Postfusional regulation of cleft glutamate concentration during LTP at ‘silent synapses’

Abstract

‘Silent synapses’ show responses from high-affinity NMDA receptors (NMDARs) but not low-affinity AMPA receptors (AMPARs), but gain AMPAR responses upon long-term potentiation (LTP). Using the rapidly reversible NMDAR antagonist l-AP5 to assess cleft glutamate concentration ([glu]cleft), we found that it peaked at 170 μM at silent neonatal synapses, but greatly increased after potentiation. Cyclothiazide (CTZ), a potentiator of AMPAR, revealed slowly rising AMPA EPSCs at silent synapses; LTP shortened their rise times. Thus, LTP at silent synapses increased rate-of-rise and peak amplitude of [glu]cleft. Release probability reported by NMDARs remained unchanged during LTP, implying that [glu]cleft increases arose from immediately presynaptic terminals. Our data suggest that changes in the dynamics of fusion-pore opening contribute to LTP.

Access optionsAccess options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Figure 1: Binding and unbinding kinetics of l-AP5 on NMDA receptors in outside-out patches.
Figure 2: l-AP5 (250 μM) completely inhibited NMDAR-mediated synaptic responses at silent synapses but lost effectiveness after pairing.
Figure 3: CTZ does not alter glutamate transporter currents recorded from astrocytes in stratum radiatum of area CA1, but enhances AMPA receptor function.
Figure 4: Potentiation of receptor function by CTZ revealed small, often slowly rising AMPAR-mediated synaptic responses from silent synapses.
Figure 5: In the presence of CTZ, pairing-induced potentiation altered EPSC kinetics.

References

  1. 1

    Bliss T. V. P. & Collingridge, G. L. A synaptic model of memory: long-term potentiation in the hippocampus. Nature 361, 31–39 (1993).

  2. 2

    Kullmann, D. M. & Siegelbaum, S. A. The site of expression of NMDA receptor-dependent LTP: new fuel for an old fire. Neuron 15, 997–1002 (1995).

  3. 3

    Malenka, R. C. & Nicoll, R. A. Long-term potentiation—a decade of progress? Science 285, 1870–1874 (1999).

  4. 4

    Liao, D., Hessler, N. A. & Malinow, R. A ctivation of postsynaptically silent synapses during pairing-induced LTP in CA1 region of hippocampal slice. Nature 375, 400–404 (1995).

  5. 5

    Isaac, J. T., Nicoll, R. A. & Malenka, R. C. Evidence for silent synapses: implications for the expression of LTP. Neuron 15, 427–434 (1995).

  6. 6

    Durand, G. M., Kovalchuk, Y. & Konnerth, A. Long-term potentiation and functional synapse induction in developing hippocampus. Nature 381, 71–75 (1996).

  7. 7

    Nüsser, Z., et al. Cell type and pathway dependence of synaptic AMPA receptor number and variability in the hippocampus. Neuron 21, 545–559 (1998).

  8. 8

    Shi, S. H. et al. Rapid spine delivery and redistribution of AMPA receptors after synaptic NMDA receptor activation. Science 284, 1811–1816 (1999).

  9. 9

    Malgaroli, A., et al. Presynaptic component of long-term potentiation visualized at individual hippocampal synapses. Science 268, 1624–1628 (1995).

  10. 10

    Ryan, T. A., Ziv, N. E. & Smith, S. J. Potentiation of evoked vesicle turnover at individually resolved synaptic boutons. Neuron 17, 125–134 (1996).

  11. 11

    Ma, L., Zablow, L., Kandel, E. R. & Siegelbaum, S. A. Cyclic AMP induces functional presynaptic boutons in hippocampal CA3–CA1 neuronal cultures. Nat. Neurosci. 2, 24–30 (1999).

  12. 12

    Patneau, D. K. & Mayer, M. L. Structure-activity relationships for amino acid transmitter candidates acting at N-methyl-d-asartate and quisqualate receptors. J. Neurosci. 10, 2385–2399 (1990).

  13. 13

    Perkel, D. J. & Nicoll, R. A. Evidence of all-or-none regulation of neurotransmitter release: implications for long-term potentiation. J. Physiol. (Lond.) 471, 481–500.

  14. 14

    Kullmann, D. M. Excitatory synapses. Neither too loud nor too quiet. Nature 13, 111–112 (1999).

  15. 15

    Clements, J. D., Lester, R. A., Tong, G., Jahr, C. E. & Westbrook, G. L. The time course of glutamate in the synaptic cleft. Science 258, 1498–1501 (1992).

  16. 16

    Kullmann, D. M. & Asztely, F. Extrasynaptic glutamate spillover in the hippocampus: evidence and implications. Trends Neurosci. 21, 8–14 (1998).

  17. 17

    Bruns, D. & Jahn, R. Real-time measurement of transmitter release from single synaptic vesicles. Nature 377, 62–65 (1995).

  18. 18

    Zhou, Z, Misler, S. & Chow, R. H. Rapid fluctuations in transmitter release from single vesicles in bovine adrenal chromaffin cells. Biophys. J. 70, 1543–1552 (1996).

  19. 19

    Spruce, A. E., Breckenridge, L. J., Lee, A. K. & Almers, W. Properties of the fusion pore that forms during exocytosis of a mast cell secretory vesicle. Neuron 4, 643–654 (1990).

  20. 20

    Lollike, K., Borregaard, N. & Lindau, M. Capacitance flickers and pseudoflickers of small granules, measured in the cell-attached configuration. Biophys. J. 75, 53–59 (1998).

  21. 21

    Tong, G. & Jahr, C. E. Multivesicular release from excitatory synapses of cultured hippocampal neurons. Neuron 12, 51–59 (1994).

  22. 22

    Clements, J. D. Transmitter timecourse in the synaptic cleft: its role in central synaptic function. Trends Neurosci. 19, 163–171 (1996).

  23. 23

    Olverman, H. J., Jones, A. W., Mewett, K. N. & Watkins, J. C. Structure/activity relations of N-methyl-d-aspartate receptor ligands as studied by their inhibition of [3H]D-2–amino-5–phosphonopentanoic acid binding in rat brain membranes. Neuroscience 26, 17–31 (1988).

  24. 24

    Rumpel, S., Hatt, H. & Gottmann, K. Silent synapses in the developing rat visual cortex: evidence for postsynaptic expression of synaptic plasticity. J. Neurosci. 18, 8863–8874 (1998).

  25. 25

    Malinow, R. & Tsien, R. W. Presynaptic enhancement shown by whole-cell recordings of long-term potentiation in hippocampal slices. Nature 346, 177–180 (1990).

  26. 26

    Diamond, J. S. & Jahr, C. E. Transporters buffer synaptically released glutamate on a submillisecond time scale. J. Neurosci. 17, 4672–4687 (1997).

  27. 27

    Rosenmund, C., Stern-Bach, Y. & Stevens, C. F. The tetrameric structure of a glutamate receptor channel. Science 280, 1596–1599 (1998).

  28. 28

    Dzubay, J. A. & Jahr, C. E. The concentration of synaptically released glutamate outside of the climbing fiber-purkinje cell synaptic cleft. J. Neurosci. 19, 5265–5274 (1999).

  29. 29

    Partin, K. M., Patneau, D. K., Winters, C. A., Mayer, M. L. & Buonanno, A. Selective modulation of desensitization at AMPA versus kainate receptors by cyclothiazide and concanavalin A. Neuron 11, 1069–1082 (1993).

  30. 30

    Yamada, K. A. & Tang, C. M Benzothiadiazides inhibit rapid glutamate receptor desensitization and enhance glutamatergic synaptic currents. J. Neurosci. 13, 3904–3915 (1993).

  31. 31

    Diamond, J. S. & Jahr, C. E. Asynchronous release of synaptic vesicles determines the time course of the AMPA receptor-mediated EPSC. Neuron 15, 1097–1107 (1995).

  32. 32

    Bellingham, M. C. & Walmsley, B. A novel presynaptic inhibitory mechanism underlies paired pulse depression at a fast central synapse. Neuron 23, 159–170 (1999).

  33. 33

    Lüscher, C. Malenka, R. C. & Nicoll, R. A. Monitoring glutamate release during LTP with glial transporter currents. Neuron 21, 435–441 (1998).

  34. 34

    Diamond, J. S., Bergles, D. E. & Jahr, C. E. Glutamate release monitored with astrocyte transporter currents during LTP. Neuron 21, 425–433 (1998).

  35. 35

    Colquhoun, D., Jonas, P. & Sakmann, B. Action of brief pulses of glutamate on AMPA/kainate receptors in patches from different neurones of rat hippocampal slices. J. Physiol. (Lond.) 458, 261–287 (1992).

  36. 36

    Bekkers, J. M. & Stevens, C. F. Cable properties of cultured hippocampal neurons determined from sucrose-evoked miniature EPSCs. J. Neurophysiol. 75, 1250–1255 (1996).

  37. 37

    Glavinovic M. I. & Rabie, H. R. Monte Carlo simulation of spontaneous miniature excitatory postsynaptic currents in rat hippocampal synapse in the presence and absence of desensitization. Pflugers Arch. 435, 193–202 (1998).

  38. 38

    Pothos, E. N., Przedborski, S., Davila, V., Schmitz, Y. & Sulzer, D. D2-like dopamine autoreceptor activation reduces quantal size in PC12 cells. J. Neurosci. 18, 4106–4118 (1998).

  39. 39

    Rossi, D. J. & Hamann, M. Spillover-mediated transmission at inhibitory synapses promoted by high affinity alpha6 subunit GABA(A) receptors and glomerular geometry. Neuron 20, 783–795 (1998).

  40. 40

    Scepek, S., Coorssen, J. R. & Lindau, M. Fusion pore expansion in horse eosinophils is modulated by Ca2+ and protein kinase C via distinct mechanisms. EMBO J. 17, 4340–4346 (1998).

  41. 41

    Malinow, R., Schulman, H. & Tsien, R. W. Inhibition of postsynaptic PKC or CaMKII blocks induction but not expression of LTP. Science 245, 862–866 (1989).

  42. 42

    Bekkers, J. M. & Stevens, C. F. Presynaptic mechanism for long-term potentiation in the hippocampus. Nature 346, 724–729 (1990).

  43. 43

    Malgaroli, A & Tsien, R. W. Glutamate-induced long-term potentiation of the frequency of miniature synaptic currents in cultured hippocampal neurons. Nature 357, 134–139 (1992).

Download references

Acknowledgements

This work was supported by the Silvio Conte-NIMH Center for Neuroscience Research at Stanford (R.W.T.), a Dean's Fellowship (S.C.) and a fellowship of the Boehringer Ingelheim Fonds (J.K.). We thank D.V. Madison and P. Mermelstein for comments on the manuscript, N. C. Harata and E. T. Kavalali for discussions and D. Wheeler and D. Profitt for technical support. We are grateful to other members of the Tsien lab for their support.

Author information

Correspondence to Richard W. Tsien.

Rights and permissions

Reprints and Permissions

About this article

Further reading