Purinergic signaling in cognitive impairment and neuropsychiatric symptoms of Alzheimer's disease.

About 10 million new cases of dementia develop worldwide each year, of which up to 70% are attributable to Alzheimer's disease (AD). In addition to the widely known symptoms of memory loss and cognitive impairment, AD patients frequently develop non-cognitive symptoms, referred to as behavioral and psychological symptoms of dementia (BPSDs). Sleep disorders are often associated with AD, but mood alterations, notably depression and apathy, comprise the most frequent class of BPSDs. BPSDs negatively affect the lives of AD patients and their caregivers, and have a significant impact on public health systems and the economy. Because treatments currently available for AD are not disease-modifying and mainly aim to ameliorate some of the cognitive symptoms, elucidating the mechanisms underlying mood alterations and other BPSDs in AD may reveal novel avenues for progress in AD therapy. Purinergic signaling is implicated in the pathophysiology of several central nervous system (CNS) disorders, such as AD, depression and sleep disorders. Here, we review recent findings indicating that purinergic receptors, mainly the A1, A2A, and P2X7 subtypes, are associated with the development/progression of AD. Current evidence suggests that targeting purinergic signaling may represent a promising therapeutic approach in AD and related conditions. This article is part of the Special Issue on "Purinergic Signaling: 50 years".

Retinal organoids from human-induced pluripotent stem cells: From studying retinal dystrophies to early diagnosis of Alzheimer's and Parkinson's disease.

Human-induced pluripotent stem cells (hiPSCs) have provided new methods to study neurodegenerative diseases. In addition to their wide application in neuronal disorders, hiPSCs technology can also encompass specific conditions, such as inherited retinal dystrophies. The possibility of evaluating alterations related to retinal disorders in 3D organoids increases the truthfulness of in vitro models. Moreover, both Alzheimer's (AD) and Parkinson's disease (PD) have been described as causing early retinal alterations, generating beta-amyloid protein accumulation, or affecting dopaminergic amacrine cells. This review addresses recent advances and future perspectives obtained from in vitro modeling of retinal diseases, focusing on retinitis pigmentosa (RP). Additionally, we depicted the possibility of evaluating changes related to AD and PD in retinal organoids obtained from potential patients long before the onset of the disease, constituting a valuable tool in early diagnosis. With this, we pointed out prospects in the study of retinal dystrophies and early diagnosis of AD and PD.

Distinct Cell-specific Roles of NOX2 and MyD88 in Epileptogenesis.

It is well established that temporal lobe epilepsy (TLE) is often related to oxidative stress and neuroinflammation. Both processes subserve alterations observed in epileptogenesis and ultimately involve distinct classes of cells, including astrocytes, microglia, and specific neural subtypes. For this reason, molecules associated with oxidative stress response and neuroinflammation have been proposed as potential targets for therapeutic strategies. However, these molecules can participate in distinct intracellular pathways depending on the cell type. To illustrate this, we reviewed the potential role of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 2 (NOX2) and myeloid differentiation primary response 88 (MyD88) in astrocytes, microglia, and neurons in epileptogenesis. Furthermore, we presented approaches to study genes in different cells, employing single-cell RNA-sequencing (scRNAseq) transcriptomic analyses, transgenic technologies and viral serotypes carrying vectors with specific promoters. We discussed the importance of identifying particular roles of molecules depending on the cell type, endowing more effective therapeutic strategies to treat TLE.

Glioma Pericytes Promote Angiogenesis by Producing Periostin.

Glioma is the prevalent aggressive primary brain tumor, with a very poor prognosis. The absence of advanced understanding of the roles played by the cells within the glioma microenvironment limits the development of effective drugs. A recent study indicates that periostin expressed by pericytes is crucial for glioma angiogenesis. Here, we describe succinctly the results and implications of this discovery in what we know about pericytes within the glioma microenvironment. The emerging knowledge from this work will benefit the development of therapies for gliomas.

Chronic Lymphocytic Leukemia (CLL): evaluation of AKT protein kinase and microRNA gene expression related to disease pathogenesis

Abstract The present study evaluated 56 patients diagnosed with Chronic Lymphocytic Leukemia (CLL) and a control group of 44 clinically healthy subjects with no previous history of leukemia. Genetic expressions of AKT and microRNAs were evaluated by quantitative PCR (qPCR). A significant increase in AKT gene expression in patients when compared to controls was observed (p = 0.017). When the patients were stratified according to Binet subgroups, a significant difference was observed between the subgroups, with this protein kinase appearing more expressed in the B+C subgroup (p = 0.013). Regarding miRNA expression, miR-let-7b and miR-26a were reduced in CLL patients, when compared to controls. However, no significant differences were observed in these microRNA expressions between the Binet subgroups (A versus B+C). By contrast, miR-21 to miR-27a oncogenes showed no expression difference between CLL patients and controls. AKT protein kinase is involved in the signaling cascade that occurs with BCR receptor activation, leading to increased lymphocyte survival and protection against the induction of cell death in CLL. Thus, increased AKT protein kinase expression and the reduction of miR-let-7b and miR-26a, both tumor suppressors, may explain increased lymphocyte survival in CLL patients and may be promising markers for the prognostic evaluation of this disease.

Flow cytometry in the analysis of hematological parameters of tilapias: applications in environmental aquatic toxicology.

Blood tissue has been used to assess animal health and the environment in which they live. This tissue is easily acquired and has the ability to respond to various adverse conditions. Several techniques have been employed in the detection of xenobiotic-induced cell damage in blood cells. In general, traditionally used technologies, such as cellular analysis in blood smears, are time-consuming and require great analytical capacity. The present study proposes flow cytometry as a method to detect changes in blood cell populations. Tilapia (Oreochromis niloticus) was selected as a model for plotting the profile of fish blood cell populations after exposure to xenobiotics without euthanizing animals or using cell markers. Populations of erythrocytes and lymphocytes were detected only by combining the techniques of FACSAria cell sorting and light microscopy. Systemic deleterious effects were found through blood analysis, such as an increased lymphocyte-rich population at 48 h of exposure followed by a subsequent decrease. Moreover, the time-dependent expression of Nrf2 suggests its participation in increased membrane disruption, indicating it has a central role in erythrocyte lifespan. The present results shed light on the viability of using flow cytometry for blood analysis of living fish.

Neurobiology of glycine transporters: From molecules to behavior.

Glycine transporters (GlyTs) are Na+/Cl--dependent neurotransmitter transporters, responsible for l-glycine uptake into the central nervous system. GlyTs are members of the solute carrier family 6 (SLC6) and comprise glycine transporter type 1 (SLC6A9; GlyT1) and glycine transporter type 2 (SLC6A5; Glyt2). GlyT1 and GlyT2 are expressed on both astrocytes and neurons, but their expression pattern in brain tissue is foremost related to neurotransmission. GlyT2 is markedly expressed in brainstem, spinal cord and cerebellum, where it is responsible for glycine uptake into glycinergic and GABAergic terminals. GlyT1 is abundant in neocortex, thalamus and hippocampus, where it is expressed in astrocytes, and involved in glutamatergic neurotransmission. Consequently, inhibition of GlyT1 transporters can modulate glutamatergic neurotransmission through NMDA receptors, suggesting an alternative therapeutic strategy. In this review, we focus on recent progress in the understanding of GlyTs role in brain function and in various diseases, such as epilepsy, hyperekplexia, neuropathic pain, drug addiction, schizophrenia and stroke, as well as in neurodegenerative disorders.

Cardiomyocyte Proteome Remodeling due to Isoproterenol-Induced Cardiac Hypertrophy during the Compensated Phase.

PURPOSE: Although the pathophysiological response of cardiac tissue to pro-hypertrophic stimulus is well characterized, a comprehensive characterization of the molecular events underlying the pathological hypertrophy in cardiomyocytes during the early compensated cardiac hypertrophy is currently lacking. EXPERIMENTAL DESIGN: A quantitative label-free proteomic analysis of cardiomyocytes isolated was conducted from mice treated subcutaneously with isoproterenol (ISO) during 7 days in comparison with cardiomyocytes from control animals (CT). RESULTS: Canonical pathway analysis of dysregulated proteins indicated that ISO-hypertrophy drives the activation of actin cytoskeleton and integrin-linked kinase (ILK) signaling, and inhibition of the sirtuin signaling. Alteration in cardiac contractile function and calcium signaling are predicted as downstream effects of ISO-hypertrophy probably due to the upregulation of key elements such as myosin-7 (MYH7). Confocal microscopy corroborated that indeed ISO-treatment led to increased abundance of MYH7. Potential early markers for cardiac hypertrophy as APBB1, GOLGA4, HOOK1, KATNA1, KIFBP, MAN2B2, and SLC16A1 are also reported. CONCLUSIONS AND CLINICAL RELEVANCE: The data consist in a complete molecular mapping of ISO-induced compensated cardiac hypertrophy model at cardiomyocyte level. Marker candidates reported may assist early diagnosis of cardiac hypertrophy and ultimately heart failure.

Metabolic Disturbances Identified in Plasma Samples from ST-Segment Elevation Myocardial Infarction Patients.

ST-segment elevation myocardial infarction (STEMI) is the most severe form of myocardial infarction (MI) and the main contributor to morbidity and mortality caused by MI worldwide. Frequently, STEMI is caused by complete and persistent occlusion of a coronary artery by a blood clot, which promotes heart damage. STEMI impairment triggers changes in gene transcription, protein expression, and metabolite concentrations, which grants a biosignature to the heart dysfunction. There is a major interest in identifying novel biomarkers that could improve the diagnosis of STEMI. In this study, the phenotypic characterization of STEMI patients (n = 15) and healthy individuals (n = 19) was performed, using a target metabolomics approach. Plasma samples were analyzed by UPLC-MS/MS (ultra-high-performance liquid chromatography-tandem mass spectrometry) and FIA-MS (MS-based flow injection analysis). The goal was to identify novel plasma biomarkers and metabolic signatures underlying STEMI. Concentrations of phosphatidylcholines, lysophosphatidylcholines, sphingomyelins, and biogenic amines were altered in STEMI patients in relation to healthy subjects. Also, after multivariate analysis, it was possible to identify alterations in the glycerophospholipids, alpha-linolenic acid, and sphingolipid metabolisms in STEMI patients.

Enhancing the Therapeutic Potential of Mesenchymal Stem Cells with the CRISPR-Cas System.

Mesenchymal stem cells (MSCs), also known as multipotent mesenchymal stromal stem cells, are found in the perivascular space of several tissues. These cells have been subject of intense research in the last decade due to their low teratogenicity, as well as their ability to differentiate into mature cells and to secrete immunomodulatory and trophic factors. However, they usually promote only a modest benefit when transplanted in experimental disease models, one of the limitations for their clinical application. The CRISPR-Cas system, in turn, is highlighted as a simple and effective tool for genetic engineering. This system was tested in clinical trials over a relatively short period of time after establishing its applicability to the edition of the mammalian cell genome. Similar to the research evolution in MSCs, the CRISPR-Cas system demonstrated inconsistencies that limited its clinical application. In this review, we outline the evolution of MSC research and its applicability, and the progress of the CRISPR-Cas system from its discovery to the most recent clinical trials. We also propose perspectives on how the CRISPR-Cas system may improve the therapeutic potential of MSCs, making it more beneficial and long lasting.

A carefully designed nanoplatform based on multi walled carbon nanotube wrapped with aptamers.

The interaction between carbon nanotubes (CNTs) and biological molecules of diagnostic and therapeutic interest, as well as the internalization of the CNTs-biomolecules complexes in different types of cell, has been extensively studied due to the potential use of these nanocomplexes as multifunctional nanoplatforms in a great variety of biomedical applications. The effective use of these nanobiotechnologies requires broad multidisciplinary studies of biocompatibility, regarding, for example, the in vitro and in vivo nanotoxicological assays, the capacity to target specific cells and the evaluation of their biomedical potential. However, the first step to be reached is the careful obtainment of the nanoplatform and the understanding of the actual surface composition and structural integrity of the complex system. In this work, we show the detailed construction of a nanoplatform created by the noncovalent interaction between oxidized multi walled carbon nanotubes (MWCNTs) and a DNA aptamer targeting tumor cells. The excess free aptamer was removed by successive washes, revealing the actual surface of the nanocomplex. The MWCNT-aptamer interaction by π-stacking was evidenced and shown to contribute in obtaining a stable nanocomplex compatible with aqueous media having good cell viability. The nucleotide sequence of the aptamer remained intact after the functionalization, allowing its use in further studies of specificity and binding affinity and for the construction of functional nanoplatforms.

Decoding resistant hypertension signalling pathways.

Resistant hypertension (RH) is a clinical condition in which the hypertensive patient has become resistant to drug therapy and is often associated with increased cardiovascular morbidity and mortality. Several signalling pathways have been studied and related to the development and progression of RH: modulation of sympathetic activity by leptin and aldosterone, primary aldosteronism, arterial stiffness, endothelial dysfunction and variations in the renin-angiotensin-aldosterone system (RAAS). miRNAs comprise a family of small non-coding RNAs that participate in the regulation of gene expression at post-transcriptional level. miRNAs are involved in the development of both cardiovascular damage and hypertension. Little is known of the molecular mechanisms that lead to development and progression of this condition. This review aims to cover the potential roles of miRNAs in the mechanisms associated with the development and consequences of RH, and explore the current state of the art of diagnostic and therapeutic tools based on miRNA approaches.

Gene delivery to Nile tilapia cells for transgenesis and the role of PI3K-c2α in angiogenesis.

Microinjection is commonly performed to achieve fish transgenesis; however, due to difficulties associated with this technique, new strategies are being developed. Here we evaluate the potential of lentiviral particles to genetically modify Nile tilapia cells to achieve transgenesis using three different approaches: spermatogonial stem cell (SSC) genetic modification and transplantation (SC), in vivo transduction of gametes (GT), and fertilised egg transduction (ET). The SC protocol using larvae generates animals with sustained production of modified sperm (80% of animals with 77% maximum sperm fluorescence [MSF]), but is a time-consuming protocol (sexual maturity in Nile tilapia is achieved at 6 months of age). GT is a faster technique, but the modified gamete production is temporary (70% of animals with 52% MSF). ET is an easier way to obtain mosaic transgenic animals compared to microinjection of eggs, but non-site-directed integration in the fish genome can be a problem. In this study, PI3Kc2α gene disruption impaired development during the embryo stage and caused premature death. The manipulator should choose a technique based on the time available for transgenic obtainment and if this generation is required to be continuous or not.

Absence of suppressor of cytokine signaling 2 turns cardiomyocytes unresponsive to LIF-dependent increases in Ca2+ levels.

Little is known regarding the role of suppressor of cytokine signaling (SOCS) in the control of cytokine signaling in cardiomyocytes. We investigated the consequences of SOCS2 ablation for leukemia inhibitory factor (LIF)-induced enhancement of intracellular Ca2+ ([Ca2+]i) transient by performing experiments with cardiomyocytes from SOCS2-knockout (ko) mice. Similar levels of SOCS3 transcripts were seen in cardiomyocytes from wild-type and SOCS2-ko mice, while SOCS1 mRNA was reduced in SOCS2-ko. Immunoprecipitation experiments showed increased SOCS3 association with gp130 receptor in SOCS2-ko myocytes. Measurements of Ca2+ in wild-type myocytes exposed to LIF showed a significant increase in the magnitude of the Ca2+ transient. This change was absent in LIF-treated SOCS2-ko cells. LIF activation of ERK and STAT3 was observed in both wild-type and SOCS2-ko cells, indicating that in SOCS2-ko, LIF receptors were functional, despite the lack of effect in the Ca2+ transient. In wild-type cells, LIF-induced increase in [Ca2+]i and phospholamban Thr17 [PLN(Thr17)] phosphorylation was inhibited by KN-93, indicating a role for CaMKII in LIF-induced Ca2+ raise. LIF-induced phosphorylation of PLN(Thr17) was abrogated in SOCS2-ko myocytes. In wild-type cardiomyocytes, LIF treatment increased L-type Ca2+ current (ICa,L), a key activator of CaMKII in response to LIF. Conversely, SOCS2-ko myocytes failed to activate ICa,L in response to LIF, providing a rationale for the lack of LIF effect on Ca2+ transient. Our data show that absence of SOCS2 turns cardiomyocytes unresponsive to LIF-induced [Ca2+] raise, indicating that endogenous levels of SOCS2 are crucial for full activation of LIF signaling in the heart.

Determining the Roles of Inositol Trisphosphate Receptors in Neurodegeneration: Interdisciplinary Perspectives on a Complex Topic.

It is well known that calcium (Ca2+) is involved in the triggering of neuronal death. Ca2+ cytosolic levels are regulated by Ca2+ release from internal stores located in organelles, such as the endoplasmic reticulum. Indeed, Ca2+ transit from distinct cell compartments follows complex dynamics that are mediated by specific receptors, notably inositol trisphosphate receptors (IP3Rs). Ca2+ release by IP3Rs plays essential roles in several neurological disorders; however, details of these processes are poorly understood. Moreover, recent studies have shown that subcellular location, molecular identity, and density of IP3Rs profoundly affect Ca2+ transit in neurons. Therefore, regulation of IP3R gene products in specific cellular vicinities seems to be crucial in a wide range of cellular processes from neuroprotection to neurodegeneration. In this regard, microRNAs seem to govern not only IP3Rs translation levels but also subcellular accumulation. Combining new data from molecular cell biology with mathematical modelling, we were able to summarize the state of the art on this topic. In addition to presenting how Ca2+ dynamics mediated by IP3R activation follow a stochastic regimen, we integrated a theoretical approach in an easy-to-apply, cell biology-coherent fashion. Following the presented premises and in contrast to previously tested hypotheses, Ca2+ released by IP3Rs may play different roles in specific neurological diseases, including Alzheimer's disease and Parkinson's disease.

Beneficial effects of angiotensin-(1-7) against deoxycorticosterone acetate-induced diastolic dysfunction occur independently of changes in blood pressure.

Mineralocorticoids have been implicated in the pathogenesis of diastolic heart failure. On the contrary, angiotensin (Ang)-(1-7) has emerged as a potential strategy for treatment of cardiac dysfunction induced by excessive mineralocorticoid receptor activation. A critical question about the cardioprotective effect of Ang-(1-7) in hypertensive models is its dependence on blood pressure (BP) reduction. Here, we addressed this question by investigating the mechanisms involved in Ang-(1-7) cardioprotection against mineralocorticoid receptor activation. Sprague-Dawley (SD) and transgenic (TG) rats that overexpress an Ang-(1-7) producing fusion protein (TG(A1-7)3292) were treated with deoxycorticosterone acetate (DOCA) for 6 weeks. After treatment, SD rats became hypertensive and developed ventricular hypertrophy. These parameters were attenuated in TG-DOCA. SD-DOCA rats developed diastolic dysfunction which was associated at the cellular level with reduced Ca(2+) transient. Oppositely, TG-DOCA myocytes presented enhanced Ca(2+) transient. Moreover, higher extracellular signal-regulated kinase phosphorylation, type 1 phosphatase, and protein kinase Cα levels were found in SD-DOCA cells. In vivo, pressor effects of DOCA can contribute to the diastolic dysfunction, raising the question of whether protection in TG was a consequence of reduced BP. To address this issue, BP in SD-DOCA was kept at TG-DOCA level by giving hydralazine or by reducing the DOCA amount given to rats (Low-DOCA). Under similar BP, diastolic dysfunction and molecular changes were still evident in DOCA-hydralazine and SD-low-DOCA, but not in TG-DOCA. In conclusion, Ang-(1-7) protective signaling against DOCA-induced diastolic dysfunction occurs independently of BP attenuation and is mediated by the activation of pathways involved in Ca(2+) handling, hypertrophy, and survival.

New method for the determination of bile acids in human plasma by liquid-phase microextraction using liquid chromatography-ion-trap-time-of-flight mass spectrometry.

Bile acids (BAs) are derived from cholesterol and produced in the liver. The most abundant bile acids in humans are usually conjugated with glycine and taurine and are divided into primary BAs such as cholic acid (CA) and chenodeoxycholic acid (CDCA) and secondary BAs like deoxycholic acid (DCA), lithocholic acid (LCA) and ursodeoxycholic acid (UDCA). The differences amongst individual bile acids (BAs) are significant in order to distinguish different pathological processes and exposure to chemical compounds. Hollow fiber based liquid-phase microextraction (HF-LPME) is a technique that combines sample cleansing, extraction and the concentration of analytes, where a hydrophobic porous capillary membrane is impregnated with an organic extraction solvent and the lumen is filled with microliters of a phase acceptor both organic by nature. The aim of this study was to develop a new method to extract bile acids from plasma through HF-LPME of two phases (octanol as the acceptor phase) using LCMS-IT-TOF. The optimized two-phased LPME procedure for the extraction of bile acids showed limits of detection 1.0 µg L(-1) and limits of quantification of 5.0 µg L(-1). The intra-assay precision ranged from 2.1 to 11.9%. The method developed was linear over the range of 5.0-200.0 µg L(-1) for all analytes. The hollow-fiber liquid-phase microextraction method was applied to human plasma from workers exposed to organic and halogenated solvents and also to unexposed volunteers. The method is simple, low cost and it does not require large amounts of organic solvents, therefore it is quite suitable for the analysis of bile acids exposed to hepatotoxic compounds.

Glycine transporters type 1 inhibitor promotes brain preconditioning against NMDA-induced excitotoxicity.

Brain preconditioning is a protective mechanism, which can be activated by sub-lethal stimulation of the NMDA receptors (NMDAR) and be used to achieve neuroprotection against stroke and neurodegenerative diseases models. Inhibitors of glycine transporters type 1 modulate glutamatergic neurotransmission through NMDAR, suggesting an alternative therapeutic strategy of brain preconditioning. The aim of this work was to evaluate the effects of brain preconditioning induced by NFPS, a GlyT1 inhibitor, against NMDA-induced excitotoxicity in mice hippocampus, as well as to study its neurochemical mechanisms. C57BL/6 mice (male, 10-weeks-old) were preconditioned by intraperitoneal injection of NFPS at doses of 1.25, 2.5 or 5.0 mg/kg, 24 h before intrahippocampal injection of NMDA. Neuronal death was evaluated by fluoro jade C staining and neurochemical parameters were evaluated by gas chromatography-mass spectrometry, scintillation spectrometry and western blot. We observed that NFPS preconditioning reduced neuronal death in CA1 region of hippocampus submitted to NMDA-induced excitotoxicity. The amino acids (glycine and glutamate) uptake and content were increased in hippocampus of animals treated with NFPS 5.0 mg/kg, which were associated to an increased expression of type-2 glycine transporter (GlyT2) and glutamate transporters (EAAT1, EAAT2 and EAAT3). The expression of GlyT1 was reduced in animals treated with NFPS. Interestingly, the preconditioning reduced expression of GluN2B subunits of NMDAR, whereas did not change the expression of GluN1 or GluN2A in all tested doses. Our study suggests that NFPS preconditioning induces resistance against excitotoxicity, which is associated with neurochemical changes and reduction of GluN2B-containing NMDAR expression.

MicroRNAs in neuronal communication.

MicroRNAs (miRNAs) are short nucleotides sequences that regulate the expression of genes in different eukaryotic cell types. A tremendous amount of knowledge on miRNAs has rapidly accumulated over the last few years, revealing the growing interest in this field of research. On the other hand, clarifying the physiological regulation of gene expression in the central nervous system is important for establishing a reference for comparison to the diseased state. It is well known that the fine tuning of neuronal networks relies on intricate molecular mechanisms, such as the adjustment of the synaptic transmission. As determined by recent studies, regulation of neuronal interactions by miRNAs has critical consequences in the development, adaptation to ambient demands, and degeneration of the nervous system. In contrast, activation of synaptic receptors triggers downstream signaling cascades that generate a vast array of effects, which includes the regulation of novel genes involved in the control of the miRNA life cycle. In this review, we have examined the hot topics on miRNA gene-regulatory activities in the broad field of neuronal communication-related processes. Furthermore, in addition to indicating the newly described effect of miRNAs on the regulation of specific neurotransmitter systems, we have pointed out how these systems affect the expression, transport, and stability of miRNAs. Moreover, we discuss newly described and under-investigation mechanisms involving the intercellular transfer of miRNAs, aided by exosomes and gap junctions. Thus, in the current review, we were able to highlight recent findings related to miRNAs that indisputably contributed towards the understanding of the nervous system in health and disease.

Development of a method for the analysis of drugs of abuse in vitreous humor by capillary electrophoresis with diode array detection (CE-DAD).

This work presents the development of an analytical method based on capillary electrophoresis with diode array detection for the analysis of drugs of abuse and biotransformation products in vitreous humor. Composition of the background electrolyte, implementation of an online pre-concentration strategy and sample preparation procedures were objects of study. The complete electrophoretic separation of 12 analytes (amphetamine, methamphetamine, 3,4-methylenedioxyamphetamine (MDA), 3,4-methylenedioxymethamphetamine (MDMA), 3,4-methylenedioxyethylamphetamine (MDEA), ketamine, cocaine, cocaethylene, lidocaine, morphine, 6-monoacetylmorphine and heroin) and the internal standard N-methyl-1-(3,4-methylenedioxyphenyl)-2-butamine (MBDB) was obtained within 13min of run. The method was validated presenting good linearity (r(2)>0.99), recovery ≥90%, precision better than 12% RSD and acceptable accuracy in the range of 86-118% at three concentration levels (50, 100 and 500ng/mL). LODs and LOQs in the order of 1-5ng/mL and 5-10ng/mL, respectively, were obtained. After validation, the method was applied to eighty-seven vitreous humor samples and the results were compared to those obtained by a liquid chromatography tandem mass spectrometry (LC-MS/MS) screening method, routinely used by the forensic toxicology laboratory of the Sao Paulo State Police, Brazil. Cocaine was detected in 7.1%, cocaethylene in 3.6%, lidocaine in 2.4% and ketamine in 1.2% of the total number of analyzed samples.