Cell Calcium

Numbers of activated receptor dictate efficacy of neurotransmitter stimulation. Many PLC coupled receptors activated by ligands elicit canonical downstream Gq/11 pathway to induce endogenous Ca2+ gated chloride channels. The coupling from receptors to effectors was analyzed in Xenopus oocytes expressing genetically modified angiotensin receptor type 1 receptor (AT1R). The latency between ATII binding and Ca2+-induced Cl- current surge was inversely correlated. AT1R activation triggered a chain of chemical reactions, of which the products were playing messengers for subsequent events. Messenger accumulation must rate-limit the agonism. For accurate quantification the speed of ATII triggered the i Cl-. The T-form AT1R-IRK1 fusion exhibits faster induction compared to the M-form. The latency of the recorded none vanished i Cl-, marking the lowest genuine calcium activation, took place at earlier time point by the timer time. The evoked i Cl- however reached similar maximal amplitudes. This kinetic effect raises the possibility to use temporal coding to complement amplitude coding (analogous to FM versus AM radio transmission) for receptor-agonist pairs.

The endoplasmic reticulum (ER) is an important regulator of Ca2+ in cells and dysregulation of ER calcium homeostasis can lead to numerous pathologies. Understanding how various pharmacological and genetic perturbations of ER Ca2+ homeostasis impacts cellular physiology would likely be facilitated by more quantitative measurements of ER Ca2+ levels that allow easier comparisons across conditions. Here, we developed a ratiometric version of our original ER-GCaMP probe that allows for more quantitative comparisons of the concentration of Ca2+ in the ER across cell types and sub-cellular compartments. Using this approach we show that the resting concentration of ER Ca2+ in primary dissociated neurons is substantially lower than that in measured in embryonic fibroblasts.

Arginine vasopressin (AVP) is well known for regulating fluid volume, osmotic balance, and vascular tone. Its role in the regulation of pancreatic and {beta} cell function has been reported, yet its effects are not fully understood, particularly regarding its interaction with plasma glucose levels. The osmotic and volume challenges posed by hyper- and hypoglycaemia in diabetes can be a significant complication of effective hormonal regulation of metabolism. In this study, we investigated the effects of AVP and synthetic peptide receptor agonists and antagonists on and {beta} cells in pancreatic tissue slices using live confocal Ca2+ imaging. Our findings demonstrate that AVP exerts glucose-dependent effects on both cell types. At low glucose concentrations, AVP, in combination with physiologically or pharmacologically increased cAMP levels, selectively activated cells without significantly affecting {beta} cells. In contrast, at higher glucose concentrations and pharmacologically elevated cAMP levels, physiological levels of AVP enhanced {beta} cell activity, leading to increased Ca2+ oscillations and insulin release. In both cell types, AVP displayed a bell-shaped concentration dependence, with lower AVP concentrations stimulating hormone release and higher concentrations leading to diminished responses, consistent with inositol trisphosphate receptor (IP3R) activation and inactivation properties. Furthermore, our results indicate that AVP acts primarily through V1b receptors in {beta} cells, with no involvement of V1a, V2 or oxytocin receptors. These findings provide new insights into the modulation of glucose-dependent release of pancreatic hormones by AVP in the context of changed blood osmolality due to hyper- or hypoglycemia in diabetes. Importantly, these results emphasize the potential of targeting AVP signaling pathways as a therapeutic approach in diabetes research, aiming to improve hormone regulation and nutrient homeostasis. HighlightsO_LIHighly spatio-temporally resolved imaging of islet Ca2+ oscillations on pancreatic tissue slices provides an in situ-like model for physiological and pharmacological approaches. C_LIO_LIPhysiological glucose stimulation triggers non-linear {beta} cell collective responses that must be taken into account when interpreting single concentration pharmacological experiments. C_LIO_LIIn a high cAMP context, AVP acts through V1b receptors on islet and {beta} cells, exhibiting a bell-shaped dependence driven by the activation-inactivation properties of IP3 receptors. C_LIO_LIAVP modulates glucose-dependent effects on and {beta} cells in a physiological concentration range, in the presence of altered blood osmolality or volume due to hyperglycemia, or to the direct effects of hypoglycemia in diabetes. C_LI

Successful Ca2+ signaling requires appropriate Ca2+ storage and buffering by endoplasmic reticulum (ER) and mitochondria. Recent research has elucidated how Ca2+ storage in ER is controlled by STIM1-mediated store-operated Ca2+ entry (SOCE). However, how cells employ mitochondrial Ca2+ buffering to maintain Ca2+ homeostasis has remained elusive. Here, with the use of mitochondria-tethered Ca2+ sensor, we noticed local Ca2+ flickering within individual mitochondria. Those Ca2+ flickers were generated on ER-mitochondrial contact sites (EMC), as indicated by their downregulation under EMC breakdown and upregulation under EMC induction. Surprisingly, EMC breakdown increased SOCE while EMC induction reduced SOCE. Further investigations revealed that EMC effect on SOCE was not through biological functions of mitochondria or through STIM1 regulators, but via IP3R-VDAC1-driven mitochondrial Ca2+ flickers on EMC. Those flickers depleted peri-EMC Ca2+ inside ER, resulting in STIM1 sequestration around EMC to cause SOCE reduction. Moreover, EMC breakdown also increased availability of STIM1 to bind with microtubule plus ends, preventing STIM1 over-activation and SOCE upregulation. Overall, ER, mitochondria and microtubules constitute a self-sufficient system to control Ca2+ homeostasis, driven by mitochondrial Ca2+ flickers to reduce SOCE and to prevent intracellular Ca2+ overload.

The neuronal intracellular chloride concentration [Cl-]i is critical for {gamma}-aminobutyric acid type A (GABAA) receptor-mediated transmission. Degradation of the extracellular matrix (ECM) is associated with raised [Cl-]I but neither the mechanisms underlying this effect nor the consequences for GABA- mediated transmission are well understood. Hitherto it has been unclear how to reconcile the effect of the ECM on [Cl-]i with the established role of cation-chloride cotransporters in setting [Cl-]I. In the present work we clarify the role of the ECM in the control of neuronal [Cl-]i. By measuring [Cl-]i in central neurons from male rats we show that the ECM affects basal [Cl-]i as well as the rate of Cl- extrusion after a high load. The mechanism is not via impermeant anions but through regulation of K+-Cl--cotransporter 2 (KCC2). ECM degradation is accompanied by an N-type Ca2+-channel- and calpain-dependent reduction in the amount of KCC2 protein, increased basal [Cl-]i, reduced Cl- extrusion capacity as well as by reduced inhibitory, or even an excitatory, effect of intense GABAA- receptor mediated trans mission. This implies a previously unrecognized pathway for the control of neuronal [Cl-]i and excitability by the ECM. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=144 SRC="FIGDIR/small/527837v1_ufig1.gif" ALT="Figure 1"> View larger version (53K): org.highwire.dtl.DTLVardef@1ee0d30org.highwire.dtl.DTLVardef@1a3eb1aorg.highwire.dtl.DTLVardef@a00b44org.highwire.dtl.DTLVardef@143b43f_HPS_FORMAT_FIGEXP M_FIG C_FIG

Neuronal populations connected by gap junctions can be revealed via dye coupling of small molecules like neurobiotin and lucifer yellow. However, the extent of dye diffusion between neurons varies with connexin subtype, loading method, and neuromodulation. Due to the increasing availability of GCaMP transgenic animals, we explore the possibility of revealing gap junctional coupling using Ca2+ imaging in the Xenopus laevis tadpole motor system. Reliable axo-axonal electrical coupling was previously found in excitatory descending interneurons (dINs) using paired recordings but not with neurobiotin dye coupling. Here, we made whole-cell patch-clamp recordings with Ca2+-supplemented intracellular solution to load Ca2+ into GCaMP6s-expressing neurons, followed by Ca2+ imaging to detect potential Ca2+ diffusion across coupled neurons. Successful membrane breakthroughs led to transient fluorescence increases in the patched neuron. However, increasing the Ca2+ concentration promoted membrane resealing and rapid loss of whole-cell recordings. Regardless of recording duration, loading-triggered fluorescence only lasted up to three minutes, suggesting rapid Ca2+ clearance. Pharmacologically blocking sarcoplasmic /endoplasmic reticulum Ca2+-ATPases and plasma membrane Na+/Ca2+ exchangers did not prolong fluorescence, although sustained fluorescence was achieved with positive current injections. Counter to our expectations, fluorescence increases in Ca2+-loaded dINs did not spread to neighboring dINs. Robust intracellular Ca2+ regulation mechanisms, membrane resealing, and long dIN axons likely hindered intercellular Ca2+ diffusion. Therefore, this approach is not appropriate for revealing electrical coupling within this system.

Due to their large scale and uniquely branched architecture, neurons critically rely on active transport of mitochondria in order to match energy production and calcium buffering to local demand. Consequently, defective mitochondrial trafficking is implicated in various neurological and neurodegenerative diseases. A key signal regulating mitochondrial transport is intracellular calcium. Elevated Ca2+ levels have been demonstrated to inhibit mitochondrial transport in many cell types, including neurons. However, it is currently unclear to what extent calcium-signaling regulates axonal mitochondrial transport during realistic neuronal activity patterns. We created a robust pipeline to quantify with high spatial resolution, absolute Ca2+ concentrations. This allows us to monitor Ca2+ dynamics with pixel precision in the axon and other neuronal compartments. We found that axonal calcium levels scale with firing frequency in the range of 0.1-1{micro}M, whereas KCl-induced depolarization generated levels almost a magnitude higher. As expected, prolonged KCl-induced depolarization did inhibit axonal mitochondrial transport in primary hippocampal neurons. However, physiologically relevant neuronal activity patterns only inhibited mitochondrial transport in axonal segments which made connections to a target neuron. In non-connecting axonal segments, we were unable to trigger this inhibitory mechanism using realistic firing patterns. Thus, we confirm that neuronal activity can indeed regulate axonal mitochondrial transport, and reveal a spatial pattern to this regulation which went previously undetected. Together, these findings indicate a potent, but localized role for activity-related calcium fluctuations in the regulation of axonal mitochondrial transport.

The relationship between Ca2+ action potential (AP) activity in immature inner hair cells (IHCs) and the spontaneous ATP-dependent intercellular Ca2+ signaling in cochlear non-sensory cells (NSCs) of the greater epithelial ridge (GER) is unclear. Here, we determined that IHCs fired asynchronous Ca2+ APs also in the absence of Ca2+ activity in the GER. Patch clamp recordings from IHCs isolated from the rest of the sensory epithelium confirmed that this firing activity is an intrinsic property of immature IHCs. However, frequency, correlation index and burst duration of IHC APs increased significantly during Ca2+ wave propagation in NSCs, and depended on wave extension in the GER. Furthermore, IHC depolarization under whole cell patch clamp conditions triggered Ca2+ signals in nearby NSCs with a delay that was proportional to the distance from the stimulated IHC. Thus the immature mammalian cochlea supports bidirectional exchange of Ca2+ signals between IHCs and NSCs.\n\nIMPACT STATEMENTIn inner hair cells of the developing mammalian cochlea, Ca2+ action potentials are both intrinsic and bidirectionally coupled to the ATP-dependent Ca2+ signaling of the surrounding non-sensory cells.

Fluctuations in external calcium concentration [Ca2+]e occur during the wake and sleep cycle and during intense neuronal activity. However, the incidence of these fluctuations on neuronal excitability is not precisely known. We show here that reducing divalent cation (Ca2+ and Mg2+) concentrations from 1.3/0.8 to 0.6/0.4 mM during 15-30 minutes induces long-term potentiation of intrinsic excitability (LTP-IE) in CA1 pyramidal neurons. LTP-IE induced by low divalent cations is associated with a hyperpolarization of the action potential threshold and constitutes a positive feed-back of brain activity. This plasticity requires Ca2+-sensing receptor (CaSR), IP3 receptor (IP3R) and calcium-calmodulin kinase II (CaMKII). In fact, LTP-IE was occluded in the presence of the calcilytic NPS-2143 and absent in CRISPR CaSR neurons. In addition, inhibiting IP3R with 2-APB and CaMKII with kin considerably reduced LTP-IE magnitude. LTP-IE and synaptic potentiation (LTP) induced by spike-timing-dependent plasticity (STDP) protocol were also found to depend on CaSR as they were totally absent in CRISPR CaSR neurons. Spontaneous excitatory synaptic activity was found to be reduced by [~]35% following LTP induced by STDP. Importantly, this drop of spontaneous activity was not observed in CRISPR CaSR neurons. Taken together, these results show that CaSR plays a critical role in LTP-IE induced by low [Ca2+]e and [Mg2+]e and as well as in LTP of synaptic transmission and intrinsic excitability induced by STDP.

Epithelia are continuously exposed to a range of biomechanical forces such as compression, stretch and shear stress arising from their dynamic microenvironments and associated to their function. Changes in tension such as stretch are known to trigger cell rearrangements and divisions, and impact cellular transcription until mechanical stress is dissipated. How cells process, adapt and respond to mechanical stress is being intensively investigated. In here we focus on epithelial folding which is the fundamental process of transformation of flatmonolayers into 3D functional tissues. By combining the innovative method for fold generation, live imaging, mechanobiology tools and chemical screening, we uncover the role of calcium waves on mechanical adaptation of folded epithelia that occurs at the tissue and nuclear level. Folding associated tensional load results in the nuclear flattening which is recovered in the time scale of minutes and is dependent on the calcium wave that spread outwards from the channel and across the epithelium. By creating a mutant overexpressing LBR that relaxed nuclear envelope, we demonstrated that despite presence of calcium waves, nuclear tension increase was essential to trigger nuclear shape recovery post folding through the activation of cellular contractility in the cPLA2 dependent manner. Overall our results identify the molecular mechanism for nuclear shape recovery and indicate that mechanical stress dissipation program is activated at the level of nuclei which serve as internal tension sensors.

Cellular crosstalk is an essential process influenced by numerous factors including secreted vesicles that transfer nucleic acids, lipids, and proteins between cells. Extracellular vesicles (EVs) have been the center of many studies focusing on neuron-to-neuron communication, but the role of EVs in progenitor-to-neuron and -astrocyte communication and whether EVs display cell-type-specific features for cellular crosstalk during neurogenesis is unknown. Here, using human-derived cerebral organoids, neural progenitors, neurons, and astrocytes, we found that many proteins coded by genes associated with neurodevelopmental disorders are transported via EVs. Thus, we characterized the protein content of EVs and showed their cell type-specific dynamics and function during brain development. Changes in the physiological crosstalk between cells can lead to neurodevelopmental disorders. EVs from patients with epilepsy were found altered in composition and function. Alterations in the intracellular and extracellular compartments highlighted a clear dysregulation of protein trafficking. This study sheds new light on the biology of EVs during brain development and neurodevelopmental disorders. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=185 SRC="FIGDIR/small/546646v1_ufig1.gif" ALT="Figure 1"> View larger version (40K): org.highwire.dtl.DTLVardef@1dad15aorg.highwire.dtl.DTLVardef@e699ccorg.highwire.dtl.DTLVardef@17b371borg.highwire.dtl.DTLVardef@5efa7b_HPS_FORMAT_FIGEXP M_FIG C_FIG Graphical abstract(left) EV uptake mechanism varies depending on the receiving cell type; NPCs transport neuron EVs (nEVs) and astrocyte EVs (aEVs) to the nucleus, astrocytes localize progenitor EVs (pEVs) to the cytoplasm, and neurons retain pEVs and aEVs along the plasma membrane. (right) Cerebral organoids (COs) from progressive Myoclonus Epilepsy Type I (EPM1) patients release EVs lacking key proteins in neurodevelopment and proteins necessary for EV biogenesis and release. Illustration created using BioRender.

Cerebrospinal fluid contacting neurons (CSFcNs) are GABAergic cells that surround the central canal (CC) of the spinal cord. Their soma is located sub-ependymally and they have a dendritic-like process that ends as a bulb (the so-called "apical process"; ApPr) inside the CC. It remains unclear how this unique anatomical organization, with the soma and the ApPr located in different extracellular environments, relates to their function as a multimodal sensor of cerebrospinal fluid (CSF) composition. One of the main physiological features of CSFcNs is a prominent spontaneous electrical activity mediated by PKD2L1 (or TRPP2) channels, a non-selective cation channel of the TRP family. PKD2L1 channels have a high single-channel conductance (around 200 pS) and can be modulated by protons and mechanical forces. In this work we investigate PKD2L1 channel sensitivity to pH and its effects on CSFcNs excitability. We demonstrate that PKD2L1 spontaneous activity generates not only phasic inward currents, but also a tonic current, both of which are modulated bidirectionally by pH with a high sensitivity around physiological values. By combining electrophysiology (direct recordings from intact and isolated ApPrs) with optical methods (laser-photolysis of protons) we further show that functional PKD2L1 channels are specifically localized in the ApPr. The spatial segregation of PKD2L1 channels, along with their biophysical properties (high single-channel conductance and pH sensitivity) and the ApPrs unique membrane properties (very high input resistance) renders CSFcN excitability exquisitely sensitive to PKD2L1 modulation. Altogether, our findings illustrate how the ApPrs properties are finely tuned to support its sensory role.

The question of whether islet neogenesis occurs in adult humans has been a subject of long-standing debate. To explore the characteristics of islet endocrine cells associated with pancreatic ducts, we employed imaging mass cytometry to examine pancreatic tissues from individuals across different age groups, including those with prediabetes or type 2 diabetes (T2D). Our analysis revealed the presence of all five pancreatic islet endocrine cell types, along with two types of non-hormone-expressing endocrine cells, located within or immediately adjacent to the ducts. These cells were most abundant in infancy, with a gradual decline observed through adulthood. Notably, ductal {beta} cells predominated in infancy, whereas ductal cells became more prevalent in adulthood, and significantly increased in the group aged over 60 years. Obesity further increased the ductal {beta} cells in the subjects aged over 60 years. Under prediabetic and T2D conditions, an increase in all duct-related endocrine cells was observed. These findings indicate that ductal cells may serve as a reservoir for new pancreatic endocrine cells, offering potential insights into the promotion of endogenous {beta} cell regeneration in diabetic patients. Highlights{bigcirc} Characterization of various islet endocrine cell types related to ducts in human pancreas. {bigcirc}The insulin-positive cells are the dominant cells among all duct-related islet endocrine cell types during the infancy period, however, the glucagon-positive cells become the dominant cells in adulthood. {bigcirc}T2D, Obesity, and aging are involved in the increase in the number of duct-related endocrine cells.

Theoretical work suggests that collective spatiotemporal behaviour of integral membrane proteins (IMPs) can be modulated by annular lipids sheathing their hydrophobic moiety. Here, we present evidence for this prediction in a natural membrane by investigating the mechanism that maintains steady amount of active isoform of Lck kinase (LckA) by Lck trans-autophosphorylation offset by the phosphatase CD45. We gauged experimental suitability by quantitation of CD45 and LckA subcellular localisation, LckA generation as a function of Lck and pharmacological perturbation. Steady LckA was challenged by swapping Lck membrane anchor with structurally divergent ones expected to substantially modify Lck annular lipids, such as that of Src or the transmembrane domains of LAT, CD4, palmitoylation-defective CD4 and CD45, respectively. The data showed only small alteration of LckA, except for CD45 hydrophobic anchor that thwarted LckA, due to excessive lateral proximity to CD45. The data are best explained by annular lipids facilitating or penalising IMPs lateral proximity, hence modulating IMPs protein-protein functional interactions. Our findings can contribute to improve the understanding of biomembranes organisation.

Synaptotagmin-1 is a vesicular protein and Ca2+ sensor for Ca2+-dependent exocytosis. Ca2+ induces synaptotagmin-1 binding to its own vesicle membrane, called the cis-interaction, thus preventing the trans-interaction of synaptotagmin-1 to the plasma membrane. However, the electrostatic regulation of the cis- and trans-membrane interaction of synaptotagmin-1 was poorly understood in different Ca2+-buffering conditions. Here we provide an assay to monitor the cis- and trans-membrane interactions of synaptotagmin-1 by using native purified vesicles and the plasma membrane-mimicking liposomes (PM-liposomes). Both ATP and EGTA similarly reverse the cis-membrane interaction of synaptotagmin-1 in free [Ca2+] of 10 to 100 M. High PIP2 concentrations in the PM-liposomes reduce the Hill coefficient of vesicle fusion and synaptotagmin-1 membrane binding; this observation suggests that local PIP2 concentrations control the Ca2+-cooperativity of synaptotagmin-1. Our data provide evidence that Ca2+ chelators, including EGTA and polyphosphate anions such as ATP, ADP, and AMP, electrostatically reverse the cis-interaction of synaptotagmin-1.

Although mice are a very instrumental model in islet beta cell research, possible phenotypic differences between strains and substrains are largely neglected in the scientific community. In this study, we show important phenotypic differences in beta cell responses to glucose between NMRI, C57BL/6J, and C57BL/6N mice, i.e., the three most commonly used strains. High-resolution multicellular confocal imaging of beta cells in acute pancreas tissue slices was used to measure and quantitatively compare the calcium dynamics in response to a wide range of glucose concentrations. Strain- and substrain-specific features were found in all three phases of beta cell responses to glucose: a shift in the dose-response curve characterizing the delay to activation and deactivation in response to stimulus onset and termination, respectively, and distinct concentration-encoding principles during the plateau phase in terms of frequency, duration, and active time changes with increasing glucose concentrations. Our results underline the significance of carefully choosing and reporting the strain to enable comparison and increase reproducibility, emphasize the importance of analyzing a number of different beta cell physiological parameters characterizing the response to glucose, and provide a valuable standard for future studies on beta cell calcium dynamics in health and disease.

Calcium (Ca{superscript 2}) release from intracellular stores, Ca{superscript 2} entry across the plasma membrane, and their coordination via store-operated Ca{superscript 2} entry (SOCE) are critical for receptor-activated Ca{superscript 2} oscillations. However, the precise mechanism of Ca{superscript 2} oscillations and whether their control loop resides at the plasma membrane or intracellularly remain unresolved. By examining the dynamics of stromal interaction molecule 1 (STIM1)--an endoplasmic reticulum (ER)-localized Ca{superscript 2} sensor that activates the Orai1 channel on the plasma membrane for SOCE--and in mast cells, we found that a significant proportion of cells exhibited STIM1 oscillations with the same periodicity as Ca{superscript 2} oscillations. These cortical oscillations, occurring in the cells cortical region and shared with ER-plasma membrane (ER-PM) contact sites proteins, were only detectable using total internal reflection fluorescence microscopy (TIRFM). Notably, STIM1 oscillations could occur independently of Ca{superscript 2} oscillations. Simultaneous imaging of cytoplasmic Ca{superscript 2} and ER Ca{superscript 2} with SEPIA-ER revealed that receptor activation does not deplete ER Ca{superscript 2}, whereas receptor activation without extracellular Ca{superscript 2} influx induces cyclic ER Ca{superscript 2} depletion. However, under such nonphysiological conditions, cyclic ER Ca{superscript 2} oscillations lead to sustained STIM1 recruitment, indicating that oscillatory Ca{superscript 2} release is neither necessary nor sufficient for STIM1 oscillations. Using optogenetic tools to manipulate ER-PM contact site dynamics, we found that persistent ER-PM contact sites reduced the amplitude of Ca{superscript 2} oscillations without alteration of oscillation frequency. Together, these findings suggest an active cortical mechanism governs the rapid dissociation of ER-PM contact sites, thereby control amplitude of oscillatory Ca{superscript 2} dynamics during receptor-induced Ca{superscript 2} oscillations.

As professional secretory cells, beta cells require adaptable mRNA translation to facilitate a rapid synthesis of proteins, including insulin, in response to changing metabolic cues. Specialized mRNA translation programs are essential drivers of cellular development and differentiation. However, in the pancreatic beta cell, the majority of factors identified to promote growth and development function primarily at the level of transcription. Therefore, despite its importance, the regulatory role of mRNA translation in the formation and maintenance of functional beta cells is not well defined. In this study, we have identified a translational regulatory mechanism in the beta cell driven by the specialized mRNA translation factor, eukaryotic initiation factor 5A (eIF5A), which facilitates beta cell maturation. The mRNA translation function of eIF5A is only active when it is post-translationally modified ("hypusinated") by the enzyme deoxyhypusine synthase (DHPS). We have discovered that the absence of beta cell DHPS in mice reduces the synthesis of proteins critical to beta cell identity and function at the stage of beta cell maturation, leading to a rapid and reproducible onset of diabetes. Therefore, our work has revealed a gatekeeper of specialized mRNA translation that permits the beta cell, a metabolically responsive secretory cell, to maintain the integrity of protein synthesis necessary during times of induced or increased demand. ARTICLE HIGHLIGHTSO_LIPancreatic beta cells are professional secretory cells that require adaptable mRNA translation for the rapid, inducible synthesis of proteins, including insulin, in response to changing metabolic cues. Our previous work in the exocrine pancreas showed that development and function of the acinar cells, which are also professional secretory cells, is regulated at the level of mRNA translation by a specialized mRNA translation factor, eIF5AHYP. We hypothesized that this translational regulation, which can be a response to stress such as changes in growth or metabolism, may also occur in beta cells. C_LIO_LIGiven that the mRNA translation function of eIF5A is only active when the factor is post-translationally modified ("hypusinated") by the enzyme deoxyhypusine synthase (DHPS), we asked the question: does DHPS/eIF5AHYP regulate the formation and maintenance of functional beta cells? C_LIO_LIWe discovered that in the absence of beta cell DHPS in mice, eIF5A is not hypusinated (activated), which leads to a reduction in the synthesis of critical beta cell proteins that interrupts pathways critical for identity and function. This translational regulation occurs at weaning age, which is a stage of cellular stress and maturation for the beta cell. Therefore without DHPS/eIF5AHYP, beta cells do not mature and mice progress to hyperglycemia and diabetes. C_LIO_LIOur findings suggest that secretory cells have a mechanism to regulate mRNA translation during times of cellular stress. Our work also implies that driving an increase in mRNA translation in the beta cell might overcome or possibly reverse the beta cell defects that contribute to early dysfunction and the progression to diabetes. C_LI

G protein-coupled receptors conformational landscape can be affected by their local, microscopic interactions within the cell plasma membrane. We employ here a pleiotropic stimulus, namely osmotic swelling, to alter the cortical environment within intact cells and monitor the response in terms of receptor function and downstream signaling. We observe that in osmotically swollen cells the {beta}2-adrenergic receptor, a prototypical GPCR, favors an active conformation, resulting in cAMP transient responses to adrenergic stimulation that have increased amplitude. The results are validated in primary cell types such as adult cardiomyocytes, a model system where swelling occurs upon ischemia-reperfusion injury. Our results suggest that receptors function is finely modulated by their biophysical context, and specifically that osmotic swelling acts as a potentiator of downstream signaling, not only for the {beta}2-adrenergic receptor, but also for other receptors, hinting at a more general regulatory mechanism.

Somatostatin (SST) is a neuropeptide known to inhibit both neuronal excitability and excitatory synaptic transmission in principal neurons. While SST is expressed in some GABAergic interneurons such as Oriens-Lacunosum Moleculare (O-LM) cells, the precise conditions governing SST release remain poorly defined. We show here that, in the presence of GABAA receptor antagonist, single O-LM interneurons of CA1 are able to inhibit pyramidal neuron excitability when O-LM cells fire at frequencies above 20 Hz. This inhibition is suppressed by the SST receptor antagonist cyclo-SST and is absent when the O-LM interneuron is located more than 150 {micro}m from the pyramidal cell. Likewise, a single O-LM cell was also able to inhibit the intrinsic excitability of another O-LM cell, provided the two cells are in proximity. Exogeneous application of SST transiently inhibits excitatory synaptic transmission and intrinsic excitability in O-LM interneurons through SST1 and SST2, respectively. Notably, this transient inhibition became sustained when clathrin-dependent endocytosis of SST receptors was blocked. Blocking of SST receptors reduced NMDA receptor-mediated responses, prevented induction of both synaptic and intrinsic potentiation in pyramidal neurons and reduced the coherence of theta oscillations. These findings reveal that SST released by O-LM interneurons constitutes an additional inhibitory mechanism, modulating both synaptic excitation and intrinsic excitability beyond the transient inhibition mediated by GABA release.