Long-term safety of brazikumab in the open-label period of a randomized phase 2a study of patients with Crohn's disease.

BACKGROUND: Short-term efficacy and safety of brazikumab (MEDI2070), a human monoclonal antibody and anti-p19 subunit inhibitor of interleukin-23, was demonstrated in a phase 2a trial in patients with moderate-to-severe active Crohn's disease (CD). We report brazikumab long-term safety and tolerability from the open-label period of this phase 2a study. METHODS: Patients who completed the 12-week, double-blind induction period were eligible for inclusion in an open-label period where all patients received subcutaneous brazikumab (210 mg) every 4 weeks for 100 weeks. Patients had moderate-to-severe active CD and had failed or were intolerant to ≥ 1 anti-tumour necrosis factor alpha (TNFα) agent. Safety assessments included treatment-emergent adverse events (TEAEs); further assessments were pharmacokinetics and immunogenicity. RESULTS: Of the 104 patients who entered the open-label period, 57 (54.8%) continued to the end of the open-label period and 47 (45.2%) discontinued brazikumab. The most common reasons for discontinuation were lack of response (14.4%), patient decision (12.5%), and TEAEs (11.5%). In total, 44 (84.6%) in the group switching from placebo to brazikumab (placebo/brazikumab) and 43 (82.7%) in the group continuing brazikumab (brazikumab/brazikumab) experienced 1 or more TEAEs. Most TEAEs were mild-to-moderate in severity. Common TEAEs included nasopharyngitis and headache. Numbers of treatment-emergent serious adverse events (TESAEs) were similar between groups. Infections occurred in 40.4% of patients in the placebo/brazikumab group and 50% in the brazikumab/brazikumab group. There were 5 TESAEs of infection, none of which were opportunistic. No major adverse cardiac events, malignancies, or deaths were reported. CONCLUSIONS: Brazikumab was well tolerated with an acceptable safety profile over a 100-week period in patients with moderate-to-severe active CD who failed or were intolerant to 1 or more anti-TNFα agents. TRIAL REGISTRATION: NCT01714726; registered October 26, 2012.

Ion concentrations in cerebrospinal fluid in wakefulness, sleep and sleep deprivation in healthy humans.

Sleep is controlled by a circadian rhythmicity, via a reduction of arousal-promoting neuromodulatory activity, and by accumulation of somnogenic factors in the interstitial fluid of the brain. Recent experiments in mice suggest that a reduced neuronal excitability caused by a reduced concentration of potassium in the brain, concomitant with an increased concentration of calcium and magnesium, constitutes an important mediator of sleep. In the present study, we examined whether such changes in ion concentrations could be detected in the cerebrospinal fluid of healthy humans. Each subject underwent cerebrospinal fluid collection at three occasions in a randomized order: at 15:00 hours-17:00 hours during waking, at 06:00 hours-07:00 hours immediately following 1 night of sleep, and at 06:00 hours-07:00 hours following 1 night of sleep deprivation. When compared with wakefulness, both sleep and sleep deprivation produced the same effect of a small (0.1 mm, about 3%), but robust and highly significant, reduction in potassium concentration. Calcium and magnesium concentrations were unchanged. Our results support a circadian modulation of neuronal excitability in the brain mediated via changes of the interstitial potassium concentration.

Alteration of NMDA receptor trafficking as a cellular hallmark of psychosis.

A dysfunction of the glutamatergic transmission, especially of the NMDA receptor (NMDAR), constitutes one of the main biological substrate of psychotic disorders, such as schizophrenia. The NMDAR signaling hypofunction, through genetic and/or environmental insults, would cause a neurodevelopmental myriad of molecular, cellular, and network alterations that persist throughout life. Yet, the mechanisms underpinning NMDAR dysfunctions remain elusive. Here, we compared the membrane trafficking of NMDAR in three gold-standard models of schizophrenia, i.e., patient's cerebrospinal fluids, genetic manipulations of susceptibility genes, and prenatal developmental alterations. Using a combination of single nanoparticle tracking, electrophysiological, biochemical, and behavioral approaches in rodents, we identified that the NMDAR trafficking in hippocampal neurons was consistently altered in all these different models. Artificial manipulations of the NMDAR surface dynamics with competing ligands or antibody-induced receptor cross-link in the developing rat brain were sufficient to regulate the adult acoustic startle reflex and compensate for an early pathological challenge. Collectively, we show that the NMDAR trafficking is markedly altered in all clinically relevant models of psychosis, opening new avenues of therapeutical strategies.

Differential acute impact of therapeutically effective and overdose concentrations of lithium on human neuronal single cell and network function.

Lithium salts are used as mood-balancing medication prescribed to patients suffering from neuropsychiatric disorders, such as bipolar disorder and major depressive disorder. Lithium salts cross the blood-brain barrier and reach the brain parenchyma within few hours after oral application, however, how lithium influences directly human neuronal function is unknown. We applied patch-clamp and microelectrode array technology on human induced pluripotent stem cell (iPSC)-derived cortical neurons acutely exposed to therapeutic (<1 mM) and overdose concentrations (>1 mM) of lithium chloride (LiCl) to assess how therapeutically effective and overdose concentrations of LiCl directly influence human neuronal electrophysiological function at the synapse, single-cell, and neuronal network level. We describe that human iPSC-cortical neurons exposed to lithium showed an increased neuronal activity under all tested concentrations. Furthermore, we reveal a lithium-induced, concentration-dependent, transition of regular synchronous neuronal network activity using therapeutically effective concentration (<1 mM LiCl) to epileptiform-like neuronal discharges using overdose concentration (>1 mM LiCl). The overdose concentration lithium-induced epileptiform-like activity was similar to the epileptiform-like activity caused by the GABAA-receptor antagonist. Patch-clamp recordings reveal that lithium reduces action potential threshold at all concentrations, however, only overdose concentration causes increased frequency of spontaneous AMPA-receptor mediated transmission. By applying the AMPA-receptor antagonist and anti-epileptic drug Perampanel, we demonstrate that Perampanel suppresses lithium-induced epileptiform-like activity in human cortical neurons. We provide insights in how therapeutically effective and overdose concentration of lithium directly influences human neuronal function at synapse, a single neuron, and neuronal network levels. Furthermore, we provide evidence that Perampanel suppresses pathological neuronal discharges caused by overdose concentrations of lithium in human neurons.

Human Cerebrospinal Fluid Promotes Neuronal Circuit Maturation of Human Induced Pluripotent Stem Cell-Derived 3D Neural Aggregates.

Human induced pluripotent stem cell (hiPSC)-derived in vitro neural and organoid models resemble fetal, rather than adult brain properties, indicating that currently applied cultivation media and supplements are insufficient to achieve neural maturation beyond the fetal stage. In vivo, cerebrospinal fluid molecules are regulating the transition of the immature fetal human brain into a mature adult brain. By culturing hiPSC-3D neural aggregates in human cerebrospinal fluid (hCSF) obtained from healthy adult individuals, we demonstrate that hCSF rapidly triggers neurogenesis, gliogenesis, synapse formation, neurite outgrowth, suppresses proliferation of residing neural stem cells, and results in the formation of synchronously active neuronal circuits in vitro within 3 days. Thus, a physiologically relevant and adult brain-like milieu triggers maturation of hiPSC-3D neural aggregates into highly functional neuronal circuits in vitro. The approach presented here opens a new avenue to identify novel physiological factors for the improvement of hiPSC neural in vitro models.

Robust Generation of Person-Specific, Synchronously Active Neuronal Networks Using Purely Isogenic Human iPSC-3D Neural Aggregate Cultures.

Reproducibly generating human induced pluripotent stem cell-based functional neuronal circuits, solely obtained from single individuals, poses particular challenges to achieve personalized and patient specific functional neuronal in vitro models. A hallmark of functional neuronal assemblies, synchronous neuronal activity, can be non-invasively studied by microelectrode array (MEA) technology, reliably capturing physiological and pathophysiological aspects of human brain function. In our here presented manuscript, we demonstrate a procedure to generate 3D neural aggregates comprising astrocytes, oligodendroglial cells, and neurons obtained from the same human tissue sample. Moreover, we demonstrate the robust ability of those neurons to create a highly synchronously active neuronal network within 3 weeks in vitro, without additionally applied astrocytes. The fusion of MEA-technology with functional neuronal circuits solely obtained from one individual's cells represent isogenic person-specific human neuronal sensor chips that pave the way for specific personalized in vitro neuronal networks as well as neurological and neuropsychiatric disease modeling.

Ionized calcium in human cerebrospinal fluid and its influence on intrinsic and synaptic excitability of hippocampal pyramidal neurons in the rat.

It is well-known that the extracellular concentration of calcium affects neuronal excitability and synaptic transmission. Less is known about the physiological concentration of extracellular calcium in the brain. In electrophysiological brain slice experiments, the artificial cerebrospinal fluid traditionally contains relatively high concentrations of calcium (2-4 mM) to support synaptic transmission and suppress neuronal excitability. Using an ion-selective electrode, we determined the fraction of ionized calcium in healthy human cerebrospinal fluid to 1.0 mM of a total concentration of 1.2 mM (86%). Using patch-clamp and extracellular recordings in the CA1 region in acute slices of rat hippocampus, we then compared the effects of this physiological concentration of calcium with the commonly used 2 mM on neuronal excitability, synaptic transmission, and long-term potentiation (LTP) to examine the magnitude of changes in this range of extracellular calcium. Increasing the total extracellular calcium concentration from 1.2 to 2 mM decreased spontaneous action potential firing, induced a depolarization of the threshold, and increased the rate of both de- and repolarization of the action potential. Evoked synaptic transmission was approximately doubled, with a balanced effect between inhibition and excitation. In 1.2 mM calcium high-frequency stimulation did not result in any LTP, whereas a prominent LTP was observed at 2 or 4 mM calcium. Surprisingly, this inability to induce LTP persisted during blockade of GABAergic inhibition. In conclusion, an increase from the physiological 1.2 mM to 2 mM calcium in the artificial cerebrospinal fluid has striking effects on neuronal excitability, synaptic transmission, and the induction of LTP. OPEN SCIENCE BADGES: This article has received a badge for *Open Materials* because it provided all relevant information to reproduce the study in the manuscript. The complete Open Science Disclosure form for this article can be found at the end of the article. More information about the Open Practices badges can be found at https://cos.io/our-services/open-science-badges/. Read the Editorial Highlight for this article on page 435.

Effects of physical exercise and stress on hippocampal CA1 and dentate gyrus synaptic transmission and long-term potentiation in adolescent and adult Wistar rats.

It is commonly recognized that physical exercise positively affects several CNS regions and improves cognitive abilities. For example, exercise is associated with an increase in neurogenesis and facilitation of long-term potentiation in the hippocampus. Conversely, animal models for depression are associated with a decrease in neurogenesis and a reduction of long-term potentiation in the hippocampus. Although exercise could be a viable option in the treatment of some forms of depression, the mechanisms responsible for such improvements have not been elucidated. In this study, we examine hippocampal function using electrophysiological field recordings in CA1 and dentate gyrus to study baseline synaptic transmission and long-term potentiation in adolescent and adult rats prenatally exposed to the glucocorticoid dexamethasone. One group of animals was allowed to run voluntarily for 10 or 21 days using an exercise wheel before the experiments, and the control group was prevented from running (i.e. the exercise wheel was locked). In adult saline-exposed animals, exercise was associated with increased long-term potentiation in the dentate gyrus. Unexpectedly, in dexamethasone-exposed animals, dentate gyrus long-term potentiation was facilitated, whereas long-term potentiation in CA1 was unaffected by prenatal dexamethasone or by 10 or 21 days of voluntary running. Irrespective of age, prenatal dexamethasone and running had limited effects on synaptic transmission and presynaptic release in CA1 and dentate gyrus. In summary, running facilitates dentate gyrus long-term potentiation in adult animals that resembles the effects of prenatal dexamethasone.

The novel antidepressant ketamine enhances dentate gyrus proliferation with no effects on synaptic plasticity or hippocampal function in depressive-like rats.

AIM: Major depressive disorder is a common and debilitating condition with substantial economic impact. Treatment options, although effective, are aimed at relieving the symptoms with limited disease modification. Ketamine, a commonly used anaesthetic, has received substantial attention as it shows rapid antidepressant effects clinically. We studied the effects of ketamine on hippocampal function and dentate gyrus proliferation in rats showing a depressive-like phenotype. METHODS: Adolescent and adult animals were pre-natally exposed to the glucocorticoid analog dexamethasone, and we verified a depressive-like phenotype using behavioural tests, such as the sucrose preference. We subsequently studied the effects of ketamine on hippocampal synaptic transmission, plasticity and dentate gyrus proliferation. In addition, we measured hippocampal glutamate receptor expression. We also tested the ketamine metabolite hydroxynorketamine for NMDA-receptor independent effects. RESULTS: Surprisingly, our extensive experimental survey revealed limited effects of ketamine or its metabolite on hippocampal function in control as well as depressive-like animals. We found no effects on synaptic efficacy or induction of long-term potentiation in adolescent and adult animals. Also there was no difference when comparing the dorsal and ventral hippocampus. Importantly, however, ketamine 24 hours prior to experimentation significantly increased the dentate gyrus proliferation, as revealed by Ki-67 immunostaining, in the depressive-like phenotype. CONCLUSION: We find limited effects of ketamine on hippocampal glutamatergic transmission. Instead, alterations in dentate gyrus proliferation could explain the antidepressant effects of ketamine.

Corrigendum: Human Cerebrospinal Fluid Promotes Neuronal Viability and Activity of Hippocampal Neuronal Circuits In Vitro.

[This corrects the article on p. 54 in vol. 10, PMID: 26973467.].

Human Cerebrospinal Fluid Promotes Neuronal Viability and Activity of Hippocampal Neuronal Circuits In Vitro.

For decades it has been hypothesized that molecules within the cerebrospinal fluid (CSF) diffuse into the brain parenchyma and influence the function of neurons. However, the functional consequences of CSF on neuronal circuits are largely unexplored and unknown. A major reason for this is the absence of appropriate neuronal in vitro model systems, and it is uncertain if neurons cultured in pure CSF survive and preserve electrophysiological functionality in vitro. In this article, we present an approach to address how human CSF (hCSF) influences neuronal circuits in vitro. We validate our approach by comparing the morphology, viability, and electrophysiological function of single neurons and at the network level in rat organotypic slice and primary neuronal cultures cultivated either in hCSF or in defined standard culture media. Our results demonstrate that rodent hippocampal slices and primary neurons cultured in hCSF maintain neuronal morphology and preserve synaptic transmission. Importantly, we show that hCSF increases neuronal viability and the number of electrophysiologically active neurons in comparison to the culture media. In summary, our data indicate that hCSF represents a physiological environment for neurons in vitro and a superior culture condition compared to the defined standard media. Moreover, this experimental approach paves the way to assess the functional consequences of CSF on neuronal circuits as well as suggesting a novel strategy for central nervous system (CNS) disease modeling.

The Bacterial (Vibrio alginolyticus) Production of Tetrodotoxin in the Ribbon Worm Lineus longissimus-Just a False Positive?

We test previous claims that the bacteria Vibrio alginolyticus produces tetrodotoxin (TTX) when living in symbiosis with the nemertean Lineus longissimus by a setup with bacteria cultivation for TTX production. Toxicity experiments on the shore crab, Carcinus maenas, demonstrated the presence of a paralytic toxin, but evidence from LC-MS and electrophysiological measurements of voltage-gated sodium channel-dependent nerve conductance in male Wistar rat tissue showed conclusively that this effect did not originate from TTX. However, a compound of similar molecular weight was found, albeit apparently non-toxic, and with different LC retention time and MS/MS fragmentation pattern than those of TTX. We conclude that C. maenas paralysis and death likely emanate from a compound <5 kDa, and via a different mechanism of action than that of TTX. The similarity in mass between TTX and the Vibrio-produced low-molecular-weight, non-toxic compound invokes that thorough analysis is required when assessing TTX production. Based on our findings, we suggest that re-examination of some published claims of TTX production may be warranted.

Tonically active NMDA receptors--a signalling mechanism critical for interneuronal excitability in the CA1 stratum radiatum.

In contrast to tonic extrasynaptic γ-aminobutyric acid (GABA)A receptor-mediated signalling, the physiological significance of tonic extrasynaptic N-methyl-D-aspartate (NMDA) receptor (NMDAR)-mediated signalling remains uncertain. In this study, reversible open-channel blockers of NMDARs, memantine and phencyclidine (PCP) were used as tools to examine tonic NMDAR-mediated signalling in rat hippocampal slices. Memantine in concentrations up to 10 µM had no effect on synaptically evoked NMDAR-mediated responses in pyramidal neurons or GABAergic interneurons. On the other hand, 10 µM memantine reduced tonic NMDAR-mediated currents in GABAergic interneurons by approximately 50%. These tonic NMDAR-mediated currents in interneurons contributed significantly to the excitability of the interneurons as 10 µM memantine reduced the disynaptic inhibitory postsynaptic current in pyramidal cells by about 50%. Moreover, 10 µM memantine, but also PCP in concentrations ≤ 1 µM, increased the magnitude of the population spike, likely because of disinhibition. The relatively higher impact of tonic NMDAR-mediated signalling in interneurons was at least partly explained by the expression of GluN2D-containing NMDARs, which was not observed in mature pyramidal cells. The current results are consistent with the idea that low doses of readily reversible NMDAR open-channel blockers preferentially inhibit tonically active extrasynaptic NMDARs, and they suggest that tonically active NMDARs contribute more prominently to the intrinsic excitation in GABAergic interneurons than in pyramidal cells. It is proposed that this specific difference between interneurons and pyramidal cells can explain the disinhibition caused by the Alzheimer's disease medication memantine.

Temperature acclimation rate of aerobic scope and feeding metabolism in fishes: implications in a thermally extreme future.

Temperature acclimation may offset the increased energy expenditure (standard metabolic rate, SMR) and reduced scope for activity (aerobic scope, AS) predicted to occur with local and global warming in fishes and other ectotherms. Yet, the time course and mechanisms of this process is little understood. Acclimation dynamics of SMR, maximum metabolic rate, AS and the specific dynamic action of feeding (SDA) were determined in shorthorn sculpin (Myoxocephalus scorpius) after transfer from 10°C to 16°C. SMR increased in the first week by 82% reducing AS to 55% of initial values, while peak postprandial metabolism was initially greater. This meant that the estimated AS during peak SDA approached zero, constraining digestion and leaving little room for additional aerobic processes. After eight weeks at 16°C, SMR was restored, while AS and the estimated AS during peak SDA recovered partly. Collectively, this demonstrated a considerable capacity for metabolic thermal compensation, which should be better incorporated into future models on organismal responses to climate change. A mathematical model based on the empirical data suggested that phenotypes with fast acclimation rates may be favoured by natural selection as the accumulated energetic cost of a slow acclimation rate increases in a warmer future with exacerbated thermal variations.

The peptide hormone cholecystokinin modulates the tonus and compliance of the bulbus arteriosus and pre-branchial vessels of the rainbow trout (Oncorhynchus mykiss).

The bulbus arteriosus is a compliant structure between the ventricle and ventral aorta of teleost fish. It serves as a "wind-kessel" that dampens pressure variations during the cardiac cycle allowing a continuous flow of blood into the gills. The bulbus arteriosus receives sympathetic innervation and is affected by several circulating substances, indicating neurohumoral control. We have previously shown that the peptide hormone, cholecystokinin (CCK), affects the hemodynamics of the cardiovascular system in rainbow trout (Oncorhynchus mykiss) by increasing flow pulse amplitude without affecting cardiac output. We hypothesized that this could be explained by an altered tonus or compliance/distensibility of the bulbus arteriosus. Our results show that there is a substantial effect of CCK on the bulbus arteriosus. Concentrations of CCK that altered the cardiac function of in situ perfused hearts also contracted the bulbus arteriosus in vitro. Pressure-volume curves revealed a change in both the tonus and the compliance/distensibility of this structure. Furthermore, the stimulatory (constricting) effect of CCK was also evident in the ventricle and vasculature leading to the gills, but absent in the atrium, efferent branchial arteries and dorsal aorta. In conclusion, CCK alters the mechanical properties of the ventricle, bulbus arteriosus, ventral aorta and afferent gill vasculature, thus maintaining adequate branchial and systemic blood flow and pressure when cardiorespiratory demands change, such as after feeding.

Surface dynamics of GluN2B-NMDA receptors controls plasticity of maturing glutamate synapses.

NMDA-type glutamate receptors (NMDAR) are central actors in the plasticity of excitatory synapses. During adaptive processes, the number and composition of synaptic NMDAR can be rapidly modified, as in neonatal hippocampal synapses where a switch from predominant GluN2B- to GluN2A-containing receptors is observed after the induction of long-term potentiation (LTP). However, the cellular pathways by which surface NMDAR subtypes are dynamically regulated during activity-dependent synaptic adaptations remain poorly understood. Using a combination of high-resolution single nanoparticle imaging and electrophysiology, we show here that GluN2B-NMDAR are dynamically redistributed away from glutamate synapses through increased lateral diffusion during LTP in immature neurons. Strikingly, preventing this activity-dependent GluN2B-NMDAR surface redistribution through cross-linking, either with commercial or with autoimmune anti-NMDA antibodies from patient with neuropsychiatric symptoms, affects the dynamics and spine accumulation of CaMKII and impairs LTP. Interestingly, the same impairments are observed when expressing a mutant of GluN2B-NMDAR unable to bind CaMKII. We thus uncover a non-canonical mechanism by which GluN2B-NMDAR surface dynamics plays a critical role in the plasticity of maturing synapses through a direct interplay with CaMKII.

Aerobic scope fails to explain the detrimental effects on growth resulting from warming and elevated CO2 in Atlantic halibut.

As a consequence of increasing atmospheric CO2, the world's oceans are becoming warmer and more acidic. Whilst the ecological effects of these changes are poorly understood, it has been suggested that fish performance including growth will be reduced mainly as a result of limitations in oxygen transport capacity. Contrary to the predictions given by the oxygen- and capacity-limited thermal tolerance hypothesis, we show that aerobic scope and cardiac performance of Atlantic halibut (Hippoglossus hippoglossus) increase following 14-16 weeks exposure to elevated temperatures and even more so in combination with CO2-acidified seawater. However, the increase does not translate into improved growth, demonstrating that oxygen uptake is not the limiting factor for growth performance at high temperatures. Instead, long-term exposure to CO2-acidified seawater reduces growth at temperatures that are frequently encountered by this species in nature, indicating that elevated atmospheric CO2 levels may have serious implications on fish populations in the future.

AMPA-silent synapses in brain development and pathology.

Synapses are constantly generated at a high rate in the developing, prepubescent brain. Newly generated glutamatergic synapses lack functional AMPA receptor-mediated transmission. Most of these 'AMPA-silent' synapses are eliminated during the developmental period, but some are specifically selected for AMPA unsilencing by correlated pre-and postsynaptic activity as the first step in a process that leads to stabilization of the synapse. Premature, or delayed, unsilencing of AMPA-silent synapses has been implicated in neurodevelopmental disorders, and abnormal generation of AMPA-silent synapses is associated with brain trauma, addiction and neurodegenerative disorders, further highlighting the importance of AMPA-silent synapses in brain pathology.

Metabolic scope and interspecific competition in sculpins of Greenland are influenced by increased temperatures due to climate change.

Ongoing climate change has led to an increase in sea surface temperatures of 2-4°C on the west coast of Greenland. Since fish are ectothermic, metabolic rate increases with ambient temperature. This makes these animals particularly sensitive to changes in temperature; subsequently any change may influence their metabolic scope, i.e. the physiological capacity to undertake aerobically challenging activities. Any temperature increase may thus disrupt species-specific temperature adaptations, at both the molecular level as well as in behavior, and concomitant species differences in the temperature sensitivity may shift the competitive balance among coexisting species. We investigated the influence of temperature on metabolic scope and competitive ability in three species of marine sculpin that coexist in Greenland coastal waters. Since these species have different distribution ranges, we hypothesized that there should be a difference in their physiological response to temperature; hence we compared their metabolic scope at three temperatures (4, 9 and 14°C). Their competitive ability at the ambient temperature of 9°C was also tested in an attempt to link physiological capacity with behaviour. The Arctic staghorn sculpin, the species with the northernmost distribution range, had a lower metabolic scope in the higher temperature range compared to the other two species, which had similar metabolic scope at the three temperatures. The Arctic staghorn sculpin also had reduced competitive ability at 9°C and may thus already be negatively affected by the current ocean warming. Our results suggest that climate change can have effects on fish physiology and interspecific competition, which may alter the species composition of the Arctic fish fauna.