adenosine-kinase and Epilepsy

Epilepsy, a complex neurological syndrome with dominant symptoms and various comorbidities, affects over 70 million people worldwide. Epilepsy-related comorbidities, including cognitive and psychiatric disorders, can impede therapy for epilepsy patients, leading to heavy burdens on patients and society. Adenosine has an anti-epileptic and anticonvulsive function in the brain. Several studies have shown that, through adenosine receptor-dependent and -independent mechanisms, adenosine can influence the development and progression (epileptogenesis) of epilepsy and its associated comorbidities. As the key enzyme for adenosine clearance, adenosine kinase (ADK) can exacerbate epileptic seizures not only by accelerating adenosine clearance, but also by increasing global DNA methylation through the transmethylation pathway. Therefore, adenosine augmentation therapies for epilepsy can have dual functions in the inhibition of epileptic seizures and the prevention of its overall progress. This review has three main purposes. First, we discuss how maladaptive changes in the adenosine pathway affect the development and progress of epilepsy in both receptor-dependent and receptor-independent ways. Second, we highlight the important influence of associated comorbidities on the prognosis of epilepsy and explore the role of adenosine in these comorbidities. Finally, we emphasize the potential of adenosine augmentation therapies in restoring normal adenosine signaling in the epileptic brain. Such treatments could effectively improve the prognosis of patients who are resistant to most antiepileptic drugs (AEDs), and thus bring new challenges and opportunities in the treatment of epilepsy patients.

Topics: Adenosine; Adenosine Kinase; Anticonvulsants; Epilepsy; Humans; Seizures

Adenosine kinase (ADK) is the key regulator of adenosine and catalyzes the metabolism of adenosine to 5'-adenosine monophosphate. The enzyme exists in two isoforms: a long isoform (ADK-long, ADK-L) and a short isoform (ADK-short, ADK-S). The two isoforms are developmentally regulated and are differentially expressed in distinct subcellular compartments with ADK-L localized in the nucleus and ADK-S localized in the cytoplasm. The nuclear localization of ADK-L and its biochemical link to the transmethylation pathway suggest a specific role for gene regulation via epigenetic mechanisms. Recent evidence reveals an adenosine receptor-independent role of ADK in determining the global methylation status of DNA and thereby contributing to epigenomic regulation. Here we summarize recent progress in understanding the biochemical interactions between adenosine metabolism by ADK-L and epigenetic modifications linked to transmethylation reactions. This review will provide a comprehensive overview of ADK-associated changes in DNA methylation in developmental, as well as in pathological conditions including brain injury, epilepsy, vascular diseases, cancer, and diabetes. Challenges in investigating the epigenetic role of ADK for therapeutic gains are briefly discussed.

Topics: Adenosine Kinase; Animals; Brain Injuries; DNA Methylation; Epigenesis, Genetic; Epilepsy; Gene Expression Regulation; Humans

Epilepsy, an ancient disease, is defined as an enduring predisposition to generate epileptic seizures and by the neurobiological, cognitive, psychological, and social consequences of this condition. Antiepileptic drugs (AEDs) are currently used as first-line treatment for patients with epilepsy; however, around 36% of patients are diagnosed with refractory epilepsy, which means two or more AEDs have been considered as failed after sufficiently correct usage. Unfortunately, it is unlikely that the improvement of the efficacy of AEDs will be easily achieved, especially since no AEDs show efficacy in ceasing epileptogenesis. Consequently, several endogenous anticonvulsants attract investigators and epileptologists, such as galanin, cannabis, and adenosine. Astrogliosis is a neuropathological hallmark of epilepsy, whatever the etiology is, and astrogliosis is frequently associated with overexpression of adenosine kinase, which means downregulation of synaptic levels of adenosine. Consequently, adenosine is negatively regulated by adenosine kinase through the astrocyte-based cycle. On the other hand, focal adenosine augmentation therapy, using adenosine kinase inhibitor, has been proved to be effective for reducing seizures in both animal models and in vitro human brain tissue resected from a variety of etiology of refractory epilepsy patients. In addition to reducing seizures, adenosine augmentation therapy can also palliate co-morbidities, like sleep, cognition, or depression. Of importance, transgenic mice with reduced ADK were resistant to epileptogenesis induced by acute brain injury. In terms of translation, based on findings of adenosinerelated epileptogenic mechanisms, the application into clinical practice seems to be feasible by molecular strategies that have been already experimentally implemented, including gene and RNA interference. In the present review, we will focus on the evidence of ADK dysfunction in the epileptic brain from human beings and animals, and review the role of ADK inhibitor in adenosine augmentation therapy and the underlying mechanism of prevention of epileptogenesis.

Topics: Adenosine; Adenosine Kinase; Animals; Anticonvulsants; Brain; Epilepsy; Humans; Mice; Protein Kinase Inhibitors

Adenosine is a well-characterized endogenous anticonvulsant and seizure terminator of the brain. Through a combination of adenosine receptor-dependent and -independent mechanisms, adenosine affects seizure generation (ictogenesis), as well as the development of epilepsy and its progression (epileptogenesis). Maladaptive changes in adenosine metabolism, in particular increased expression of the astroglial enzyme adenosine kinase (ADK), play a major role in epileptogenesis. Increased expression of ADK has dual roles in both reducing the inhibitory tone of adenosine in the brain, which consequently reduces the threshold for seizure generation, and also driving an increased flux of methyl-groups through the transmethylation pathway, thereby increasing global DNA methylation. Through these mechanisms, adenosine is uniquely positioned to link metabolism with epigenetic outcome. Therapeutic adenosine augmentation therefore not only holds promise for the suppression of seizures in epilepsy, but moreover the prevention of epilepsy and its progression overall. This review will focus on adenosine-related mechanisms implicated in ictogenesis and epileptogenesis and will discuss therapeutic opportunities and challenges.

Topics: Adenosine; Adenosine Kinase; Animals; Anticonvulsants; Astrocytes; Brain; DNA Methylation; Epigenesis, Genetic; Epilepsy; Humans; Receptor, Adenosine A1; Seizures

Adenine nucleotides and adenosine are signaling molecules that activate purinergic receptors P1 and P2. Activation of A1 adenosine receptors has an anticonvulsant action, whereas activation of A2A receptors might initiate seizures. Therefore, a significant limitation to the use of A1 receptor agonists as drugs in the CNS might be their peripheral side effects. The anti-epileptic activity of adenosine is related to its increased concentration outside the cell. This increase might result from the inhibition of the equilibrative nucleoside transporters (ENTs). Moreover, the implantation of implants or stem cells into the brain might cause slow and persistent increases in adenosine concentrations in the extracellular spaces of the brain. The role of adenosine in seizure inhibition has been confirmed by results demonstrating that in patients with epilepsy, the adenosine kinase (ADK) present in astrocytes is the only purine-metabolizing enzyme that exhibits increased expression. Increased ADK activity causes intensified phosphorylation of adenosine to 5'-AMP, which therefore lowers the adenosine level in the extracellular spaces. These changes might initiate astrogliosis and epileptogenesis, which are the manifestations of epilepsy. Seizures might induce inflammatory processes and vice versa. Activation of P2X7 receptors causes intensified release of pro-inflammatory cytokines (IL-1β and TNF-α) and activates metabolic pathways that induce inflammatory processes in the CNS. Therefore, antagonists of P2X7 and the interleukin 1β receptor might be efficient drugs for recurring seizures and prolonged status epilepticus. Inhibitors of ADK would simultaneously inhibit the seizures, prevent the astrogliosis and epileptogenesis processes and prevent the formation of new epileptogenic foci. Therefore, these drugs might become beneficial seizure-suppressing drugs.

Topics: Adenosine; Adenosine Kinase; Animals; Epilepsy; Humans; Purinergic P2X Receptor Antagonists; Receptor, Adenosine A2A; Receptors, Purinergic; Receptors, Purinergic P2X7; Signal Transduction

Despite the introduction of at least 20 new antiepileptic drugs (AEDs) into clinical practice over the past decades, about one third of all epilepsies remain refractory to conventional forms of treatment. In addition, currently used AEDs have been developed to suppress neuronal hyperexcitability, but not necessarily to address pathogenic mechanisms involved in epilepsy development or progression (epileptogenesis). For those reasons endogenous seizure control mechanisms of the brain may provide alternative therapeutic opportunities. Adenosine is a well characterized endogenous anticonvulsant and seizure terminator of the brain. Several lines of evidence suggest that endogenous adenosine-mediated seizure control mechanisms fail in chronic epilepsy, whereas therapeutic adenosine augmentation effectively prevents epileptic seizures, even those that are refractory to conventional AEDs. New findings demonstrate that dysregulation of adenosinergic mechanisms are intricately involved in the development of epilepsy and its comorbidities, whereas adenosine-associated epigenetic mechanisms may play a role in epileptogenesis. The first goal of this review is to discuss how maladaptive changes of adenosinergic mechanisms contribute to the expression of seizures (ictogenesis) and the development of epilepsy (epileptogenesis) by focusing on pharmacological (adenosine receptor dependent) and biochemical (adenosine receptor independent) mechanisms as well as on enzymatic and transport based mechanisms that control the availability (homeostasis) of adenosine. The second goal of this review is to highlight innovative adenosine-based opportunities for therapeutic intervention aimed at reconstructing normal adenosine function and signaling for improved seizure control in chronic epilepsy. New findings suggest that transient adenosine augmentation can have lasting epigenetic effects with disease modifying and antiepileptogenic outcome. This article is part of the Special Issue entitled 'Purines in Neurodegeneration and Neuroregeneration'.

Topics: 5'-Nucleotidase; Adenosine; Adenosine Kinase; Animals; Anticonvulsants; Astrocytes; Epilepsy; Genetic Therapy; Homeostasis; Humans; Nucleoside Transport Proteins; Receptors, Purinergic P1; Signal Transduction

Experimental research over the past decade has supported the critical role of astrocytes activated by different types of injury and the pathophysiological processes that underlie the development of epilepsy. In both experimental and human epileptic tissues astrocytes undergo complex changes in their physiological properties, which can alter glio-neuronal communication, contributing to seizure precipitation and recurrence. In this context, understanding which of the molecular mechanisms are crucially involved in the regulation of glio-neuronal interactions under pathological conditions associated with seizure development is important to get more insight into the role of astrocytes in epilepsy. This article reviews current knowledge regarding the role of glial adenosine kinase as a neuropathological marker of the epileptic brain. Both experimental findings in clinically relevant models, as well as observations in drug-resistant human epilepsies will be discussed, highlighting the link between astrogliosis, dysfunction of adenosine homeostasis and seizure generation and therefore suggesting new strategies for targeting astrocyte-mediated epileptogenesis.

Topics: Adenosine Kinase; Animals; Biomarkers; Brain; Epilepsy; Humans; Neuroglia

Extracellular levels of the brain's endogenous anticonvulsant and neuroprotectant adenosine largely depend on an astrocyte-based adenosine cycle, comprised of ATP release, rapid degradation of ATP into adenosine, and metabolic reuptake of adenosine through equilibrative nucleoside transporters and phosphorylation by adenosine kinase (ADK). Changes in ADK expression and activity therefore rapidly translate into changes of extracellular adenosine, which exerts its potent anticonvulsive and neuroprotective effects by activation of pre- and postsynaptic adenosine A(1) receptors. Increases in ADK increase neuronal excitability, whereas decreases in ADK render the brain resistant to seizures and injury. Importantly, ADK was found to be overexpressed and associated with astrogliosis and spontaneous seizures in rodent models of epilepsy, as well as in human specimen resected from patients with hippocampal sclerosis and temporal lobe epilepsy. Several lines of evidence indicate that overexpression of astroglial ADK and adenosine deficiency are pathological hallmarks of the epileptic brain. Consequently, adenosine augmentation therapies constitute a powerful approach for seizure prevention, which is effective in models of epilepsy that are resistant to conventional antiepileptic drugs. The adenosine kinase hypothesis of epileptogenesis suggests that adenosine dysfunction in epilepsy undergoes a biphasic response: an acute surge of adenosine that can be triggered by any type of injury might contribute to the development of astrogliosis via adenosine receptor-dependent and -independent mechanisms. Astrogliosis in turn is associated with overexpression of ADK, which was shown to be sufficient to trigger spontaneous recurrent electrographic seizures. Thus, ADK emerges as a promising target for the prediction and prevention of epilepsy.

Topics: Adenosine; Adenosine Kinase; Animals; Disease Models, Animal; Epilepsy; Humans; Metabolic Diseases; Phosphorylation

Topics: Adenosine; Adenosine Kinase; Adenosine Triphosphate; Animals; Brain; Diet, Carbohydrate-Restricted; Diet, Ketogenic; Energy Metabolism; Epilepsy; Humans

Nucleoside receptors are known to be important targets for a variety of brain diseases. However, the therapeutic modulation of their endogenous agonists by inhibitors of nucleoside metabolism represents an alternative therapeutic strategy that has gained increasing attention in recent years. Deficiency in endogenous nucleosides, in particular of adenosine, may causally be linked to a variety of neurological diseases and neuropsychiatric conditions ranging from epilepsy and chronic pain to schizophrenia. Consequently, augmentation of nucleoside function by inhibiting their metabolism appears to be a rational therapeutic strategy with distinct advantages: (i) in contrast to specific receptor modulation, the increase (or decrease) of the amount of a nucleoside will affect several signal transduction pathways simultaneously and therefore have the unique potential to modify complex neurochemical networks; (ii) by acting on the network level, inhibitors of nucleoside metabolism are highly suited to fine-tune, restore, or amplify physiological functions of nucleosides; (iii) therefore inhibitors of nucleoside metabolism have promise for the "soft and smart" therapy of neurological diseases with the added advantage of reduced systemic side effects. This review will first highlight the role of nucleoside function and dysfunction in physiological and pathophysiological situations with a particular emphasis on the anticonvulsant, neuroprotective, and antinociceptive roles of adenosine. The second part of this review will cover pharmacological approaches to use inhibitors of nucleoside metabolism, with a special emphasis on adenosine kinase, the key regulator of endogenous adenosine. Finally, novel gene-based therapeutic strategies to inhibit nucleoside metabolism and focal treatment approaches will be discussed.

Topics: Adenosine; Adenosine Kinase; Analgesics; Animals; Anticonvulsants; Brain; Brain Diseases; Epilepsy; Gene Expression; Humans; Metabolic Networks and Pathways; Mice; Mice, Knockout; Neuroprotective Agents; Pain; Protein Kinase Inhibitors; Purinergic P1 Receptor Agonists; Purinergic P1 Receptor Antagonists; Rats; Receptor, Adenosine A1; Receptor, Adenosine A2A; Schizophrenia; Signal Transduction; Sleep Wake Disorders

Since its discovery a decade ago, RNA interference (RNAi) has been developed not only into powerful experimental tools but also into promising novel therapeutics. In contrast to conventional antiepileptic drugs (AEDs) that target specific proteins such as ion channels or receptors, RNAi-based therapeutics exploit an endogenous regulatory mechanism of gene expression and thereby are poised to prevent or reverse pathogenetic mechanisms involved in seizure development. Therapeutic RNAi has been widely explored for dominant targets involved in neurodegenerative diseases; however, their use for epilepsy therapy has received less attention. This review discusses potential RNAi-based targets that are of interest for epilepsy therapy, including adenosine kinase (ADK), the key negative regulator of the brain's endogenous anticonvulsant adenosine. Overexpression of ADK, and the resulting adenosine deficiency, are pathologic hallmarks of the sclerotic epileptic brain, and have been implicated in seizure generation. Therefore, RNAi-strategies aimed at reducing ADK (and increasing adenosine) are based on a direct neurochemical rationale that has recently been explored experimentally using ex vivo and in vivo gene therapy approaches. Technical issues and challenges remain before those promising tools can be developed into future therapeutics for epilepsy.

Topics: Adenosine; Adenosine Kinase; Animals; Anticonvulsants; Brain; Epilepsy; Gene Expression Regulation; Genetic Therapy; Humans; Mice; MicroRNAs; RNA Interference

Epilepsy is a common seizure disorder affecting approximately 70 million people worldwide. Current pharmacotherapy is neuron-centered, frequently accompanied by intolerable side effects, and fails to be effective in about one third of patients. Therefore, new therapeutic concepts are needed. Recent research suggests an astrocytic basis of epilepsy, presenting the possibility of novel therapeutic targets. In particular, dysfunction of the astrocyte-controlled, endogenous, adenosine-based seizure control system of the brain is implicated in seizure generation. Thus, astrogliosis - a pathological hallmark of the epileptic brain - is associated with upregulation of the adenosine-removing enzyme adenosine kinase (ADK), resulting in focal adenosine deficiency. Both astrogliotic upregulation of ADK in epilepsy and transgenic overexpression of ADK are associated with seizures, and inhibition of ADK prevents seizures in a mouse model of pharmacoresistant epilepsy. These findings link adenosine deficiency with seizures and predict that adenosine augmentation therapies (AATs) will likely be effective in preventing seizures. Given the wide-spread systemic and central side effects of systemically administered AATs, focal AATs (i.e., limited to the astrogliotic lesion) are a necessity. This Commentary will discuss the pharmacological rationale for the development of focal AATs. Additionally, several AAT strategies will be discussed: (1) adenosine released from silk-based brain implants; (2) adenosine released from locally implanted encapsulated cells; (3) adenosine released from stem cell-derived brain implants; and (4) adenosine augmenting gene therapies. Finally, new developments and therapeutic challenges in using focal AATs for epilepsy therapy will critically be evaluated.

Topics: Adenosine; Adenosine Kinase; Animals; Anticonvulsants; Astrocytes; Brain; Drug Delivery Systems; Drug Implants; Epilepsy; Genetic Therapy; Humans; Mice; Neuroglia; Rats; Stem Cells

Adenosine is a modulator of brain function uniquely positioned to integrate excitatory and inhibitory neurotransmission. The past few years brought a wealth of new data fostering our understanding of how the adenosine system is involved in the pathogenesis of neurological diseases. Thus, dysregulation of the adenosine system is implicated in epileptogenesis and cell therapies have been developed to locally augment adenosine in an approach to prevent seizures. While activation of inhibitory adenosine A(1) receptors is beneficial in epilepsy, chronic pain and cerebral ischemia, inhibition of facilitatory A(2A) receptors has profound neuroprotective effects, which are currently exploited in clinical trials in Parkinson's disease. A new era of adenosine-based therapies has begun, with the prospect to cover a wide range of neurological diseases.

Topics: Adenosine; Adenosine Kinase; Alzheimer Disease; Animals; Brain Diseases; Brain Ischemia; Epilepsy; Humans; Huntington Disease; Pain; Parkinson Disease; Schizophrenia; Synaptic Transmission

Current therapies for epilepsy are largely symptomatic and do not affect the underlying mechanisms of disease progression, i.e. epileptogenesis. Given the large percentage of pharmacoresistant chronic epilepsies, novel approaches are needed to understand and modify the underlying pathogenetic mechanisms. Although different types of brain injury (e.g. status epilepticus, traumatic brain injury, stroke) can trigger epileptogenesis, astrogliosis appears to be a homotypic response and hallmark of epilepsy. Indeed, recent findings indicate that epilepsy might be a disease of astrocyte dysfunction. This review focuses on the inhibitory neuromodulator and endogenous anticonvulsant adenosine, which is largely regulated by astrocytes and its key metabolic enzyme adenosine kinase (ADK). Recent findings support the "ADK hypothesis of epileptogenesis": (i) Mouse models of epileptogenesis suggest a sequence of events leading from initial downregulation of ADK and elevation of ambient adenosine as an acute protective response, to changes in astrocytic adenosine receptor expression, to astrocyte proliferation and hypertrophy (i.e. astrogliosis), to consequential overexpression of ADK, reduced adenosine and - finally - to spontaneous focal seizure activity restricted to regions of astrogliotic overexpression of ADK. (ii) Transgenic mice overexpressing ADK display increased sensitivity to brain injury and seizures. (iii) Inhibition of ADK prevents seizures in a mouse model of pharmacoresistant epilepsy. (iv) Intrahippocampal implants of stem cells engineered to lack ADK prevent epileptogenesis. Thus, ADK emerges both as a diagnostic marker to predict, as well as a prime therapeutic target to prevent, epileptogenesis.

Topics: Adenosine Kinase; Animals; Epilepsy; Humans; Models, Biological

Despite recent medical advances pharmacoresistant epilepsy continues to be a major health problem. The knowledge of endogenous protective mechanisms of the brain may lead to the development of rational therapies tailored to a patient's needs. Adenosine has been identified as an endogenous neuromodulator with antiepileptic and neuroprotective properties. However, the therapeutic use of adenosine or its receptor agonists is largely precluded by strong peripheral and central side effects. Thus, local delivery of adenosine to a critical site of the brain may provide a solution for the therapeutic use of adenosine. The following rationale for the local augmentation of the adenosine system as a novel therapeutic principle in the treatment of epilepsy has been established: (1) Deficits in the adenosinergic system are associated with epileptogenesis and these deficits promote seizures. Thus, reconstitution of an inhibitory adenosinergic tone is a rational therapeutic approach. (2) The focal paracrine delivery of adenosine from encapsulated cells suppresses seizures in kindled rats without overt side effects. (3) The anticonvulsant activity of locally released adenosine is maintained in models of epilepsy which are resistant to major antiepileptic drugs. This review summarizes the rationale and recent approaches for adenosine-based cell therapies for pharmacoresistant epilepsies.

Topics: Adenosine; Adenosine Kinase; Animals; Anticonvulsants; Cell- and Tissue-Based Therapy; Drug Tolerance; Epilepsy; Humans

Adenosine is an inhibitory modulator of brain activity with neuroprotective and anticonvulsant properties. Adenosine levels are regulated mainly by adenosine kinase (ADK), an enzyme that is responsible for the removal of adenosine via phosphorylation to AMP. Recent evidence indicates that expression of ADK undergoes rapid coordinated changes during brain development and following brain injury, such as after epileptic seizures and stroke. Thus, transient downregulation of ADK after acute brain injury protects the brain from seizures and cell death. Conversely, chronic overexpression of ADK causes seizures in epilepsy and promotes cell death in epilepsy and stroke. These findings have direct implications for the rational definition of ADK as a therapeutic target. In recent years, novel treatment strategies have been developed that make use of the intracerebral transplantation of cells that are ADK deficient and, thus, release adenosine. A new era of cell-based delivery of adenosine has begun, which holds great promise for novel therapies for epilepsy and stroke.

Topics: Adenosine; Adenosine Kinase; Animals; Brain; Drug Delivery Systems; Enzyme Inhibitors; Epilepsy; Humans; Stroke

Adenosine (ADO) acts as an inhibitory neuromodulator throughout the central and peripheral nervous system and can regulate seizure and nociceptive activity. However, the positive actions of systemically administered ADO are usually accompanied by undesirable side effects such as hypomobility and cardio-suppression. Adenosine kinase (AK) is the primary metabolic enzyme regulating intra- and extracellular concentrations of ADO. We review the recent development of structurally novel nucleoside and nonnucleoside AK inhibitors that demonstrate high specificity for the AK enzyme. Several of these compounds have shown significant beneficial effects in animal models of epilepsy and pain with an improved preclinical therapeutic window over direct acting ADO receptor agonists.

Topics: Adenosine Kinase; Analgesics; Animals; Anticonvulsants; Enzyme Inhibitors; Epilepsy; Humans; Pain; Structure-Activity Relationship

Adenosine (ADO) is an endogenous inhibitory neuromodulator that increases nociceptive thresholds in response to tissue trauma and inflammation. Adenosine kinase (AK) is a key intracellular enzyme regulating intra- and extracellular concentrations of ADO. AK inhibition selectively amplifies extracellular ADO levels at cell and tissue sites where accelerated release of ADO occurs. AK inhibitors have been shown to provide effective antinociceptive, antiinflammatory and anticonvulsant activity in animal models, thus suggesting their potential therapeutic utility for pain, inflammation, epilepsy and possibly other central and peripheral nervous system diseases associated with cellular trauma and inflammation. This beneficial outcome may potentially lack nonspecific effects associated with the systemic administration of ADO receptor agonists. Until recently all of the reported AK inhibitors contained adenosine-like structural motif. The present review will discuss design, synthesis and analgesic and antiinflammatory properties of the novel nonnucleoside AK inhibitors that do not have close structural resemblance with the natural substrate ADO. Two classes of the nonnucleoside AK inhibitors are built on pyridopyrimidine and alkynylpyrimidine cores.

Topics: Adenosine Kinase; Animals; Drug Design; Enzyme Inhibitors; Epilepsy; Humans; Inflammation; Molecular Structure; Pain; Pyrimidines; Structure-Activity Relationship

Adenosine kinase (AK; EC 2.7.1.20) is a key intracellular enzyme regulating intra and extracellular concentrations of adenosine (ADO), an endogenous modulator of intercellular signalling that reduces cell excitability during tissue stress and trauma. The inhibitory effects of ADO are mediated by interactions with specific cell-surface G-protein coupled receptors (GPCR), which regulate membrane cation flux, membrane polarisation and the release of excitatory neurotransmitters. Inhibition of AK potentiates local extracellular ADO levels at cell and tissue sites which are undergoing accelerated ADO release. Thus, AK inhibition represents a mechanism to selectively enhance the endogenous protective actions of ADO during cellular stress while potentially minimising the non-specific effects associated with the systemic administration of ADO receptor agonists. Novel, potent AK inhibitors have recently been synthesised that demonstrate high specificity for this particular enzyme as compared to other ADO metabolic enzymes, transporters and receptors. AK inhibitors have been shown to increase ADO concentrations in various systems in vitro, as well as in an in vivo model of neurotoxicity. In addition, AK inhibitors have demonstrated efficacy in animal models of epilepsy, cerebral ischaemia as well as pain and inflammation, thus suggesting their potential therapeutic utility for these conditions.

Topics: Adenosine Kinase; Animals; Brain Ischemia; Disease Models, Animal; Enzyme Inhibitors; Epilepsy; Humans; Inflammation; Pain; Purinergic P1 Receptor Antagonists; Receptors, Cell Surface

Deep brain stimulation (DBS) of the anterior nucleus of the thalamus, is an effective therapy for patients with drug-resistant epilepsy, yet, its mechanism of action remains elusive. Adenosine kinase (ADK), a key negative regulator of adenosine, is a potential modulator of epileptogenesis. DBS has been shown to increase adenosine levels, which may suppress seizures via A1 receptors (A. Control group, SE (status epilepticus) group, SE-DBS group, and SE-sham-DBS group were included in this study. One week after a pilocarpine-induced status epilepticus, rats in the SE-DBS group were treated with DBS for 4 weeks. The rats were monitored by video-EEG. ADK and A. Compared with the SE group and SE-sham-DBS group, DBS could reduce the frequency of spontaneous recurrent seizures (SRS) and the number of interictal epileptic discharges. The DPCPX, an A. The findings indicate that DBS can reduce SRS in epileptic rats via inhibition of ADK and activation of A

Topics: Adenosine Kinase; Animals; Disease Progression; Epilepsy; Male; Pilocarpine; Rats; Rats, Sprague-Dawley; Receptor, Adenosine A1; Seizures; Status Epilepticus

Resistance to traditional antiepileptic drugs is a major challenge in chronic epilepsy treatment. MicroRNA-based gene therapy is a promising alternative but has demonstrated limited efficacy due to poor blood-brain barrier permeability, cellular uptake, and targeting efficiency. Adenosine is an endogenous antiseizure agent deficient in the epileptic brain due to elevated adenosine kinase (ADK) activity in reactive A1 astrocytes. We designed a nucleic acid nanoantiepileptic drug (tFNA-ADK

Topics: Adenosine; Adenosine Kinase; Animals; Anticonvulsants; Astrocytes; Epilepsy; Mice; Nucleic Acids

Vagus nerve stimulation (VNS) is an alternative treatment option for individuals with refractory epilepsy, with nearly 40% of patients showing no benefit after VNS and only 6%-8% achieving seizure freedom. It is presently unclear why some patients respond to treatment and others do not. Therefore, identification of biomarkers to predict efficacy of VNS is of utmost importance. The objective of this study was to explore whether genetic variations in genes involved in adenosine kinase (ADK), ecto-5'-nucleotidase (NT5E), and adenosine A1 receptor (A1R) are linked to outcome of VNS in patients with refractory epilepsy.. Thirty single-nucleotide polymorphisms (SNPs), including 9 in genes encoding ADK, 3 in genes encoding NT5E, and 18 in genes encoding A1R, were genotyped in 194 refractory epilepsy patients who underwent VNS. The chi-square test and binary logistic regression were used to determine associations between genetic differences and VNS efficacy.. A significant association between ADK SNPs rs11001109, rs7899674, and rs946185 and seizure reduction with VNS was found. Regardless of sex, age, seizure frequency and type, antiseizure drug use, etiology, and prior surgical history, all patients (10/10 patients [100%]) with minor allele homozygosity at rs11001109 (genotype AA) or rs946185 (AA) achieved > 50% seizure reduction and 4 patients (4/10 [40%]) achieved seizure freedom. VNS therapy demonstrated higher efficacy among carriers of minor allele rs7899674 (CG + GG) (68.3% vs 48.8% for patients with major allele homozygosity).. Homozygous ADK SNPs rs11001109 (AA) and rs946185 (AA), as well as minor allele rs7899674 (CG + GG), may serve as useful biomarkers for prediction of VNS therapy outcome.

Topics: Adenosine Kinase; Biomarkers; Drug Resistant Epilepsy; Epilepsy; Humans; Polymorphism, Single Nucleotide; Seizures; Treatment Outcome; Vagus Nerve; Vagus Nerve Stimulation

Topics: Adenosine Kinase; Animals; Caspase 1; Epilepsy; Inflammasomes; Interleukin-1beta; Mice; NLR Family, Pyrin Domain-Containing 3 Protein

Adenosine kinase (ADK) deficiency is characterized by liver disease, dysmorphic features, epilepsy and developmental delay. This defect disrupts the adenosine/AMP futile cycle and interferes with the upstream methionine cycle. We report the clinical, histological and biochemical courses of three ADK children carrying two new mutations and presenting with neonatal cholestasis and neurological disorders. One of them died of liver failure whereas the other two recovered from their liver damage. As the phenotype was consistent with a mitochondrial disorder, we studied liver mitochondrial respiratory chain activities in two patients and revealed a combined defect of several complexes. In addition, we retrospectively analyzed methionine plasma concentration, a hallmark of ADK deficiency, in a cohort of children and showed that methionine level in patients with ADK deficiency was strongly increased compared with patients with other liver diseases. ADK deficiency is a cause of neonatal or early infantile liver disease that may mimic primary mitochondrial disorders. In this context, an elevation of methionine plasma levels over twice the upper limit should not be considered as a nonspecific finding. ADK deficiency induced-liver dysfunction is most often transient, but could be life-threatening.

Topics: Adenosine; Adenosine Kinase; Amino Acid Metabolism, Inborn Errors; Child; Developmental Disabilities; Epilepsy; Female; Genetic Predisposition to Disease; Glycine N-Methyltransferase; Humans; Infant; Infant, Newborn; Liver Diseases; Male; Retrospective Studies

Over one-third of all patients with epilepsy are refractory to treatment and there is an urgent need to develop new drugs that can prevent the development and progression of epilepsy. Epileptogenesis is characterized by distinct histopathologic and biochemical changes, which include astrogliosis and increased expression of the adenosine-metabolizing enzyme adenosine kinase (ADK; EC 2.7.1.20). Increased expression of ADK contributes to epileptogenesis and is therefore a target for therapeutic intervention. We tested the prediction that the transient use of an ADK inhibitor administered during the latent phase of epileptogenesis can mitigate the development of epilepsy.. We used the intrahippocampal kainic acid (KA) mouse model of temporal lobe epilepsy, which is characterized by ipsilateral hippocampal sclerosis with granule cell dispersion and the development of recurrent hippocampal paroxysmal discharges (HPDs). KA-injected mice were treated with the ADK inhibitor 5-iodotubercidin (5-ITU, 1.6 mg/kg, b.i.d., i.p.) during the latent phase of epileptogenesis from day 3-8 after injury; the period when gradual increases in hippocampal ADK expression begin to manifest. HPDs were assessed at 6 and 9 weeks after KA administration followed by epilepsy histopathology including assessment of granule cell dispersion, astrogliosis, and ADK expression.. 5-ITU significantly reduced the percent time in seizures by at least 80% in 56% of mice at 6 weeks post-KA. This reduction in seizure activity was maintained in 40% of 5-ITU-treated mice at 9 weeks. 5-ITU also suppressed granule cell dispersion and prevented maladaptive ADK increases in these protected mice.. Our results show that the transient use of a small-molecule ADK inhibitor, given during the early stages of epileptogenesis, has antiepileptogenic disease-modifying properties, which provides the rationale for further investigation into the development of a novel class of antiepileptogenic ADK inhibitors with increased efficacy for epilepsy prevention.

Topics: Adenosine Kinase; Animals; Anticonvulsants; Brain; Enzyme Inhibitors; Epilepsy; Male; Mice; Mice, Inbred C57BL; Tubercidin

Adenosine kinase (ADK) is supposed to be a schizophrenia susceptibility gene based on the findings that ADK is an enzyme that catalyzes transfer of the gamma-phosphate from ATP to adenosine, which interacts with dopamine and glutamate neurotransmitters. However, no reports of schizophrenia cases with loss of function variants in the ADK region have been published. In our previous study investigating copy number variants in schizophrenia, we detected a copy number variant in the ADK region in 1 of 1699 schizophrenia patients.. We validated the ADK deletion by determining the breakpoint. Then, we compared the relative expression of ADK in 32 schizophrenia patients, including a schizophrenia patient with deletion of ADK, with 29 healthy controls using lymphoblastoid cell lines. Furthermore, we evaluated the clinical phenotypes of the schizophrenia with ADK deletion.. We validated the copy number variants with Sanger sequencing and predicted that this copy number variant results in loss of function of ADK. Furthermore, expression analysis of mRNA from peripheral blood in this schizophrenia patient with the ADK deletion showed an extremely low level of ADK. Here we describe a case report of a patient with ADK deletion with phenotypes (schizophrenia, parkinsonism, epilepsy) that are predicted when ADK function is disrupted.. Considering that the patient had a low ADK mRNA level and showed a phenotype that may be related to ADK deficiency, the copy number variants in the region of ADK may be strongly related to the phenotypes described here, such as schizophrenia, Parkinsonism, and epilepsy.

Topics: Adenosine Kinase; DNA Copy Number Variations; Epilepsy; Female; Humans; Middle Aged; Parkinson Disease; Phenotype; RNA, Messenger; Schizophrenia

Focal cortical dysplasia type IIB (FCDIIB) is a developmental malformation of the cerebral cortex that is associated with pharmacoresistant epilepsy. Overexpression of adenosine kinase (ADK) has been regarded as a pathologic hallmark of epilepsy. We hypothesized that the epileptogenic mechanisms underlying FCDIIB are related to abnormal ADK expression. We used immunohistochemistry to examine the expression of ADK and of heterogeneous cell population markers of astrocytes (glial fibrillary acidic protein), immature glia (vimentin), immature neurons (neuronal class III beta-tubulin, TUJ1), multipotential progenitor cells (nestin), mature neurons (microtubule-associated protein 2), and antiapoptotic gene products (Bcl-2) in surgically resected human epileptic cortical specimens from FCDIIB patients (n = 20). Expression patterns were compared with those in normal autopsy (n = 6) and surgical control (n = 6) brain samples. Balloon cells in FCDII lesions were immunoreactive for ADK (77%) and balloon cells expressing the different cell markers expressing different degrees of ADK. Adenosine kinase expression assessed by Western blot and enzymatic activity were also greater in FCD versus control samples. These results suggest that upregulation of ADK is a common pathologic component of FCDIIB. Adenosine kinase might, therefore, be a target in the treatment of epilepsy associated with FCD.

Topics: Adenosine Kinase; Adolescent; Adult; Cerebral Cortex; Child; Child, Preschool; Epilepsy; Female; Gene Expression Regulation, Enzymologic; Glial Fibrillary Acidic Protein; Humans; Male; Malformations of Cortical Development, Group I; Microtubule-Associated Proteins; Proto-Oncogene Proteins c-bcl-2; Tubulin; Vimentin; Young Adult

The aim of the present study was to investigate adenosine deaminase (ADA) and adenosine kinase (ADK) expression in human glioma and to explore its correlation with glioma‑associated epilepsy. Tumor tissues (n=45) and peritumoral tissues (n=14) were obtained from glioma patients undergoing surgery. Normal control tissues (n=8) were obtained from brain trauma patients. The disease grade was determined by histological evaluation and the degree of tumor invasion was evaluated using immunofluorescence analyses. mRNA and protein expression of ADA and ADK were evaluated using reverse transcription quantitative polymerase chain reaction or western blot analysis, respectively. Based on histological evaluations, four cases were classified as Grade I gliomas, 18 cases as Grade II, 17 cases as Grade III and six cases were considered Grade IV. Increased ADA and ADK expression was observed in tumor tissues. ADA was predominantly distributed in the cytoplasm of tumor cells, whereas ADK was detected in the cytoplasm as well as in the nuclei. ADA and ADK levels were upregulated in patients with Grade II and Grade III gliomas compared to those in control subjects (p<0.05). In addition, tumor invasion was detected in peritumoral tissues. The number of ADA‑positive or ADK‑positive cells in tumor tissues was similar between glioma patients with and without epilepsy (p>0.05). However, ADA and ADK expression was upregulated in peritumoral tissues derived from patients with epilepsy compared to that in glioma patients without epilepsy. The results of the present study suggested that ADA and ADK are involved in glioma progression, and that increased ADA and ADK levels in peritumoral tissues may be associated with epilepsy in glioma patients.

Topics: Adenosine Deaminase; Adenosine Kinase; Adolescent; Adult; Aged; Brain Neoplasms; Case-Control Studies; Cell Nucleus; Child; Child, Preschool; Cytoplasm; Disease Progression; Epilepsy; Female; Gene Expression Regulation, Neoplastic; Glioma; Humans; Male; Middle Aged; Neoplasm Grading

Epigenetic modifications, including changes in DNA methylation, lead to altered gene expression and thus may underlie epileptogenesis via induction of permanent changes in neuronal excitability. Therapies that could inhibit or reverse these changes may be highly effective in halting disease progression. Here we identify an epigenetic function of the brain's endogenous anticonvulsant adenosine, showing that this compound induces hypomethylation of DNA via biochemical interference with the transmethylation pathway. We show that inhibition of DNA methylation inhibited epileptogenesis in multiple seizure models. Using a rat model of temporal lobe epilepsy, we identified an increase in hippocampal DNA methylation, which correlates with increased DNA methyltransferase activity, disruption of adenosine homeostasis, and spontaneous recurrent seizures. Finally, we used bioengineered silk implants to deliver a defined dose of adenosine over 10 days to the brains of epileptic rats. This transient therapeutic intervention reversed the DNA hypermethylation seen in the epileptic brain, inhibited sprouting of mossy fibers in the hippocampus, and prevented the progression of epilepsy for at least 3 months. These data demonstrate that pathological changes in DNA methylation homeostasis may underlie epileptogenesis and reversal of these epigenetic changes with adenosine augmentation therapy may halt disease progression.

Topics: Adenosine; Adenosine Kinase; Animals; Anticonvulsants; Azacitidine; Base Sequence; Brain; CpG Islands; Decitabine; DNA (Cytosine-5-)-Methyltransferases; DNA Methylation; Drug Implants; Epigenesis, Genetic; Epilepsy; Male; Mice; Mossy Fibers, Hippocampal; Rats; Rats, Sprague-Dawley; Sequence Analysis, DNA

Adenosine kinase (ADK), a largely astrocyte-based metabolic enzyme, regulates adenosine homeostasis in the brain. Overexpression of ADK decreases extracellular adenosine and consequently leads to seizures. We hypothesized that dysfunction in the metabolism of tumor astrocytes is related to changes in ADK expression and that those changes might be associated with the development of epilepsy in patients with tumors.. We compared ADK expression and cellular distribution in surgically removed tumor tissue (n = 45) and peritumoral cortex (n = 20) of patients with glial and glioneuronal tumors to normal control tissue obtained at autopsy (n = 11). In addition, we compared ADK expression in tumor patients with and without epilepsy. To investigate ADK expression, we used immunohistochemistry and Western blot analysis. ADK activity measurement was performed in surgical specimens of astrocytomas World Health Organization (WHO) grade III (n = 3), peritumoral cortex (n = 3), and nonepileptic cortex (n = 3).. Immunohistochemistry predominantly showed cytoplasmic labeling in tumors and peritumoral tissue containing infiltrating tumor cells. ADK immunoreactivity was significantly stronger in tumor and peritumoral tissue compared to normal white matter and normal cortex, especially in astrocytoma WHO grade III, as confirmed by Western blot analysis and ADK activity measurements. Importantly, we found a significantly higher expression of ADK in the peritumoral infiltrated tissue of patients with epilepsy than in patients without epilepsy.. These results suggest a dysregulation of ADK in astrocytic brain tumors. Moreover, the upregulation of ADK observed in peritumoral infiltrated tissue of glioma patients with epilepsy supports the role of this enzyme in tumor-associated epilepsy.

Topics: Adenosine Kinase; Adolescent; Adult; Aged; Astrocytoma; Brain Neoplasms; Cerebral Cortex; Child; Epilepsy; Female; Humans; Immunohistochemistry; Male; Middle Aged; Up-Regulation; Young Adult

Given the high incidence of refractory epilepsy, novel therapeutic approaches and concepts are urgently needed. To date, viral-mediated delivery and endogenous expression of antisense sequences as a strategy to prevent seizures have received little attention in epilepsy therapy development efforts. Here we validate adenosine kinase (ADK), the astrocyte-based key negative regulator of the brain's endogenous anticonvulsant adenosine, as a potential therapeutic target for antisense-mediated seizure suppression.. We developed adenoassociated virus 8 (AAV8)-based gene therapy vectors to selectively modulate ADK expression in astrocytes. Cell type selectivity was achieved by expressing an Adk-cDNA in sense or antisense orientation under the control of an astrocyte-specific gfaABC1D promoter. Viral vectors where injected into the CA3 of wild-type mice or spontaneously epileptic Adk-tg transgenic mice that overexpress ADK in brain. After virus injection, ADK expression was assessed histologically and biochemically. In addition, intracranial electroencephalography (EEG) recordings were obtained.. We demonstrate in wild-type mice that viral overexpression of ADK within astrocytes is sufficient to trigger spontaneous recurrent seizures in the absence of any other epileptogenic event, whereas ADK downregulation via AAV8-mediated RNA interference almost completely abolished spontaneous recurrent seizures in Adk-tg mice.. Our data demonstrate that modulation of astrocytic ADK expression can trigger or prevent seizures, respectively. This is the first study to use an antisense approach to validate ADK as a rational therapeutic target for the treatment of epilepsy and suggests that gene therapies based on the knock down of ADK might be a feasible approach to control seizures in refractory epilepsy.

Topics: Adenosine Kinase; Animals; Anticonvulsants; Astrocytes; DNA, Antisense; DNA, Complementary; Electroencephalography; Epilepsy; Gene Knockdown Techniques; Genetic Therapy; Genetic Vectors; Glial Fibrillary Acidic Protein; HEK293 Cells; Humans; Mice; Mice, Transgenic; Promoter Regions, Genetic; Signal Processing, Computer-Assisted

A ketogenic diet (KD) is a high-fat, low-carbohydrate metabolic regimen; its effectiveness in the treatment of refractory epilepsy suggests that the mechanisms underlying its anticonvulsive effects differ from those targeted by conventional antiepileptic drugs. Recently, KD and analogous metabolic strategies have shown therapeutic promise in other neurologic disorders, such as reducing brain injury, pain, and inflammation. Here, we have shown that KD can reduce seizures in mice by increasing activation of adenosine A1 receptors (A1Rs). When transgenic mice with spontaneous seizures caused by deficiency in adenosine metabolism or signaling were fed KD, seizures were nearly abolished if mice had intact A1Rs, were reduced if mice expressed reduced A1Rs, and were unaltered if mice lacked A1Rs. Seizures were restored by injecting either glucose (metabolic reversal) or an A1R antagonist (pharmacologic reversal). Western blot analysis demonstrated that the KD reduced adenosine kinase, the major adenosine-metabolizing enzyme. Importantly, hippocampal tissue resected from patients with medically intractable epilepsy demonstrated increased adenosine kinase. We therefore conclude that adenosine deficiency may be relevant to human epilepsy and that KD can reduce seizures by increasing A1R-mediated inhibition.

Topics: Adenosine Kinase; Adolescent; Adult; Animals; Anticonvulsants; Diet, Ketogenic; Electroencephalography; Epilepsy; Hippocampus; Humans; Mice; Mice, Knockout; Mice, Transgenic; Receptor, Adenosine A1; Seizures; Young Adult

Sudden unexpected death in epilepsy (SUDEP) is a significant cause of mortality in people with epilepsy. Two postulated causes for SUDEP, cardiac and respiratory depression, can both be explained by overstimulation of adenosine receptors. We hypothesized that SUDEP is a consequence of a surge in adenosine as a result of prolonged seizures combined with deficient adenosine clearance; consequently, blockade of adenosine receptors should prevent SUDEP. Here we induced impaired adenosine clearance in adult mice by pharmacologic inhibition of the adenosine-removing enzymes, adenosine kinase and deaminase. Combination of impaired adenosine clearance with kainic acid-induced seizures triggered sudden death in all animals. Most importantly, the adenosine receptor antagonist caffeine, when given after seizure onset, increased survival from 23.75 +/- 1.35 min to 54.86 +/- 6.59 min (p < 0.01). Our data indicate that SUDEP is due to overactivation of adenosine receptors and that caffeine treatment after seizure onset might be beneficial.

Topics: Adenine; Adenosine; Adenosine Deaminase Inhibitors; Adenosine Kinase; Animals; Caffeine; Cause of Death; Death, Sudden; Disease Models, Animal; Enzyme Inhibitors; Epilepsy; Kainic Acid; Mice; Purinergic P1 Receptor Antagonists; Receptors, Purinergic P1; Risk Factors; Seizures; Survival Analysis; Tubercidin

Intracerebral delivery of anti-epileptic compounds represents a novel strategy for the treatment of refractory epilepsy. Adenosine is a possible candidate for local delivery based on its proven anti-epileptic effects. Neural stem cells constitute an ideal cell source for intracerebral transplantation and long-term drug delivery. In order to develop a cell-based system for the long-term delivery of adenosine, we isolated neural progenitor cells from adenosine kinase deficient mice (Adk(-/-)) and compared their differentiation potential and adenosine release properties with corresponding wild-type cells.. Fetal neural progenitor cells were isolated from the brains of Adk(-/-) and C57BL/6 mice fetuses and expanded in vitro. Before and after neural differentiation, supernatants were collected and assayed for adenosine release using liquid chromatography-tandem mass spectrometry (LC-MS/MS).. Adk(-/-) cells secreted significantly more adenosine compared to wild-type cells at any time point of differentiation. Undifferentiated Adk(-/-) cells secreted 137+/-5 ng adenosine per 10(5) cells during 24 h in culture, compared to 11+/-1 ng released from corresponding wild-type cells. Adenosine release was maintained after differentiation as differentiated Adk(-/-) cells continued to release significantly more adenosine per 24 h (47+/-1 ng per 10(5) cells) compared to wild-type cells (3+/-0.2 ng per 10(5) cells).. Fetal neural progenitor cells isolated from Adk(-/-) mice--but not those from C57BL/6 mice--release amounts of adenosine considered to be of therapeutic relevance.

Topics: Adenosine; Adenosine Kinase; Animals; Astrocytes; Blotting, Western; Cell Differentiation; Chromatography, Liquid; Epilepsy; Female; Fetal Stem Cells; Immunohistochemistry; Injections, Intraventricular; Male; Mass Spectrometry; Mice; Mice, Inbred C57BL; Mice, Knockout; Polymerase Chain Reaction; Stem Cell Transplantation

The astrocytic enzyme adenosine kinase (ADK) is a key negative regulator of the brain's endogenous anticonvulsant adenosine. Astrogliosis with concomitant upregulation of ADK is part of the epileptogenic cascade and contributes to seizure generation. To molecularly dissect the respective roles of astrogliosis and ADK-expression for seizure generation, we used a transgenic approach to uncouple ADK-expression from astrogliosis: in Adk-tg mice the endogenous Adk-gene was deleted and replaced by a ubiquitously expressed Adk-transgene with novel ectopic expression in pyramidal neurons, resulting in spontaneous seizures. Here, we followed a unique approach to selectively injure the CA3 of these Adk-tg mice. Using this strategy, we had the opportunity to study astrogliosis and epileptogenesis in the absence of the endogenous astrocytic Adk-gene. After triggering epileptogenesis we demonstrate astrogliosis without upregulation of ADK, but lack of seizures, whereas matching wild-type animals developed astrogliosis with upregulation of ADK and spontaneous recurrent seizures. By uncoupling ADK-expression from astrogliosis, we demonstrate that global expression levels of ADK rather than astrogliosis per se contribute to seizure generation.

Topics: Adenosine Kinase; Animals; Astrocytes; Brain; Cell Death; Chronic Disease; Epilepsy; Gliosis; Kainic Acid; Male; Mice; Mice, Knockout; Mice, Transgenic; Pyramidal Cells; Recurrence; Seizures; Severity of Illness Index; Status Epilepticus; Time Factors; Tissue Distribution; Transgenes; Up-Regulation

Epilepsy therapy is largely symptomatic and no effective therapy is available to prevent epileptogenesis. We therefore analysed the potential of stem cell-derived brain implants and of paracrine adenosine release to suppress the progressive development of seizures in the rat kindling-model. Embryonic stem (ES) cells, engineered to release the inhibitory neuromodulator adenosine by biallelic genetic disruption of the adenosine kinase gene (Adk-/-), and respective wild-type (wt) cells, were differentiated into neural precursor cells (NPs) and injected into the hippocampus of rats prior to kindling. Therapeutic effects of NP-derived brain implants were compared with those of wt baby hamster kidney cells (BHK) and adenosine releasing BHK cell implants (BHK-AK2), which were previously shown to suppress seizures by paracrine adenosine release. Wild-type NP-graft recipients were characterized by an initial delay of seizure development, while recipients of adenosine releasing NPs displayed sustained protection from developing generalized seizures. In contrast, recipients of wt BHK cells failed to display any effects on kindling development, while recipients of BHK-AK2 cells were only moderately protected from seizure development. The therapeutic effect of Adk(-/-)-NPs was due to graft-mediated adenosine release, since seizures could transiently be provoked after blocking adenosine A1 receptors. Histological analysis of NP-implants at day 26 revealed cell clusters within the infrahippocampal cleft as well as intrahippocampal location of graft-derived cells expressing mature neuronal markers. In contrast, BHK and BHK-AK2 cell implants only formed cell clusters within the infrahippocampal cleft. We conclude that ES cell-derived adenosine releasing brain implants are superior to paracrine adenosine release from BHK-AK2 cell implants in suppressing seizure progression in the rat kindling-model. These findings may indicate a potential antiepileptogenic function of stem cell-mediated adenosine delivery.

Topics: Adenosine; Adenosine Kinase; Animals; Brain; Cells, Cultured; Embryonic Stem Cells; Epilepsy; Genetic Engineering; Kindling, Neurologic; Male; Models, Animal; Paracrine Communication; Rats; Rats, Sprague-Dawley

Adenosine kinase deficient (Adk-/-) embryonic stem cells (ESCs) encapsulated in synthetic polymers have previously been shown to provide therapeutic adenosine release and transient seizure suppression in epileptic rats. Here we explored the utility of biopolymer-substrates to promote long-term adenosine release from Adk-/- ESCs. Three different substrates were studied: (1) type I collagen (Col-1), (2) silk-fibroin (SF), and (3) poly(L-ornithine) (PO) coated tissue culture plastic. Adk-/- or wild type (wt) ESC-derived glial precursor cells were seeded on the substrates and cultured either in proliferation medium containing growth factors or in differentiation medium devoid of growth factors. In proliferation medium cell proliferation was higher and metabolic activity lower on Col-1 and PO substrates as compared to SF. Cells from both genotypes readily differentiated into astrocytes after growth factor removal on all substrates. Adk-/- cells cultured on biopolymers released significantly more adenosine than their wt counterparts at all developmental stages. Adenosine release was similar on SF and PO substrates and the amounts released from Adk-/- cells (>20 ng/ml) were considered to be of therapeutic relevance. Taken together, these results suggest that silk matrices are particularly suitable biomaterials for ESC encapsulation and for the design of adenosine releasing bioincubators for the treatment of epilepsy.

Topics: Adenosine; Adenosine Kinase; Animals; Biocompatible Materials; Capsules; Cell Differentiation; Cell Proliferation; Cells, Cultured; Collagen Type I; Delayed-Action Preparations; Embryo, Mammalian; Epilepsy; Fibroins; Glucose; Hydrophobic and Hydrophilic Interactions; Mice; Mutation; Neuroglia; Peptides; Stem Cells