the idea that graphitized inclusions in metal could be formed by metal catalytic graphitization of matrix IOM. A detailed comparison between the IOM extracted from a fresh part and a terrestrially weathered part of SAH 97096 shows the similarity between both IOM samples in spite of the high degree of mineral alteration in the latter. The isolated IOM exhibits a heterogeneous polyaromatic macromolecular structure, sometimes highly graphitized, without any detectable free radicals and deuterium-heterogeneity and having mean H- and N-isotopic compositions in the range of values observed for carbonaceous chondrites. It contains some submicrometer-sized areas highly enriched in $^{15}N$ ($\delta^{15}N$ up to 1600 permil). These observations reinforce the idea that the IOM found in carbonaceous chondrites is a common component widespread in the solar system. Most of the features of SAH 97096 IOM could be explained by the thermal modification of this main component.

International audience; Quantification of water content is relevant in various topics in geology and planetary sciences. NanoSIMS has capabilities for high spatial resolution imaging and offers opportunities to accurately quantify water contents at fine scale on small surface areas. The main concern using ion microprobe techniques is to estimate and minimize contribution of water contamination, from residual gas in the sample chamber, sticking onto the surface of the sample. Here we tackle a set of sputtering/analytical parameters and we evaluate their relative influence on theOH−/Si− ratio.We demonstrate that a high erosion rate, reached using a primary beamintensity of ~25 pA, is sufficient to lower this OH contamination for basaltic glass. This leads us to describe a procedure to correct for OH contamination and thus determine accurate values of OH/Si ratio in order to quantify water contents in silicate materials using NanoSIMS imaging.

International audience; We describe a procedure to determine precise and accurate elemental abundance by means of quantitative imaging using secondary ion mass spectrometry (on images covering 10 10 mm2) applied to natural Insoluble Organic Matter (IOM). Dynamic SIMS conditions are reached for a 16 keV Cs+ fluence of >2.0 1017 Cs+ cm 2 implanted at the surface of the IOM. Once the sample surface is saturated in cesium, steady-state implantation and sputtering yield are reached: constant secondary ion count rates are then observed. Two calibrations of the nitrogen abundance are presented. The nitrogen abundance is expressed either as [N] or N/C atomic ratio calibrated by means of 12C14N /12C or 12C14N /12C2 ionic ratios respectively. The 12C14N /12C uncertainty is always larger than the uncertainty of 12C14N /12C2 . At a smaller scale inside an image ( 1 1 mm2), a little larger than the primary beam size, processes induced by topographic irregularity of the IOM powder result in a variation of the 12C14N ,12C2 and 12C count rates and increase the variability of the ionic ratios. The chemical contrast produced by a rough sample surface exposed to the Cs+ rastering is evaluated with a scratched epoxy resin for 12C14N /12C and 12C14N /12C2 elemental ratios. Owing to similarities in emission parameters, the determination of the N/C ratio in rough organic surfaces is improved by using 12C 14N /12C2 instead of 12C 14N /12C .

NanoSIMS is a relatively new technology that is being applied to ancient carbonaceous structures to gain insight into their biogenicity and syngeneity. NanoSIMS studies of well preserved organic microfossils from the Neoproterozoic (similar to 0.8Ga) Bitter Springs Formation have established elemental distributions in undisputedly biogenic structures. Results demonstrate that sub-micron scale maps of metabolically important elements (carbon [C], nitrogen [measured as CN ion], and sulfur [S]) can be correlated with kerogenous structures identified by optical microscopy. Spatial distributions of C, CN, and S in individual microfossils are nearly identical, and variations in concentrations of these elements parallel one another. In elemental maps, C, CN, and S appear as globules, aligned to form remnant walls or sheaths of fossiliferous structures. The aligned character and parallel variation of C and CN are the strongest indicators of biogenicity.