Now showing 1 - 10 of 22
  • Publication
    Preconditioning effects of tumor necrosis factor-α and glutamate on calcium dynamics in rat organotypic hippocampal cultures
    During cerebral ischemia, elevation of TNF-α and glutamate to pathophysiological levels in the hippocampus may induce dysregulation of normal synaptic processes, leading ultimately to cell death. Previous studies have shown that patients subjected to a mild transient ischemic attack within a critical time window prior to a more severe ischemic episode may show attenuation in the clinical severity of the stroke and result in a more positive functional outcome. In this study we have investigated the individual contribution of pre-exposure to TNF-α or glutamate in the development of ‘ischemic tolerance’ to a subsequent insult, using organotypic hippocampal cultures. At 6 days in vitro (DIV), cultures were exposed to an acute concentration of glutamate (30 μM) or TNF-α (5 ng/ml) for 30 min, followed by 24 h recovery period. We then examined the effect of the pretreatments on calcium dynamics of the cells within the CA region. We found that pretreatment with TNF-α or glutamate caused in a significant reduction in subsequent glutamate-induced Ca2+ influx 24 h later (control: 100.0 ± 0.8%, n = 7769 cells; TNF-α: 76.8 ± 1.0%, n = 5543 cells; glutamate: 75.3 ± 1.4%, n = 3859 cells; p < 0.001). Antagonism of circulating TNF-α (using infliximab, 25 μg/ml), and inhibition of the p38 MAP kinase pathway (using SB 203580, 10 μM) completely reversed this effect. However glutamate preconditioning did not appear to be mediated by p38 MAP kinase signalling, or NMDAR activation as neither SB 203580 nor D-AP5 (100 μM) altered this effect. Glutamate and TNF-α preconditioning resulted in small yet significant alterations in resting Ca2+ levels (control: 100.0 ± 0.9%, n = 2994 cells; TNF-α: 109.7 ± 1.0%, n = 2884 cells; glutamate; 93.3 ± 0.8%, n = 2899 cells; p < 0.001), TNF-α's effect reversed by infliximab and SB 203580. Both TNF-α and glutamate also resulted in the reduction of the proportion (P) of responsive cells within the CA region of the hippocampus (control; P = 0.459, 0.451 ≤ x ≥ 0.467, n = 14,968 cells, TNF-α; P = 0.40, 0.392 ≤ x ≥ 0.407, n = 15,218; glutamate; P = 0.388, 0.303 ≤ x≥ 0.396, n = 13,919 cells), and in the depression of the frequency of spontaneous Ca2+ events (vs. control: TNF-α: p > 0.00001, D = 0.0454; glutamate: p > 0.0001, D = 0.0534). Our results suggest that attenuation in resting Ca2+ activity and Ca2+ related responsiveness of cells within the CA region as a result of glutamate or TNF-α pre-exposure, may contribute to the development of ischemic tolerance.
    Scopus© Citations 12  356
  • Publication
    A role for prolyl hydroxylase domain proteins in hippocampal synaptic plasticity
    Hypoxia-inducible factors (HIFs) are key transcriptional regulators that play a major role in oxygen homeostasis. HIF activity is tightly regulated by oxygen-dependent hydroxylases, which additionally require iron and 2-oxoglutarate as cofactors. Inhibition of these enzymes has become a novel target to modulate the hypoxic response for therapeutic benefit. Inhibition of prolyl-4-hydroxylase domains (PHDs) have been shown to delay neuronal cell death and protect against ischemic injury in the hippocampus. In this study we have examined the effects of prolyl hydroxylase inhibition on synaptic transmission and plasticity in the hippocampus. Field excitatory postsynaptic potentials (fEPSPs) and excitatory postsynaptic currents (EPSCs) were elicited by stimulation of the Schaffer collateral pathway in the CA1 region of the hippocampus. Treatment of rat hippocampal slices with low concentrations (10 µM) of the iron chelator deferosoxamine (DFO) or the 2-oxoglutarate analogue dimethyloxalyl glycine (DMOG) had no effect on fEPSP. In contrast, application of 1 mM DMOG resulted in a significant decrease in fEPSP slope. Antagonism of the NMDA receptor attenuated the effects of DMOG on baseline synaptic signalling. In rat hippocampal slices pretreated with DMOG and DFO the induction of long-term potentiation (LTP) by tetanic stimulation was strongly impaired. Similarly, neuronal knockout of the single PHD family member PHD2 prevented murine hippocampal LTP. Preconditioning of PHD2 deficient hippocampi with either DMOG, DFO, or the PHD specific inhibitor JNJ-42041935, did not further decrease LTP suggesting that DMOG and DFO influences synaptic plasticity primarily by inhibiting PHDs rather than unspecific effects. These findings provide striking evidence for a modulatory role of PHD proteins on synaptic plasticity in the hippocampus.
      731Scopus© Citations 18
  • Publication
    The effect of nicotine induced behavioral sensitization on dopamine D1 receptor pharmacology: An in vivo and ex vivo study in the rat
    Behavioral sensitization is a phenomenon which can develop following repeated intermittent administration of a wide range of psychostimulants. The mechanism underlying this phenomenon is putatively involved in many neuropsychiatric disorders (incl. drug addiction) and therefore, understanding of its mechanism is of great importance. Although these different drugs have diverse molecular targets located across the brain, they share the ability to increase the release of dopamine. The aim of the present study is to investigate the effect of the dopamine D1 ligands on nicotine induced behavioral sensitization and their molecular consequences in the striatum. Wistar rats were chronically intermittent (5 days) treated with vehicle or nicotine (0.4 mg/kg; s.c.) and locomotor activity was measured. After the establishment of sensitization, rats were withdrawn for a period of 5 days followed by administration of a nicotine challenge, with or without a pre-challenge of SCH-23390 (0.03 mg/kg; i.p., a dopamine D1 antagonist) given 30 min prior to the nicotine. Either 45 min or 24 h post-challenge, the dorsal striatum was isolated and used to assess the effect of forskolin, dopamine, and dopamine D1 ligands on intracellular cAMP accumulation ex vivo. The cAMP levels were measured using LC-MS/MS. Acute nicotine administration significantly increased locomotor activity and subsequent chronic nicotine administration induced the development of locomotor sensitization. Moreover, following the 5 days withdrawal period, a nicotine challenge produced a robust sensitized response (i.e. expression of sensitization) and was antagonized by SCH-23390. Nicotine induced sensitization had no effect on forskolin stimulated cAMP accumulation however, it increased the efficacy of dopamine for the dopamine D1 receptor, and decreased the potency of D1 agonists. These effects were antagonized by in vivo pre-challenge with SCH-23390. Moreover, a time dependent effect was observed between tissue taken at 45 min and 24 h post-challenge. In conclusion, the present findings provide a connection between behavioral sensitization and intracellular cAMP accumulation through the dopamine D1 receptor. Therefore D1 signaling in the dorsal striatum may play an important role in the underlying mechanism of the expression of nicotine induced behavioral sensitization.
      549Scopus© Citations 12
  • Publication
    Frequency-dependent modulation of dopamine release by nicotine and dopamine D1 ligands: an in vitro fast cyclic voltammetry study in rat striatum
    Nicotine is a highly addictive drug and exerts this effect partially through the modulation of dopamine release and increasing extracellular dopamine in regions such as the brain reward systems. Nicotine acts in these regions on nicotinic acetylcholine receptors. The effect of nicotine on the frequency dependent modulation of dopamine release is well established and the purpose of this study was to investigate whether dopamine D1 receptor (D1R) ligands have an influence on this. Using fast cyclic voltammetry and rat corticostriatal slices, we show that D1R ligands are able to modulate the effect of nicotine on dopamine release. Nicotine (500nM) induced a decrease in dopamine efflux at low frequency (single pulse or 5 pulses at 10Hz) and an increase at high frequency (100Hz) electrical field stimulation. The D1R agonist SKF-38393, whilst having no effect on dopamine release on its own or on the effect of nicotine upon multiple pulse evoked dopamine release, did significantly prevent and reverse the effect of nicotine on single pulse dopamine release. Interestingly similar results were obtained with the D1R antagonist SCH-23390. In this study we have demonstrated that the modulation of dopamine release by nicotine can be altered by D1R ligands, but only when evoked by single pulse stimulation, and are likely working via cholinergic interneuron driven dopamine release.
      255Scopus© Citations 7
  • Publication
    A role for tumor necrosis factor-alpha in ischemia and ischemic preconditioning
    (BioMed Central, 2011-08-02) ;
    During cerebral ischemia, elevation of TNF-α and glutamate to pathophysiological levels may induce dysregulation of normal synaptic processes, leading ultimately to cell death. Previous studies have shown that patients subjected to a mild transient ischemic attack within a critical time window prior to a more severe ischemic episode may show attenuation in the clinical severity of the stroke and result in a more positive functional outcome. Studies with organotypic hippocampal cultures and mixed primary hippocampal cultures have shown that prior incubation with low concentrations of glutamate and TNF-α increase the resistance of neurones to a subsequent insult from glutamate, AMPA and NMDA, while co-exposure of TNF-α and for example AMPA may have neuroprotective effects compared to cultures exposed to excitotoxic agents alone. In addition our work has shown that although glutamate and TNF-α pretreatment induces analogous levels of desensitisation of the intracellular calcium dynamics of neurons under resting conditions and in response to acute glutamate stimulation, their downstream signalling pathways involved in this response do not converge. Glutamate and TNF-α would appear to have opposing effects on resting Ca2+ levels which supports the proposal that they have distinct modes of preconditioning
      219Scopus© Citations 79
  • Publication
    Neuroimmunology and synaptic function
    (Elsevier, 2015-09)
    This Special Issue of Neuropharmacology is devoted to specific aspects of neuroimmunology and synaptic function. It contains 12 invited reviews from eminent scientists from all around the globe. These distinguished experts in the field of neuroimmunology and neuroscience provide exciting reviews on a range of topics, which include pro-inflammatory cytokines, cannabinoids, obesity, and neurodegenerative diseases. These articles demonstrate that there is a diverse impact of the immune system in synaptic function. Whilst it has been many years since it was first recognized that the immune and central nervous system communicate, it has only been in the last 20 years or so that detailed knowledge has begun to appear about this interaction. Neurons and glia and indeed other cell types are involved in many neuro-modulatory cross talk mechanisms. For example cytokines can modulate both directly and indirectly neuronal activity in both the central and peripheral nervous systems. We also now know that cells in the large and small intestine can directly communicate with many peripheral and central neurons and glial cells. Indeed research into how immune molecules modulate synaptic function could be considered a relatively new field of scientific investigation.
      445Scopus© Citations 2
  • Publication
    The hypoxia mimetic protocatechuic acid ethyl ester inhibits synaptic signaling and plasticity in the rat hippocampus
    (Elsevier, 2018-01-15) ;
    During hypoxia a number of physiological changes occur within neurons including the stabilization of hypoxia-inducible factors (HIFs). The activity of these proteins is regulated by O2, Fe2+, 2-OG and ascorbate-dependant hydroxylases which contain prolyl-4-hydroxylase domains (PHDs). PHD inhibitors have been widely used and have been shown to have a preconditioning and protective effect against a later and more severe hypoxic insult. In this study we have investigated the neuroprotective effects of the PHD inhibitor, protocatechuic acid ethyl ester (ethyl 3,4, dihydroxybenzoate: EDHB) as well as its effects on synaptic transmission and plasticity in the rat hippocampus using electrophysiological techniques. We report for the first time, an acute concentration-dependent and reversible inhibitory effect of EDHB (10–100 μM) on synaptic transmission in the dentate gyrus but not Cornu Ammonis 1 (CA1) region which does not affect cell viability. This effect was attenuated through the application of the NMDA or GABAA receptor antagonists, AP-5 and picrotoxin in the dentate gyrus. There were no changes in the ratio of paired responses after EDHB application suggesting a post-synaptic mechanism of action. EDHB (100 μM), was found to inhibit synaptic plasticity in both the dentate gyrus and CA1 regions. Application of exogenous Fe2+ (100 μM) or digoxin (100 nM) did not reverse EDHB’s inhibitory effect on synaptic transmission or plasticity in both regions, suggesting that its effects may be HIF-independent. These results highlight a novel modulatory role for the PHD inhibitor EDHB in hippocampal synaptic transmission and plasticity. A novel post-synaptic mechanism of action may be involved possibly involving NMDA and GABAA receptor activation.
      552Scopus© Citations 7
  • Publication
    Hypoxia-inducible factor signaling mechanisms in the central nervous system
    (John Wiley & Sons, 2013-06-14) ;
    In the CNS neurons are highly sensitive to the availability of oxygen. In conditions where oxygen availability is decreased neuronal function can be altered, leading to injury and cell death. Hypoxia has been implicated in a number of central nervous system pathologies including stroke, head trauma, and neurodegenerative diseases. Depending on the duration and severity of the oxygen deprivation, cellular oxygen-sensor responses activate a variety of short- and long-term energy saving and cellular protection mechanisms.  Failure of synaptic transmission can be observed within minutes following this hypoxia. The acute affects of hypoxia on synaptic transmission are primarily mediated by altering ion fluxes across membranes, presynaptic effects of adenosine and other actions at glutamatergic receptors. A more long-term feature of the response of neurons to hypoxia is the activation of transcription factors such as hypoxia inducible factor. The activation of hypoxia inducible factor is governed by a family of dioxygenases called hypoxia inducible factor prolyl 4 hydroxylases (PHDs). Under hypoxic conditions, PHD activity is inhibited, thereby allowing hypoxia inducible factor to accumulate and translocate to the nucleus, where it binds to the hypoxia-responsive element sequences of target gene promoters. Inhibition of PHD activity stabilizes hypoxia inducible factor and other proteins thus acting as a neuroprotective agent. This review will focus on the response of neuronal cells to hypoxia inducible factor and its targets, including the prolyl hydroxylases. We also present evidence for acute effects of PHD inhibition on synaptic transmission and plasticity in the hippocampus.
      1000Scopus© Citations 41
  • Publication
    Theta-Burst Stimulation of Hippocampal Slices Induces Network-Level Calcium Oscillations and Activates Analogous Gene Transcription to Spatial Learning
    Over four decades ago, it was discovered that high-frequency stimulation of the dentate gyrus induces long-term potentiation (LTP) of synaptic transmission. LTP is believed to underlie how we process and code external stimuli before converting it to salient information that we store as 'memories'. It has been shown that rats performing spatial learning tasks display theta-frequency (3–12 Hz) hippocampal neural activity. Moreover, administering theta-burst stimulation (TBS) to hippocampal slices can induce LTP. TBS triggers a sustained rise in intracellular calcium [Ca2+]i in neurons leading to new protein synthesis important for LTP maintenance. In this study, we measured TBS-induced [Ca2+]i oscillations in thousands of cells at increasing distances from the source of stimulation. Following TBS, a calcium wave propagates radially with an average speed of 5.2 µm/s and triggers multiple and regular [Ca2+]i oscillations in the hippocampus. Interestingly, the number and frequency of [Ca2+]i fluctuations post-TBS increased with respect to distance from the electrode. During the post-tetanic phase, 18% of cells exhibited 3 peaks in [Ca2+]i with a frequency of 17 mHz, whereas 2.3% of cells distributed further from the electrode displayed 8 [Ca2+]i oscillations at 33 mHz. We suggest that these observed [Ca2+]i oscillations could lead to activation of transcription factors involved in synaptic plasticity. In particular, the transcription factor, NF-κB, has been implicated in memory formation and is up-regulated after LTP induction. We measured increased activation of NF-κB 30 min post-TBS in CA1 pyramidal cells and also observed similar temporal up-regulation of NF-κB levels in CA1 neurons following water maze training in rats. Therefore, TBS of hippocampal slice cultures in vitro can mimic the cell type-specific up-regulations in activated NF-κB following spatial learning in vivo. This indicates that TBS may induce similar transcriptional changes to spatial learning and that TBS-triggered [Ca2+]i oscillations could activate memory-associated gene expression.
      333Scopus© Citations 10
  • Publication
    Acute hypoxic exposure and prolyl-hydroxylase inhibition improves synaptic transmission recovery time from a subsequent hypoxic insult in rat hippocampus
    In the CNS short episodes of acute hypoxia can result in a decrease in synaptic transmission which may be fully reversible upon re-oxygenation. Stabilization of hypoxia-inducible factor (HIF) by inhibition of prolyl hydroxylase domain (PHD) enzymes has been shown to regulate the cellular response to hypoxia and confer neuroprotection both in vivo and in vitro. Hypoxic preconditioning has become a novel therapeutic target to induce neuroprotection during hypoxic insults. However, there is little understanding of the effects of repeated hypoxic insults or pharmacological PHD inhibition on synaptic signalling. In this study we have assessed the effects of hypoxic exposure and PHD inhibition on synaptic transmission in the rat CA1 hippocampus. Field excitatory postsynaptic potentials (fEPSPs) were elicited by stimulation of the Schaffer collatoral pathway. 30 min hypoxia (gas mixture 95% N2/5% CO2) resulted in a significant and fully reversible decrease in fEPSP slope associated with decreases in partial pressures of tissue oxygen. 15-30 min of hypoxia was sufficient to induce stabilization of HIF in hippocampal slices. Exposure to a second hypoxic insult after 60 min resulted in a similar depression of fEPSP slope but with a significantly greater rate of recovery of the fEPSP. Prior single treatment of slices with the PHD inhibitor, dimethyloxalylglycine (DMOG) also resulted in a significantly greater rate of recovery of fEPSP post hypoxia. These results suggest that hypoxia and ‘pseudohypoxia’ preconditioning may improve the rate of recovery of hippocampal neurons to a subsequent acute hypoxia.
      555Scopus© Citations 8