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Functional contributions of glutamate transporters at the parallel fibre to Purkinje neuron synapse-relevance for the progression of cerebellar ataxia.

Power EM, Empson RM - Cerebellum Ataxias (2014)

Bottom Line: The enhanced PN excitability also recruited a presynaptic mGluR4 dependent mechanism that modified short term plasticity at the PF synapse.Our findings indicate that reduced glutamate transporter activity, as occurs in the early stages of some forms of human cerebellar ataxias, excessively excites PNs and disrupts the timing of their output.Our findings raise the possibility that sustaining cerebellar glutamate uptake may provide a therapeutic approach to prevent this disruption and the glutamate excitotoxicity-induced PN death that signals the end point of the disease.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, Brain Health Research Centre, University of Otago School of Medical Sciences, PO Box 56, 9054 Dunedin, New Zealand.

ABSTRACT

Background: Rapid uptake of glutamate by neuronal and glial glutamate transporters (EAATs, a family of excitatory amino acid transporters) is critical for shaping synaptic responses and for preventing excitotoxicity. Two of these transporters, EAAT4 in Purkinje neurons (PN) and EAAT1 in Bergmann glia are both enriched within the cerebellum and altered in a variety of human ataxias.

Results: PN excitatory synaptic responses and firing behaviour following high frequency parallel fibre (PF) activity commonly encountered during sensory stimulation in vivo were adversely influenced by acute inhibition of glutamate transporters. In the presence of a non-transportable blocker of glutamate transporters we observed very large amplitude and duration excitatory postsynaptic currents accompanied by excessive firing of the PNs. A combination of AMPA and mGluR1, but not NMDA, type glutamate receptor activation powered the hyper-excitable PN state. The enhanced PN excitability also recruited a presynaptic mGluR4 dependent mechanism that modified short term plasticity at the PF synapse.

Conclusions: Our findings indicate that reduced glutamate transporter activity, as occurs in the early stages of some forms of human cerebellar ataxias, excessively excites PNs and disrupts the timing of their output. Our findings raise the possibility that sustaining cerebellar glutamate uptake may provide a therapeutic approach to prevent this disruption and the glutamate excitotoxicity-induced PN death that signals the end point of the disease.

No MeSH data available.


Related in: MedlinePlus

The mGluR4 (type III) receptor antagonist MPPG specifically prevented the change in paired pulse facilitation (PPF) upon inhibition of glutamate uptake.A shows that application of 50 μM TBOA in the presence of the mGluR4 antagonist MPPG no longer changed the PPF and that PPF was also not significantly changed by 25 μM MPPG alone. The graph shows the individual and mean values of the changes in PPF, bars are mean values and error bars are SEM, filled circles show values from individual neurons and connecting grey lines show response changes in individual neurons. The statistical comparison used a repeated measures oneway ANOVA where ns denotes not significant, F2,23 = 1.1, P = 0.35. Importantly the first EPSC was also unchanged (data not shown) in these experiments, F2,23 = 4.4, P = 0.08. TBOA still increased the time course of the recovery of the second of the pair of EPSCs, consistent with its postsynaptic action independent of the presence of MPPG, mean values changed from 14.7 ± 1.6 ms in control cells, to 14.2 ± 1.5 ms in MPPG alone treated cells to 21.7 ± 3.4 ms in MPPG and TBOA treated cells (F2,23 = 11.8, P < 0.05 repeated measures one way ANOVA). B shows for the same example cell as in A that the amplitude of the 200 Hz EPSC also increased in the presence of TBOA and MPPG, the graph is as for A above with a repeated measures oneway ANOVA where F2,14 = 8.7, P < 0.05 and multiple comparisons where * represents P < 0.05 and ns denotes not significant.
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Fig5: The mGluR4 (type III) receptor antagonist MPPG specifically prevented the change in paired pulse facilitation (PPF) upon inhibition of glutamate uptake.A shows that application of 50 μM TBOA in the presence of the mGluR4 antagonist MPPG no longer changed the PPF and that PPF was also not significantly changed by 25 μM MPPG alone. The graph shows the individual and mean values of the changes in PPF, bars are mean values and error bars are SEM, filled circles show values from individual neurons and connecting grey lines show response changes in individual neurons. The statistical comparison used a repeated measures oneway ANOVA where ns denotes not significant, F2,23 = 1.1, P = 0.35. Importantly the first EPSC was also unchanged (data not shown) in these experiments, F2,23 = 4.4, P = 0.08. TBOA still increased the time course of the recovery of the second of the pair of EPSCs, consistent with its postsynaptic action independent of the presence of MPPG, mean values changed from 14.7 ± 1.6 ms in control cells, to 14.2 ± 1.5 ms in MPPG alone treated cells to 21.7 ± 3.4 ms in MPPG and TBOA treated cells (F2,23 = 11.8, P < 0.05 repeated measures one way ANOVA). B shows for the same example cell as in A that the amplitude of the 200 Hz EPSC also increased in the presence of TBOA and MPPG, the graph is as for A above with a repeated measures oneway ANOVA where F2,14 = 8.7, P < 0.05 and multiple comparisons where * represents P < 0.05 and ns denotes not significant.

Mentions: We therefore turned to mGluR4, a metabotropic glutamate receptor known to be expressed presynaptically at PFs [37] and known to depress their glutamate release probability [38]. As seen in Figure 5A, 25 μM MPPG, an antagonist of mGluR4 at PFs [39] abolished the TBOA-enhanced PPF. MPPG did not influence the TBOA-induced slower recovery of the second of the pair of EPSCs nor did it prevent the enhanced amplitude of the 200 Hz evoked large EPSC (both P > 0.05 in multiple comparisons from one way ANOVAs), see also Figure 5B. This strongly supports the action of excess glutamate at PF presynaptic mGluR4 receptors. MPPG also did not itself influence PPF, see Figure 5A, or the amplitude of the first EPSC (mean values were unchanged from control 204 ± 26 pA to 217 ± 32 in MPPG and 239 ± 39 pA in TBOA and MPPG, not significant in a one way ANOVA). Although KA-R also contribute to enhanced facilitation following high frequency PF stimulation [40] we did not observe any remaining PPF enhancement in the presence of TBOA and MPPG suggesting that presynaptic KA-R were unlikely to be recruited under our conditions.Figure 5


Functional contributions of glutamate transporters at the parallel fibre to Purkinje neuron synapse-relevance for the progression of cerebellar ataxia.

Power EM, Empson RM - Cerebellum Ataxias (2014)

The mGluR4 (type III) receptor antagonist MPPG specifically prevented the change in paired pulse facilitation (PPF) upon inhibition of glutamate uptake.A shows that application of 50 μM TBOA in the presence of the mGluR4 antagonist MPPG no longer changed the PPF and that PPF was also not significantly changed by 25 μM MPPG alone. The graph shows the individual and mean values of the changes in PPF, bars are mean values and error bars are SEM, filled circles show values from individual neurons and connecting grey lines show response changes in individual neurons. The statistical comparison used a repeated measures oneway ANOVA where ns denotes not significant, F2,23 = 1.1, P = 0.35. Importantly the first EPSC was also unchanged (data not shown) in these experiments, F2,23 = 4.4, P = 0.08. TBOA still increased the time course of the recovery of the second of the pair of EPSCs, consistent with its postsynaptic action independent of the presence of MPPG, mean values changed from 14.7 ± 1.6 ms in control cells, to 14.2 ± 1.5 ms in MPPG alone treated cells to 21.7 ± 3.4 ms in MPPG and TBOA treated cells (F2,23 = 11.8, P < 0.05 repeated measures one way ANOVA). B shows for the same example cell as in A that the amplitude of the 200 Hz EPSC also increased in the presence of TBOA and MPPG, the graph is as for A above with a repeated measures oneway ANOVA where F2,14 = 8.7, P < 0.05 and multiple comparisons where * represents P < 0.05 and ns denotes not significant.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Fig5: The mGluR4 (type III) receptor antagonist MPPG specifically prevented the change in paired pulse facilitation (PPF) upon inhibition of glutamate uptake.A shows that application of 50 μM TBOA in the presence of the mGluR4 antagonist MPPG no longer changed the PPF and that PPF was also not significantly changed by 25 μM MPPG alone. The graph shows the individual and mean values of the changes in PPF, bars are mean values and error bars are SEM, filled circles show values from individual neurons and connecting grey lines show response changes in individual neurons. The statistical comparison used a repeated measures oneway ANOVA where ns denotes not significant, F2,23 = 1.1, P = 0.35. Importantly the first EPSC was also unchanged (data not shown) in these experiments, F2,23 = 4.4, P = 0.08. TBOA still increased the time course of the recovery of the second of the pair of EPSCs, consistent with its postsynaptic action independent of the presence of MPPG, mean values changed from 14.7 ± 1.6 ms in control cells, to 14.2 ± 1.5 ms in MPPG alone treated cells to 21.7 ± 3.4 ms in MPPG and TBOA treated cells (F2,23 = 11.8, P < 0.05 repeated measures one way ANOVA). B shows for the same example cell as in A that the amplitude of the 200 Hz EPSC also increased in the presence of TBOA and MPPG, the graph is as for A above with a repeated measures oneway ANOVA where F2,14 = 8.7, P < 0.05 and multiple comparisons where * represents P < 0.05 and ns denotes not significant.
Mentions: We therefore turned to mGluR4, a metabotropic glutamate receptor known to be expressed presynaptically at PFs [37] and known to depress their glutamate release probability [38]. As seen in Figure 5A, 25 μM MPPG, an antagonist of mGluR4 at PFs [39] abolished the TBOA-enhanced PPF. MPPG did not influence the TBOA-induced slower recovery of the second of the pair of EPSCs nor did it prevent the enhanced amplitude of the 200 Hz evoked large EPSC (both P > 0.05 in multiple comparisons from one way ANOVAs), see also Figure 5B. This strongly supports the action of excess glutamate at PF presynaptic mGluR4 receptors. MPPG also did not itself influence PPF, see Figure 5A, or the amplitude of the first EPSC (mean values were unchanged from control 204 ± 26 pA to 217 ± 32 in MPPG and 239 ± 39 pA in TBOA and MPPG, not significant in a one way ANOVA). Although KA-R also contribute to enhanced facilitation following high frequency PF stimulation [40] we did not observe any remaining PPF enhancement in the presence of TBOA and MPPG suggesting that presynaptic KA-R were unlikely to be recruited under our conditions.Figure 5

Bottom Line: The enhanced PN excitability also recruited a presynaptic mGluR4 dependent mechanism that modified short term plasticity at the PF synapse.Our findings indicate that reduced glutamate transporter activity, as occurs in the early stages of some forms of human cerebellar ataxias, excessively excites PNs and disrupts the timing of their output.Our findings raise the possibility that sustaining cerebellar glutamate uptake may provide a therapeutic approach to prevent this disruption and the glutamate excitotoxicity-induced PN death that signals the end point of the disease.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, Brain Health Research Centre, University of Otago School of Medical Sciences, PO Box 56, 9054 Dunedin, New Zealand.

ABSTRACT

Background: Rapid uptake of glutamate by neuronal and glial glutamate transporters (EAATs, a family of excitatory amino acid transporters) is critical for shaping synaptic responses and for preventing excitotoxicity. Two of these transporters, EAAT4 in Purkinje neurons (PN) and EAAT1 in Bergmann glia are both enriched within the cerebellum and altered in a variety of human ataxias.

Results: PN excitatory synaptic responses and firing behaviour following high frequency parallel fibre (PF) activity commonly encountered during sensory stimulation in vivo were adversely influenced by acute inhibition of glutamate transporters. In the presence of a non-transportable blocker of glutamate transporters we observed very large amplitude and duration excitatory postsynaptic currents accompanied by excessive firing of the PNs. A combination of AMPA and mGluR1, but not NMDA, type glutamate receptor activation powered the hyper-excitable PN state. The enhanced PN excitability also recruited a presynaptic mGluR4 dependent mechanism that modified short term plasticity at the PF synapse.

Conclusions: Our findings indicate that reduced glutamate transporter activity, as occurs in the early stages of some forms of human cerebellar ataxias, excessively excites PNs and disrupts the timing of their output. Our findings raise the possibility that sustaining cerebellar glutamate uptake may provide a therapeutic approach to prevent this disruption and the glutamate excitotoxicity-induced PN death that signals the end point of the disease.

No MeSH data available.


Related in: MedlinePlus