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The Journal of Neuroscience, September 1, 2001, 21(17):6666-6672
Three-Dimensional Comparison of Ultrastructural Characteristics
at Depressing and Facilitating Synapses onto Cerebellar Purkinje
Cells
Matthew A.
Xu-Friedman1,
Kristen M.
Harris2, and
Wade G.
Regehr1
1 Department of Neurobiology, Harvard Medical School,
Boston, Massachusetts 02115, and 2 Department of Biology,
Boston University, Boston, Massachusetts 02215
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ABSTRACT |
Cerebellar Purkinje cells receive two distinctive types of
excitatory inputs. Climbing fiber (CF) synapses have a high probability of release and show paired-pulse depression (PPD), whereas parallel fiber (PF) synapses facilitate and have a low probability of release. We examined both types of synapses using serial electron microscopic reconstructions in 15-d-old rats to look for anatomical correlates of
these differences. PF and CF synapses were distinguishable by their
overall ultrastructural organization. There were differences between PF
and CF synapses in how many release sites were within 1 µm of a
mitochondrion (67 vs 84%) and in the degree of astrocytic ensheathment
(67 vs 94%). However, the postsynaptic density sizes for both types of
synapses were similar (0.13-0.14 µm2). For both
types of synapses, we counted the number of docked vesicles per release
site to test whether this number determines the probability of
release and synaptic plasticity. PF and CF synapses had the same number
of anatomically docked vesicles (7-8). The number of docked vesicles
at the CF does not support a simple model of PPD in which release of a
single vesicle during the first pulse depletes the anatomically docked
vesicle pool at a synapse. Alternatively, only a fraction of
anatomically docked vesicles may be release ready, or PPD could result
from multivesicular release at each site. Similarities in the number of
docked vesicles for PF and CF synapses indicate that differences in
probability of release are unrelated to the number of anatomically
docked vesicles at these synapses.
Key words:
climbing fiber; parallel fiber; probability of release; docked vesicles; electron microscope; serial reconstruction
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INTRODUCTION |
Purkinje cells receive two kinds of
excitatory synaptic inputs (Palay and Chan-Palay, 1974 ), each of which
exhibits different short-term plasticity and initial probability of
release. The first originates in the inferior olive and is known as the
climbing fiber (CF) (Ramón y Cajal, 1995). The CF shows prominent
paired-pulse depression (PPD) (Eccles et al., 1966; Hashimoto and Kano,
1998) and has a high probability of release, which has been estimated at >40% (Dittman and Regehr, 1998; Silver et al., 1998). The second input is from cerebellar granule cells, which send their axons into the
cerebellar molecular layer as parallel fibers (PFs). PFs show prominent
paired-pulse facilitation (Konnerth et al., 1990; Perkel et al., 1990)
and have a low probability of release, estimated to be <5% (Dittman
et al., 2000).
The differences in short-term plasticity and initial probability of
release between PFs and CFs are important because they are the only
excitatory inputs to Purkinje cells, which are themselves the only
outputs from the cerebellar cortex. However, the mechanisms underlying
short-term plasticity and the probability of release are not well
understood. CF and PF synapses present a useful system with which to
examine these issues because they have different properties while
synapsing onto the same target neuron.
One approach to understanding physiological differences between
synapses has been to compare their anatomical structures (Herrera et
al., 1985; Walrond et al., 1993; Msghina et al., 1998; Schikorski and
Stevens, 1999). One hypothesis for the basis of the probability of
release is that it is proportional to the number of docked vesicles.
This hypothesis predicts that the number of docked vesicles at CF
release sites would be markedly greater than at PF release sites,
because of the extreme differences between them in probability of
release. Studies at other synapses have found a correlation between the
probability of release and the size of the readily releasable pool
(RRP) of vesicles, as quantified using physiological stimulation
(Stevens and Tsujimoto, 1995; Dobrunz and Stevens, 1997),
FM1-43 staining (Murthy et al., 1997), and by counting docked
vesicles (Harris and Sultan, 1995; Schikorski and Stevens, 1997, 1999,
2001).
Another aspect of short-term plasticity to be explained is the
mechanism underlying PPD. PPD has been proposed to result from a
reduction in the number of release-ready synapses through depletion of
docked vesicles (Takeuchi, 1958; Elmqvist and Quastel, 1965; Betz,
1970; Dobrunz and Stevens, 1997). CFs show strong PPD, so measurements
of the number of docked vesicles will provide useful information for
depletion models.
We used serial electron microscopy (EM) to make detailed
reconstructions of both CF and PF synapses onto Purkinje cells. Our primary focus was on features that may affect short-term plasticity, but we have also documented characteristics relevant to other aspects
of synaptic physiology. Although we found some differences, PFs and CFs
had similar numbers of docked vesicles, sizes of postsynaptic density
(PSD), and postsynaptic spine volumes. Both fibers ended exclusively on
spines, and the release sites were predominantly ensheathed by
astrocytes. These data constrain models concerning the mechanisms
underlying PPD. In addition, they suggest that the difference in the
probability of release between PFs and CFs cannot be accounted for by
the number of docked vesicles.
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MATERIALS AND METHODS |
Two 15-d-old Long-Evans hooded rats (weights of 28 and 31 gm)
were used for the reconstructions. Each animal was heavily anesthetized using Nembutal (0.8 mg/10 gm) and was perfused transcardially for 30 min with a 2% paraformaldehyde-2.5% glutaraldehyde mix in 0.1 M cacodylate buffer (CB), pH 7.3, warmed to 34-37°C.
After 1 hr, the brain was removed and stored in the same fixative at room temperature overnight. Parasagittal thick sections (400 µm) were
prepared on a tissue chopper, fixed with 1%
OsO4-1.5% KFeCN in CB, subsequently fixed with
1% OsO4 in CB, and stained with 1% uranyl
acetate before dehydrating through acetone and embedding in 1:1
Epon/Spurr's resin. Serial thin sections were cut on a Reichert
Ultracut S (Leica, Nussloch, Germany), collected on
Formvar-coated slot grids, dipped in saturated uranyl acetate in
acetone, rinsed in water, dipped in 0.2% lead citrate, rinsed, and dried.
The effects of tissue preparation on synaptic ultrastructure are not
completely known. Studies comparing aldehyde fixation and rapid
freezing do report differences in vesicle and mitochondrion shape
(Nakajima and Reese, 1983; Brewer and Lynch, 1986), but importantly,
differences were not reported for the synaptic characteristics we
measured here. There are conflicting reports of the effects of
aldehyde fixation on synaptic release (Smith and Reese, 1980; Rosenmund
and Stevens, 1997), but its easier applicability in vivo
made it the preferable method. We subsequently did not observe evidence
of significant vesicular release, such as large numbers of  profiles at the synapses of interest.
Three series of sections were prepared and photographed on a Jeol
(Peabody, MA) 1200EX electron microscope with the grids mounted in a
rotating holder (SRH 10mod). The accelerating voltage was 80-120 kV at
a magnification of 5000-10,000×. Magnification was calibrated using
diffraction grating replica number 60021 (Ernest F. Fullam Inc.,
Latham, NY). Section thickness was calibrated using cylindrical objects
such as dendrites and mitochondria (Fiala and Harris, 2001b).
Photographic negatives were scanned at 1000 dots per inch on an
AGFA (Ridgefield Park, NJ) Duoscan 2500, and the resulting images were
aligned using SEM Align (available from the "Tools" section
of website www.synapses.bu.edu), by identifying several pairs
of matching points in adjacent sections and using a least-squares
linear fit (Fiala and Harris, 2001a).
Synapses were selected for reconstruction on the basis of their
completeness and optimal angle of sectioning (i.e., perpendicular to
the synaptic cleft). Relevant structures were traced and measured with
IGL Trace (www.synapses.bu.edu). When identifying docked vesicles, image contrast and brightness were optimized to detect cytoplasmic space between the vesicle and presynaptic membranes. For
display purposes, object surfaces were exported from IGL Trace to
trueSpace 4.3 (Caligari Corp., Mountain View, CA) for three-dimensional (3-D) rendering. To examine an extensive segment of a CF, up to four
photographs per section were arranged into a composite using Adobe
Photoshop 5.5 (Adobe Systems, San Jose, CA) before aligning. Figures 2
and 5 were colorized using Igor 4 (WaveMetrics Inc., Lake Oswego, OR).
We also labeled PFs and CFs with fluorescent dyes. To label PFs, we
prepared transverse 300 µm slices from rat cerebellar vermis and
directed a jet of dye (0.06% Fast Green, 0.06% Triton X-100, and
0.09% 10,000 molecular weight Texas Red dextran in H2O) at the molecular layer, with a large suction
pipette placed nearby to restrict the area labeled. We waited at least
1 hr for the dye to diffuse before imaging fibers >100 µm from the
fill site. To label CFs, we injected 0.5 µl of dye (saturated DiI in dimethylformamide-DMSO) into the inferior olive of postnatal
day 11 rats, waited 4 d for the dye to diffuse, and cut
sagittal sections. We imaged slices on an Olympus Optical (Tokyo,
Japan) Fluoview confocal microscope.
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RESULTS |
Figure 1 illustrates anatomical
differences between CFs and PFs at the light microscope level.
By 15 d of age, each Purkinje cell receives a single
climbing fiber input (Crepel et al., 1976; Ito, 1984), which wraps
around the major dendritic branches of the Purkinje cell, making
hundreds of contacts on somatic and dendritic spines (Fig.
1A). In contrast, PFs run perpendicular to the
Purkinje cell dendritic arbor, with synaptic varicosities at ~5 µm
intervals along their length, and they make few contacts with each
Purkinje cell (Fig. 1B) (Palay and Chan-Palay,
1974).
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Figure 1.
Overview of anatomical and physiological
differences between cerebellar CFs and PFs. A,
Top, Image of an individual CF labeled by injections of
DiI in the inferior olive. The fiber is shown in a sagittal slice taken
from cerebellar vermis, and a single optical section was imaged using a
confocal microscope. The CF enters from the bottom and makes a complex
arborization, with multiple processes running parallel to each other
along the Purkinje cell dendrite, but the synapses themselves are not
discernable. A, Bottom, Paired-pulse
depression at the CF. A CF was stimulated twice at an interval of 20 msec, and the EPSCs were recorded in the Purkinje cell. The
second EPSC is 60% of the size of the first. B,
Top, Image of a band of PFs labeled with Texas Red
dextran in a transverse slice (imaged using a confocal microscope).
Each PF runs perpendicular to the Purkinje cell dendritic arbor.
Varicosities, which correspond to synaptic contacts, are scattered
along the length of the PF. B, Bottom,
Paired-pulse facilitation at the PF. A band of PFs was stimulated twice
at an interval of 20 msec, and the EPSCs were recorded in the Purkinje
cell. The second EPSC is 2.5 times larger than the first. Physiology
traces are adapted from Dittman et al. (2000).
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Short-term plasticity in CFs and PFs are compared in Figure 1
(bottom). The CF shows paired-pulse depression, indicating a high probability of release, whereas the PF shows paired-pulse facilitation, indicating a low probability of release. To look for an
anatomical basis for the difference in short-term
plasticity and probability of release between CF and PF synapses, we
examined CF and PF release sites using serial electron microscopy and
quantified a number of synaptic characteristics that could play a role.
Climbing fibers
We prepared two series (of 108 and 143 sections) for examining
release sites made by two CFs. We followed one CF in each series and
identified the synaptic contacts made by each axon using the presence of presynaptic active zones and PSDs. Figure
2 shows four serial sections taken from
two example CF synapses, in which we have identified and traced the CF
(blue), the postsynaptic spines (pink),
and ensheathing astrocytes (yellow). Both segments of
axon have mitochondria present and a high density of vesicles. The
release site in Figure 2A is larger than average,
whereas the release site in Figure 2B is smaller than
average. The spine in Figure 2B
(pink) connects through a very narrow neck to the parent dendrite (top). These two synapses were ensheathed by
astrocytic processes (yellow) at both ends of the
synaptic cleft. We classified vesicles as docked if they were located
opposite PSDs in a spine and if the vesicle membrane directly touched
the presynaptic membrane (Fig. 2C, green); if
there was any cytoplasmic space between the two membranes, they were
classified as nondocked (Fig. 2C, arrows). This
strict criterion was used because only these docked vesicles are in a
position to be readily released.
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Figure 2.
Serial EM sections of CFs. A,
B, Two sample release sites from the same CF. CF axons
are shaded blue, Purkinje cell dendrites and spines are
shaded pink, and astrocytes are shaded
yellow. The release site in A is larger
than average, whereas the release site in B is smaller
than average. In A, the synaptic cleft and PSD are
clearly identifiable in the top three panels. In
B, two spines are visible emerging from the parent
dendrite at the top of each panel. The
PSD for one spine is visible in the bottom three panels,
and the second makes a PSD in later sections. C,
Close-ups of the active zone in A. The presynaptic
terminal is on the left, and the postsynaptic spine,
with PSD clearly visible, is on the right. A vesicle was
classified as docked (green) if it was located
opposite a PSD and directly touched the presynaptic membrane. Nondocked
vesicles close to the membrane are indicated by
arrows.
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We made a partial reconstruction of one CF (Fig.
3B,C)
and its postsynaptic Purkinje cell (Fig.
3A,C). Over this ~60 µm stretch of dendrite, we identified 67 release sites (Fig. 3, red),
all of which were onto spines, and each spine received only
one synapse. The CF wraps around the Purkinje cell dendrite, branching
frequently and producing processes running back and forth over its
surface to make clusters of contacts (Fig. 3C), similar to
the light microscope view in Figure 1A.
The location of synapses is not uniform over the Purkinje
cell dendrite, because spines appear to form in clusters, and synapses
do not occur at regular intervals along the CF.
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Figure 3.
3-D reconstruction of a CF terminating on a
Purkinje cell from serial EM. A, Purkinje cell proximal
dendrite. The soma is off the bottom, and distal into
the molecular layer is toward the top. Spines that make
contact with a CF are included, and their PSDs are labeled in
red. The dendrite is viewed at an angle, so as an indication of
scale, the dendrite is 3.8 µm in diameter at the position
indicated by the arrow. B, CF
axon, colored in blue. PSDs on the Purkinje cell
are labeled in red. Labels A-D in
B correspond to the segments of CF reconstructed
in more detail in Figure 4. C, CF and Purkinje cell
superimposed.
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We identified CF release sites using the presence of an active zone
across from a PSD on a Purkinje cell spine. We made detailed reconstructions of 13 release sites in the first CF and 18 in the
second. The reconstructed axon segments in Figure
4 illustrate various characteristics of
these release sites. Some release sites were made en passant (Fig.
4A-C), and others were in distinct boutons (Fig.
4A,D). Multiple release sites
frequently were found very near each other (Fig. 4C).
Vesicle density near release sites was high, and often the density was
lower in parts of the axon with no release sites (Fig.
A,D). Most release sites had at
least one mitochondrion nearby, and some had several (Fig.
4A-D). Docked vesicles were distributed across the
whole active zone (Fig. 4).
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Figure 4.
3-D reconstructions of sample CF release sites.
Reconstructions of each PSD with associated docked vesicles are shown
unsmoothed next to the axon. Axons are light blue,
mitochondria are tan, vesicles are dark
blue (uniformly represented as 40 nm spheres), and PSDs on the
opposing spine are transparent red. A,
Two release sites. The first is located on the main branch of the CF
axon, with a locally high density of vesicles and many mitochondria.
The second is located in a bouton with no mitochondria.
B, Two release sites, made en passant, with a high
density of vesicles and three mitochondria nearby. The rightmost part
(with a large density of vesicles and mitochondria) branches toward
additional release sites (data not shown). C, Three
release sites made en passant, with a high density of vesicles and two
mitochondria (partially cut off). D, Two release sites,
both in boutons, only one of which has a mitochondrion.
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At these reconstructed release sites, we quantified the number of
docked vesicles, PSD surface area, and spine volume. Because the
averages from these series were nearly the same, we combined them
(Table 1; see Fig. 7). We found that the
number of docked vesicles ranged from 1 to 14 with an average of
7.3 ± 3.0 (mean ± SD) and that this number of docked
vesicles at each site correlated with the PSD area (r = 0.80) and spine volume (r = 0.73).
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Table 1.
Quantitative comparison of ultrastructural characteristics
of climbing fiber and parallel fiber synapses
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To quantify the presence of mitochondria on the presynaptic
side, we examined the 67 release sites in the reconstructed CF of
Figure 3. When we measured the distance between the active zone and the
nearest mitochondrion, we found that 84% of the release sites had a
mitochondrion within 1 µm (median, 0.32 µm), and some release sites
had as many as six mitochondria within 1 µm. Astrocytes are prominent
over the proximal segment of the Purkinje cell dendrite (Fig. 2). We
restricted our attention to the presence of astrocytic ensheathment
around the perimeter of the synaptic cleft, quantifying the percentage
of the perimeter that was bounded by astrocytic processes (same method
as Ventura and Harris, 1999). A total of 58% of synapses were at least
90% ensheathed, and the median degree of ensheathment was 94%
(average 87 ± 18%).
Parallel fibers
We prepared two series (of 89 and 82 sections) for examining the
synapses made by PFs. PF synapses were easily identified as
varicosities (Fig.
5A,B,
blue), with a large increase in vesicle density around an
active zone that contacted a spine with a postsynaptic density
(pink). Astrocytic processes
(yellow) appear to occupy less of the neuropil near
PF synapses than near CF synapses (compare with Fig. 2), although as
quantified below, most PF synapses have at least some astrocytic
ensheathment. The synapses in Figure 5, A and B,
had mitochondria in other sections that are not pictured. We classified
vesicles as docked using the same strict criteria as for CFs (Fig.
5C, green).
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Figure 5.
Serial EM sections of PFs. A,
B, Two example series. PFs are shaded
blue, Purkinje cell spines are shaded
pink, and astrocytes are shaded yellow.
Every panel contains a clearly defined synaptic cleft
and PSD. The release site in A is larger than average,
whereas the release site in B is smaller than average.
Although neither PF has a mitochondrion in the sections shown here, the
release site in A has a mitochondrion in a section that
is 1.5 µm away, and the site in B has one in a section
that is 0.2 µm away. C, Close-up of the active zone in
A. The presynaptic terminal is on the
left, and the postsynaptic spine, with PSD clearly
visible, is on the right. Docked vesicles are labeled in
green, and nondocked vesicles close to the membrane are
indicated by arrows.
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We chose a set of 10 PFs in each series to reconstruct in detail. We
identified release sites using the presence of an active zone across
from a PSD on a Purkinje cell spine. Each spine received a single
synaptic contact. All release sites were located in axonal varicosities, with high local vesicle densities (Fig.
6). Some release sites had nearby
mitochondria (Fig. 6A-C), but others did not (Fig.
6D,E). Occasionally, we saw more
than one release site per varicosity (Fig. 6F).
Docked vesicles were distributed across the entire active zone (Fig.
6).
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Figure 6.
3-D reconstructions of sample PF release sites.
Reconstructions of each PSD with associated docked vesicles are shown
unsmoothed under the axon. The color scheme is as described in the
legend to Figure 4. A-C, PFs with one to two
mitochondria near the release site. D, E,
PFs without mitochondria. F, PF with two release sites
onto different spines in the same varicosity.
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For these reconstructed synapses, we quantified the total number of
vesicles in a varicosity, the number of docked vesicles, the PSD
surface area, and the spine volume. The two series were not
significantly different, so their results were combined (Table 1). The
total number of vesicles in each varicosity was large (480 ± 160, range of 237-770), similar to a previous study (Harris and Stevens,
1988). We did not perform a count of total vesicles at CF release
sites, because their geometry is much more complex and they are not as
readily isolated from each other. The PSD surface area was on average
0.13 µm2 at the PF, which was similar to
the average of 0.14 µm2 at the
CF, but the variability was lower than at the CF (SD, 0.04 vs 0.08 µm2) (Fig.
7A). In addition, the average
number of docked vesicles was 8.1 ± 2.9 at PFs, which was close
to the number at CF release sites (7.3 ± 3.0), and the
distribution of the number of docked vesicles was also very
similar for these two synapse types (Fig. 7B). The number of
docked vesicles correlated with PSD area (r = 0.75).
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Figure 7.
Comparison of ultrastructural characteristics at
PF and CF release sites. Data are presented as cumulative histograms.
PF distributions are shown as thin lines, and CF
distributions are shown as thick lines.
A, PSD area. B, Docked vesicle number.
C, Distance from active zone to nearest mitochondrion.
D, Degree of ensheathment.
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All PFs that had release sites within the middle 20 sections (1.2 µm)
of the second series were reconstructed across the entire series to
quantify the presence of presynaptic mitochondria and the degree of
astrocytic ensheathment. Of the 39 parallel fibers in this population,
four (10%) had two release sites per varicosity, which were made onto
different spines. This is considerably less than the 27% of
varicosities with more than one release site reported in older rats
(Harris and Stevens, 1988). Only one PF showed a second varicosity with
a release site (2.2 µm away), consistent with the observation from
fluorescent and Golgi staining that varicosities are spaced 5 µm
apart on average. Of the 39 PFs, 25 (64%) had a mitochondrion within 1 µm of the release site (Fig. 7C) and 11 (28%) had no
mitochondrion within the series. The median distance to the nearest
mitochondrion was 0.37 µm, which was not greatly different from the
CF (0.32 µm). For 12 of 42 synapses (29%), 90% of the perimeter of
the synaptic cleft was ensheathed by astrocytes (median, 67%
ensheathed; average, 65 ± 29%) (Fig. 7D), which was
less than at the CF.
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DISCUSSION |
These serial reconstructions of CF and PF synapses onto Purkinje
cells revealed that, despite their distinctive overall anatomical structures, they were primarily similar in a number of ultrastructural characteristics related to synaptic physiology. Foremost among these
similarities, these synapses have the same number of morphologically docked vesicles. Thus, the number of docked vesicles cannot account for
the differences in the initial probability of release and short-term
plasticity exhibited by CF and PF synapses.
Probability of release
We compared the numbers of docked vesicles and the
electrophysiological properties of different synapses. Our measurements of seven to eight docked vesicles per active zone at the PF and CF are
less than those reported for a number of cells in sensory pathways: 22 in goldfish bipolar neurons (von Gersdorff et al., 1996), 32 in frog
saccular hair cells (Lenzi et al., 1999), and 130 in cat rod
photoreceptors (Rao-Mirotznik et al., 1995). The large number of docked
vesicles at these active zones may help these cells to respond rapidly
and reliably to the sensory signal driving them. Each of these cells
releases transmitter in response to graded or receptor potentials, so
their probability of release cannot be compared.
A more direct comparison can be made with CNS neurons that fire action
potentials and have a well defined probability of release. At
hippocampal CA1 neurons, the number of docked vesicles per active zone
can range from 2 to 36 (Harris and Sultan, 1995), with an average of 10 (Schikorski and Stevens, 1997). In layers 1a and 1b of pyriform cortex,
the average number of docked vesicles is 16 and 27, respectively
(Schikorski and Stevens, 1999). These measurements, together with
physiological studies of hippocampal and pyriform cortical cells, have
suggested that the number of docked vesicles correlates with the
probability of release (Bower and Haberly, 1986; Harris and Sultan,
1995; Dobrunz and Stevens, 1997; Murthy et al., 1997; Schikorski and
Stevens, 1997, 2001). However, although there is a large difference in
the probability of release between CF and PF synapses, they both have
approximately the same number of docked vesicles, and this number is
somewhat less that what has been observed at these other CNS synapses.
There are several possible explanations for why the number of docked
vesicles at PF and CF synapses does not correlate with the probability
of release. One possibility is that not all of the anatomically docked
vesicles are release ready, which could arise from a priming step after
docking that differs between PFs and CFs. Alternatively, there are many
proteins involved in synaptic release, many of which have multiple
isoforms or phosphorylation sites, which could affect probability of
release (Sudhof, 1995). In addition, the calcium signal that drives
release could differ between the two synapses as a result of
differences in calcium influx, buffering, or diffusion distance to the
release-ready vesicle. Such differences in release machinery or calcium
signal would not be apparent ultrastructurally.
Depletion model of depression
Our finding of multiple docked vesicles at CF release sites
constrains models of the mechanisms underlying paired-pulse depression. One prominent model of PPD proposes that it is attributable to depletion of release-ready sites (Takeuchi, 1958; Elmqvist and Quastel,
1965; Betz, 1970). In the simplest case, a single vesicle is released
from the RRP per release site, and the RRP corresponds to anatomically
docked vesicles (Stevens and Wang, 1995). Our measurements of docked
vesicles at the CF are inconsistent with this simple model, because the
loss of a single vesicle from a pool of approximately eight vesicles
cannot account for the observed PPD of 40% at the CF (Fig.
1A). One possibility is that not all anatomically
docked vesicles are release ready, and our measurements of anatomically
docked vesicles overestimate the RRP. Another possibility is that the
RRP could be depleted more rapidly after the first pulse because of
multivesicular release, as recent experiments at the CF suggest
(Wadiche and Jahr, 2000).
Other ultrastructural characteristics
Although the primary focus of these studies was on docked
vesicles, short-term plasticity, and probability of release, we also
documented characteristics of these synapses that could affect other
aspects of synaptic function: PSD area, mitochondrial distribution, and
astrocytic ensheathment. PSD area was similar on average for both CF
and PF synapses (0.14 vs 0.13), but variability was larger at CF than
PF synapses. This variability likely contributes to the distribution of
miniature EPSC amplitudes at these synapses. For both types of
synapses, a minority of active zones were >1 µm from the nearest
mitochondrion (33% of PFs, 16% of CFs). This may indicate
heterogeneity in calcium handling and energy production, as has been
suggested for CA3 synapses onto CA1 pyramidal cells, for which half of
the varicosities lack mitochondria (Shepherd and Harris, 1998). We also
found that both PF and CF synapses were ensheathed by astrocytic
processes (average degree of ensheathment of 87% for CFs and 65% for
PFs) to a greater extent than synapses in the stratum radiatum in area
CA1 of the hippocampus (average of 43%) (Ventura and Harris, 1999).
Variability in cerebellar ensheathment has also been reported by Nusser
et al. (2000). Because astrocytes play a large role in neurotransmitter
uptake, these findings suggest that CF synapses are likely to be
particularly insensitive to cross talk and desensitization because of
spillover of glutamate.
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FOOTNOTES |
Received May 7, 2001; revised June 15, 2001; accepted June 19, 2001.
This work was supported by National Institutes of Health Grants NS07112
(W.G.R.) and NS21184 and MH57351 (K.M.H.). We thank M. Feinberg,
M. Ericsson, and E. Benecchi for invaluable help with the electron
microscopy and J. Fiala for help using the SEM Align and IGL Trace
programs. K. Foster and A. Kreitzer provided help with the fluorescent
labeling. We thank D. Blitz, A. Carter, K. Foster, A. Kreitzer, and K. Vogt for comments on this manuscript.
Correspondence should be addressed to Wade G. Regehr, Department of
Neurobiology, Harvard Medical School, 220 Longwood Avenue, Boston, MA
02115. E-mail: wade_regehr{at}hms.harvard.edu.
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REFERENCES |
-
Betz WJ
(1970)
Depression of transmitter release at the neuromuscular junction of the frog.
J Physiol (Lond)
206:629-644[Abstract/Free Full Text].
-
Bower JM,
Haberly LB
(1986)
Facilitating and nonfacilitating synapses on pyramidal cells: a correlation between physiology and morphology.
Proc Natl Acad Sci USA
83:1115-1119[Abstract/Free Full Text].
-
Brewer PA,
Lynch K
(1986)
Stimulation-associated changes in frog neuromuscular junctions. A quantitative ultrastructural comparison of rapid-frozen and chemically fixed nerve terminals.
Neuroscience
17:881-895[Medline].
-
Crepel F,
Mariani J,
Delhaye-Bouchaud N
(1976)
Evidence for a multiple innervation of Purkinje cells by climbing fibers in the immature rat cerebellum.
J Neurobiol
7:567-578[Web of Science][Medline].
-
Dittman JS,
Regehr WG
(1998)
Calcium dependence and recovery kinetics of presynaptic depression at the climbing fiber to Purkinje cell synapse.
J Neurosci
18:6147-6162[Abstract/Free Full Text].
-
Dittman JS,
Kreitzer AC,
Regehr WG
(2000)
Interplay between facilitation, depression, and residual calcium at three presynaptic terminals.
J Neurosci
20:1374-1385[Abstract/Free Full Text].
-
Dobrunz LE,
Stevens CF
(1997)
Heterogeneity of release probability, facilitation, and depletion at central synapses.
Neuron
18:995-1008[Web of Science][Medline].
-
Eccles JC,
Llinas R,
Sasaki K,
Voorhoeve PE
(1966)
Interaction experiments on the responses evoked in Purkinje cells by climbing fibres.
J Physiol (Lond)
182:297-315[Abstract/Free Full Text].
-
Elmqvist D,
Quastel DMJ
(1965)
A quantitative study of end-plate potentials in isolated human muscle.
J Physiol (Lond)
178:505-529[Free Full Text].
-
Fiala JC,
Harris KM
(2001a)
Extending unbiased stereology of brain ultrastructure to three-dimensional volumes.
J Am Med Inform Assoc
8:1-16[Abstract/Free Full Text].
-
Fiala JC,
Harris KM
(2001b)
Cylindrical diameters method for calibrating section thickness in serial electron microscopy.
J Microsc
202:468-472[Web of Science][Medline].
-
Harris KM,
Stevens JK
(1988)
Dendritic spines of rat cerebellar Purkinje cells: serial electron microscopy with reference to their biophysical characteristics.
J Neurosci
8:4455-4469[Abstract].
-
Harris KM,
Sultan P
(1995)
Variation in the number, location, and size of synaptic vesicles provides an anatomical basis for the nonuniform probability of release at hippocampal CA1 synapses.
Neuropharmacology
34:1387-1395[Web of Science][Medline].
-
Hashimoto K,
Kano M
(1998)
Presynaptic origin of paired-pulse depression at climbing fibre-Purkinje cell synapses in the rat cerebellum.
J Physiol (Lond)
506:391-405[Abstract/Free Full Text].
-
Herrera AA,
Grinnell AD,
Wolowske B
(1985)
Ultrastructural correlates of naturally occurring differences in transmitter release efficacy in frog motor nerve terminals.
J Neurocytol
14:193-202[Web of Science][Medline].
-
Ito M
(1984)
In: The cerebellum and neural control. New York: Raven.
-
Konnerth A,
Llano I,
Armstrong CM
(1990)
Synaptic currents in cerebellar Purkinje cells.
Proc Natl Acad Sci USA
87:2662-2665[Abstract/Free Full Text].
-
Lenzi D,
Runyeon JW,
Crum J,
Ellisman MH,
Roberts WM
(1999)
Synaptic vesicle populations in saccular hair cells reconstructed by electron tomography.
J Neurosci
19:119-132[Abstract/Free Full Text].
-
Msghina M,
Govind CK,
Atwood HL
(1998)
Synaptic structure and transmitter release in crustacean phasic and tonic motor neurons.
J Neurosci
18:1374-1382[Abstract/Free Full Text].
-
Murthy VN,
Sejnowski TJ,
Stevens CF
(1997)
Heterogeneous release properties of visualized individual hippocampal synapses.
Neuron
18:599-612[Web of Science][Medline].
-
Nakajima Y,
Reese TS
(1983)
Inhibitory and excitatory synapses in crayfish stretch receptor organs studied with direct rapid-freezing and freeze-substitution.
J Comp Neurol
213:66-73[Web of Science][Medline].
-
Nusser Z,
Roth A,
Schorge S,
Hausser M
(2000)
Simulations of synaptic transmission in 3-D reconstructions of cerebellar neuropil.
Soc Neurosci Abstr
26:1122.
-
Palay SL,
Chan-Palay V
(1974)
In: Cerebellar cortex: cytology and organization. New York: Springer.
-
Perkel DJ,
Hestrin S,
Sah P,
Nicoll RA
(1990)
Excitatory synaptic currents in Purkinje cells.
Proc R Soc Lond B Biol Sci
241:116-121[Medline].
-
Ramón y Cajal S
(1995)
In: Histology of the nervous system. New York: Oxford UP.
-
Rao-Mirotznik R,
Harkins AB,
Buchsbaum G,
Sterling P
(1995)
Mammalian rod terminal: architecture of a binary synapse.
Neuron
14:561-569[Web of Science][Medline].
-
Rosenmund C,
Stevens CF
(1997)
The rate of aldehyde fixation of the exocytotic machinery in cultured hippocampal synapses.
J Neurosci Methods
76:1-5[Web of Science][Medline].
-
Schikorski T,
Stevens CF
(1997)
Quantitative ultrastructural analysis of hippocampal excitatory synapses.
J Neurosci
17:5858-5867[Abstract/Free Full Text].
-
Schikorski T,
Stevens CF
(1999)
Quantitative fine-structural analysis of olfactory cortical synapses.
Proc Natl Acad Sci USA
96:4107-4112[Abstract/Free Full Text].
-
Schikorski T,
Stevens CF
(2001)
Morphological correlates of functionally defined synaptic vesicle populations.
Nat Neurosci
4:391-395[Web of Science][Medline].
-
Shepherd GM,
Harris KM
(1998)
Three-dimensional structure and composition of CA3
 CA1 axons in rat hippocampal slices: implications for presynaptic connectivity and compartmentalization.
J Neurosci
18:8300-8310[Abstract/Free Full Text]. -
Silver RA,
Momiyama A,
Cull-Candy SG
(1998)
Locus of frequency-dependent depression identified with multiple-probability fluctuation analysis at rat climbing fibre-Purkinje cell synapses.
J Physiol (Lond)
510:881-902[Abstract/Free Full Text].
-
Smith JE,
Reese TS
(1980)
Use of aldehyde fixatives to determine the rate of synaptic transmitter release.
J Exp Biol
89:19-29[Abstract/Free Full Text].
-
Stevens CF,
Tsujimoto T
(1995)
Estimates for the pool size of releasable quanta at a single central synapse and for the time required to refill the pool.
Proc Natl Acad Sci USA
92:846-849[Abstract/Free Full Text].
-
Stevens CF,
Wang Y
(1995)
Facilitation and depression at single central synapses.
Neuron
14:795-802[Web of Science][Medline].
-
Sudhof TC
(1995)
The synaptic vesicle cycle: a cascade of protein-protein interactions.
Nature
375:645-653[Medline].
-
Takeuchi A
(1958)
The long-lasting depression in neuromuscular transmission of frog.
Jpn J Physiol
12:102-113.
-
Ventura R,
Harris KM
(1999)
Three-dimensional relationships between hippocampal synapses and astrocytes.
J Neurosci
19:6897-6906[Abstract/Free Full Text].
-
von Gersdorff H,
Vardi E,
Matthews G,
Sterling P
(1996)
Evidence that vesicles on the synaptic ribbon of retinal bipolar neurons can be rapidly released.
Neuron
16:1221-1227[Web of Science][Medline].
-
Wadiche JJ,
Jahr CE
(2000)
Elevated levels of glutamate at cerebellar synapses.
Soc Neurosci Abstr
26:1385.
-
Walrond JP,
Govind CK,
Huestis SE
(1993)
Two structural adaptations for regulating transmitter release at lobster neuromuscular synapses.
J Neurosci
13:4831-4845[Abstract].
Copyright © 2001 Society for Neuroscience 0270-6474/01/21176666-07$05.00/0
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[Full Text]
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[Full Text]
[PDF]
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|
|
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|
|
|
|
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The density of AMPA receptors activated by a transmitter quantum at the climbing fibre-Purkinje cell synapse in immature rats
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549(1):
75 - 92.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
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23(8):
3154 - 3163.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
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J. Neurosci.,
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23(6):
2182 - 2192.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
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J Neurophysiol,
February 1, 2003;
89(2):
979 - 988.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
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SNAREs and control of synaptic release probabilities
FASEB J,
February 1, 2003;
17(2):
130 - 135.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
K. Satzler, L. F. Sohl, J. H. Bollmann, J. G. G. Borst, M. Frotscher, B. Sakmann, and J. H. R. Lubke
Three-Dimensional Reconstruction of a Calyx of Held and Its Postsynaptic Principal Neuron in the Medial Nucleus of the Trapezoid Body
J. Neurosci.,
December 15, 2002;
22(24):
10567 - 10579.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
A. G. Millar, H. Bradacs, M. P. Charlton, and H. L. Atwood
Inverse Relationship between Release Probability and Readily Releasable Vesicles in Depressing and Facilitating Synapses
J. Neurosci.,
November 15, 2002;
22(22):
9661 - 9667.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
P. Isope and B. Barbour
Properties of Unitary Granule Cellright-arrowPurkinje Cell Synapses in Adult Rat Cerebellar Slices
J. Neurosci.,
November 15, 2002;
22(22):
9668 - 9678.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
C. Saviane, L. P Savtchenko, G. Raffaelli, L. L Voronin, and E. Cherubini
Frequency-dependent shift from paired-pulse facilitation to paired-pulse depression at unitary CA3-CA3 synapses in the rat hippocampus
J. Physiol.,
October 15, 2002;
544(2):
469 - 476.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
R. Fernandez-Chacon, O.-H. Shin, A. Konigstorfer, M. F. Matos, A. C. Meyer, J. Garcia, S. H. Gerber, J. Rizo, T. C. Sudhof, and C. Rosenmund
Structure/Function Analysis of Ca2+ Binding to the C2A Domain of Synaptotagmin 1
J. Neurosci.,
October 1, 2002;
22(19):
8438 - 8446.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
R. Ichikawa, T. Miyazaki, M. Kano, T. Hashikawa, H. Tatsumi, K. Sakimura, M. Mishina, Y. Inoue, and M. Watanabe
Distal Extension of Climbing Fiber Territory and Multiple Innervation Caused by Aberrant Wiring to Adjacent Spiny Branchlets in Cerebellar Purkinje Cells Lacking Glutamate Receptor delta 2
J. Neurosci.,
October 1, 2002;
22(19):
8487 - 8503.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
J. Waters and S. J Smith
Vesicle pool partitioning influences presynaptic diversity and weighting in rat hippocampal synapses
J. Physiol.,
June 15, 2002;
541(3):
811 - 823.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
V. Scheuss, R. Schneggenburger, and E. Neher
Separation of Presynaptic and Postsynaptic Contributions to Depression by Covariance Analysis of Successive EPSCs at the Calyx of Held Synapse
J. Neurosci.,
February 1, 2002;
22(3):
728 - 739.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
H. Varoqui, M. K.-H. Schafer, H. Zhu, E. Weihe, and J. D. Erickson
Identification of the Differentiation-Associated Na+/PI Transporter as a Novel Vesicular Glutamate Transporter Expressed in a Distinct Set of Glutamatergic Synapses
J. Neurosci.,
January 1, 2002;
22(1):
142 - 155.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
TOP
ABSTRACT
INTRODUCTION
