Valproate decreases transgenerationally blood pressure by affecting thyrotropin-releasing hormone promoter DNA methylation and gene expression in spontaneously hypertensive rat.

Central TRH, a neuropeptide, is involved in cardiovascular regulation. We demonstrated that the overexpression of diencephalic TRH (dTRH) in SHR rats can be prevented by antisense treatment, normalizing blood pressure (BP). Valproate (VPA) is an inhibitor of histone deacetylases (HDAC) which modulates gene expression through epigenetic modifications such as DNA methylation. AIMS: Study the role of HDAC inhibition in the regulation of dTRH gene expression and its effect on the pathogenesis of hypertension. MAIN METHODS: We treated 7-weeks-old male and female SHR and WKY rats with VPA for 10 weeks and evaluated BP, dTRH mRNA and methylation gene status. KEY FINDINGS: VPA attenuated the elevated BP and dTRH mRNA expression characteristic of SHR. Indeed, we found a significant 62% reduction in dTRH mRNA expression in the SHR + VPA group compared to control SHR. The decrease TRH mRNA expression induced by VPA was confirmed "in vitro" in a primary neuron culture using trichostatin A. With methylation specific PCR we demonstrated a significant increase in TRH promoter DNA methylation level in SHR + VPA group compared to control SHR. After 2 weeks of treatment interruption, rats were mated. Although they did not receive any treatment, the offspring born from VPA-treated SHR parents showed similar changes in BP, dTRH expression and methylation status, implying a transgenerational inheritance. Our findings suggest that dTRH modulation by epigenetics mechanism affects BP and could be inherited by the next generation in SHR rats.

Turn-on mode diarylethenes for bioconjugation and fluorescence microscopy of cellular structures.

The use of photoswitchable fluorescent diarylethenes (fDAEs) as protein labels in fluorescence microscopy and nanoscopy has been limited by labeling inhomogeneity and the need for ultraviolet light for fluorescence activation (on-switching). To overcome these drawbacks, we prepared "turn-on mode" fDAEs featuring thienyl substituents, multiple polar residues, and a reactive maleimide group in the core structure. Conjugates with antibodies and nanobodies displayed complete on-switching and excitation with violet (405 nm) and yellow-green (<565 nm) light, respectively. Besides, they afforded high signal-to-noise ratios and low unspecific labeling in fluorescence imaging. Irradiation with visible light at 532 nm or 561 nm led to transient on-off switching ("blinking") of the fDAEs of double-labeled nanobodies so that nanoscale superresolution images were readily attained through switching and localization of individual fluorophores.

Virtual Experience Toolkit: An End-to-End Automated 3D Scene Virtualization Framework Implementing Computer Vision Techniques.

Virtualization plays a critical role in enriching the user experience in Virtual Reality (VR) by offering heightened realism, increased immersion, safer navigation, and newly achievable levels of interaction and personalization, specifically in indoor environments. Traditionally, the creation of virtual content has fallen under one of two broad categories: manual methods crafted by graphic designers, which are labor-intensive and sometimes lack precision; traditional Computer Vision (CV) and Deep Learning (DL) frameworks that frequently result in semi-automatic and complex solutions, lacking a unified framework for both 3D reconstruction and scene understanding, often missing a fully interactive representation of the objects and neglecting their appearance. To address these diverse challenges and limitations, we introduce the Virtual Experience Toolkit (VET), an automated and user-friendly framework that utilizes DL and advanced CV techniques to efficiently and accurately virtualize real-world indoor scenarios. The key features of VET are the use of ScanNotate, a retrieval and alignment tool that enhances the precision and efficiency of its precursor, supported by upgrades such as a preprocessing step to make it fully automatic and a preselection of a reduced list of CAD to speed up the process, and the implementation in a user-friendly and fully automatic Unity3D application that guides the users through the whole pipeline and concludes in a fully interactive and customizable 3D scene. The efficacy of VET is demonstrated using a diversified dataset of virtualized 3D indoor scenarios, supplementing the ScanNet dataset.

ß-Galactosidase- and Photo-Activatable Fluorescent Probes for Protein Labeling and Super-Resolution STED Microscopy in Living Cells.

We report on the synthesis of two fluorescent probes which can be activated by ß-Galactosidase (ß-Gal) enzymes and/or light. The probes contained 2-nitro-4-oxybenzyl and 3-nitro-4-oxybenzyl fragments, with ß-Gal residues linked to C-4. We performed the enzymatic and photoactivation of the probes in a cuvette and compared them, prior to the labeling of Vimentin-Halo fusion protein in live cells with overexpressed ß-galactosidase. The dye fluorescence afforded the observation of enzyme activity by means of confocal and super-resolution optical microscopy based on stimulated emission depletion (STED). The tracing of enzymatic activity with the retention of activated fluorescent products inside cells was combined with super-resolution imaging as a tool for use in biomedicine and life science.

Supramolecular Complex of Cucurbit[7]uril with Diketopyrrolopyrole Dye: Fluorescence Boost, Biolabeling and Optical Microscopy.

New photostable and bright supramolecular complexes based on cucurbit[7]uril (CB7) host and diketopyrrolopyrole (DPP) guest dyes having two positively charged 4-(trimethylammonio)phenyl groups were prepared and characterized. The dye core displays large Stokes shift (in H2O, abs./emission max. 480/550 nm; ϵ~19 000, τfl>4 ns), strong binding with the host (~560 nM Kd) and a linker affording fluorescence detection of bioconjugates with antibody and nanobody. Combination of protein-functionalized DPP dye with CB7 improves photostability and affords up to 12-fold emission gain. Two-color confocal and stimulated emission depletion (STED) microscopy with 595 nm or 655 nm STED depletion lasers shows that the presence of CB7 not only leads to improved brightness and image quality, but also results in DPP becoming cell-permeable.

Photoactivatable Large Stokes Shift Fluorophores for Multicolor Nanoscopy.

We designed caging-group-free photoactivatable live-cell permeant dyes with red fluorescence emission and ∼100 nm Stokes shifts based on a 1-vinyl-10-silaxanthone imine core structure. The proposed fluorophores undergo byproduct-free one- and two-photon activation, are suitable for multicolor fluorescence microscopy in fixed and living cells, and are compatible with super-resolution techniques such as STED (stimulated emission depletion) and PALM (photoactivated localization microscopy). Use of photoactivatable labels for strain-promoted tetrazine ligation and self-labeling protein tags (HaloTag, SNAP-tag), and duplexing of an imaging channel with another large Stokes shift dye have been demonstrated.

Accelerated MINFLUX Nanoscopy, through Spontaneously Fast-Blinking Fluorophores.

The introduction of MINFLUX nanoscopy allows single molecules to be localized with one nanometer precision in as little as one millisecond. However, current applications have so far focused on increasing this precision by optimizing photon collection, rather than minimizing the localization time. Concurrently, commonly used fluorescent switches are specifically designed for stochastic methods (e.g., STORM), optimized for a high photon yield and rather long on-times (tens of milliseconds). Here, accelerated MINFLUX nanoscopy with up to a 30-fold gain in localization speed is presented. The improvement is attained by designing spontaneously blinking fluorescent markers with remarkably fast on-times, down to 1-3 ms, matching the iterative localization process used in a MINFLUX microscope. This design utilizes a silicon rhodamine amide core, shifting the spirocyclization equilibrium toward an uncharged closed form at physiological conditions and imparting intact live cell permeability, modified with a fused (benzo)thiophene spirolactam fragment. The best candidate for MINFLUX microscopy (also suitable for STORM imaging) is selected through detailed characterization of the blinking behavior of single fluorophores, bound to different protein tags. Finally, optimization of the localization routines, customized to the fast blinking times, renders a significant speed improvement on a commercial MINFLUX microscope.

Photoactivatable Carbo- and Silicon-Rhodamines and Their Application in MINFLUX Nanoscopy.

New photoactivatable fluorescent dyes (rhodamine, carbo- and silicon-rhodamines [SiR]) with emission ranging from green to far red have been prepared, and their photophysical properties studied. The photocleavable 2-nitrobenzyloxycarbonyl unit with an alpha-carboxyl group as a branching point and additional functionality was attached to a polycyclic and lipophilic fluorescent dye. The photoactivatable probes having the HaloTagTM amine (O2) ligand bound with a dye core were obtained and applied for live-cell staining in stable cell lines incorporating Vimentin (VIM) or Nuclear Pore Complex Protein NUP96 fused with the HaloTag. The probes were applied in 2D (VIM, NUP96) and 3D (VIM) MINFLUX nanoscopy, as well as in superresolution fluorescence microscopy with single fluorophore activation (VIM, live-cell labeling). Images of VIM and NUPs labeled with different dyes were acquired and their apparent dimensions and shapes assessed on a lower single-digit nanometer scale. Applicability and performance of the photoactivatable dye derivatives were evaluated in terms of photoactivation rate, labeling and detection efficiency, number of detected photons per molecule and other parameters related to MINFLUX nanoscopy.

Supramolecular Complex of Photochromic Diarylethene and Cucurbit[7]uril: Fluorescent Photoswitching System for Biolabeling and Imaging.

Photoswitchable fluorophores─proteins and synthetic dyes─whose emission is reversibly switched on and off upon illumination, are powerful probes for bioimaging, protein tracking, and super-resolution microscopy. Compared to proteins, synthetic dyes are smaller and brighter, but their photostability and the number of achievable switching cycles in aqueous solutions are lower. Inspired by the robust photoswitching system of natural proteins, we designed a supramolecular system based on a fluorescent diarylethene (DAE) and cucurbit[7]uril (CB7) (denoted as DAE@CB7). In this assembly, the photoswitchable DAE molecule is encapsulated by CB7 according to the host-guest principle, so that DAE is protected from the environment and its fluorescence brightness and fatigue resistance in pure water improved. The fluorescence quantum yield (Φfl) increased from 0.40 to 0.63 upon CB7 complexation. The photoswitching of the DAE@CB7 complex, upon alternating UV and visible light irradiations, can be repeated 2560 times in aqueous solution before half-bleaching occurs (comparable to fatigue resistance of the reversibly photoswitchable proteins), while free DAE can be switched on and off only 80 times. By incorporation of reactive groups [maleimide and N-hydroxysuccinimidyl (NHS) ester], we prepared bioconjugates of DAE@CB7 with antibodies and demonstrated both specific labeling of intracellular proteins in cells and the reversible on/off switching of the probes in cellular environments under irradiations with 355 nm/485 nm light. The bright emission and robust photoswitching of DAE-Male3@CB7 and DAE-NHS@CB7 complexes (without exclusion of air oxygen and addition of any stabilizing/antifading reagents) enabled confocal and super-resolution RESOLFT (reversible saturable optical fluorescence transitions) imaging with apparent 70-90 nm optical resolution.

Cleavable Linker Incorporation into a Synthetic Dye-Nanobody-Fluorescent Protein Assembly: FRET, FLIM and STED Microscopy.

A bright and photostable fluorescent dye with a disulfide (S-S) linker and maleimide group (Rho594-S2-mal), as cleavable and reactive sites, was synthesized and conjugated with anti-GFP nanobodies (NB). The binding of EGFP (FRET donor) with anti-GFP NB labeled with one or two Rho594-S2-mal residues was studied in vitro and in cellulo. The linker was cleaved with dithiothreitol recovering the donor (FP) signal. The bioconjugates (FP-NB-dye) were applied in FRET-FLIM assays, confocal imaging, and superresolution STED microscopy.

Bis-Rhodamines Bridged with a Diazoketone Linker: Synthesis, Structure, and Photolysis.

Two fluorophores bound with a short photoreactive bridge are fascinating structures and remained unexplored. To investigate the synthesis and photolysis of such dyes, we linked two rhodamine dyes via a diazoketone bridge (-COCN2-) attached to position 5' or 6' of the pendant phenyl rings. For that, the mixture of 5'- or 6'-bromo derivatives of the parent dye was prepared, transformed into 1,2-diarylacetylenes, hydrated to 1,2-diarylethanones, and converted to diazoketones Ar1COCN2Ar2. The high performance liquid chromatography (HPLC) separation gave four individual regioisomers of Ar1COCN2Ar2. Photolysis of the model compound─C6H5COCN2C6H5─in aqueous acetonitrile at pH 7.3 and under irradiation with 365 nm light provided diphenylacetic acid amide (Wolff rearrangement). However, under the same conditions, Ar1COCN2Ar2 gave mainly α-diketones Ar1COCOAr2. The migration ability of the very bulky dye residues was low, and the Wolff rearrangement did not occur. We observed only moderate fluorescence increase, which may be explained by the insufficient quenching ability of diazoketone bridge (-COCN2-) and its transformation into another (weaker) quencher, 1,2-diarylethane-1,2-dione.

Cardiac Thyrotropin-releasing Hormone Inhibition Improves Ventricular Function and Reduces Hypertrophy and Fibrosis After Myocardial Infarction in Rats.

BACKGROUND: Cardiac thyrotropin-releasing hormone (TRH) is a tripeptide with still unknown functions. We demonstrated that the left ventricle (LV) TRH system is hyperactivated in spontaneously hypertensive rats and its inhibition prevented cardiac hypertrophy and fibrosis. Therefore, we evaluated whether in vivo cardiac TRH inhibition could improve myocardial function and attenuate ventricular remodeling in a rat model of myocardial infarction (MI). METHODS AND RESULTS: In Wistar rats, MI was induced by a permanent left anterior descending coronary artery ligation. A coronary injection of a specific small interfering RNA against TRH was applied simultaneously. The control group received a scrambled small interfering RNA. Cardiac remodeling variables were evaluated one week later. In MI rats, TRH inhibition decreased LV end-diastolic (1.049 ± 0.102 mL vs 1.339 ± 0.102 mL, P < .05), and end-systolic volumes (0.282 ± 0.043 mL vs 0.515 ± 0.037 mL, P < .001), and increased LV ejection fraction (71.89 ± 2.80% vs 65.69 ± 2.85%, P < .05). Although both MI groups presented similar infarct size, small interfering RNA against TRH treatment attenuated the cardiac hypertrophy index and myocardial interstitial collagen deposition in the peri-infarct myocardium. These effects were accompanied by attenuation in the rise of transforming growth factor-ß, collagen I, and collagen III, as well as the fetal genes (atrial natriuretic peptide, B-type natriuretic peptide, and beta myosin heavy chain) expression in the peri-infarct region. In addition, the expression of Hif1α and vascular endothelial growth factor significantly increased compared with all groups. CONCLUSIONS: Cardiac TRH inhibition improves LV systolic function and attenuates ventricular remodeling after MI. These novel findings support the idea that TRH inhibition may serve as a new therapeutic strategy against the progression of heart failure.

Triarylmethane Fluorophores Resistant to Oxidative Photobluing.

Spectral stability of small-molecule fluorescent probes is required for correct interpretation and reproducibility of multicolor fluorescence imaging data, in particular under high (de)excitation light intensities of super-resolution imaging or in single-molecule applications. We propose a synthetic approach to a series of spectrally stable rhodamine fluorophores based on sequential Ru- and Cu-catalyzed transformations, evaluate their stability against photobleaching and photoconversion in the context of other fluorophores using chemometric analysis, and demonstrate chemical reactivity of fluorophore photoproducts. The substitution patterns providing the photoconversion-resistant triarylmethane fluorophores have been identified, and the applicability of nonbluing labels in live-cell STED nanoscopy is demonstrated.

Reversibly Photoswitchable Fluorescent Diarylethenes Resistant against Photobleaching in Aqueous Solutions.

Low photostability in aqueous solutions is the main drawback of synthetic photochromic dyes, which limits their switching performance and utility in biology, medicine, and life sciences. Even the most promising photochromes-reversibly photoswitchable diarylethenes (DAEs) with fluorescent "closed" forms-are known to undergo only several tens (typically 20-30) of switching cycles in aqueous solutions. In this work, we introduce water-soluble and highly photostable 1,2-[bis(2-ethyl/2-isobutyl-1-benzothiophene-1,1-dioxide-6-phenyl-3-yl)]perfluorocyclopentenes decorated with four -CONHC(CH2R)3 residues capped with 12 carboxylic acid groups (R = CH2CO2H, O(CH2)2CO2H). Under irradiation with UV (365 nm) and visible light (470 nm), they provide several hundred (for the "rapid" DAEs with isobutyl groups, up to 1000) full switching cycles in aqueous solutions without exclusion of air oxygen (outperforming the photoswitchable and fluorescent protein Dreiklang). The new branching approach based on secondary carboxamides with N-tert-alkyl residues decorated with polar groups may serve as a blueprint for the design of highly fatigue resistant and reversibly photoswitchable fluorescent probes applicable in life sciences as aqueous solutions.

[Use of virtual reality for neurodevelopmental disorders. A review of the state of the art and future agenda]. / Uso de entornos virtuales para trastornos del neurodesarrollo: una revisión del estado del arte y agenda futura.

To date, the diagnostic tools for autism spectrum disorder (ASD) have been mostly based on qualitative criteria from observational information in contexts with low ecological validity. We are witnessing a growing scientific activity that proposes the use of implicit measures for the evaluation and diagnosis of ASD. These measures are based on processes of a biological and unconscious nature, underlying the capacity of human cognition, and are obtained through the acquisition and treatment of brain, physiological and behavioral responses in order to obtain the behavioral structure of the ASD patient facing a stimulus. The complex relationship between physiological responses and the behavioral structure of the ASD patient requires the use of advanced techniques of signal processing based on cognitive computation. Artificial intelligence (AI) techniques, such as machine learning and neurocomputing applied to the analysis of psychophysiological signals, have demonstrated their robustness for the classification of complex cognitive constructs. Virtual reality (VR) is a tool that allows recreating real-life situations with high sensory fidelity, but at the same time individually controlling each of the situations and stimuli that influence human behavior. It also allows the measurement in real time of human reactions to such stimuli. This document analyzes the latest scientific and technological advances relevant to its applications in the diagnosis of ASD. We conclude that VR is a very valuable tool for ASD research, especially for the evaluation and diagnosis of complex skills and competencies.

Hasta ahora las herramientas diagnósticas de los trastornos del espectro autista (TEA) se basan mayoritariamente en criterios cualitativos de información observacional en contextos con baja validez ecológica. Una creciente actividad científica propone medidas implícitas para la evaluación y diagnóstico del TEA. Dichas medidas se basan en procesos de carácter biológico e inconsciente, subyacentes a la capacidad de cognición humana y son obtenidas a través de la adquisición y tratamiento de respuestas cerebrales, fisiológicas y comportamentales, con el objetivo de obtener la estructura comportamental del paciente TEA ante un estímulo. La compleja relación existente entre respuestas fisiológicas y la estructura comportamental del paciente TEA ante un estímulo, obliga a utilizar técnicas avanzadas de tratamiento de la señal basadas en computación cognitiva. Las técnicas de inteligencia artificial, tales como aprendizaje automático (machine learning) y neurocomputación aplicadas al análisis de señales psicofisiológicas, han demostra do su robustez para la clasificación de complejos constructos cognitivos. La realidad virtual (RV) es una herramienta que permite recrear situaciones de la vida real con una alta fidelidad sensorial, pero al mismo tiempo controlar individualmente cada una de las situaciones y estímulos que influyen en el comportamiento humano. También permite la medición en tiempo real de las reacciones humanas ante tales estímulos. Este documento analiza los últimos avances científicos y tecnológicos relevantes para sus aplicaciones en el diagnóstico del TEA. Afirmamos que la RV es una herramienta muy valiosa para la investigación del TEA, especialmente para la evaluación y diagnóstico de habilidades y competencias complejas.

Mono- and bithiophene-substituted diarylethene photoswitches with emissive open or closed forms.

We present a new series of photochromic 1,2-bis(2-ethylbenzo[b]thiophen-3-yl)perfluorocyclopentenes with an oxidized benzothiophene core (O) or a nonoxidized one, decorated with mono- (Th1) and bithiophene (Th2) units attached to positions 6 and 6' (Sy = symmetric) or only to position 6 (As = asymmetric). "Oxidized" compounds have highly fluorescent closed forms emitting in the visible region (yellow to red). The dyes with nonoxidized benzothiophenes possess fluorescent open forms with rather low emission efficiency. The photoswitching kinetics was studied at several wavelengths with UV and visible light. New diarylethenes underwent ring-closure reactions by irradiation with UV light (365 nm, 405 nm), and the reversible ring-opening by irradiation with visible light (470 nm, 530 nm). The on-switching of fluorescence due to the ring-closure reaction was observed also with visible light of 470 nm (to an extent of 10% for compound SyOTh 1 ) and attributed to the Urbach tail effect. Due to a high degree of fluorescence modulation (>270), good fatigue resistance and large fluorescence quantum yield, compound SyOTh 1 emerged as a candidate for single-molecule based super-resolution fluorescence microscopy.

Photoactivatable Rhodamine Spiroamides and Diazoketones Decorated with "Universal Hydrophilizer" or Hydroxyl Groups.

Photoactivatable rhodamine spiroamides and spirocyclic diazoketones emerged recently as synthetic markers applicable in multicolor super-resolution microscopy. However, their applicability in single molecule localization microscopy (SMLM) is often limited by aggregation, unspecific adhesion, and low reactivity caused by insufficient solubility and precipitation from aqueous solutions. We report here two synthetic modifications increasing the polarity of compact polycyclic and hydrophobic labels decorated with a reactive group: attachment of 3-sulfo-l-alanyl-beta-alanine dipeptide (a "universal hydrophilizer") or allylic hydroxylation in photosensitive rhodamine diazoketones (and spiroamides). The super-resolution images of tubulin and keratin filaments in fixed and living cells exemplify the performance of "blinking" spiroamides derived from N, N, N', N'-tetramethyl rhodamine.

Fluorescent Photoswitchable Diarylethenes for Biolabeling and Single-Molecule Localization Microscopies with Optical Superresolution.

A modular assembly of water-soluble diarylethenes (DAEs), applicable as biomarkers for optical nanoscopy, is reported. Reversibly photoswitchable 1,2-bis(2-alkyl-6-phenyl-1-benzothiophene-1,1-dioxide-3-yl)perfluorocyclopentenes possessing a fluorescent "closed" form were decorated with one or two methoxy group(s) attached to the para-position(s) of phenyl ring(s) and two, four, or eight carboxylic acid groups. Antibody conjugates of these DAEs feature low aggregation, efficient photoswitching in aqueous buffers, specific staining of cellular structures, and photophysical properties (high emission efficiencies and low cycloreversion quantum yields) enabling their application in superresolution microscopy. Images of tubulin, vimentin, and nuclear pore complexes are presented. The superresolution images can also be acquired by using solely 488 nm light without additional photoactivation with UV light. These DAEs exhibit reversible photoswitching without requiring any additives to the imaging media and open new paths toward the modular design of fluorescent dyes for bioimaging with optical superresolution.

Bichromophoric Compounds with Orthogonally and Parallelly Arranged Chromophores Separated by Rigid Spacers.

Electronic energy transfer (EET) between chromophores is of fundamental importance for many biological processes and optoelectronic devices. However, common models fall short in fully describing the process, especially in bichromophoric model systems with a donor and acceptor connected by a rigid linker providing perpendicular geometries. Herein, we report a novel strategy for preparing bichromophores containing adamantane or 2-(2-adamantylidene)adamantane as rigid spacers, providing a fixed distance between chromophores, and their parallel or perpendicular arrangement without chromophore rotation. New fluorophores were developed and linked via spiroatoms. Bichromophores with identical (blue-blue) or different (blue-red) chromophores were synthesized, either in orthogonal or parallel geometry. These were characterized by absorption/fluorescence spectroscopy, time-resolved fluorescence anisotropy, and fluorescence antibunching measurements. Based on the Förster point-dipole approximation, EET efficiencies were estimated by using geometrical parameters from (time-dependent) density functional calculations. For bichromophores with parallel geometry, the predicted EET efficiencies were near unity and fit the measurements. In spite of estimated values around 0.4 and 0.5, 100 % efficiency was observed also for bichromophores with orthogonal geometry. The new rigid scaffolds presented here open new possibilities for the synthesis of bichormophores with well-defined parallel or perpendicular geometry.

First-Ever Stroke and Transient Ischemic Attack Incidence and 30-Day Case-Fatality Rates in a Population-Based Study in Argentina.

BACKGROUND AND PURPOSE: Epidemiological data about stroke are scarce in low- and middle-income Latin-American countries. We investigated annual incidence of first-ever stroke and transient ischemic attack (TIA) and 30-day case-fatality rates in a population-based setting in Tandil, Argentina. METHODS: We prospectively identified all first-ever stroke and TIA cases from overlapping sources between January 5, 2013, and April 30, 2015, in Tandil, Argentina. We calculated crude and standardized incidence rates. We estimated 30-day case-fatality rates. RESULTS: We identified 334 first-ever strokes and 108 TIAs. Age-standardized incidence rate per 100 000 for Segi's World population was 76.5 (95% confidence interval [CI], 67.8-85.9) for first-ever stroke and 25.1 (95% CI, 20.2-30.7) for first-ever TIA, 56.1 (95% CI, 48.8-64.2) for ischemic stroke, 13.5 (95% CI, 9.9-17.9) for intracerebral hemorrhage, and 4.9 (95% CI, 2.7-8.1) for subarachnoid hemorrhage. Stroke incidence was slightly higher for men (87.8; 95% CI, 74.6-102.6) than for women (73.2; 95% CI, 61.7-86.1) when standardized for the Argentinean population. Thirty-day case-fatality rate was 14.7% (95% CI, 10.8-19.5) for ischemic stroke, 24.1% (95% CI, 14.2-36.6) for intracerebral hemorrhage, and 1.9% (95% CI, 0.4-5.8) for TIA. CONCLUSIONS: This study provides the first prospective population-based stroke and TIA incidence and case-fatality estimate in Argentina. First-ever stroke incidence was lower than that reported in previous Latin-American studies, but first-ever TIA incidence was higher. Thirty-day case-fatality rates were similar to those of other population-based Latin-American studies.