J. Cell Sci. 18, 257-269 (1975)
Printed in Great Britain
257
CILIARY GRANULE PLAQUES: MEMBRANEINTERCALATED PARTICLE AGGREGATES
ASSOCIATED WITH Ca2+-BINDING SITES
IN PARAMECIUM
H. PLATTNER
InstitutfUr Zellbiologie, Universitat Miinchen,
Goethestrafle 33, D-8 Miinchen 2, W-Germany
SUMMARY
In Paramecium nine rectangular aggregates of membrane-intercalated particles surround
the freeze-cleaved membrane of the ciliary base. These 'ciliary granule plaques' occur
independently of the 'ciliary necklace' which is observed in a more basal position on some
cilia. In each individual plaque the ~ 10 nm large granules are arranged in a square grid
pattern: the granules of one plaque invariably form 3 vertical rows (number of horizontally
arranged particles per row: JV\ = 3) and 3-6 horizontal rows (mean value for N, = 48); the
centre-to-centre spacing of granules is dh = 22 nm in the horizontal and d, = 24 in the vertical
direction. The ciliary granule plaques coincide with electron-dense deposits which can be
produced in a position adjacent to the inner side of the basal ciliary shaft membrane by
3 procedures: (a) when cells are fixed in glutaraldehyde after incubation with Ca1+ concentrations (up to 25 mM) which are higher than normally present in the medium (about 0-5 mM)
but which do not immobilize the animals prior to fixation in glutaraldehyde; (6) when
Ca1+ (up to 75 mM) was added to glutaraldehyde; (c) when cells were incubated with Ca11"
as in (a) and then fixed in OsO4 solutions to which oxalate ions were added in concentrations
equivalent to Ca1+. Comparable deposits were observed also with Ca1+ concentrations
normally present in the medium, but they occurred much more rarely. The occurrence
of electron-dense deposits was not enhanced by addition of Na+, Mg s+ or La 3+ instead of
CaI+. In cross-sectioned ciliary bases the electron-dense Cas+-dependent deposits face the 9
doublet microtubules. Occasionally the substructure of Ca'+-dependent deposits is resolved in
median sections; their periodicity in the vertical direction (d, = z$ran;Nc = 5-2) corresponds
to the periodicity of membrane-intercalated particles of freeze-cleaved plaques. Other parameters determined for Ca1+-dependent electron-dense deposits coincide also with those determined for freeze-cleaved ciliary granule plaques (values in parentheses are for cells with Ca2+
added before fixation with glutaraldehyde and ultrathin sectioning): The height of plaques
(or deposits) is 84(ii5)nm; the distance from their lower end to the ciliary base is 138
(121) nm. Plaques (or deposits) are 45 (44) nm broad and the distance separating adjacent
plaques (deposits) is 48 (50) nm. Plaques (deposits) are arranged with a horizontal periodicity
of mean 93-102 (92) nm. It is concluded that the ciliary granule plaques are identical to
or associated with Ca*+-binding sites involved in ciliary activity or other functions.
INTRODUCTION
In previous work the occurrence of regular arrangements of membrane-intercalated
particles in the freeze-cleaved cortical membrane complex of Paramecium was
described (Plattner, Miller & Bachmann, 1973). Most of these structures were
258
H. Plattner
shown to represent membrane-junctions. In contrast, one type of the intramembranous particle aggregates, described by us as 'd-type' structures and called
'ciliary granule plaques' in this study is associated only with one single membrane,
namely the membrane of the ciliary shaft base. The present investigation attempts
to elucidate functional aspects of this conspicuous membrane specialization. It is also
shown that ciliary granule plaques are morphological elements clearly distinct from
'ciliary necklaces' (Gilula & Satir, 1972).
MATERIALS AND METHODS
Parainecium aurelia (mating type VIII) was cultured in a bacterized salad infusion medium.
The normal Ca 1+ -content was about 0 5 mM (Plattner, 1974). For the localization of Ca i +
up to 25 mM CaCl, .2H1O was added to cultures 10 min before fixation. The reactions of
paramecia upon addition of Ca 1+ were observed under a phase-contrast microscope. Ca J + was
localized essentially according to Oschman & Wall (1972) by fixation with glutaraldehyde,
or according to Constantin, Franzini-Armstrong & Podolsky (1965) by fixation with OsO 4 oxalate solutions.
Aliquots with different Ca 1+ concentrations in the medium were fixed in suspension for
1 h in 2 5 or 625 % glutaraldehyde, either without or with postfixation in 1 % OsO 4 ; for
controls cells were fixed in 1 % OsO 4 only. Other samples were treated with Ca 1+ (up to
25 rain) and then fixed in 1 % OsO 4 with dipotassium oxalate added in concentrations equivalent to Ca*+. Other cells were fixed (without previous addition of Ca 1+ to the culture medium)
in a 625 % glutaraldehyde solution to which up to 75 mM Ca 1+ was added. For further
controls similar experiments were made but comparable concentrations (up to 50 mM) of
either Na + , Mg a + (as chlorides) or La 3 + (as nitrate in cacodylate buffer p H 6-8) were added to
glutaraldehyde instead of Ca 1+ . In other controls fixation by glutaraldehyde preceded the
addition of Ca 1+ .
Double-distilled glutaraldehyde was used, which was obtained from Merck-Schuchardt
(Darmstadt and Munich, W. Germany) and stored at — 20 °C before use. All experiments
were conducted at room temperature. All fixatives were in o-i M cacodylate buffer, P H 7 2 ,
except for La 3 + solutions which were buffered at p H 6-8. After fixation cells were washed
with o-i M cacodylate buffer, p H 7-2, dehydrated in graded series of acetone solutions and
embedded in Epon. Sections of 50—80 run (silver interference colour) were analysed in a
Siemens Elmiskop I at 80 kV, either unstained or after double staining (20 min 7-5 %
magnesium uranyl acetate; 10 min alkaline lead citrate). For freeze-cleaving, which was
performed at — 100 °C in a Balzers unit (Balzers, Liechtenstein), the cells were frozen without
fixation or antifreeze treatment as described previously (Plattner et al. 1973).
Quantitative evaluation of micrographs was performed at final magnifications of 30000—
100000 depending on the parameters measured (see Fig. 1, p. 261). Cilia from all regions
of the cell body were evaluated, provided that the orientation of the sectioning plane or of
the freeze-fracture was appropriate; on ultrathin sections only longitudinally and crosssectioned cilia were evaluated; on freeze-cleaving replicas measurements were performed
only on longitudinal fractures.
RESULTS
When paramecia cultured in the normal medium which contains about 0-5 mM Ca2+
were fixed in glutaraldehyde only, single deposits of intense electron density occurred
occasionally adjacent to the inside of the basal portion of the ciliary membrane. The
Caz+ concentration could be raised up to 25 mM without deleterious effects on the
integrity of paramecia. When cells were fixed only in glutaraldehyde after previous
Ciliary granule plaques
2+
259
exposure to increased Ca concentrations, similar electron-dense deposits occurred
at the same sites, i.e. on the basal ciliary shaft membrane, but they occurred much
more regularly and were more conspicuous (Fig. 3) than with the normal Ca2+
content. The same deposits occurred when Ca2+ was added (up to 75 ITIM) to the
glutaraldehyde fixative. When cell suspensions to which up to 25 HIM Ca2"1" had been
added werefixedwith OsO4 in the presence of oxalate ions in concentrations equivalent
to Ca2*, similar electron-dense deposits occurred at the same sites (Figs. 5, 9) as
with glutaraldehyde. Five millimolar Ca2"*" was the minimum concentration necessary
for maximal deposit formation; increase in Ca2+ concentration above 10 mM had
little effect.
In all cases the electron density of the Ca^-dependent deposits was not enhanced
after section staining (Fig. 10 versus 11). No deposits occurred when Ca2"1" was
added after previous fixation with glutaraldehyde. When Na+, Mg2"*" or La3"1" were
added instead of Ca2+ but in similar concentrations to glutaraldehyde, electron-dense
deposits were rare or absent. When OsO4 was used as the only fixative, no deposits
were found, even in the presence of increased Ca2+ concentrations before or
during fixation. Fixation with OsO4 following glutaraldehyde had no influence on
the occurrence of Ca2+-dependent electron-dense deposits. Addition of oxalate ions
before fixation in OsO4 provoked considerable membrane damage which rendered
further analysis impossible. With OsO4-oxalate solutions less membrane damage
was found and quantitative evaluation could be performed.
The Ca2+-dependent electron-dense deposits coincide with the location of the
ciliary granule plaques which one observes on the base of freeze-cleaved Paramecium
cilia (compare Figs. 2, 4, 8). The ciliary granule plaques consist of groups of triple
vertical rows formed by individual membrane-intercalated particles (Figs. 4, 8). The
freeze-cleaved plaques are composed of granules only when viewed from the outside
of the cell (A-face), whereas on the B-face (i.e. the outer split half of the cell membrane
viewed from inside) plaques are represented by corresponding holes. Sometimes a
ciliary necklace occurs in addition to and basally from the plaques (Fig. 4). The
necklace consists of a single or double horizontal zig-zag row of membrane-intercalated
granules and corresponds to the structures described in more detail by Gilula &
Satir (1972).
Both the ciliary granule plaques as well as the Ca2+-dependent electron-dense
deposits are present on single or double cilia occurring on the outer cell surface as
well as on cilia of the cytopharynx. However, they might both also be absent from
any type of cilia; this phenomenon, however, and any possible differences in cilia
along the extension of the body surface were not analysed quantitatively.
To investigate in more detail the possible relationship between Ca2+-dependent
deposits and ciliary granule plaques, morphometric analyses, presented in Table 1,
were carried out. Structural details are documented in micrographs of Figs. 4-11.
Evidently all parameters tested are identical for deposits and plaques (Table 1).
They both occur at practically the same distance from the ciliary base. They have
comparable dimensions and the periodicity around the ciliary shaft is nearly the
same. Calculations based on the period between individual freeze-cleaved plaques
H
111(18)
±46
122(17)
±54
104 (4)
±53
—
±70
84 (22)
±27
"5(39)
±49
118(15)
H'
54d3)
+ 10
127 (5)
±34
101 (3)
±9
±24
136(17)
±33
110(18)
±40
109 (4)
B'
50(15)
±27
±22
P
±30
"8(13)
±25
102 (16)
±23
92 (20)
86(6)
±26
94 ('4)
±24
±30
105 (21)
For symbols see Fig. 1.
±26
51(6)
±29
—
59(2i)
48(16)
± 12
50(21)
±27
67 ('3)
±31
36(6)
±5
48(14)
—
45 (20)
± 24
±23
±30
±62
121 (38)
B
45 (22)
±6
44(20)
138(16)
Nh
—
—
—
3 0 (26)
± O'O
—
Nv
—
5 2 (5)
± i*6
_
—
_—
52(5)
±i-6
±09
48(23)
dh
—
—
—
—
±2
—
22 (22)
—
26(7)
±4
—
—
24(18)
±3
29(8)
±6
—
arranged particles. ± Values indicate standard deviations. Numbers in parentheses indicate numbers of structures analysed.
No data are given if only less than 3 observations could be made.
• Values pooled from data presented in detail in the last 3 entries of the table.
10 min 5, 10 or 25 mM Ca 2+
followed by glutaraldehyde*
10 mtn 10 mM Ca 2+ followed
by 1 % O s O 4 + i o m M d i - K oxalate
25 mM Ca a+ +glutaraldehyde,
simultaneously
10 min 5 mM Ca a+ followed
by gtutaraldchyde
10 min 10 mM Ca2+ followed
by glutaraldehyde
10 min 25 mM Ca2+ followed
by glutaraldehyde
Freezc-cleaving
Treatment of cells
Table i. Morphometric characteristics of ciliary granule plaques (derived from freeze-c leaving results) as compared with data
from Ca2+-dependent electron-dense deposits (derived from ultrathin sections)
Ciliary granule plaques
261
in relation to the actual diameter of the ciliary shaft (which can be only partly
surveyed) indicate that 8-10 plaques could be accommodated theoretically around
the ciliary shaft. Similarly a 9-fold periodicity could be frequently derived from the
spacings along a sequence (which is mostly incomplete) of individual electron-dense
deposits along a cross-sectioned ciliary base. The Ca^-dependent deposits are
sometimes seen to face the 9 doublet microtubules.
'100 nm
Fig. 1. Scheme of ciliary granule plaques, drawn approximately to scale according
to the morphometric analyses; parameters indicated are quantified in Table 1. Each
plaque consists invariably of 3 vertical rows of membrane-intercalated particles.
Individual plaques are separated by smooth membrane regions from each other and
from the ciliary necklace, which only sometimes is recognized simultaneously with
the ciliary granule plaques.
Sometimes granular elements are faintly visible (especially on grazing sections
tangential to the ciliary membrane) and fine filaments radiate from there to the
outer microtubules (on median sections) on cells fixed conventionally, i.e. with
glutaraldehyde and OsO4 without the addition of Ca2* (Fig. 7); the ciliary membrane
mostly bulges out in this basal region, which coincides with the region which otherwise contains the Ca2+-dependent deposits or the plaques, respectively. As seen in
Figs. 5, 6 and 9-11 the Ca2+-dependent deposits lie on the inner side of the ciliary
membrane, which they do not penetrate completely; for particles composing plaques
(Figs. 4, 8) one has to assume that they penetrate the ciliary membrane (cf. Plattner
et al. 1973), in contrast to deposits. Therefore it cannot be stated whether the deposits
are identical to or only associated indirectly with the plaque granules. However, some
262
H. Plattner
direct or indirect relation to plaque granules is very likely also for the reason that
deposits are always attached closely to the inner side of the very same region of
the basal ciliary membrane. Sometimes Ca^-dependent deposits display a periodic
SM&structure, especially on cilia cut in median positions (Figs. 5, 6); in most cases,
however, deposits were too heavy to allow for the identification of substructures.
According to the morphometric analysis, these substructures correspond probably
to the periodic arrangement of individual granules which constitute freeze-cleaved
plaques.
The various parameters of the Ca^-dependent deposits - obtained by different
methods - are all very much alike (Table 1): upon addition of 5, 10 or 25 mM Ca24"
before glutaraldehyde fixation; upon addition of 25 mM Ca H to glutaraldehyde;
upon exposure to iomM Ca2+ and subsequent fixation in OsO4 with 10 mM oxalate.
No difference was found between cells fixed in 2-0 or 6-25 % glutaraldehyde,
respectively.
With all methods used, electron-dense deposits were associated also with other
sites on the cell membrane, e.g. along the surface membrane including more apical
regions of the ciliary membrane. The occurrence of these deposits was less frequent
and less regular, however, and they were structurally less defined than the Ca2+dependent deposits which coincide with the ciliary granule plaques. With the
glutaraldehyde, as well as with the OsO4-oxalate method, electron-dense deposits
also occur sometimes along the outer microtubules and the fibrillar substructures
connecting the peripheral 9 and the central 2 microtubules of cilia. The axosome
was intensely electron-dense only when fixation was performed with OsO4-oxalate
solutions.
Electron-dense deposits disappeared more and more from all these sites, including
the basal ciliary membrane, as the Ca2+ concentration in cultures was raised to levels
deleterious to the cells (> 25 mM). With Ca2+ added to the glutaraldehyde fixative
this effect became evident only at higher concentrations (> 75 HIM).
DISCUSSION
Recently several authors have described the occurrence of conspicuous regular
arrangements of membrane-intercalated particles consisting of 1-6 (or more) parallel,
horizontal rows of granules within the freeze-cleaved basal portion of ciliary membranes; these structures were termed ciliary necklaces (cf. Gilula & Satir, 1972). A
ciliary necklace surrounds the basal portion of freeze-cleaved cilia in a variety of
protozoan, invertebrate and vertebrate cells (Satir & Gilula, 1970; Flower, 1971;
Gilula & Satir, 1972; Wunderlich & Speth, 1972; Matsusaka, 1974; Tani, Ikeda,
Nishiura & Higashi, 1974). In contrast, the occurrence of ciliary granule plaques
has been reported only from Tetrahymena (Wunderlich & Speth, 1972) and from
Paramecium, where we called these membrane specializations 'd-type' structures
(Plattner et al. 1973). In both species the plaques may occur simultaneously with the
ciliary necklace; however, the occurrence of a necklace is not readily recognizable
in all cilia of Paramecium, even when ciliary granule plaques are present. The location
Ciliary granule plaques
263
of plaques apical from the necklace and the different morphology clearly establish
these structures as separate entities.
With appropriate fixation procedures electron-dense deposits could be localized
on the inside of the membrane of the ciliary shaft base, when living P. aurelia cells
were exposed to Ca2+ concentrations which on the one hand exceeded the concentrations normally present in cultures and which on the other hand are known not
to interfere with the maintenance of the physiological, Ca^-dependent surface
potential in P. caudatum (Naitoh & Eckert, 1974). Both the oxalate precipitation
method (Constantin et al. 1965) and the method of Oschman & Wall (1972) produced
Ca^-dependent electron-dense deposits. The same dense deposits occurred when
Ca2"1" was added to the glutaraldehyde fixative in concentrations identical with those
in in vivo experiments or higher (up to 75 mM). They were absent when Ca2+ was
substituted by Na+, Mg2+ or La3"1". These observations support the assumption that
the deposits contain Ca2+. Recently, it was shown by X-ray microanalysis that
electron-dense deposits obtained with both methods contain Ca2+. After fixation
with glutaraldehyde in the presence of Ca2+ the main constituents of electron-dense
deposits formed are calcium and phosphorus (Hillman & Llinds, 1974; Oschman,
Hall, Peters & Wall, 1974; Skaer, Peters & Emmines, 1974) but not magnesium; as
in the present study, Mg2+ also failed to produce electron-dense deposits in other
systems (Hillman & Llinas, 1974). Deposits obtained with oxalate in the fixative
contain primarily Ca2+ (Diculescu & Popescu, 1973) but not Mg24" or phosphorus
(Coleman & Terepka, 1972a, b\ Braatz & Komnick, 1973). Since the results obtained
by X-ray microanalysis with either the Oschman & Wall method or with the oxalate
method appear quite similar in all cell types tested so far, they seem to be more
generally applicable. Both methods are evidently more specific for the localization of
Ca2+ than the pyroantimonate method (Garfield, Henderson & Daniel, 1972).
Several parameters were determined for Ca2+-dependent deposits on ultrathin
sections and for the freeze-cleaved ciliary granule plaques. The number of cilia
available for quantitation was restricted by the necessity of selecting appropriate
sectioning planes. It appears from Table 1 that all parameters determined for the
Ca^-dependent deposits (obtained under various experimental conditions) always
coincide with those of ciliary granule plaques. Morphological equivalents were
faintly visible also on conventionally (in the absence of additional Ca2+) fixed and
cut cilia (Fig. 7). One can, therefore, conclude that all these structural elements
correspond to each other.
Among several mechanisms, which were discussed recently by Oschman et al.
(1974) as being possibly involved in the formation of Ca2+-dependent deposits
(occurring upon glutaraldehyde fixation), two aspects appear important in the present
case: enhanced permeability of membranes for Ca2+ and trapping of Ca2+ by inorganic
phosphate liberated at sites of ATPase activity. The first aspect has to be seriously
considered, since on the one hand Ca2"1" is much less concentrated within paramecia
(icr 7 M; Naitoh & Eckert, 1974) than outside (0-5 mM or experimentally increased
concentrations), and since on the other hand both glutaraldehyde (Elbers, 1966;
Morel, Baker & Wayland, 1971; Vassar, Hards, Brooks, Hagenberger & Seaman,
17
C E L iS
264
H. Plattner
1972) and OsO4 (Elbers, 1966; Bone & Denton, 1971) greatly enhance the cationpermeability of biomembranes. In consequence, even if Ca2+-dependent deposits
were artifacts, their reproducible occurrence with widely different methods makes it
very likely that they are functional equivalents identical to or associated with the
specific morphological substructures with which they coincide. The second aspect
discussed by Oschman et al. (1974) has also to be considered; however, it appears
less important due to the absence of phosphorus in deposits produced by oxalate
precipitation (Coleman & Terepka, 1972a, b; Braatz & Komnick, 1973) and because
OsO4 would probably reduce any ATPase-activity in the present experiments;
nevertheless, Ca2+-binding sites could coincide with regions also containing sites of
ATPase activity.
Although in Tetrahymena the ATPase contained in the dynein arms, which are
attached to the doublet microtubules, binds Ca2+ as well as Mg2+ (Gibbons, 1966)
the role of Ca24" in ciliary activity is not yet clear (cf. Stephens, 1974)- In Paramecium
the cyclic bending of cilia is regulated by a Mg^-dependent ATPase; in contrast,
Ca2+ is a cofactor for another ATPase system, which is responsible for the orientation
of the effective stroke (Naitoh, 1969; Naitoh & Eckert, 1974). In model systems
obtained by Triton extraction of Paramecium cells, Ca2+ has also some influence
on the beat frequency (Naitoh & Kaneko, 1972). Since ciliary activity originates at
and probably depends upon the integrity of the ciliary base (Goldstein, 1974), it
would be reasonable to assume the location of Ca2+-binding sites close to the ciliary
base. In paramecia, such binding sites would be saturated with Ca2+ concentrations
of about 5 ran, since the Ca2+-dependent deposits at the ciliary base are not
remarkably enhanced by higher Ca2+ concentrations; the same concentration was
reported to saturate Ca^-binding sites of synaptic vesicles (Politoff, Rose & Pappas,
J
974)The absence of ciliary granule plaques in many systems might indicate that the
plaques would not necessarily be involved directly in cyclic bending of cilia in
general. This question could be further clarified by comparative ultrastructural
analyses. It remains to be seen whether the Ca2+-binding sites, which were shown
to coincide or to be associated with the ciliary granule plaques, are involved in
Ca2+-directed regulations of ciliary activity - such as direction or frequency - at
least in ciliates. One also certainly has to consider possible functions of Ca2+-binding
sites other than in connexion with motility - for example, mating agglutination
occurring on isolated Paramecium cilia in the presence of Ca2+ (Takahashi, Takeuchi
& Hiwatashi, 1974). The present study provides at the very least some evidence that
ciliary granule plaques (as well as probably ciliary necklaces) are not merely
ornamental structural elements decorating the necks of protozoan cilia.
I gratefully acknowledge the support provided by Professor L. Bachmann in the course
of some experiments, the critical reviewing of the manuscript by Professor F. Miller, the
technical assistance of Mrs D. Wolfram, the performance of the photographic work by
Miss E. M. Praetorius, and financial support from the Deutsche Forschungsgemeinschaft
(grant PI 78/1).
Ciliary granule plaques
265
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(Received 16 December 1974)
Fig. 2. Platinum-carbon replica from a freeze-cleaved cell. Arrows point to ciliary
granule plaques contained within the base of the split ciliary membranes. Note
coincidence of plaques with Ca1+-dependent electron-dense deposits in Fig. 3. The
shadow-casting direction is from bottom to top. x 36200.
Fig. 3. Cell exposed to 10 mM Cas+ and subsequently fixed in glutaraldehyde only;
section staining. Electron-dense deposits occur on the basal portion of cilia (arrows),
i.e. at the same sites as ciliary granule plaques are located (compare Fig. 2); deposits
are closely attached to the inside of the ciliary membrane. Some deposits are found
also in more apical regions of cilia and one deposit (upper part of figure) is associated
with the membrane complex formed by the cell membrane and the outer alveolar
membrane, x 36200.
Ciliary granule plaques
268
H. Planner
Figs. 4-7. Lateral projection of basal ciliary regions, x 100000.
Fig. 4. Freeze-cleaved ciliary base exhibiting several ciliary granule plaques (bars
and arrows) which are composed each of 3 vertical lines of membrane-intercalated
particles. Further down a ciliary necklace is faintly visible as a single zig-zag row
of particles. Shadow casting is from top to bottom. From Plattner et al. (1973).
Fig. 5. Exposure to 10 mM Ca t+ followed by fixation with 1 % OsO4 and 10 mM
oxalate. The electron-dense deposits occurring along the inner side of the basal
ciliary membrane (bar and arrows) are quite similar to those obtained with glutaraldehyde fixation in Fig. 6. Note the periodic arrangement of deposits.
Fig. 6. Cell with added 5 mM Caa+ and subsequently fixed in glutaraldehyde only;
section staining. Electron-dense deposits are attached to the cell membrane; they
are concentrated at a certain distance from the lowest part of the cilium. Although
deposits occur also in more apical regions, a periodic arrangement of deposits (bar
and arrows) can be recognized only in regions comparable to specialized membrane
regions seen in Figs. 4, 5 and 7.
Fig. 7. Pair of cilia after consecutive fixation in glutaraldehyde and OsO4 without
addition of Ca l+ . The basal ciliary membrane is devoid of electron-dense deposits.
In a position marked by bars and arrows, i.e. where periodically arranged electrondense deposits occur in Figs. 5 and 6 and where periodically arranged membraneintercaJated particles are located in Figs. 4 and 8, this figure displays granular
elements along a sectioning plane tangential to the cell membrane (right); furthermore, fibrillar elements radiating from there are faintly recognizable in a median
section (left).
Figs. 8-11. Vertical projections of the basal ciliary region, x 100000.
Fig. 8. This freeze-cleaved ciliary base displays 3 ciliary granule plaques consisting
each of 3 vertical rows of membrane-intercalated particles (arrows). Some particles
occur also outside the plaques. Compare the coincidence of plaque-containing regions
with equivalent structures in Figs. 5—7 and 9—11. A ciliary necklace cannot be
recognized on this cilium (compare Fig. 4). Shadow casting from bottom left to
top right.
Fig. 9. Exposure to 10 mM Ca1+ with subsequent fixation in 1 % OsO4 with
10 mM oxalate added; no section staining. This cross-cut ciliary base displays
electron-dense deposits (arrows) associated with the inner side of the basal ciliary
membrane.
Fig. 10. Fixation in glutaraldehyde supplemented with 75 mM Ca1+; section
staining. Electron-dense deposits (arrows) are similar to those in Fig. 9.
Fig. 11. Unstained control to Fig. 10.
Ciliary granule plaques
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