1. Neuroscience
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Human pyramidal to interneuron synapses are mediated by multi-vesicular release and multiple docked vesicles

  1. Gábor Molnár
  2. Márton Rózsa
  3. Judith Baka
  4. Noémi Holderith
  5. Pál Barzó
  6. Zoltan Nusser
  7. Gábor Tamás  Is a corresponding author
  1. University of Szeged, Hungary
  2. Hungarian Academy of Sciences, Hungary
Short Report
Cite this article as: eLife 2016;5:e18167 doi: 10.7554/eLife.18167
3 figures


Figure 1 with 1 supplement
Monosynaptic excitatory connections from pyramidal cells to interneurons in the human and rat cerebral cortex.

(A) Firing pattern of a presynaptic pyramidal cell (black) and a postsynaptic basket cell (blue) simultaneously recorded in an acute slice of the human cerebral cortex. (B) LM reconstruction of the recorded pyramidal cell (left, soma and dendrites: black, axons: green) and basket cell (right, soma and dendrites: blue, axons: red). Scale bars apply to both cells. Reconstructions of the cells are separated for clarity with relative positions of the somata (pyramidal cell: black, basket cell: blue) indicated on both panels. Roman numbers represent cortical layers. (C) Presynaptic action potentials of the pyramidal cell (black) elicited unitary EPSCs in the basket cell held at −70 mV (individual traces: blue, average: top blue trace). (D) Route of the presynaptic axon (green) from the PC soma (black) to putative synaptic contacts (arrows) on the basket cell dendrites (blue). (E–H) A rat pyramidal cell to basket cell connection is presented similar to panels A–D. (I) Human and rat unitary EPSCs have similar latencies, rise and decay times, but the human EPSC charge is significantly larger. Data presented as mean ± SD.

Figure 1—figure supplement 1
Human EPSC parameters according to age and gender.

No correlation was found between the age of patients and the charge, rise time, latency and decay time of human EPSCs (Spearman correlation). Parameters of EPSCs measured in female and male patients were not significantly different.

Higher number of functional release sites in human excitatory synapses is revealed by multiple probability fluctuation analysis.

(A) Firing pattern of a human pyramidal cell (black) monosynaptically connected to a postsynaptic basket cell (blue). Bottom, Pyramidal cell action potential-evoked EPSCs in the basket cell recorded in 0.5, 1.5, 2 and 4 mM [Ca2+]o (grey: 10 consecutive traces; blue: averages). (B) Left, Charge transferred by the unitary EPSCs recorded in different [Ca2+]o. Inset, The shaded blue area illustrates the charge of an EPSC. Middle, mean ± SD of the EPSC charge (in the same experiment). Right, parabolic fit to the variance versus mean of EPSC charge recorded in different [Ca2+]o, revealing an estimate for the number of functional release sites (Nfrs = 11.3) and the quantal size (q = 45.9 fC). (C–D) A similar experiment is shown for a rat pyramidal cell (black) to basket cell (red) connection (Nfrs = 4.9; q = 25.0 fC). (E) Left, Dependence of the EPSC charge in human (blue) and rat (red) on the [Ca2+]o (1 mM: 87.3 ± 66.7 fC vs. 9.6 ± 12.7 fC; 1.5 mM: 218.7 ± 129.7 fC vs. 30.2 ± 37.6 fC; 2 mM: 349.7 ± 299.0 fC vs. 54.0 ± 66.1 fC; 4 mM: 563.2 ± 365.1 fC vs. 94.2 ± 113.8 fC). Right, Parabolic fits to the variance versus mean plots in human (blue) and rat (red). (F) Multiple probability fluctuation analysis of human and rat unitary EPSCs fails to show a significant difference in quantal size (q, human: 40.2 ± 14.9 fC, n = 10; rat: 29.6 ± 13.3 fC, n = 12; p = 0.11, MW U-test), but the Nfrs in human is substantially larger compared to rats (human: 20.5 ± 15.4, n = 10; rat: 4.7 ± 2.3, n = 12; p < 0.001, MW U-test). However, the number of LM detected contact sites (Nlm, right, human: 3.3 ± 1.5, n = 9; rat: 2.9 ± 1.5, n = 15) between presynaptic axons and postsynaptic dendrites is similar in human and rat (p = 0.36, MW U-test). Reconstructions with and without MPFA are shown by circles and diamonds, respectively. Data presented as mean ± SD.

Figure 3 with 1 supplement
Active zones (AZs) of excitatory synapses are twice as large and harbor 4 times more docked vesicles in human compared to rats.

(A, H) Electron microscopic images of 20 nm thick sections show axon terminals forming asymmetrical synapses on a human (A) and a rat (H) FS cell dendrite (intracellularly filled interneuron visualized with peroxidase: dark precipitate). (B, I) Higher magnification views of the boxed areas in A and H, showing docked vesicles (*) at the AZs. (C, J) 3D reconstructions of the terminals shown in B and I. The human terminal (c, grey semitransparent contour) contains an AZ (blue) of 0.09 μm2 with 4 docked vesicles (white spheres), whereas the rat AZ (red in J) is 0.04 μm2 and has 2 docked vesicles. (D, K–L) Electron tomographic subvolumes (0.6 nm thick) of human (D) and rat (KL) axon terminals (t) that establish asymmetrical synaptic contacts on FS cell dendrites (dark precipitate). (EG, M–O) Boxed areas from D and K-L show docked vesicles (*). Panels G and O show membrane proximal vesicles with distances smaller than 5 nm (distance between arrowheads). (P) The area of AZs, determined from 3D reconstructions from 20 nm serial sections, is twice as large in humans than in rats (0.077 ± 0.051 μm2, n = 22, from 3 separate human samples; rat: 0.041 ± 0.017 μm2, n = 19 from 3 animals; p = 0.002, MW U-test). (Q) The number of the docked vesicles, identified in fully reconstructed AZs from 20 nm serial sections, is 4-times larger in human compared to rats (human: 4.2 ± 2.2, n = 21; rat: 1.3 ± 0.8, n = 18, p<0.001, MW U-test). (R) The density of docked vesicles, measured either in 20 nm reconstructions (human: 58.5 ± 24.6 / μm2, n = 21, rat: 32.5 ± 19.9 / μm2, n = 18, p < 0.001, unpaired t-test) or in EM tomographic volumes (human: 49.5 ± 23.8 / μm2, n = 33, rat: 24.3 ± 16.0 / μm2, n = 31, P < 0.001, MW U-test), is significantly different between the two species. (S) The number of the docked vesicles shows a positive correlation with the AZ area (Spearman correlation). Scale bars: A, D, H, K, L: 200 nm, B, E-G, I, M-O: 100 nm, C, J: side of the cubes: 200 nm. Data presented as mean ± SD.

Figure 3—figure supplement 1
Ratio of docked vesicles in the presynaptic vesicle pool and bouton volumes in human and rat axon terminals.

(A) The density of docked vesicles and the size of the proximal vesicle pool normalized to the AZ area in axon terminals showed no significant correlation both in human (ρ = 0.15, p = 0.59, Spearman correlation), and rat samples (ρ = –0.09, p = 0.76). (B) Axonal bouton volumes were bigger in humans compared to rats (p<0.01, MW U-test).


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