The quality of cortical network function recovery depends on localization and degree of axonal demyelination.

Myelin loss is a severe pathological hallmark common to a number of neurodegenerative diseases, including multiple sclerosis (MS). Demyelination in the central nervous system appears in the form of lesions affecting both white and gray matter structures. The functional consequences of demyelination on neuronal network and brain function are not well understood. Current therapeutic strategies for ameliorating the course of such diseases usually focus on promoting remyelination, but the effectiveness of these approaches strongly depends on the timing in relation to the disease state. In this study, we sought to characterize the time course of sensory and behavioral alterations induced by de- and remyelination to establish a rational for the use of remyelination strategies. By taking advantage of animal models of general and focal demyelination, we tested the consequences of myelin loss on the functionality of the auditory thalamocortical system: a well-studied neuronal network consisting of both white and gray matter regions. We found that general demyelination was associated with a permanent loss of the tonotopic cortical organization in vivo, and the inability to induce tone-frequency-dependent conditioned behaviors, a status persisting after remyelination. Targeted, focal lysolecithin-induced lesions in the white matter fiber tract, but not in the gray matter regions of cortex, were fully reversible at the morphological, functional and behavioral level. These findings indicate that remyelination of white and gray matter lesions have a different functional regeneration potential, with the white matter being able to regain full functionality while cortical gray matter lesions suffer from permanently altered network function. Therefore therapeutic interventions aiming for remyelination have to consider both region- and time-dependent strategies.

14-3-3 Proteins regulate K2P 5.1 surface expression on T lymphocytes.

K2P 5.1 channels (also called TASK-2 or Kcnk5) have already been shown to be relevant in the pathophysiology of autoimmune disease because they are known to be upregulated on peripheral and central T lymphocytes of multiple sclerosis (MS) patients. Moreover, overexpression of K2P 5.1 channels in vitro provokes enhanced T-cell effector functions. However, the molecular mechanisms regulating intracellular K2P 5.1 channel trafficking are unknown so far. Thus, the aim of the study is to elucidate the trafficking of K2P 5.1 channels on T lymphocytes. Using mass spectrometry analysis, we have identified 14-3-3 proteins as novel binding partners of K2P 5.1 channels. We show that a non-classical 14-3-3 consensus motif (R-X-X-pT/S-x) at the channel's C-terminus allows the binding between K2P 5.1 and 14-3-3. The mutant K2P 5.1/S266A diminishes the protein-protein interaction and reduces the amplitude of membrane currents. Application of a non-peptidic 14-3-3 inhibitor (BV02) significantly reduces the number of wild-type channels in the plasma membrane, whereas the drug has no effect on the trafficking of the mutated channel. Furthermore, blocker application reduces T-cell effector functions. Taken together, we demonstrate that 14-3-3 interacts with K2P 5.1 and plays an important role in channel trafficking.

Human T cells in silico: Modelling their electrophysiological behaviour in health and disease.

Although various types of ion channels are known to have an impact on human T cell effector functions, their exact mechanisms of influence are still poorly understood. The patch clamp technique is a well-established method for the investigation of ion channels in neurons and T cells. However, small cell sizes and limited selectivity of pharmacological blockers restrict the value of this experimental approach. Building a realistic T cell computer model therefore can help to overcome these kinds of limitations as well as reduce the overall experimental effort. The computer model introduced here was fed off ion channel parameters from literature and new experimental data. It is capable of simulating the electrophysiological behaviour of resting and activated human CD4(+) T cells under basal conditions and during extracellular acidification. The latter allows for the very first time to assess the electrophysiological consequences of tissue acidosis accompanying most forms of inflammation.

Sodium chloride promotes pro-inflammatory macrophage polarization thereby aggravating CNS autoimmunity.

The increasing incidence in Multiple Sclerosis (MS) during the last decades in industrialized countries might be linked to a change in dietary habits. Nowadays, enhanced salt content is an important characteristic of Western diet and increased dietary salt (NaCl) intake promotes pathogenic T cell responses contributing to central nervous system (CNS) autoimmunity. Given the importance of macrophage responses for CNS disease propagation, we addressed the influence of salt consumption on macrophage responses in CNS autoimmunity. We observed that EAE-diseased mice receiving a NaCl-high diet showed strongly enhanced macrophage infiltration and activation within the CNS accompanied by disease aggravation during the effector phase of EAE. NaCl treatment of macrophages elicited a strong pro-inflammatory phenotype characterized by enhanced pro-inflammatory cytokine production, increased expression of immune-stimulatory molecules, and an antigen-independent boost of T cell proliferation. This NaCl-induced pro-inflammatory macrophage phenotype was accompanied by increased activation of NF-kB and MAPK signaling pathways. The pathogenic relevance of NaCl-conditioned macrophages is illustrated by the finding that transfer into EAE-diseased animals resulted in significant disease aggravation compared to untreated macrophages. Importantly, also in human monocytes, NaCl promoted a pro-inflammatory phenotype that enhanced human T cell proliferation. Taken together, high dietary salt intake promotes pro-inflammatory macrophages that aggravate CNS autoimmunity. Together with other studies, these results underline the need to further determine the relevance of increased dietary salt intake for MS disease severity.

CD8(+) T Cell-Mediated Neuronal Dysfunction and Degeneration in Limbic Encephalitis.

Autoimmune inflammation of the limbic gray matter structures of the human brain has recently been identified as major cause of mesial temporal lobe epilepsy with interictal temporal epileptiform activity and slowing of the electroencephalogram, progressive memory disturbances, as well as a variety of other behavioral, emotional, and cognitive changes. Magnetic resonance imaging exhibits volume and signal changes of the amygdala and hippocampus, and specific anti-neuronal antibodies binding to either intracellular or plasma membrane neuronal antigens can be detected in serum and cerebrospinal fluid. While effects of plasma cell-derived antibodies on neuronal function and integrity are increasingly becoming characterized, potentially contributing effects of T cell-mediated immune mechanisms remain poorly understood. CD8(+) T cells are known to directly interact with major histocompatibility complex class I-expressing neurons in an antigen-specific manner. Here, we summarize current knowledge on how such direct CD8(+) T cell-neuron interactions may impact neuronal excitability, plasticity, and integrity on a single cell and network level and provide an overview on methods to further corroborate the in vivo relevance of these mechanisms mainly obtained from in vitro studies.

The two-pore domain K2 P channel TASK2 drives human NK-cell proliferation and cytolytic function.

Natural killer (NK) cells are a subset of cytotoxic lymphocytes that recognize and kill tumor- and virus-infected cells without prior stimulation. Killing of target cells is a multistep process including adhesion to target cells, formation of an immunological synapse, and polarization and release of cytolytic granules. The role of distinct potassium channels in this orchestrated process is still poorly understood. The current study reveals that in addition to the voltage-gated KV 1.3 and the calcium-activated KCa 3.1 channels, human NK cells also express the two-pore domain K2 P channel TASK2 (TWIK-related acid-sensitive potassium channel). Expression of Task2 varies among NK-cell subsets and depends on their differentiation and activation state. Despite its different expression in TASK2(high) CD56(bright) CD16(-) and TASK2(low) CD56(dim) CD16(+) NK cells, TASK2 is involved in cytokine-induced proliferation and cytolytic function of both subsets. TASK2 is crucial for leukocyte functional antigen (LFA-1) mediated adhesion of both resting and cytokine-activated NK cells to target cells, an early step in killing of target cells. With regard to the following mechanism, TASK2 plays a role in release of cytotoxic granules by resting, but not IL-15-induced NK cells. Taken together, our data exhibit two-pore potassium channels as important players in NK-cell activation and effector function.

Distinct Neurodegenerative Changes in an Induced Pluripotent Stem Cell Model of Frontotemporal Dementia Linked to Mutant TAU Protein.

Frontotemporal dementia (FTD) is a frequent form of early-onset dementia and can be caused by mutations in MAPT encoding the microtubule-associated protein TAU. Because of limited availability of neural cells from patients' brains, the underlying mechanisms of neurodegeneration in FTD are poorly understood. Here, we derived induced pluripotent stem cells (iPSCs) from individuals with FTD-associated MAPT mutations and differentiated them into mature neurons. Patient iPSC-derived neurons demonstrated pronounced TAU pathology with increased fragmentation and phospho-TAU immunoreactivity, decreased neurite extension, and increased but reversible oxidative stress response to inhibition of mitochondrial respiration. Furthermore, FTD neurons showed an activation of the unfolded protein response, and a transcriptome analysis demonstrated distinct, disease-associated gene expression profiles. These findings indicate distinct neurodegenerative changes in FTD caused by mutant TAU and highlight the unique opportunity to use neurons differentiated from patient-specific iPSCs to identify potential targets for drug screening purposes and therapeutic intervention.

Differential phospholipase C-dependent modulation of TASK and TREK two-pore domain K+ channels in rat thalamocortical relay neurons.

KEY POINTS: During the behavioural states of sleep and wakefulness thalamocortical relay neurons fire action potentials in high frequency bursts or tonic sequences, respectively. The modulation of specific K(+) channel types, termed TASK and TREK, allows these neurons to switch between the two modes of activity. In this study we show that the signalling lipids phosphatidylinositol 4,5-bisphosphate (PIP2) and diacylglycerol (DAG), which are components of their membrane environment, switch on and shut off TREK and TASK channels, respectively. These channel modulations contribute to a better understanding of the molecular basis of the effects of neurotransmitters such as ACh which are released by the brainstem arousal system. The present report introduces PIP2 and DAG as new elements of signal transduction in the thalamus. The activity of two-pore domain potassium channels (K2P ) regulates the excitability and firing modes of thalamocortical (TC) neurons. In particular, the inhibition of two-pore domain weakly inwardly rectifying K(+) channel (TWIK)-related acid-sensitive K(+) (TASK) channels and TWIK-related K(+) (TREK) channels, as a consequence of the stimulation of muscarinic ACh receptors (MAChRs) which are coupled to phosphoinositide-specific phospholipase C (PLCß), induces a shift from burst to tonic firing. By using a whole cell patch-clamp approach, the contribution of the membrane-bound second messenger molecules phosphatidylinositol 4,5-bisphosphate (PIP2 ) and diacylglycerol (DAG) acting downstream of PLCß was probed. The standing outward current (ISO ) was used to monitor the current through TASK and TREK channels in TC neurons. By exploiting different manoeuvres to change the intracellular PIP2 level in TC neurons, we here show that the scavenging of PIP2 (by neomycin) results in an increased muscarinic effect on ISO whereas increased availability of PIP2 (inclusion to the patch pipette; histone-based carrier) decreased muscarinic signalling. The degree of muscarinic inhibition specifically depends on phosphatidylinositol phosphate (PIP) and PIP2 but no other phospholipids (phosphatidic acid, phosphatidylserine). The use of specific blockers revealed that PIP2 is targeting TREK but not TASK channels. Furthermore, we demonstrate that the inhibition of TASK channels is induced by the application of the DAG analogue 1-oleoyl-2-acetyl-sn-glycerol (OAG). Under current clamp conditions the activation of MAChRs and PLCß as well as the application of OAG resulted in membrane depolarization, while PIP2 application via histone carrier induced a hyperpolarization. These results demonstrate a differential role of PIP2 and DAG in K2P channel modulation in native neurons which allows a fine-tuned inhibition of TREK (via PIP2 depletion) and TASK (via DAG) channels following MAChR stimulation.

First report on an inotropic peptide activating tetrodotoxin-sensitive, "neuronal" sodium currents in the heart.

BACKGROUND: New therapeutic approaches to improve cardiac contractility without severe risk would improve the management of acute heart failure. Increasing systolic sodium influx can increase cardiac contractility, but most sodium channel activators have proarrhythmic effects that limit their clinical use. Here, we report the cardiac effects of a novel positive inotropic peptide isolated from the toxin of the Black Judean scorpion that activates neuronal tetrodotoxin-sensitive sodium channels. METHODS AND RESULTS: All venoms and peptides were isolated from Black Judean Scorpions (Buthotus Hottentotta) caught in the Judean Desert. The full scorpion venom increased left ventricular function in sedated mice in vivo, prolonged ventricular repolarization, and provoked ventricular arrhythmias. An inotropic peptide (BjIP) isolated from the full venom by chromatography increased cardiac contractility but did neither provoke ventricular arrhythmias nor prolong cardiac repolarization. BjIP increased intracellular calcium in ventricular cardiomyocytes and prolonged inactivation of the cardiac sodium current. Low concentrations of tetrodotoxin (200 nmol/L) abolished the effect of BjIP on calcium transients and sodium current. BjIP did not alter the function of Nav1.5, but selectively activated the brain-type sodium channels Nav1.6 or Nav1.3 in cellular electrophysiological recordings obtained from rodent thalamic slices. Nav1.3 (SCN3A) mRNA was detected in human and mouse heart tissue. CONCLUSIONS: Our pilot experiments suggest that selective activation of tetrodotoxin-sensitive neuronal sodium channels can safely increase cardiac contractility. As such, the peptide described here may become a lead compound for a new class of positive inotropic agents.

The CNS under pathophysiologic attack--examining the role of K2p channels.

Members of the two-pore domain K(+) channel (K2P) family are increasingly recognized as being potential targets for therapeutic drugs and could play a role in the diagnosis and treatment of neurologic disorders. Their broad and diverse expression pattern in pleiotropic cell types, importance in cellular function, unique biophysical properties, and sensitivity toward pathophysiologic parameters represent the basis for their involvement in disorders of the central nervous system (CNS). This review will focus on multiple sclerosis (MS) and stroke, as there is growing evidence for the involvement of K2P channels in these two major CNS disorders. In MS, TASK1-3 channels are expressed on T lymphocytes and are part of a signaling network regulating Ca(2+)- dependent pathways that are mandatory for T cell activation, differentiation, and effector functions. In addition, TASK1 channels are involved in neurodegeneration, resulting in autoimmune attack of CNS cells. On the blood-brain barrier, TREK1 channels regulate immune cell trafficking under autoinflammatory conditions. Cerebral ischemia shares some pathophysiologic similarities with MS, including hypoxia and extracellular acidosis. On a cellular level, K2P channels can have both proapoptotic and antiapoptotic effects, either promoting neurodegeneration or protecting neurons from ischemic cell death. TASK1 and TREK1 channels have a neuroprotective effect on stroke development, whereas TASK2 channels have a detrimental effect on neuronal survival under ischemic conditions. Future research in preclinical models is needed to provide a more detailed understanding of the contribution of K2P channel family members to neurologic disorders, before translation to the clinic is an option.

The role of two-pore-domain background K⁺ (K2p) channels in the thalamus.

The thalamocortical system is characterized by two fundamentally different activity states, namely synchronized burst firing and tonic action potential generation, which mainly occur during the behavioral states of sleep and wakefulness, respectively. The switch between the two firing modes is crucially governed by the bidirectional modulation of members of the K2P channel family, namely tandem of P domains in a weakly inward rectifying K(+) (TWIK)-related acid-sensitive K(+) (TASK) and TWIK-related K(+) (TREK) channels, in thalamocortical relay (TC) neurons. Several physicochemical stimuli including neurotransmitters, protons, di- and multivalent cations as well as clinically used drugs have been shown to modulate K2P channels in these cells. With respect to modulation of these channels by G-protein-coupled receptors, PLCß plays a unique role with both substrate breakdown and product synthesis exerting important functions. While the degradation of PIP2 leads to the closure of TREK channels, the production of DAG induces the inhibition of TASK channels. Therefore, TASK and TREK channels were found to be central elements in the control of thalamic activity modes. Since research has yet focused on identifying the muscarinic pathway underling the modulation of TASK and TREK channels in TC neurons, future studies should address other thalamic cell types and members of the K2P channel family.

Regionally specific expression of high-voltage-activated calcium channels in thalamic nuclei of epileptic and non-epileptic rats.

The polygenic origin of generalized absence epilepsy results in dysfunction of ion channels that allows the switch from physiological asynchronous to pathophysiological highly synchronous network activity. Evidence from rat and mouse models of absence epilepsy indicates that altered Ca(2+) channel activity contributes to cellular and network alterations that lead to seizure activity. Under physiological circumstances, high voltage-activated (HVA) Ca(2+) channels are important in determining the thalamic firing profile. Here, we investigated a possible contribution of HVA channels to the epileptic phenotype using a rodent genetic model of absence epilepsy. In this study, HVA Ca(2+) currents were recorded from neurons of three different thalamic nuclei that are involved in both sensory signal transmission and rhythmic-synchronized activity during epileptic spike-and-wave discharges (SWD), namely the dorsal part of the lateral geniculate nucleus (dLGN), the ventrobasal thalamic complex (VB) and the reticular thalamic nucleus (NRT) of epileptic Wistar Albino Glaxo rats from Rijswijk (WAG/Rij) and non-epileptic August Copenhagen Irish (ACI) rats. HVA Ca(2+) current densities in dLGN neurons were significantly increased in epileptic rats compared with non-epileptic controls while other thalamic regions revealed no differences between the strains. Application of specific channel blockers revealed that the increased current was carried by L-type Ca(2+) channels. Electrophysiological evidence of increased L-type current correlated with up-regulated mRNA and protein expression of a particular L-type channel, namely Cav1.3, in dLGN of epileptic rats. No significant changes were found for other HVA Ca(2+) channels. Moreover, pharmacological inactivation of L-type Ca(2+) channels results in altered firing profiles of thalamocortical relay (TC) neurons from non-epileptic rather than from epileptic rats. While HVA Ca(2+) channels influence tonic and burst firing in ACI and WAG/Rij differently, it is discussed that increased Cav1.3 expression may indirectly contribute to increased robustness of burst firing and thereby the epileptic phenotype of absence epilepsy.

Ischemia-induced cell depolarization: does the hyperpolarization-activated cation channel HCN2 affect the outcome after stroke in mice?

BACKGROUND: Brain ischemia is known to include neuronal cell death and persisting neurological deficits. A lack of oxygen and glucose are considered to be key mediators of ischemic neurodegeneration while the exact mechanisms are yet unclear. In former studies the expression of two different two-pore domain potassium (K2P) channels (TASK1, TREK1) were shown to ameliorate neuronal damage due to cerebral ischemia. In neurons, TASK channels carrying hyperpolarizing K+ leak currents, and the pacemaker channel HCN2, carrying depolarizing Ih, stabilize the membrane potential by a mutual functional interaction. It is assumed that this ionic interplay between TASK and HCN2 channels enhances the resistance of neurons to insults accompanied by extracellular pH shifts. METHODS: In C57Bl/6 (wildtype, WT), hcn2+/+ and hcn2-/- mice we used an in vivo model of cerebral ischemia (transient middle cerebral artery occlusion (tMCAO)) to depict a functional impact of HCN2 in stroke formation. Subsequent analyses comprise behavioural tests and hcn2 gene expression assays. RESULTS: After 60 min of tMCAO induction in WT mice, we collected tissue samples at 6, 12, and 24 h after reperfusion. In the infarcted neocortex, hcn2 expression analyses revealed a nominal peak of hcn2 expression 6 h after reperfusion with a tendency towards lower expression levels with longer reperfusion times. Hcn2 gene expression levels in infarcted basal ganglia did not change after 6 h and 12 h. Only at 24 h after reperfusion, hcn2 expression significantly decreases by ~55%. However, 30 min of tMCAO in hcn2-/- as well as hcn2+/+ littermates induced similar infarct volumes. Behavioural tests for global neurological function (Bederson score) and motor function/coordination (grip test) were performed at day 1 after surgery. Again, we found no differences between the groups. CONCLUSIONS: Here, we hypothesized that the absence of HCN2, an important functional counter player of TASK channels, affects neuronal survival during stroke-induced tissue damage. However, together with a former study on TASK3 these results implicate that both TASK3 and HCN2 which were supposed to be neuroprotective due to their pH-dependency, do not influence ischemic neurodegeneration during stroke in the tMCAO model.

Endothelial TWIK-related potassium channel-1 (TREK1) regulates immune-cell trafficking into the CNS.

The blood-brain barrier (BBB) is an integral part of the neurovascular unit (NVU). The NVU is comprised of endothelial cells that are interconnected by tight junctions resting on a parenchymal basement membrane ensheathed by pericytes, smooth muscle cells and a layer of astrocyte end feet. Circulating blood cells, such as leukocytes, complete the NVU. BBB disruption is common in several neurological diseases, but the molecular mechanisms involved remain largely unknown. We analyzed the role of TWIK-related potassium channel-1 (TREK1, encoded by KCNK2) in human and mouse endothelial cells and the BBB. TREK1 was downregulated in endothelial cells by treatment with interferon-γ (IFN-γ) and tumor necrosis factor-α (TNF-α). Blocking TREK1 increased leukocyte transmigration, whereas TREK1 activation had the opposite effect. We identified altered mitogen-activated protein (MAP) kinase signaling, actin remodeling and upregulation of cellular adhesion molecules as potential mechanisms of increased migration in TREK1-deficient (Kcnk2(-/-)) cells. In Kcnk2(-/-) mice, brain endothelial cells showed an upregulation of the cellular adhesion molecules ICAM1, VCAM1 and PECAM1 and facilitated leukocyte trafficking into the CNS. Following the induction of experimental autoimmune encephalomyelitis (EAE) by immunization with a myelin oligodendrocyte protein (MOG)35-55 peptide, Kcnk2(-/-) mice showed higher EAE severity scores that were accompanied by increased cellular infiltrates in the central nervous system (CNS). The severity of EAE was attenuated in mice given the amyotrophic lateral sclerosis drug riluzole or fed a diet enriched with linseed oil (which contains the TREK-1 activating omega-3 fatty acid α-linolenic acid). These beneficial effects were reduced in Kcnk2(-/-) mice, suggesting TREK-1 activating compounds may be used therapeutically to treat diseases related to BBB dysfunction.

Thalamic involvement in patients with neurologic impairment due to Shiga toxin 2.

OBJECTIVE: The outbreak of hemolytic-uremic syndrome and diarrhea caused by Shiga toxin-producing Escherichia coli O104:H4 in Germany during May to July 2011 involved severe and characteristic neurologic manifestations with a strong female preponderance. Owing to these observations, we designed a series of experimental studies to evaluate the underlying mechanism of action of this clinical picture. METHODS: A magnetic resonance imaging and electroencephalographic study of patients was performed to evaluate the clinical picture in detail. Thereafter, combinations of different experimental settings, including electrophysiological and histological analyses, as well as calcium imaging in brain slices of rats, were conducted. RESULTS: We report on 7 female patients with neurologic symptoms and signs including bilateral thalamic lesions and encephalopathic changes indicative of a predominant involvement of the thalamus. Experimental studies in rats revealed an enhanced expression of the Shiga toxin receptor globotriaosylceramide on thalamic neurons in female rats as compared to other brain regions in the same rats and to male animals. Incubation of brain slices with Shiga toxin 2 evoked a strong membrane depolarization and intracellular calcium accumulation in neurons, associated with neuronal apoptosis, predominantly in the thalamic area. INTERPRETATION: These findings suggest that the direct cytotoxic effect of Shiga toxin 2 in the thalamus might contribute to the pathophysiology of neuronal complications in hemolytic-uremic syndrome.

Identification of two-pore domain potassium channels as potent modulators of osmotic volume regulation in human T lymphocytes.

Many functions of T lymphocytes are closely related to cell volume homeostasis and regulation, which utilize a complex network of membrane channels for anions and cations. Among the various potassium channels, the voltage-gated K(V)1.3 is well known to contribute greatly to the osmoregulation and particularly to the potassium release during the regulatory volume decrease (RVD) of T cells faced with hypotonic environment. Here we address a putative role of the newly identified two-pore domain (K(2P)) channels in the RVD of human CD4(+) T lymphocytes, using a series of potent well known channel blockers. In the present study, the pharmacological profiles of RVD inhibition revealed K(2P)5.1 and K(2P)18.1 as the most important K(2P) channels involved in the RVD of both naïve and stimulated T cells. The impact of chemical inhibition of K(2P)5.1 and K(2P)18.1 on the RVD was comparable to that of K(V)1.3. K(2P)9.1 also notably contributed to the RVD of T cells but the extent of this contribution and its dependence on the activation status could not be unambiguously resolved. In summary, our data provide first evidence that the RVD-related potassium efflux from human T lymphocytes relies on K(2P) channels.

Ca(2+)-dependent large conductance K(+) currents in thalamocortical relay neurons of different rat strains.

Mutations in genes coding for Ca(2+) channels were found in patients with childhood absence epilepsy (CAE) indicating a contribution of Ca(2+)-dependent mechanisms to the generation of spike-wave discharges (SWD) in humans. Since the involvement of Ca(2+) signals remains unclear, the aim of the present study was to elucidate the function of a Ca(2+)-dependent K(+) channel (BKCa) under physiological conditions and in the pathophysiological state of CAE. The activation of BKCa channels is dependent on both voltage and intracellular Ca(2+) concentrations. Moreover, these channels exhibit an outstandingly high level of regulatory heterogeneity that builds the basis for the influence of BKCa channels on different aspects of neuronal activity. Here, we analyse the contribution of BKCa channels to firing of thalamocortical relay neurons, and we test the hypothesis that BKCa channel activity affects the phenotype of a genetic rat model of CAE. We found that the activation of the ß2-adrenergic receptor/protein kinase A pathway resulted in BKCa channel inhibition. Furthermore, BKCa channels affect the number of action potentials fired in a burst and produced spike frequency adaptation during tonic activity. The latter result was confirmed by a computer modelling approach. We demonstrate that the ß2-adrenergic inhibition of BKCa channels prevents spike frequency adaptation and, thus, might significantly support the tonic firing mode of thalamocortical relay neurons. In addition, we show that BKCa channel functioning differs in epileptic WAG/Rij and thereby likely contributes to highly synchronised, epileptic network activity.

The TASK1 channel inhibitor A293 shows efficacy in a mouse model of multiple sclerosis.

The two-pore domain potassium channel TASK1 (KCNK3) has recently emerged as an important modulator in autoimmune CNS inflammation. Previously, it was shown that T lymphocytes obtained from TASK1(-/-) mice display impaired T cell effector functions and that TASK1(-/-) mice show a significantly reduced disease severity in myelin oligodendrocyte glycoprotein (MOG(35-55)) peptide induced experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis. We here evaluate a potent and specific TASK1 channel inhibitor, A293, which caused a dose-dependent reduction of T cell effector functions (cytokine production and proliferation). This effect was abolished in CD4(+) T cells from TASK1(-/-) mice but not in cells from TASK3(-/-) mice. In electrophysiological measurements, A293 application induced a significant reduction of the outward current of wildtype T lymphocytes, while there was no effect in TASK1(-/-) cells. Preventive and therapeutic application of A293 significantly ameliorated the EAE disease course in wildtype mice while it had no significant effect in TASK1(-/-) mice and was still partly effective in TASK3(-/-) mice. In summary, our findings support the concept of TASK1 as an attractive drug target for autoimmune disorders.

Adenylyl cyclases: expression in the developing rat thalamus and their role in absence epilepsy.

Adenylyl cyclases (ACs) synthesize the second messenger cyclic AMP (cAMP) which influences the function of multiple ion channels. Former studies point to a malfunction of cAMP-dependent ion channel regulation in thalamocortical relay neurons that contribute to the development of the absence epileptic phenotype of a rat genetic model (WAG/Rij). Here, we provide detailed information about the thalamic gene and protein expression of Ca(2+)/calmodulin-activated AC isoforms in rat thalamus. Data from WAG/Rij were compared to those from non-epileptic controls (August-Copenhagen Irish rats) to elucidate whether differential expression of ACs contributes to the dysregulation of thalamocortical activity. At one postnatal stage (P21), we found the gene expression of two specific Ca(2+)-activated AC isoforms (AC-1 and AC-3) to be significantly down-regulated in epileptic tissue, and we identified the isoform AC-1 to be the most prominent one in both strains. However, Western blot data and analysis of enzymatic AC activity revealed no differences between the two strains. While basal AC activity was low, cAMP production was boosted by application of a forskolin derivative up to sevenfold. Despite previous hints pointing to a major contribution of ACs, the presented data show that there is no apparent causality between AC activity and the occurrence of the epileptic phenotype.