An account on the history of pharmacology in Spain.

Here we present an account on the history of pharmacology in Spain. Pharmacology as an independent science in Europe began with the creation of university chairs. Of particular relevance was the appointment in 1872 of Osswald Shmiedeberg as chairman of an Institute of Pharmacology at the University of Strassbourg, Germany. Teófilo Hernando pioneered in Spain the new emerging pharmacology at the beginning of the XX Century. He made a posdoctoral stay in the laboratory of Schmiedeberg, working on digitalis. In 1912 he won the chair of "Materia Médica y Arte de Recetar" at "Universidad Central of Madrid" (today, "Universidad Complutense de Madrid", UCM). He soon decided to transform such subject to the emerging modern pharmacology, with the teaching of experimental pharmacology in the third course of medical studies and clinical therapeutics (today clinical pharmacology) in the sixth course. This was the status of pharmacology in 1920, supporting the view that Hernando was a pioneer of clinical pharmacology. However, the Spanish Civil War and the II Word War interropted this division of preclinical and clinical pharmacology; only in the 1980's was clinical pharmacolgy partially developed in Spain. From a scientific point of view, Hernando directly trained various young pharmacologists that extended the new science to various Spanish universities. Some of his direct disciples were Benigno Lorenzo Velázquez, Francisco García Valdecasas, Rafael Méndez, Tomás Alday, Gabriel Sánchez de la Cuesta, Dámaso Gutiérrez or Ramón P é rez-Cirera. One of the central research subject was the analysis of the effects of digitalis on the cat and frog heart. In the initiation of the 1970 s pharmacologists trained by those Hernando's students grew throughout various universities and the "Consejo Superior de Investigaciones Científicas" (CSIC). And hence, in 1972 the "Sociedad Española de Farmacología" (SEF) emerged. Later on, in the 1990's the "Sociedad Española de Farmacología Clínica (SEFC) also emerged. The relationship between the two societies is still weak. Out of the vast scope of the pharmacological sciences, Spanish pharmacologists have made relevant contributions in two areas namely, neuropsychopharmacology and cardiovacular pharmacology. Nonetheless, in other areas such as smooth muscle, gastroenterology, pharmacogenetics and hepatic toxicity, Spanish pharmacologists have also made relevant contributions. A succint description of such contributions is made. Finally, some hints on perspectives for the further development of preclinical and clinical pharmacology in Spain, are offered.

A Permutation Entropy analysis to determine significant daily intervals to improve risk stratification tasks from COVID patients.

SARS-CoV-2 infection has a wide range of clinical manifestations making its diagnosis difficult, which is an important problem to solve. We evaluated heart rate data extracted from the Stanford University database. The data set considers heart rate and step records of 118 patients, where 90 correspond to healthy individuals and 28 patients with COVID. Each daily record was divided into 5-minute segments, providing 288 data per patient. The date of symptom onset was considered as a reference point to extract subsets of data whose variability was considerable, such as 30 days before the date and 30 days after it. Each of the 60 segments of 288 data per patient was treated using Permutation Entropy, Approximate Entropy, Spectral Entropy and Singular Value Decomposition Entropy. The average of the data from each group was used to construct the circadian profiles which were analyzed using the Mann-Whitney-Wilcoxon test, determining the most relevant 5-minute segments, whose p-value was less than 0.05. In this way, the Spectral Entropy was discarded as it did not show any significantly different segment. The efficiency of the method was reflected in the performance of a logistic model for binary classification proposed in this work, which reflected an accuracy of 94.12% in the PE case, 88% in the ApEn case and 94% in the SVDE case. The proposed analysis turns out to be highly efficient when detecting significant segments that allow improving the classification tasks carried out by Machine Learning models, which provides a basis for the study of statistics such as entropy to delimit databases and improve the performance of classifier models.

Towards automatic EEG cyclic alternating pattern analysis: a systematic review.

This study conducted a systematic review to determine the feasibility of automatic Cyclic Alternating Pattern (CAP) analysis. Specifically, this review followed the 2020 Preferred Reporting Items for Systematic reviews and Meta-Analyses (PRISMA) guidelines to address the formulated research question: is automatic CAP analysis viable for clinical application? From the identified 1,280 articles, the review included 35 studies that proposed various methods for examining CAP, including the classification of A phase, their subtypes, or the CAP cycles. Three main trends were observed over time regarding A phase classification, starting with mathematical models or features classified with a tuned threshold, followed by using conventional machine learning models and, recently, deep learning models. Regarding the CAP cycle detection, it was observed that most studies employed a finite state machine to implement the CAP scoring rules, which depended on an initial A phase classifier, stressing the importance of developing suitable A phase detection models. The assessment of A-phase subtypes has proven challenging due to various approaches used in the state-of-the-art for their detection, ranging from multiclass models to creating a model for each subtype. The review provided a positive answer to the main research question, concluding that automatic CAP analysis can be reliably performed. The main recommended research agenda involves validating the proposed methodologies on larger datasets, including more subjects with sleep-related disorders, and providing the source code for independent confirmation.

Combining Heart Rate Variability and Oximetry to Improve Apneic Event Screening in Non-Desaturating Patients.

In this paper, we thoroughly analyze the detection of sleep apnea events in the context of Obstructive Sleep Apnea (OSA), which is considered a public health problem because of its high prevalence and serious health implications. We especially evaluate patients who do not always show desaturations during apneic episodes (non-desaturating patients). For this purpose, we use a database (HuGCDN2014-OXI) that includes desaturating and non-desaturating patients, and we use the widely used Physionet Apnea Dataset for a meaningful comparison with prior work. Our system combines features extracted from the Heart-Rate Variability (HRV) and SpO2, and it explores their potential to characterize desaturating and non-desaturating events. The HRV-based features include spectral, cepstral, and nonlinear information (Detrended Fluctuation Analysis (DFA) and Recurrence Quantification Analysis (RQA)). SpO2-based features include temporal (variance) and spectral information. The features feed a Linear Discriminant Analysis (LDA) classifier. The goal is to evaluate the effect of using these features either individually or in combination, especially in non-desaturating patients. The main results for the detection of apneic events are: (a) Physionet success rate of 96.19%, sensitivity of 95.74% and specificity of 95.25% (Area Under Curve (AUC): 0.99); (b) HuGCDN2014-OXI of 87.32%, 83.81% and 88.55% (AUC: 0.934), respectively. The best results for the global diagnosis of OSA patients (HuGCDN2014-OXI) are: success rate of 95.74%, sensitivity of 100%, and specificity of 89.47%. We conclude that combining both features is the most accurate option, especially when there are non-desaturating patterns among the recordings under study.

Forty years of the adrenal chromaffin cell through ISCCB meetings around the world.

This historical review focuses on the evolution of the knowledge accumulated during the last two centuries on the biology of the adrenal medulla gland and its chromaffin cells (CCs). The review emerged in the context of a series of meetings that started on the Spanish island of Ibiza in 1982 with the name of the International Symposium on Chromaffin Cell Biology (ISCCB). Hence, the review is divided into two periods namely, before 1982 and from this year to 2022, when the 21st ISCCB meeting was just held in Hamburg, Germany. The first historical period extends back to 1852 when Albert Kölliker first described the fine structure and function of the adrenal medulla. Subsequently, the adrenal staining with chromate salts identified the CCs; this was followed by the establishment of the embryological origin of the adrenal medulla, and the identification of adrenaline-storing vesicles. By the end of the nineteenth century, the basic morphology, histochemistry, and embryology of the adrenal gland were known. The twentieth century began with breakthrough findings namely, the experiment of Elliott suggesting that adrenaline was the sympathetic neurotransmitter, the isolation of pure adrenaline, and the deciphering of its molecular structure and chemical synthesis in the laboratory. In the 1950s, Blaschko isolated the catecholamine-storing vesicles from adrenal medullary extracts. This switched the interest in CCs as models of sympathetic neurons with an explosion of studies concerning their functions, i.e., uptake of catecholamines by chromaffin vesicles through a specific coupled transport system; the identification of several vesicle components in addition to catecholamines including chromogranins, ATP, opioids, and other neuropeptides; the calcium-dependence of the release of catecholamines; the underlying mechanism of exocytosis of this release, as indicated by the co-release of proteins; the cross-talk between the adrenal cortex and the medulla; and the emission of neurite-like processes by CCs in culture, among other numerous findings. The 1980s began with the introduction of new high-resolution techniques such as patch-clamp, calcium probes, marine toxins-targeting ion channels and receptors, confocal microscopy, or amperometry. In this frame of technological advances at the Ibiza ISCCB meeting in 1982, 11 senior researchers in the field predicted a notable increase in our knowledge in the field of CCs and the adrenal medulla; this cumulative knowledge that occurred in the last 40 years of history of the CC is succinctly described in the second part of this historical review. It deals with cell excitability, ion channel currents, the exocytotic fusion pore, the handling of calcium ions by CCs, the kinetics of exocytosis and endocytosis, the exocytotic machinery, and the life cycle of secretory vesicles. These concepts together with studies on the dynamics of membrane fusion with super-resolution imaging techniques at the single-protein level were extensively reviewed by top scientists in the field at the 21st ISCCB meeting in Hamburg in the summer of 2022; this frontier topic is also briefly reviewed here. Many of the concepts arising from those studies contributed to our present understanding of synaptic transmission. This has been studied in physiological or pathophysiological conditions, in CCs from animal disease models. In conclusion, the lessons we have learned from CC biology as a peripheral model for brain and brain disease pertain more than ever to cutting-edge research in neurobiology. In the 22nd ISCCB meeting in Israel in 2024 that Uri Asheri is organizing, we will have the opportunity of seeing the progress of the questions posed in Ibiza, and on other questions that undoubtedly will arise.

Mitochondrial dysfunction in chromaffin cells from the R6/1 mouse model of Huntington's disease: Impact on exocytosis and calcium current regulation.

From a pathogenic perspective, Huntington's disease (HD) is being considered as a synaptopathy. As such, alterations in brain neurotransmitter release occur. As the activity of the sympathoadrenal axis is centrally controlled, deficits in the exocytotic release of catecholamine release may also occur. In fact, in chromaffin cells (CCs) of the adrenal medulla of the R6/1 model of HD, decrease of secretion and altered kinetics of the exocytotic fusion pore have been reported. Those alterations could be linked to mitochondrial deficits occurring in peripheral CCs, similar to those described in brain mitochondria. Here we have inquired about alterations in mitochondrial structure and function and their impact on exocytosis and calcium channel currents (ICa). We have monitored various parameters linked to those events, in wild type (WT) and the R6/1 mouse model of HD at a pre-disease stage (2 months age, 2 m), and when motor deficits are present (7 months age, 7 m). In isolated CCs from 7 m and in the adrenal medulla of R6/1 mice, we found the following alterations (with respect 7 m WT mice): (i) augmented fragmented mitochondria and oxidative stress with increased oxidized glutathione; (ii) decreased basal and maximal respiration; (iii) diminution of ATP cell levels; (iv) mitochondrial depolarization; (v) drastic decrease of catecholamine release with poorer potentiation by protonophore FCCP; (vi) decreased ICa inhibition by FCCP; and (vii) lesser potentiation by BayK8644 of ICa and smaller prolongation of current deactivation. Of note was the fact several of these alterations were already manifested in CCs from 2 m R6/1 mice at pre-disease stages. Based on those results, a plausible hypothesis can be raised in the sense that altered mitochondrial function seems to be an early primary event in HD pathogenesis. This is in line with an increasing number of mitochondrial, metabolic, and inflammatory alterations being recently reported in various HD peripheral tissues.

Small Synthetic Hyaluronan Disaccharide BIS014 Mitigates Neuropathic Pain in Mice.

Neuropathic pain (NP) is a challenging condition to treat, as the need for new drugs to treat NP is an unmet goal. We investigated the analgesic potential of a new sulfated disaccharide compound, named BIS014. Oral administration (p.o.) of this compound induced ameliorative effects in formalin-induced nociception and capsaicin-induced secondary mechanical hypersensitivity in mice, but also after partial sciatic nerve transection (spared nerve injury), chemotherapy (paclitaxel)-induced NP, and diabetic neuropathy induced by streptozotocin. Importantly, BIS014, at doses active on neuropathic hypersensitivity (60 mg/kg/p.o.), did not alter exploratory activity or motor coordination (in the rotarod test), unlike a standard dose of gabapentin (40 mg/kg/p.o.) which although inducing antiallodynic effects on the NP models, it also markedly decreased exploration and motor coordination. In docking and molecular dynamic simulation studies, BIS014 interacted with TRPV1, a receptor involved in pain transmission where it behaved as a partial agonist. Additionally, similar to capsaicin, BIS014 increased cytosolic Ca2+ concentration ([Ca2+]c) in neuroblastoma cells expressing TRPV1 receptors; these elevations were blocked by ruthenium red. BIS014 did not block capsaicin-elicited [Ca2+]c transients, but inhibited the increase in the firing rate of action potentials in bradykinin-sensitized dorsal root ganglion neurons stimulated with capsaicin. Perspective: We report that the oral administration of a new sulfated disaccharide compound, named BIS014, decreases neuropathic pain from diverse etiology in mice. Unlike the comparator gabapentin, BIS014 does not induce sedation. Thus, BIS014 has the potential to become a new efficacious non-sedative oral medication for the treatment of neuropathic pain.

A-phase index: an alternative view for sleep stability analysis based on automatic detection of the A-phases from the cyclic alternating pattern.

STUDY OBJECTIVES: Sleep stability can be studied by evaluating the cyclic alternating pattern (CAP) in electroencephalogram (EEG) signals. The present study presents a novel approach for assessing sleep stability, developing an index based on the CAP A-phase characteristics to display a sleep stability profile for a whole night's sleep. METHODS: Two ensemble classifiers were developed to automatically score the signals, one for "A-phase" and the other for "non-rapid eye movement" estimation. Both were based on three one-dimension convolutional neural networks. Six different inputs were produced from the EEG signal to feed the ensembles' classifiers. A proposed heuristic-oriented search algorithm individually tuned the classifiers' structures. The outputs of the two ensembles were combined to estimate the A-phase index (API). The models can also assess the A-phase subtypes, their API, and the CAP cycles and rate. RESULTS: Four dataset variations were considered, examining healthy and sleep-disordered subjects. The A-phase average estimation's accuracy, sensitivity, and specificity range was 82%-87%, 72%-80%, and 82%-88%, respectively. A similar performance was attained for the A-phase subtype's assessments, with an accuracy range of 82%-88%. Furthermore, in the examined dataset's variations, the API metric's average error varied from 0.15 to 0.25 (with a median range of 0.11-0.24). These results were attained without manually removing wake or rapid eye movement periods, leading to a methodology suitable to produce a fully automatic CAP scoring algorithm. CONCLUSIONS: Metrics based on API can be understood as a new view for CAP analysis, where the goal is to produce and examine a sleep stability profile.

Online Detection of Catecholamine Release from the Perfused Rat Adrenal Gland.

Retrogradely perfused adrenal glands have historically served for establishing many of our current knowledge on the stimulus-secretion coupling process. Although the use of intact adrenals has largely been switched to isolated chromaffin cells, adrenal glands are still a very valuable tool to characterize physiological and pharmacological questions. Even more, this is an excellent preparation for studying the splanchnic nerve/chromaffin cell interaction. In this chapter, we will provide the ways to (i) perform retrograde perfusion of isolated rat adrenals, (ii) the method to apply electrical splanchnic nerve stimulation, and (iii) the preparation of adrenals to conduct online electrochemical detection of catecholamine release.

Purinergic Receptors P2X7 and P2X4 as Markers of Disease Progression in the rd10 Mouse Model of Inherited Retinal Dystrophy.

The purinergic receptor P2X7 (P2X7R) is implicated in all neurodegenerative diseases of the central nervous system. It is also involved in the retinal degeneration associated with glaucoma, age-related macular degeneration, and diabetic retinopathy, and its overexpression in the retina is evident in these disorders. Retinitis pigmentosa is a progressive degenerative disease that ultimately leads to blindness. Here, we investigated the expression of P2X7R during disease progression in the rd10 mouse model of RP. As the purinergic receptor P2X4 is widely co-expressed with P2X7R, we also studied its expression in the retina of rd10 mice. The expression of P2X7R and P2X4R was examined by immunohistochemistry, flow cytometry, and western blotting. In addition, we analyzed retinal functionality by electroretinographic recordings of visual responses and optomotor tests and retinal morphology. We found that the expression of P2X7R and P2X4R increased in rd10 mice concomitant with disease progression, but with different cellular localization. Our findings suggest that P2X7R and P2X4R might play an important role in RP progression, which should be further analyzed for the pharmacological treatment of inherited retinal dystrophies.

Multiple Time Series Fusion Based on LSTM: An Application to CAP A Phase Classification Using EEG.

The Cyclic Alternating Pattern (CAP) is a periodic activity detected in the electroencephalogram (EEG) signals. This pattern was identified as a marker of unstable sleep with several possible clinical applications; however, there is a need to develop automatic methodologies to facilitate real-world applications based on CAP assessment. Therefore, a deep learning-based EEG channels' feature level fusion was proposed in this work and employed for the CAP A phase classification. Two optimization algorithms optimized the channel selection, fusion, and classification procedures. The developed methodologies were evaluated by fusing the information from multiple EEG channels for patients with nocturnal frontal lobe epilepsy and patients without neurological disorders. Results showed that both optimization algorithms selected a comparable structure with similar feature level fusion, consisting of three electroencephalogram channels (Fp2-F4, C4-A1, F4-C4), which is in line with the CAP protocol to ensure multiple channels' arousals for CAP detection. Moreover, the two optimized models reached an area under the receiver operating characteristic curve of 0.82, with average accuracy ranging from 77% to 79%, a result in the upper range of the specialist agreement and best state-of-the-art works, despite a challenging dataset. The proposed methodology also has the advantage of providing a fully automatic analysis without requiring any manual procedure. Ultimately, the models were revealed to be noise-resistant and resilient to multiple channel loss, being thus suitable for real-world application.

Combined drug triads for synergic neuroprotection in retinal degeneration.

This review focuses on retina degeneration occurring during glaucoma, age-related macular degeneration (AMD), diabetic retinopathy (DR), and retinitis pigmentosa (RP), and on the potential therapeutic use of triads of repositioned medicines, addressed to distinct but complementary targets, to prevent, delay or stop retina cell death. Although myriad pathogenic mechanisms have been implicated in these disorders, common signaling pathways leading to apoptotic cell death to all of them, and to all neurodegenerative diseases are (i) calcium dyshomeostasis/excitotoxicity; (ii) oxidative stress/mitochondrial dysfunction, and (iii) neuroinflammation/P2X7 receptor activation. From a therapeutic point of view, it is relevant to consider the multitarget approach based on the use of combined medicines acting on complementary pathogenic mechanisms that has been highly successful in the treatment of chronic diseases such as cancer, AIDS, pain, hypertension, Parkinson's disease, cardiac failure, depression, or the epilepsies as the basic mechanisms of cell death do not differ between the different CNS degenerative diseases. We suggest the multi-target therapy approach could be more effective compared with single-drug treatments. Used at doses lower than standard, these triads may also be safer and more efficient. After the establishment of a proof-of-concept in animal models of retinal degeneration, potential successful preclinical trials of such combinations may eventually drive to test this concept in clinical trials in patients, first to evaluate the safety and efficacy of the drug combinations in humans and then their therapeutic advantages, if any, seeking the prevention and/or the delay of retina degeneration and blindness.

Heuristic Optimization of Deep and Shallow Classifiers: An Application for Electroencephalogram Cyclic Alternating Pattern Detection.

Methodologies for automatic non-rapid eye movement and cyclic alternating pattern analysis were proposed to examine the signal from one electroencephalogram monopolar derivation for the A phase, cyclic alternating pattern cycles, and cyclic alternating pattern rate assessments. A population composed of subjects free of neurological disorders and subjects diagnosed with sleep-disordered breathing was studied. Parallel classifications were performed for non-rapid eye movement and A phase estimations, examining a one-dimension convolutional neural network (fed with the electroencephalogram signal), a long short-term memory (fed with the electroencephalogram signal or with proposed features), and a feed-forward neural network (fed with proposed features), along with a finite state machine for the cyclic alternating pattern cycle scoring. Two hyper-parameter tuning algorithms were developed to optimize the classifiers. The model with long short-term memory fed with proposed features was found to be the best, with accuracy and area under the receiver operating characteristic curve of 83% and 0.88, respectively, for the A phase classification, while for the non-rapid eye movement estimation, the results were 88% and 0.95, respectively. The cyclic alternating pattern cycle classification accuracy was 79% for the same model, while the cyclic alternating pattern rate percentage error was 22%.

Availability and performance of face based non-contact methods for heart rate and oxygen saturation estimations: A systematic review.

BACKGROUND: Consumer-level cameras have provided an advantage of designing cost-effective, non-contact physiological parameters estimation approaches which is not possible with gold standard estimation techniques. This encourages the development of non-contact estimation methods using camera technology. Therefore, this work aims to present a systematic review summarizing the currently existing face-based non-contact methods along with their performance. METHODS: This review includes all heart rate (HR) and oxygen saturation (SpO2) studies published in journals and a few reputed conferences, which have compared the proposed estimation methods with one or more standard reference devices. The articles were collected from the following research databases: Institute of Electrical and Electronics Engineers (IEEE), PubMed, Web of Science (WoS), Science Direct, and Association of Computer Machinery (ACM) digital library. All database searches were completed on May 20, 2021. Each study was assessed using a finite set of identified factors for reporting bias. RESULTS: Out of 332 identified studies, 32 studies were selected for the final review. Additionally, 18 studies were included by thoroughly checking these studies. 3 out of 50 (6%) studies were performed in clinical conditions, while the remaining studies were carried out on a healthy population. 42 out of 50 (84%) studies have estimated HR, while 5/50 (10%) studies have measured SpO2 only. The remaining three studies have estimated both parameters. The majority of the studies have used 1-3 min videos for estimation. Among the estimation methods, Deep Learning and Independent component analysis (ICA) were used by 11/42 (26.19%) and 9/42 (21.42%) studies, respectively. According to the Bland-Altman analysis, only 8/45 (17.77%) HR studies achieved the clinically accepted error limits whereas, for SpO2, 4/5 (80%) studies have matched the industry standards (±3%). DISCUSSION: Deep Learning and ICA have been predominantly used for HR estimations. Among deep learning estimation methods, convolutional neural networks have been employed till date due to their good generalization ability. Most non-contact HR estimation methods need significant improvements to implement these methods in a clinical environment. Furthermore, these methods need to be tested on the subjects suffering from any related disease. SpO2 estimation studies are challenging and need to be tested by conducting hypoxemic events. The authors would encourage reporting the detailed information about the study population, the use of longer videos, and appropriate performance metrics and testing under abnormal HR and SpO2 ranges for future estimation studies.

A Motion and Illumination Resistant Non-Contact Method Using Undercomplete Independent Component Analysis and Levenberg-Marquardt Algorithm.

Heart Rate (HR) estimation is of utmost importance due to its applicability in diverse fields. Conventional methods for HR estimation require skin contact and are not suitable in certain scenarios such as sensitive skin or prolonged unobtrusive HR monitoring. Therefore remote photoplethysmography (rPPG) methods have become an active area of research. These methods utilize the facial videos acquired using a camera followed by extracting the Blood Volume Pulse (BVP) signal for heart rate calculation. The existing rPPG methods either utilized a single color channel or weighted color differences, which has certain limitations dealing with motion and illumination artifacts. This study considered BVP extraction as an undercomplete problem and proposed a method resistant to motion and illumination variation artifacts. This method is based on an undercomplete independent component analysis, aiming to estimate the unmixing matrix using a non-linear Cumulative Density Function (CDF) that has been optimized using the customized Levenberg-Marquardt algorithm. Therefore, the method is named U-LMA. The proposed method was tested under three scenarios: constrained, motion, and illumination variations scenarios. High Pearson correlation coefficient values and smaller lower-upper statistical limits of Bland-Altman plots justified the outstanding performance of the proposed U-LMA. Furthermore, its comparative analysis with the state-of-the-art methods demonstrated its efficacy and reliability, which was proven by the lowest error and highest correlation values (0.01 significance level). Additionally, higher accuracy satisfying the clinically accepted error differences also justified its clinical relevance.

Increased expression of the ATP-gated P2X7 receptor reduces responsiveness to anti-convulsants during status epilepticus in mice.

BACKGROUND AND PURPOSE: Refractory status epilepticus is a clinical emergency associated with high mortality and morbidity. Increasing evidence suggests neuroinflammation contributes to the development of drug-refractoriness during status epilepticus. Here, we have determined the contribution of the ATP-gated P2X7 receptor, previously linked to inflammation and increased hyperexcitability, to drug-refractory status epilepticus and its therapeutic potential. EXPERIMENTAL APPROACH: Status epilepticus was induced via a unilateral microinjection of kainic acid into the amygdala in adult mice. Severity of status epilepticus was compared in animals with overexpressing or knock-out of the P2X7 receptor, after inflammatory priming by pre-injection of bacterial lipopolysaccharide (LPS) and in mice treated with P2X7 receptor-targeting and anti-inflammatory drugs. KEY RESULTS: Mice overexpressing P2X7 receptors were unresponsive to several anticonvulsants (lorazepam, midazolam, phenytoin and carbamazepine) during status epilepticus. P2X7 receptor expression increased in microglia during status epilepticus, at times when responses to anticonvulsants were reduced. Overexpression of P2X7 receptors induced a pro-inflammatory phenotype in microglia during status epilepticus and the anti-inflammatory drug minocycline restored normal responses to anticonvulsants in mice overexpressing P2X7 receptors. Pretreatment of wild-type mice with LPS increased P2X7 receptor levels in the brain and reduced responsiveness to anticonvulsants during status epilepticus, which was overcome by either genetic deletion of P2X7 receptors or treatment with the P2X7 receptor antagonists, AFC-5128 or ITH15004. CONCLUSION AND IMPLICATIONS: Our results demonstrate that P2X7 receptor-induced pro-inflammatory effects contribute to resistance to pharmacotherapy during status epilepticus. Therapies targeting P2X7 receptors could be novel adjunctive treatments for drug-refractory status epilepticus.

Intracellular calcium and inflammatory markers, mediated by purinergic stimulation, are differentially regulated in monocytes of patients with major depressive disorder.

The P2X7 receptor (P2X7R) is a ligand-gated ion channel that is being recognized as a major player in neuropsychiatric disorders such as Major Depressive Disorder (MDD). P2X7R activation is triggered by high extracellular ATP concentrations, leading to channel opening and inducing an increase in cytosolic calcium concentration ([Ca2+]c), that activates the inflammatory pathway. Those receptors are expressed not only in CNS cells but also in peripheral blood cells, where they are activated in response to inflammatory molecules such as bacterial lipopolysaccharide (LPS). LPS induced-tissue damage promotes an elevation of extracellular ATP, triggering the NRLP3-inflammasome assembly and activation that, sequentially, induces caspase-1 cleavage and IL-1ß processing and secretion. In this context, we attempt to understand the role of P2X7R in [Ca2+]c homeostasis regulation, inflammasome expression and its pharmacological modulation in MDD. For this purpose, monocytes were isolated from peripheral blood of MDD patients and [Ca2+]c was monitored with the intracellular probe Fura-2. Our results point out to P2X7R as the responsible of the Ca2+ imbalance, as well as TNF-α-dependent activation of caspase-1 in MDD patients. In addition, P2X7R blockade with its specific antagonist, JNJ-47965567, reduces the Ca2+ entry upon Bz-ATP exposure. Altogether, our results point that MDD patients have both, Ca2+ homeostasis alteration and an inflammatory status, which promote an independent-inflammasome activation of caspase-1. Therefore, we propose the pharmacological modulation of P2X7R as a therapeutic approach against MDD symptoms.

Progressive Mitochondrial SOD1G93A Accumulation Causes Severe Structural, Metabolic and Functional Aberrations through OPA1 Down-Regulation in a Mouse Model of Amyotrophic Lateral Sclerosis.

In recent years, the "non-autonomous motor neuron death" hypothesis has become more consolidated behind amyotrophic lateral sclerosis (ALS). It postulates that cells other than motor neurons participate in the pathology. In fact, the involvement of the autonomic nervous system is fundamental since patients die of sudden death when they become unable to compensate for cardiorespiratory arrest. Mitochondria are thought to play a fundamental role in the physiopathology of ALS, as they are compromised in multiple ALS models in different cell types, and it also occurs in other neurodegenerative diseases. Our study aimed to uncover mitochondrial alterations in the sympathoadrenal system of a mouse model of ALS, from a structural, bioenergetic and functional perspective during disease instauration. We studied the adrenal chromaffin cell from mutant SOD1G93A mouse at pre-symptomatic and symptomatic stages. The mitochondrial accumulation of the mutated SOD1G93A protein and the down-regulation of optic atrophy protein-1 (OPA1) provoke mitochondrial ultrastructure alterations prior to the onset of clinical symptoms. These changes affect mitochondrial fusion dynamics, triggering mitochondrial maturation impairment and cristae swelling, with increased size of cristae junctions. The functional consequences are a loss of mitochondrial membrane potential and changes in the bioenergetics profile, with reduced maximal respiration and spare respiratory capacity of mitochondria, as well as enhanced production of reactive oxygen species. This study identifies mitochondrial dynamics regulator OPA1 as an interesting therapeutic target in ALS. Additionally, our findings in the adrenal medulla gland from presymptomatic stages highlight the relevance of sympathetic impairment in this disease. Specifically, we show new SOD1G93A toxicity pathways affecting cellular energy metabolism in non-motor neurons, which offer a possible link between cell specific metabolic phenotype and the progression of ALS.

One hundred years from Otto Loewi experiment, a dream that revolutionized our view of neurotransmission.

One hundred years ago, a 4-page paper published in the Pflüger's Archiv fur die Gesamte Physiologie des Menschen und der Tiere dramatically changed our view on synaptic transmission. The paper reported an ingenious, yet straightforward experiment made by Professor Otto Loewi in 1920 and published in 1921, which constitutes the first clear-cut proof for the chemical nature of transmission of the nerve impulse from nerve to muscle. The approach to this experiment was, however, tortuous and long.

Ultrasound evaluation of adrenal gland size in clinically healthy dogs and in dogs with hyperadrenocorticism.

BACKGROUND: Precise reference intervals of adrenal gland thickness are required for detection of adrenomegaly in dogs with hyperadrenocorticism (HAC). METHODS: Eighty-six clinically healthy dogs were prospectively included, and 91 dogs with untreated HAC were retrospectively evaluated. Dorso-ventral adrenal gland thickness was ultrasonographically measured on the sagittal plane. Dogs were classified into four body weight categories, and those with HAC were also ultrasonographically classified as consistent with pituitary-dependent HAC (PDH), adrenal-dependent HAC (FAT), equivocal adrenal asymmetry (EAA), or normal adrenal thickness. RESULTS: The upper limits for left adrenal gland in clinically healthy dogs were 5.1 mm (≥2.5-5 kg), 5.5 mm (>5-10 kg), 6.4 mm (>10-20 kg), and 7.3 mm (>20-40 kg), and for right adrenal gland the upper limits were 5.3 mm (≥2.5-5 kg), 6.8 mm (>5-10 kg), 7.5 mm (>10-20 kg), and 8.7 mm (>20-40 kg). The sensitivity of ultrasound to detect adrenomegaly in dogs with HAC was 95.6%. Most dogs with HAC (56.0%) had ultrasound findings consistent with either PDH or FAT; however, EAA was commonly occurring in 39.6% of dogs with HAC. CONCLUSIONS: The sensitivity of ultrasonography to detect adrenomegaly in dogs with HAC is high when using four weight categories. EAA is common in dogs with HAC.