Inhibition of astroglial hemichannels prevents synaptic transmission decline during spreading depression.

BACKGROUND: Spreading depression (SD) is an intriguing phenomenon characterized by massive slow brain depolarizations that affect neurons and glial cells. This phenomenon is repetitive and produces a metabolic overload that increases secondary damage. However, the mechanisms associated with the initiation and propagation of SD are unknown. Multiple lines of evidence indicate that persistent and uncontrolled opening of hemichannels could participate in the pathogenesis and progression of several neurological disorders including acute brain injuries. Here, we explored the contribution of astroglial hemichannels composed of connexin-43 (Cx43) or pannexin-1 (Panx1) to SD evoked by high-K+ stimulation in brain slices. RESULTS: Focal high-K+ stimulation rapidly evoked a wave of SD linked to increased activity of the Cx43 and Panx1 hemichannels in the brain cortex, as measured by light transmittance and dye uptake analysis, respectively. The activation of these channels occurs mainly in astrocytes but also in neurons. More importantly, the inhibition of both the Cx43 and Panx1 hemichannels completely prevented high K+-induced SD in the brain cortex. Electrophysiological recordings also revealed that Cx43 and Panx1 hemichannels critically contribute to the SD-induced decrease in synaptic transmission in the brain cortex and hippocampus. CONCLUSIONS: Targeting Cx43 and Panx1 hemichannels could serve as a new therapeutic strategy to prevent the initiation and propagation of SD in several acute brain injuries.

SARS-CoV-2 spike protein S1 activates Cx43 hemichannels and disturbs intracellular Ca2+ dynamics.

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes the ongoing coronavirus disease 2019 (COVID-19). An aspect of high uncertainty is whether the SARS-CoV-2 per se or the systemic inflammation induced by viral infection directly affects cellular function and survival in different tissues. It has been postulated that tissue dysfunction and damage observed in COVID-19 patients may rely on the direct effects of SARS-CoV-2 viral proteins. Previous evidence indicates that the human immunodeficiency virus and its envelope protein gp120 increase the activity of connexin 43 (Cx43) hemichannels with negative repercussions for cellular function and survival. Here, we evaluated whether the spike protein S1 of SARS-CoV-2 could impact the activity of Cx43 hemichannels. RESULTS: We found that spike S1 time and dose-dependently increased the activity of Cx43 hemichannels in HeLa-Cx43 cells, as measured by dye uptake experiments. These responses were potentiated when the angiotensin-converting enzyme 2 (ACE2) was expressed in HeLa-Cx43 cells. Patch clamp experiments revealed that spike S1 increased unitary current events with conductances compatible with Cx43 hemichannels. In addition, Cx43 hemichannel opening evoked by spike S1 triggered the release of ATP and increased the [Ca2+]i dynamics elicited by ATP. CONCLUSIONS: We hypothesize that Cx43 hemichannels could represent potential pharmacological targets for developing therapies to counteract SARS-CoV-2 infection and their long-term consequences.

Effect of positive end-expiratory pressure on lung injury and haemodynamics during experimental acute respiratory distress syndrome treated with extracorporeal membrane oxygenation and near-apnoeic ventilation.

BACKGROUND: Lung rest has been recommended during extracorporeal membrane oxygenation (ECMO) for severe acute respiratory distress syndrome (ARDS). Whether positive end-expiratory pressure (PEEP) confers lung protection during ECMO for severe ARDS is unclear. We compared the effects of three different PEEP levels whilst applying near-apnoeic ventilation in a model of severe ARDS treated with ECMO. METHODS: Acute respiratory distress syndrome was induced in anaesthetised adult male pigs by repeated saline lavage and injurious ventilation for 1.5 h. After ECMO was commenced, the pigs received standardised near-apnoeic ventilation for 24 h to maintain similar driving pressures and were randomly assigned to PEEP of 0, 10, or 20 cm H2O (n=7 per group). Respiratory and haemodynamic data were collected throughout the study. Histological injury was assessed by a pathologist masked to PEEP allocation. Lung oedema was estimated by wet-to-dry-weight ratio. RESULTS: All pigs developed severe ARDS. Oxygenation on ECMO improved with PEEP of 10 or 20 cm H2O, but did not in pigs allocated to PEEP of 0 cm H2O. Haemodynamic collapse refractory to norepinephrine (n=4) and early death (n=3) occurred after PEEP 20 cm H2O. The severity of lung injury was lowest after PEEP of 10 cm H2O in both dependent and non-dependent lung regions, compared with PEEP of 0 or 20 cm H2O. A higher wet-to-dry-weight ratio, indicating worse lung injury, was observed with PEEP of 0 cm H2O. Histological assessment suggested that lung injury was minimised with PEEP of 10 cm H2O. CONCLUSIONS: During near-apnoeic ventilation and ECMO in experimental severe ARDS, 10 cm H2O PEEP minimised lung injury and improved gas exchange without compromising haemodynamic stability.

Current concepts in acute liver failure.

Acute liver failure (ALF) is a severe condition secondary to a myriad of causes associated with poor outcomes. The prompt diagnosis and identification of the aetiology allow the administration of specific treatments plus supportive strategies and to define the overall prognosis, the probability of developing complications and the need for liver transplantation. Pivotal issues are adequate monitoring and the institution of prophylactic strategies to reduce the risk of complications, such as progressive liver failure, cerebral oedema, renal failure, coagulopathies or infections. In this article, we review the main aspects of ALF, including the definition, diagnosis and complications. Also, we describe the standard-of-care strategies and recent advances in the treatment of ALF. Finally, we include our experience of care patients with ALF.

Role of astrocyte connexin hemichannels in cortical spreading depression.

Cortical spreading depression (CSD) is an intriguing phenomenon consisting of massive slow brain depolarizations that affects neurons and glial cells. It has been recognized since 1944, but its pathogenesis has only been uncovered during the last decade. Acute brain injuries can be further complicated by CSD in >50% of severe cases. This phenomenon is repetitive and produces a metabolic overload that increments secondary damage. Propagation of CSD is known to be linked to excitotoxicity, but the mechanisms associated with its initiation remain less understood. It has been shown that CSD can be initiated by increases in extracellular [K+] ([K+]e), and animal models use high [K+]e to promote CSD. Connexin hemichannel activity increases due to high [K+]e and low extracellular [Ca2+], conditions that occur after brain injury. Moreover, glial cell gap junction channels are fundamental in controlling extracellular medium composition, particularly in maintaining normal extracellular glutamate and K+ concentrations through "spatial buffering". However, the role of astrocytic gap junctions under tissue stress can change to damage spread in the acute damage zone whereas the reduced communication in adjacent zone would reduce cell dead propagation. Here, we review the main findings associated with CSD, and discuss the possible involvement of astrocytic connexin-based channels in secondary damage propagation. This article is part of a Special Issue entitled: Gap Junction Proteins edited by Jean Claude Herve.

Age-related NMDA signaling alterations in SOD2 deficient mice.

Oxidative stress affects the survival and function of neurons. Hence, they have a complex and highly regulated machinery to handle oxidative changes. The dysregulation of this antioxidant machinery is associated with a wide range of neurodegenerative conditions. Therefore, we evaluated signaling alterations, synaptic properties and behavioral performance in 2 and 6-month-old heterozygous manganese superoxide dismutase knockout mice (SOD2+/- mice). We found that their low antioxidant capacity generated direct oxidative damage in proteins, lipids, and DNA. However, only 6-month-old heterozygous knockout mice presented behavioral impairments. On the other hand, synaptic plasticity, synaptic strength and NMDA receptor (NMDAR) dependent postsynaptic potentials were decreased in an age-dependent manner. We also analyzed the phosphorylation state of the NMDAR subunit GluN2B. We found that while the levels of GluN2B phosphorylated on tyrosine 1472 (synaptic form) remain unchanged, we detected increased levels of GluN2B phosphorylated on tyrosine 1336 (extrasynaptic form), establishing alterations in the synaptic/extrasynaptic ratio of GluN2B. Additionally, we found increased levels of two phosphatases associated with dephosphorylation of p-1472: striatal-enriched protein tyrosine phosphatase (STEP) and phosphatase and tensin homolog deleted on chromosome Ten (PTEN). Moreover, we found decreased levels of p-CREB, a master transcription factor activated by synaptic stimulation. In summary, we describe mechanisms by which glutamatergic synapses are altered under oxidative stress conditions. Our results uncovered new putative therapeutic targets for conditions where NMDAR downstream signaling is altered. This work also contributes to our understanding of processes such as synapse formation, learning, and memory in neuropathological conditions.

Connexin43 hemichannels mediate secondary cellular damage spread from the trauma zone to distal zones in astrocyte monolayers.

The mechanism of secondary damage spread after brain trauma remains unsolved. In this work, we redirected the attention to astrocytic communication pathways. Using an in vitro trauma model that consists of a scratch injury applied to an astrocyte monolayer, we found a significant and transient induction of connexin43 (Cx43) hemichannel activity in regions distal from the injury, which was maximal ∼1 h after scratch. Two connexin hemichannel blockers, La(3+) and the peptide Gap26, abolished the increased activity, which was also absent in Cx43 KO astrocytes. In addition, the scratch-induced increase of hemichannel activity was prevented by inhibition of P2 purinergic receptors. Changes in hemichannel activity took place with a particular spatial distribution, with cells located at ∼17 mm away from the scratch presenting the highest activity (dye uptake). In contrast, the functional state of gap junction channels (dye coupling) was not significantly affected. Cx43 hemichannel activity was also enhanced by the acute extracellular application of 60 mM K(+) . The increase in hemichannel activity was associated with an increment in apoptotic cells at 24 h after scratch that was totally prevented by Gap26 peptide. These findings suggest that Cx43 hemichannels could be a new approach to prevent or reduce the secondary cell damage of brain trauma.

Moderate Traumatic Brain Injury in Adult Population: The Latin American Brain Injury Consortium Consensus for Definition and Categorization.

Moderate traumatic brain injury (TBI) is a diagnosis that describes diverse patients with heterogeneity of primary injuries. Defined by a Glasgow Coma Scale between 9 and 12, this category includes patients who may neurologically worsen and require increasing intensive care resources and/or emergency neurosurgery. Despite the unique characteristics of these patients, there have not been specific guidelines published before this effort to support decision-making in these patients. A Delphi consensus group from the Latin American Brain Injury Consortium was established to generate recommendations related to the definition and categorization of moderate TBI. Before an in-person meeting, a systematic review of the literature was performed identifying evidence relevant to planned topics. Blinded voting assessed support for each recommendation. A priori the threshold for consensus was set at 80% agreement. Nine PICOT questions were generated by the panel, including definition, categorization, grouping, and diagnosis of moderate TBI. Here, we report the results of our work including relevant consensus statements and discussion for each question. Moderate TBI is an entity for which there is little published evidence available supporting definition, diagnosis, and management. Recommendations based on experts' opinion were informed by available evidence and aim to refine the definition and categorization of moderate TBI. Further studies evaluating the impact of these recommendations will be required.

Physiological effects of high-flow nasal cannula oxygen therapy after extubation: a randomized crossover study.

BACKGROUND: Prophylactic high-flow nasal cannula (HFNC) oxygen therapy can decrease the risk of extubation failure. It is frequently used in the postextubation phase alone or in combination with noninvasive ventilation. However, its physiological effects in this setting have not been thoroughly investigated. The aim of this study was to determine comprehensively the effects of HFNC applied after extubation on respiratory effort, diaphragm activity, gas exchange, ventilation distribution, and cardiovascular biomarkers. METHODS: This was a prospective randomized crossover physiological study in critically ill patients comparing 1 h of HFNC versus 1 h of standard oxygen after extubation. The main inclusion criteria were mechanical ventilation for at least 48 h due to acute respiratory failure, and extubation after a successful spontaneous breathing trial (SBT). We measured respiratory effort through esophageal/transdiaphragmatic pressures, and diaphragm electrical activity (ΔEAdi). Lung volumes and ventilation distribution were estimated by electrical impedance tomography. Arterial and central venous blood gases were analyzed, as well as cardiac stress biomarkers. RESULTS: We enrolled 22 patients (age 59 ± 17 years; 9 women) who had been intubated for 8 ± 6 days before extubation. Respiratory effort was significantly lower with HFNC than with standard oxygen therapy, as evidenced by esophageal pressure swings (5.3 [4.2-7.1] vs. 7.2 [5.6-10.3] cmH2O; p < 0.001), pressure-time product (85 [67-140] vs. 156 [114-238] cmH2O*s/min; p < 0.001) and ΔEAdi (10 [7-13] vs. 14 [9-16] µV; p = 0.022). In addition, HFNC induced increases in end-expiratory lung volume and PaO2/FiO2 ratio, decreases in respiratory rate and ventilatory ratio, while no changes were observed in systemic hemodynamics, Troponin T, or in amino-terminal pro-B-type natriuretic peptide. CONCLUSIONS: Prophylactic application of HFNC after extubation provides substantial respiratory support and unloads respiratory muscles. Trial registration January 15, 2021. NCT04711759.

Connexin- and pannexin-based channels in normal skeletal muscles and their possible role in muscle atrophy.

Precursor cells of skeletal muscles express connexins 39, 43 and 45 and pannexin1. In these cells, most connexins form two types of membrane channels, gap junction channels and hemichannels, whereas pannexin1 forms only hemichannels. All these channels are low-resistance pathways permeable to ions and small molecules that coordinate developmental events. During late stages of skeletal muscle differentiation, myofibers become innervated and stop expressing connexins but still express pannexin1 hemichannels that are potential pathways for the ATP release required for potentiation of the contraction response. Adult injured muscles undergo regeneration, and connexins are reexpressed and form membrane channels. In vivo, connexin reexpression occurs in undifferentiated cells that form new myofibers, favoring the healing process of injured muscle. However, differentiated myofibers maintained in culture for 48 h or treated with proinflammatory cytokines for less than 3 h also reexpress connexins and only form functional hemichannels at the cell surface. We propose that opening of these hemichannels contributes to drastic changes in electrochemical gradients, including reduction of membrane potential, increases in intracellular free Ca(2+) concentration and release of diverse metabolites (e.g., NAD(+) and ATP) to the extracellular milieu, contributing to multiple metabolic and physiologic alterations that characterize muscles undergoing atrophy in several acquired and genetic human diseases. Consequently, inhibition of connexin hemichannels expressed by injured or denervated skeletal muscles might reduce or prevent deleterious changes triggered by conditions that promote muscle atrophy.

[Prolonged hypothermia in refractory intracranial hypertension. Report of one case]. / Hipotermia intravascular prolongada en un paciente con hipertensión endocraneana refractaria.

The use of hypothermia after cardiac arrest caused by ventricular fibrillation is a standard clinical practice, however its use for neuroprotection has been extended to other conditions. We report a 23-year-old male with intracranial hypertension secondary to a parenchymal hematoma associated to acute hydrocephalus. An arterial malformation was found and embolized. Due to persistent intracranial hypertension, moderate hypothermia with a target temperature of 33°C was started. After 12 hours of hypothermia, intracranial pressure was controlled. After 13 days of hypothermia a definitive control of intracranial pressure was achieved. The patient was discharged 40 days after admission, remains with a mild hemiparesia and is reassuming his university studies.

Spontaneous breathing promotes lung injury in an experimental model of alveolar collapse.

Vigorous spontaneous breathing has emerged as a promotor of lung damage in acute lung injury, an entity known as "patient self-inflicted lung injury". Mechanical ventilation may prevent this second injury by decreasing intrathoracic pressure swings and improving regional air distribution. Therefore, we aimed to determine the effects of spontaneous breathing during the early stage of acute respiratory failure on lung injury and determine whether early and late controlled mechanical ventilation may avoid or revert these harmful effects. A model of partial surfactant depletion and lung collapse was induced in eighteen intubated pigs of 32 ±4 kg. Then, animals were randomized to (1) SB-group: spontaneous breathing with very low levels of pressure support for the whole experiment (eight hours), (2) Early MV-group: controlled mechanical ventilation for eight hours, or (3) Late MV-group: first half of the experiment on spontaneous breathing (four hours) and the second half on controlled mechanical ventilation (four hours). Respiratory, hemodynamic, and electric impedance tomography data were collected. After the protocol, animals were euthanized, and lungs were extracted for histologic tissue analysis and cytokines quantification. SB-group presented larger esophageal pressure swings, progressive hypoxemia, lung injury, and more dorsal and inhomogeneous ventilation compared to the early MV-group. In the late MV-group switch to controlled mechanical ventilation improved the lung inhomogeneity and esophageal pressure swings but failed to prevent hypoxemia and lung injury. In a lung collapse model, spontaneous breathing is associated to large esophageal pressure swings and lung inhomogeneity, resulting in progressive hypoxemia and lung injury. Mechanical ventilation prevents these mechanisms of patient self-inflicted lung injury if applied early, before spontaneous breathing occurs, but not when applied late.

Cardiac function in critically ill patients with severe COVID: A prospective cross-sectional study in mechanically ventilated patients.

PURPOSE: To evaluate cardiac function in mechanically ventilated patients with COVID-19. MATERIALS AND METHODS: Prospective, cross-sectional multicenter study in four university-affiliated hospitals in Chile. All consecutive patients with COVID-19 ARDS requiring mechanical ventilation admitted between April and July 2020 were included. We performed systematic transthoracic echocardiography assessing right and left ventricular function within 24 h of intubation. RESULTS: 140 patients aged 57 ± 11, 29% female were included. Cardiac output was 5.1 L/min [IQR 4.5-6.2] and 86% of the patients required norepinephrine. ICU mortality was 29% (40 patients). Fifty-four patients (39%) exhibited right ventricle dilation out of whom 20 patients (14%) exhibited acute cor pulmonale (ACP). Eight out of the twenty patients with ACP exhibited pulmonary embolism (40%). Thirteen patients (9%) exhibited left ventricular systolic dysfunction (ejection fraction <45%). In the multivariate analysis acute cor pulmonale and PaO2/FiO2 ratio were independent predictors of ICU mortality. CONCLUSIONS: Right ventricular dilation is highly prevalent in mechanically ventilated patients with COVID-19 ARDS. Acute cor pulmonale was associated with reduced pulmonary function and, in only 40% of patients, with co-existing pulmonary embolism. Acute cor pulmonale is an independent risk factor for ICU mortality.

The effects of arterial CO2 on the injured brain: Two faces of the same coin.

Serum levels of carbon dioxide (CO2) closely regulate cerebral blood flow (CBF) and actively participate in different aspects of brain physiology such as hemodynamics, oxygenation, and metabolism. Fluctuations in the partial pressure of arterial CO2 (PaCO2) modify the aforementioned variables, and at the same time influence physiologic parameters in organs such as the lungs, heart, kidneys, and the gastrointestinal tract. In general, during acute brain injury (ABI), maintaining normal PaCO2 is the target to be achieved. Both hypercapnia and hypocapnia may comprise secondary insults and should be avoided during ABI. The risks of hypocapnia mostly outweigh the potential benefits. Therefore, its therapeutic applicability is limited to transient and second-stage control of intracranial hypertension. On the other hand, inducing hypercapnia could be beneficial when certain specific situations require increasing CBF. The evidence supporting this claim is very weak. This review attempts providing an update on the physiology of CO2, its risks, benefits, and potential utility in the neurocritical care setting.

Pannexin-1 channel opening is critical for COVID-19 pathogenesis.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) rapidly rampaged worldwide, causing a pandemic of coronavirus disease (COVID -19), but the biology of SARS-CoV-2 remains under investigation. We demonstrate that both SARS-CoV-2 spike protein and human coronavirus 229E (hCoV-229E) or its purified S protein, one of the main viruses responsible for the common cold, induce the transient opening of Pannexin-1 (Panx-1) channels in human lung epithelial cells. However, the Panx-1 channel opening induced by SARS-CoV-2 is greater and more prolonged than hCoV-229E/S protein, resulting in an enhanced ATP, PGE2, and IL-1ß release. Analysis of lung lavages and tissues indicate that Panx-1 mRNA expression is associated with increased ATP, PGE2, and IL-1ß levels. Panx-1 channel opening induced by SARS-CoV-2 spike protein is angiotensin-converting enzyme 2 (ACE-2), endocytosis, and furin dependent. Overall, we demonstrated that Panx-1 channel is a critical contributor to SARS-CoV-2 infection and should be considered as an alternative therapy.

Impact of emergency intubation on central venous oxygen saturation in critically ill patients: a multicenter observational study.

INTRODUCTION: Central venous oxygen saturation (ScvO2) has emerged as an important resuscitation goal for critically ill patients. Nevertheless, growing concerns about its limitations as a perfusion parameter have been expressed recently, including the uncommon finding of low ScvO2 values in patients in the intensive care unit (ICU). Emergency intubation may induce strong and eventually divergent effects on the physiologic determinants of oxygen transport (DO2) and oxygen consumption (VO2) and, thus, on ScvO2. Therefore, we conducted a study to determine the impact of emergency intubation on ScvO2. METHODS: In this prospective multicenter observational study, we included 103 septic and non-septic patients with a central venous catheter in place and in whom emergency intubation was required. A common intubation protocol was used and we evaluated several parameters including ScvO2 before and 15 minutes after emergency intubation. Statistical analysis included chi-square test and t test. RESULTS: ScvO2 increased from 61.8 +/- 12.6% to 68.9 +/- 12.2%, with no difference between septic and non-septic patients. ScvO2 increased in 84 patients (81.6%) without correlation to changes in arterial oxygen saturation (SaO2). Seventy eight (75.7%) patients were intubated with ScvO2 less than 70% and 21 (26.9%) normalized the parameter after the intervention. Only patients with pre-intubation ScvO2 more than 70% failed to increase the parameter after intubation. CONCLUSIONS: ScvO2 increases significantly in response to emergency intubation in the majority of septic and non-septic patients. When interpreting ScvO2 during early resuscitation, it is crucial to consider whether the patient has been recently intubated or is spontaneously breathing.

Therapeutic hypothermia for acute brain injuries.

Therapeutic hypothermia, recently termed target temperature management (TTM), is the cornerstone of neuroprotective strategy. Dating to the pioneer works of Fay, nearly 75 years of basic and clinical evidence support its therapeutic value. Although hypothermia decreases the metabolic rate to restore the supply and demand of O2, it has other tissue-specific effects, such as decreasing excitotoxicity, limiting inflammation, preventing ATP depletion, reducing free radical production and also intracellular calcium overload to avoid apoptosis. Currently, mild hypothermia (33°C) has become a standard in post-resuscitative care and perinatal asphyxia. However, evidence indicates that hypothermia could be useful in neurologic injuries, such as stroke, subarachnoid hemorrhage and traumatic brain injury. In this review, we discuss the basic and clinical evidence supporting the use of TTM in critical care for acute brain injury that extends beyond care after cardiac arrest, such as for ischemic and hemorrhagic strokes, subarachnoid hemorrhage, and traumatic brain injury. We review the historical perspectives of TTM, provide an overview of the techniques and protocols and the pathophysiologic consequences of hypothermia. In addition, we include our experience of managing patients with acute brain injuries treated using endovascular hypothermia.

Respiratory responses to pH in the absence of pontine and dorsal medullary areas in the newborn mouse in vitro.

The contribution of pons and dorsal medulla in establishing the pattern of fictive respiration and in mediating the respiratory response to acidification was studied using the isolated brainstem-spinal cord preparation from neonatal mouse. About 40% of ponto-medullary preparations (retaining pons) showed spontaneous, but irregular respiratory-like rhythm. In the other 60%, the elimination of the pons often was followed by the initiation of a respiratory-like rhythm. Medullary preparations, derived from either inactive or rhythmic ponto-medullary preparations, showed a regular respiratory-like rhythm, which was also of a higher frequency and a bigger amplitude than that observed in ponto-medullary preparations. In contrast, ventral medullary preparations, derived from medullary preparations by eliminating the dorsal medulla, showed an irregular rhythm with a reduced amplitude of the integrated inspiratory burst. In ponto-medullary and ventral medullary preparations, acidification of the superfusion medium increased the respiratory frequency, while in medullary preparations, it increased the frequency and reduced the amplitude of the inspiratory burst. Our results suggest that pontine structures influence negatively the rate and depth of the respiratory-like rhythm, while dorsal medullary structures influence positively the depth of the rhythm. They also suggest that the pattern of response to pH supported by the ventral medulla is modified by the input provided from pons and dorsal medulla.