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. 1997 Mar;72(3):1458–1471. doi: 10.1016/S0006-3495(97)78792-7

Calcium dynamics associated with a single action potential in a CNS presynaptic terminal.

F Helmchen 1, J G Borst 1, B Sakmann 1
PMCID: PMC1184528  PMID: 9138591

Abstract

Calcium dynamics associated with a single action potential were studied quantitatively in the calyx of Held, a large presynaptic terminal in the rat brainstem. Terminals were loaded with different concentrations of high- or low-affinity Ca2+ indicators via patch pipettes. Spatially averaged Ca2+ signals were measured fluorometrically and analyzed on the basis of a single compartment model. A single action potential led to a total Ca2+ influx of 0.8-1 pC. The accessible volume of the terminal was about 0.4 pl; thus the total calcium concentration increased by 10-13 microM. The Ca(2+)-binding ratio of the endogenous buffer was about 40, as estimated from the competition with Fura-2, indicating that 2.5% of the total calcium remained free. This is consistent with the peak increase in free calcium concentration of about 400 nM, which was measured directly with MagFura-2. The decay of the [Ca2+]i transients was fast, with time constants of 100 ms at 23 degrees C and 45 ms at 35 degrees C, indicating Ca2+ extrusion rates of 400 and 900 s-1, respectively. The combination of the relatively low endogenous Ca(2+)-binding ratio and the high rate of Ca2+ extrusion provides an efficient mechanism for rapidly removing the large Ca2+ load of the terminal evoked by an action potential.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Adler E. M., Augustine G. J., Duffy S. N., Charlton M. P. Alien intracellular calcium chelators attenuate neurotransmitter release at the squid giant synapse. J Neurosci. 1991 Jun;11(6):1496–1507. doi: 10.1523/JNEUROSCI.11-06-01496.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Agard D. A., Hiraoka Y., Shaw P., Sedat J. W. Fluorescence microscopy in three dimensions. Methods Cell Biol. 1989;30:353–377. doi: 10.1016/s0091-679x(08)60986-3. [DOI] [PubMed] [Google Scholar]
  3. Augustine G. J., Charlton M. P. Calcium dependence of presynaptic calcium current and post-synaptic response at the squid giant synapse. J Physiol. 1986 Dec;381:619–640. doi: 10.1113/jphysiol.1986.sp016347. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Augustine G. J., Neher E. Neuronal Ca2+ signalling takes the local route. Curr Opin Neurobiol. 1992 Jun;2(3):302–307. doi: 10.1016/0959-4388(92)90119-6. [DOI] [PubMed] [Google Scholar]
  5. Barnes-Davies M., Forsythe I. D. Pre- and postsynaptic glutamate receptors at a giant excitatory synapse in rat auditory brainstem slices. J Physiol. 1995 Oct 15;488(Pt 2):387–406. doi: 10.1113/jphysiol.1995.sp020974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Blaustein M. P. Calcium transport and buffering in neurons. Trends Neurosci. 1988 Oct;11(10):438–443. doi: 10.1016/0166-2236(88)90195-6. [DOI] [PubMed] [Google Scholar]
  7. Borst J. G., Helmchen F., Sakmann B. Pre- and postsynaptic whole-cell recordings in the medial nucleus of the trapezoid body of the rat. J Physiol. 1995 Dec 15;489(Pt 3):825–840. doi: 10.1113/jphysiol.1995.sp021095. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Borst J. G., Sakmann B. Calcium influx and transmitter release in a fast CNS synapse. Nature. 1996 Oct 3;383(6599):431–434. doi: 10.1038/383431a0. [DOI] [PubMed] [Google Scholar]
  9. Brinley F. J., Jr Calcium buffering in squid axons. Annu Rev Biophys Bioeng. 1978;7:363–392. doi: 10.1146/annurev.bb.07.060178.002051. [DOI] [PubMed] [Google Scholar]
  10. Escobar A. L., Monck J. R., Fernandez J. M., Vergara J. L. Localization of the site of Ca2+ release at the level of a single sarcomere in skeletal muscle fibres. Nature. 1994 Feb 24;367(6465):739–741. doi: 10.1038/367739a0. [DOI] [PubMed] [Google Scholar]
  11. Fogelson A. L., Zucker R. S. Presynaptic calcium diffusion from various arrays of single channels. Implications for transmitter release and synaptic facilitation. Biophys J. 1985 Dec;48(6):1003–1017. doi: 10.1016/S0006-3495(85)83863-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Forsythe I. D. Direct patch recording from identified presynaptic terminals mediating glutamatergic EPSCs in the rat CNS, in vitro. J Physiol. 1994 Sep 15;479(Pt 3):381–387. doi: 10.1113/jphysiol.1994.sp020303. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Friauf E. Transient appearance of calbindin-D28k-positive neurons in the superior olivary complex of developing rats. J Comp Neurol. 1993 Aug 1;334(1):59–74. doi: 10.1002/cne.903340105. [DOI] [PubMed] [Google Scholar]
  14. Groden D. L., Guan Z., Stokes B. T. Determination of Fura-2 dissociation constants following adjustment of the apparent Ca-EGTA association constant for temperature and ionic strength. Cell Calcium. 1991 Apr;12(4):279–287. doi: 10.1016/0143-4160(91)90002-v. [DOI] [PubMed] [Google Scholar]
  15. Grynkiewicz G., Poenie M., Tsien R. Y. A new generation of Ca2+ indicators with greatly improved fluorescence properties. J Biol Chem. 1985 Mar 25;260(6):3440–3450. [PubMed] [Google Scholar]
  16. Helmchen F., Imoto K., Sakmann B. Ca2+ buffering and action potential-evoked Ca2+ signaling in dendrites of pyramidal neurons. Biophys J. 1996 Feb;70(2):1069–1081. doi: 10.1016/S0006-3495(96)79653-4. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Jackson M. B., Konnerth A., Augustine G. J. Action potential broadening and frequency-dependent facilitation of calcium signals in pituitary nerve terminals. Proc Natl Acad Sci U S A. 1991 Jan 15;88(2):380–384. doi: 10.1073/pnas.88.2.380. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Jaffe D. B., Johnston D., Lasser-Ross N., Lisman J. E., Miyakawa H., Ross W. N. The spread of Na+ spikes determines the pattern of dendritic Ca2+ entry into hippocampal neurons. Nature. 1992 May 21;357(6375):244–246. doi: 10.1038/357244a0. [DOI] [PubMed] [Google Scholar]
  19. Kao J. P., Tsien R. Y. Ca2+ binding kinetics of fura-2 and azo-1 from temperature-jump relaxation measurements. Biophys J. 1988 Apr;53(4):635–639. doi: 10.1016/S0006-3495(88)83142-4. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Konishi M., Berlin J. R. Ca transients in cardiac myocytes measured with a low affinity fluorescent indicator, furaptra. Biophys J. 1993 Apr;64(4):1331–1343. doi: 10.1016/S0006-3495(93)81494-2. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Llinás R., Sugimori M., Silver R. B. Microdomains of high calcium concentration in a presynaptic terminal. Science. 1992 May 1;256(5057):677–679. doi: 10.1126/science.1350109. [DOI] [PubMed] [Google Scholar]
  22. Llinás R., Sugimori M., Simon S. M. Transmission by presynaptic spike-like depolarization in the squid giant synapse. Proc Natl Acad Sci U S A. 1982 Apr;79(7):2415–2419. doi: 10.1073/pnas.79.7.2415. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Lohmann C., Friauf E. Distribution of the calcium-binding proteins parvalbumin and calretinin in the auditory brainstem of adult and developing rats. J Comp Neurol. 1996 Mar 25;367(1):90–109. doi: 10.1002/(SICI)1096-9861(19960325)367:1<90::AID-CNE7>3.0.CO;2-E. [DOI] [PubMed] [Google Scholar]
  24. Martinez-Serrano A., Blanco P., Satrústegui J. Calcium binding to the cytosol and calcium extrusion mechanisms in intact synaptosomes and their alterations with aging. J Biol Chem. 1992 Mar 5;267(7):4672–4679. [PubMed] [Google Scholar]
  25. Mintz I. M., Sabatini B. L., Regehr W. G. Calcium control of transmitter release at a cerebellar synapse. Neuron. 1995 Sep;15(3):675–688. doi: 10.1016/0896-6273(95)90155-8. [DOI] [PubMed] [Google Scholar]
  26. Morest D. K. The growth of synaptic endings in the mammalian brain: a study of the calyces of the trapezoid body. Z Anat Entwicklungsgesch. 1968 Nov 4;127(3):201–220. doi: 10.1007/BF00526129. [DOI] [PubMed] [Google Scholar]
  27. Neher E., Augustine G. J. Calcium gradients and buffers in bovine chromaffin cells. J Physiol. 1992 May;450:273–301. doi: 10.1113/jphysiol.1992.sp019127. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Neher E. The use of fura-2 for estimating Ca buffers and Ca fluxes. Neuropharmacology. 1995 Nov;34(11):1423–1442. doi: 10.1016/0028-3908(95)00144-u. [DOI] [PubMed] [Google Scholar]
  29. Neher E., Zucker R. S. Multiple calcium-dependent processes related to secretion in bovine chromaffin cells. Neuron. 1993 Jan;10(1):21–30. doi: 10.1016/0896-6273(93)90238-m. [DOI] [PubMed] [Google Scholar]
  30. Oliva C., Cohen I. S., Mathias R. T. Calculation of time constants for intracellular diffusion in whole cell patch clamp configuration. Biophys J. 1988 Nov;54(5):791–799. doi: 10.1016/S0006-3495(88)83017-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Regehr W. G., Atluri P. P. Calcium transients in cerebellar granule cell presynaptic terminals. Biophys J. 1995 May;68(5):2156–2170. doi: 10.1016/S0006-3495(95)80398-X. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Regehr W. G., Delaney K. R., Tank D. W. The role of presynaptic calcium in short-term enhancement at the hippocampal mossy fiber synapse. J Neurosci. 1994 Feb;14(2):523–537. doi: 10.1523/JNEUROSCI.14-02-00523.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Regehr W. G., Tank D. W. The maintenance of LTP at hippocampal mossy fiber synapses is independent of sustained presynaptic calcium. Neuron. 1991 Sep;7(3):451–459. doi: 10.1016/0896-6273(91)90297-d. [DOI] [PubMed] [Google Scholar]
  34. Reuter H., Porzig H. Localization and functional significance of the Na+/Ca2+ exchanger in presynaptic boutons of hippocampal cells in culture. Neuron. 1995 Nov;15(5):1077–1084. doi: 10.1016/0896-6273(95)90096-9. [DOI] [PubMed] [Google Scholar]
  35. Roberts W. M. Localization of calcium signals by a mobile calcium buffer in frog saccular hair cells. J Neurosci. 1994 May;14(5 Pt 2):3246–3262. doi: 10.1523/JNEUROSCI.14-05-03246.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Sala F., Hernández-Cruz A. Calcium diffusion modeling in a spherical neuron. Relevance of buffering properties. Biophys J. 1990 Feb;57(2):313–324. doi: 10.1016/S0006-3495(90)82533-9. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Schneggenburger R., Zhou Z., Konnerth A., Neher E. Fractional contribution of calcium to the cation current through glutamate receptor channels. Neuron. 1993 Jul;11(1):133–143. doi: 10.1016/0896-6273(93)90277-x. [DOI] [PubMed] [Google Scholar]
  38. Schweizer F. E., Betz H., Augustine G. J. From vesicle docking to endocytosis: intermediate reactions of exocytosis. Neuron. 1995 Apr;14(4):689–696. doi: 10.1016/0896-6273(95)90213-9. [DOI] [PubMed] [Google Scholar]
  39. Simon S. M., Llinás R. R. Compartmentalization of the submembrane calcium activity during calcium influx and its significance in transmitter release. Biophys J. 1985 Sep;48(3):485–498. doi: 10.1016/S0006-3495(85)83804-2. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Sinha S. R., Wu L. G., Saggau P. Presynaptic calcium dynamics and transmitter release evoked by single action potentials at mammalian central synapses. Biophys J. 1997 Feb;72(2 Pt 1):637–651. doi: 10.1016/s0006-3495(97)78702-2. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Smith S. J., Augustine G. J. Calcium ions, active zones and synaptic transmitter release. Trends Neurosci. 1988 Oct;11(10):458–464. doi: 10.1016/0166-2236(88)90199-3. [DOI] [PubMed] [Google Scholar]
  42. Stuart G. J., Dodt H. U., Sakmann B. Patch-clamp recordings from the soma and dendrites of neurons in brain slices using infrared video microscopy. Pflugers Arch. 1993 Jun;423(5-6):511–518. doi: 10.1007/BF00374949. [DOI] [PubMed] [Google Scholar]
  43. Stuenkel E. L. Regulation of intracellular calcium and calcium buffering properties of rat isolated neurohypophysial nerve endings. J Physiol. 1994 Dec 1;481(Pt 2):251–271. doi: 10.1113/jphysiol.1994.sp020436. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Tank D. W., Regehr W. G., Delaney K. R. A quantitative analysis of presynaptic calcium dynamics that contribute to short-term enhancement. J Neurosci. 1995 Dec;15(12):7940–7952. doi: 10.1523/JNEUROSCI.15-12-07940.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Ukhanov K. Y., Flores T. M., Hsiao H. S., Mohapatra P., Pitts C. H., Payne R. Measurement of cytosolic Ca2+ concentration in Limulus ventral photoreceptors using fluorescent dyes. J Gen Physiol. 1995 Jan;105(1):95–116. doi: 10.1085/jgp.105.1.95. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Wagner J., Keizer J. Effects of rapid buffers on Ca2+ diffusion and Ca2+ oscillations. Biophys J. 1994 Jul;67(1):447–456. doi: 10.1016/S0006-3495(94)80500-4. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Wu L. G., Saggau P. Pharmacological identification of two types of presynaptic voltage-dependent calcium channels at CA3-CA1 synapses of the hippocampus. J Neurosci. 1994 Sep;14(9):5613–5622. doi: 10.1523/JNEUROSCI.14-09-05613.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Zhao M., Hollingworth S., Baylor S. M. Properties of tri- and tetracarboxylate Ca2+ indicators in frog skeletal muscle fibers. Biophys J. 1996 Feb;70(2):896–916. doi: 10.1016/S0006-3495(96)79633-9. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Zhou Z., Misler S. Action potential-induced quantal secretion of catecholamines from rat adrenal chromaffin cells. J Biol Chem. 1995 Feb 24;270(8):3498–3505. [PubMed] [Google Scholar]

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