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Structure of the satellite cell sheath around the cell body axon hillock and initial segment of frog dorsal root ganglion cells.

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THE ANATOMICAL RECORD 215: 182-191 (1986)
Structure of the Satellite Cell Sheath Around the
Cell Body, Axon Hillock, and Initial Segment of Frog
Dorsal Root Ganglion Cells
EMIKO MATSUMOTO AND JACK ROSENBLUTH
Departments of Physiology and Rehabilitation Medicine, New York University School of
Medicine, New York, N Y 10016
ABSTRACT
The structure of the satellite cell sheath of frog dorsal root ganglion
cells was studied in thin sections and freeze-fracture replicas. The sheath around
the cell body is composed of thin satellite cell lamellae closely applied to the
neuronal plasma membrane. At the axon hillock the sheath divides into outer and
inner components separated by a broad space containing a distinctive extracellular
matrix and occasional flattened satellite cell processes. The sheath around the initial
segment is usually multilayered but less compact than that around the cell body,
and in some places it exhibits node-like interruptions. Apart from occasional particle
groupings characteristic of tight junctions and gap junctions, the satellite cells
display homogeneously distributed intramembranous particles in both fracture faces
in all regions of the sheath.
Our previous study of frog dorsal root ganglion cells
(Matsumoto and Rosenbluth, 1985) has shown that the
plasma membrane of the axon hillock (AH) and initial
segment (IS) regions of these neurons has high concentration of E-face intramembranous particles compared
with that of the cell body. Histochemical data (Devor
and Obermayer, 1984) derived by the ferric ion-ferrocyanide method (Quick and Waxman, 1977) also suggest
that the AH and IS of peripheral sensory neurons, like
the IS of spinal cord neurons (Waxman and Quick, 1978),
have properties similar to those of the node of Ranvier.
Physiological studies of bullfrog ganglia (Ito, 1957, 1959)
concluded that the IS has a lower threshold for spike
generation than the cell body, in this respect too resembling the node of Ranvier. Since the satellite cell sheath
investing the sensory neuron may play a n important
role in regulating its microenvironment, and therefore
its electrical activity (Pannese, 1981),the possibility exists that the sheath around the AH and IS may be
structurally different from that around the cell body
(CB),corresponding to differences in the functions of the
respective regions.
Some evidence for regional differentiation of the
sheath was noted in our previous report on the plasma
membrane of the neuron. This paper is concerned primarily with the structure of the sheath and shows that
although the plasmalemma of the satellite cell is similar
in all regions of the sheath, the gross structure of the
AH and IS sheath is quite different from that of the CB
sheath but similar to that of the sheath around the
nodal region of myelinated axons. These results suggest
that the satellite cell in the AH and IS regions is differentiated in accordance with the specialized functions of
these regions of the ganglion cell.
0 1986 ALAN R. LISS, INC.
MATERIALS AND METHODS
Grass frogs (Ranapipiens)were anesthetized with 10%
ethyl carbamate and perfused through the heart with a
mixture of 2%formaldehyde and 2%glutaraldehyde in
0.1 M sodium cacodylate buffer at pH 7.4. Dorsal root
ganglia were dissected out and kept in the same fixative
for one to several days.
For electron microscopy of thin sections, the tissues
were rinsed with frog Ringer’s solution and postfixed
with ferrocyanide-reduced osmium tetroxide (Karnovsky, 1971) in 0.1 M cacodylate buffer for 1-2 hr. Then
the specimens were dehydrated in methanol and embedded in Araldite. Thin sections were stained with potassium permanganate and uranyl acetate as described
elsewhere (Rosenbluth, 1974).
For freeze-fracture, the tissues were cut into small
pieces, glycerinated in lo%, 20%, and 30% glycerol in
Ringer’s solution, mounted on gold specimen disks, and
frozen in Freon 22 cooled by liquid nitrogen. The specimens were fractured with a microtome knife in a Balzers BAF 400D freeze-fracture unit at -115°C and
shadowed with platinum-carbon and carbon. For complementary replication, specimens were vibratomed into
50 pm slices, glycerinated, sandwiched between gold
disks, frozen, fractured in a double-replica holder, and
shadowed as above. The replicas were processed as described previously (Tao-Cheng and Rosenbluth, 1984)
and examined in a Philips EM 300 electron microscope.
Received April 23, 1985; accepted November 18, 1985
Dr. Matsumoto’s present address is Department of Fine Morphology, The Institute of Medical Science, The University of Tokyo, Japan.
SATELLITE CELL STRUCTURE
183
visible at arrows in the P-face of a satellite cell membrane. C, neuronal
cytoplasm; N, satellite cell nuclear membrane. x 30,000.
Fig. 3, Enlargement of the rectangle in Figure 2, ps, P-face of a
satellite cell membrane. x 64,000.
Fig. 4. A gap junction (arrow) in the E-face of the satellite cell
Fig. 2. Freeze-fracture replica of a satellite cell sheath around the
CB. Several layers of the sheath are exposed. Short tight junctions are membrane (ES). X270,000.
Fig. 1. Thin section showing a satellite cell sheath around the nerve
cell body (CB). Protrusions of the neuron into the sheath are visible at
asterisks. Arrow, coated vesicle; arrowhead, coated pit; F, fibroblast;
BL, basal lamina. X33,OOO.
Fig. 5. Thin section showing profiles of initial segments (IS),a heminode (HN), and the axon hillock (AH) of a ganglion cell (CB). The
satellite cell (SC) sheath over the AH is split into inner (I) and outer
(0)components enclosing an interior space (Sf. E, endoneurium. ~9,000.
Fig. 6. High-power view showing the distal end of the outer compo-
nent. Collagen fibrils penetrate a short distance into the space between
two sheaths adjacent to the end, but not beyond the basal lamina that
bridges across the space (S) between the outer (0)
and inner (I) cornponents of the sheath (at arrow). E, endoneurial space; IS, initial segment; BL, basal lamina lining inner and outer components. ~42,000.
Fig. 7. Thin section showing a satellite cell sheath around the proxima1 IS. Inner (I
and
)outer (0)
components are separated by an interior
space (S) containing a flocculent material. Ax, axon; CB, nerve cell
body. ~ 9 , 0 0 0 .
Fig. 8. Trabecular processes in the space between inner and outer
components. The processes are partially coated with flocculent material. Arrows, pinocytotic pits; Ax, axoplasm; arrowheads, focal densities; I, inner component. ~ 4 0 , 0 0 0 .
186
E. MATSUMOTO AND J. ROSENBLUTH
Fig, 9. Thin section showing a node-like gap (between arrows) in the cell sheath around the IS. E,
endoneurial space containing collagen fibrilis; I, interior space containing flocculent material (cf. Fig. 14
in Matsumoto and Rosenbluth, 1985). x 30,000.
Electron micrographs of the replicas were printed at a
final magnification of 130,000 and the intramembranous particles were counted. The area used for counting
was measured with a planimeter and the particle concentration was determined.
RESULTS
The satellite cell sheath around the neuronal cell body
is composed of one to several lamellae 0.02-0.5 pm in
thickness separated from each other by extracellular
clefts 10-50 nm in width (Fig. 1).The satellite cell cytoplasm is rich in filaments, microtubules, and mitochondria, and pinocytotic pits and vesicles are also abundant
especially in the outermost layer, which is covered by a
basal lamina (Fig. 1).In replicas, the satellite cells display a surface that is smooth except for groups of pinocytotic pits (Fig. 2). Short strands (0.2-0.6 pm) of tight
junctions and small gap junctions between adjacent layers can be seen occasionally as well (Figs. 2-4). Intramembranous particles are homogeneously distributed
over the plasmalemma of each layer. Their concentration in the P-face is 1,509 f 189/pm2 (9 cells, area
counted 5.06 pm2) and in the E-face 1,032 & 97/pm2 (5
cells, area counted 2.75 pm2).
As noted previously (Matsumoto and Rosenbluth,
19851, over the axon hillock and over the beginning of
the initial segment the satellite cell sheath is split into
outer and inner components (Figs. 5-7, 10). The inner
-
component enwraps the neuron closely, but the outer
component is separated by a n extracellular space several microns wide (Figs. 5, 7, 10). Trabecular processes
extend from both inner and outer components into this
space (Figs. 5-8), which also contains a distinctive flocculent material. The outer surface of the outer component is covered by a basal lamina, which separates the
interior space between the outer and inner components
from the connective tissue space outside. Collagen fibrils
do not penetrate past this basal lamina into the interior
space even though the latter is continuous with the
connective tissue space where the outer component ends
over the initial segment (Fig. 6). Focal densities are
sometimes present at the tips of satellite cell processes
and in the plasmalemma of the outer layer of the inner
component (Fig. 8).
Such complex sheaths are present around all spinal
ganglion cells, but large neurons have larger interior
spaces containing more trabecular processes, whereas
small neurons have smaller interior spaces with only a
few finger-like processes (Fig. 5). The nucleus of the
satellite cell often occurs near the axon hillock, and
trabecular processes may extend from the satellite cell
perikaryon as well.
In replicas, the outer component of the sheath displays
many undulations and pinocytotic pits (Figs. 10,16).The
particle density in the inner and outer components
around the axon hillock (Figs. 11-13) is comparable to
SATELLITE CELL STRUCTURE
Fig. 10. Freeze-fracture replica of the satellite cell sheath around the proximal IS. The axon is invested
by the irregular inner component of the satellite cell sheath (I) consisting of multiple lamellae, a space
(S),and the outer component of the sheath (0).Ax, axoplasm; CB, nerve cell body; E, endoneurial
connective tissue (cf. Fig. 15 in Matsumoto and Rosenhluth, 1985). x 10,000.
187
Fig. 11. Freeze-fracture replica of the inner component of the sheath
around the AH. Ax, axon; PS, P-face of the satellite cell membrane;
ES, E-face of another satellite cell membrane. ~24,000.
Fig. 13. Enlargement of the rectangle in Figure 11showing particles
in the P-face of the satellite cell membrane. ~ 6 0 , 0 0 0 .
Fig. 12. Plasma membrane of the inner satellite cell component (ES)
showing E-face particles. SC, satellite cell lamellae; S, space between
Figs. 14, 15. Complementary pair showing P-face (Fig. 14)of a satellite cell membrane around the IS containing a tight junction and
closely associated gap junction (arrow). The E-face (Fig. 15)shows the
same structures. x 105,000.
inner and outer components. ~ 6 0 , 0 0 0 .
SATELLITE CELL STRUCTURE
189
Fig. 16. Freeze-fracture replica of the outer component of the sheath around the AH. The P-face of the
satellite cell membrane (PSI displays numerous intramembranous particles and pinocytotic pits (arrows).
BL, basal lamina; Co, collagen fibrils; E, endoneurial space; S, space between outer and inner components.
x 30,000.
that of the sheath around the remainder of the cell bod
(P-face, 1,532 f 201/pm2, 3 cells, area counted 1.69 pm ,
E-face 844 f 222/pm2, 6 cells, area counted 1.83 pm2.
The sheath around the initial segment is also composed of thin layers of satellite cell processes. The number of these layers varies from 1to 50 along the length
of the initial segment. The inner few layers invest the
axon closely, but the outer layers are much looser (Fig.
9). Gaps occur in the satellite cell sheath (Matsumoto
and Rosenbluth, 1985) so that the axolemma is directly
exposed to extracellular space in some places (Fig. 9).
Such interruptions, which are 1-1.5 pm long, occur along
the entire length of the initial segment. The complexity
of the sheath around the AH and initial portion of the
IS is illustrated in a tracing shown in Figure 17.
Distally, the initial segment terminates at a heminode
(Fig. 5), where the satellite cell and the adjacent
Schwann cell both extend multiple villous projections
toward the axon. The region covered by these villi varies
in length from -1.2 pm to more than 3 pm, and in
fortunate sections a node-like undercoating may be seen
along this entire extent.
Freeze-fracture replicas of the satellite cell sheath
around the initial segment reveal that the inner layers
occasionally display gap junctions and tight junctions in
close association (Figs. 14,15). Otherwise the intramembranous article concentrations in this region are 1,421
85ipm!. (8 cells, area counted 3.41 pm2) in the P-face
and 1,064 f 151/pm2(5 cells, area counted 1.76 pm2) in
E
the E-face. Both concentrations are similar to those of
the sheath around the cell body and axon hillock.
DISCUSSION
This study shows that the satellite cells investing dorsal root ganglion cells have similar concentrations of
intramembranous particles in CB, AH, and IS regions.
The E-face concentration is relatively high, as it is in
the plasmalemma of oligodendrocytes (Massa and Mugnaini, 1982). It has been reported that the satellite cell
membrane around chick (Pannese et al., 19771, adult
fowl (Pannese et al., 19781, and rat (Gotow et al., 1985)
dorsal root ganglion cells and rat sympathetic ganglion
cells (Elfvin and Forsman, 1978) exhibits the orthogonal
arrays of intramembranous particles characteristic of
mammalian astrocyte membranes (Dermietzel, 1973,
1974; Landis and Reese, 1974; Brightman and Reese,
1975; Nabeshima et al., 1975; Hanna et al., 1976; Anders
and Brightman, 1979; Wujek and Reier, 1984). Such
assemblies were not found in the frog satellite cell membrane, however. This is in accord with previous reports
that frog ependymal cells (Korte and Rosenbluth, 1981)
and astrocytes (Wujek and Reier, 1984) lack such assemblies as well. The only particle groupings seen were
short tight junctions and gap junctions distributed between layers of satellite cells around both the CB and
IS. Such junctions have been reported previously only
in the sheath around the CB (Pannese et al., 1978; Gotow et al., 1985). Although gap junctions are rare here,
190
E. MATSUMOTO AND J. ROSENBLUTH
Fig. 17. Tracing of a satellite cell sheath in the AH and IS regions. BL, basal lamina; S, interior space
between outer and inner components of the AH sheath containing flocculent material and trabecular
processes; arrows, gaps in the satellite cell around the IS (cf. Fig. 13 in Matsumoto and Rosenbluth, 1985).
191
SATELLITE CELL STRUCTURE
compared with satellite cells of sympathetic ganglia
(Elfvin and Forsman, 1978), their existence suggests
some degree of metabolic or electrical coupling of satellite cell layers that may help in regulating the ionic
environment of the neuron. The short tight junctions
reported here and by Pannese et al. (1978) are of uncertain significance because they are too short to form
diffusion barriers. Indeed, tracers in the extracellular
space can reach the neuronal plasma membrane (Rosenbluth and Wissig, 1964).
Although the satellite cell plasmalemma is not regionally differentiated, the sheath around the AH and IS is
different from that around the CB on a grosser level. In
the AH and IS regions satellite cells form trabecular
structures. Similar structures have been also observed
in bullfrog spinal ganglia (Pannese, 1981), frog (Pick,
1963;Yamamoto, 1963; Uchizono, 1964;Taxi, 1976),toad
(Uchizono, 19641, and lizard (Pannese, 1981)sympathetic
ganglia. However, such structures have not been reported in other vertebrates (see Pannese, 1981, for review). In mammalian spinal ganglia, the satellite cell
sheath around the AH is thicker than that around the
CB, and the space between the satellite cell processes is
narrow (Pannese, 1960; Moses et al., 1965; Pineda et al.,
1967; Pannese, 19811, although larger “cisternae” also
occur (Zenker and Hogel, 1976).The flocculent material
around the satellite cell processes resembles the material around the Schwann cell microvilli at the node of
Ranvier (see Rosenbluth, 1983, for review) and may also
be involved in regulation of the ionic environment
around the AH and IS, especially in view of the interruptions in the IS satellite cell sheath, which resemble
nodal gaps. It is not known to what extent such interruptions also occur in the sheath surrounding the IS of
adult mammalian spinal ganglia, but at least during
developmental stages, some regions of the axolemma
have been shown t o be exposed t o the extracellular space
(Yamadori, 1970).
Our previous study (Matsumoto and Rosenbluth, 1985)
has demonstrated that the axolemma exhibits patches
of undercoating in the IS and high concentrations of Eface particles in the IS and AH. The configuration of the
sheath covering the IS and AH regions further supports
the similarity between them and the node of Ranvier
and suggests that the respective regions are functionally
similar as well.
In summary, the present study shows that the satellite
cell sheath around the AH and IS regions of frog dorsal
root ganglion cells displays a complex structure, resembling that at the node of Ranvier.
ACKNOWLEDGMENTS
This study was supported by grants from the National
Institutes of Health (NS-07495) and Muscular Dystrophy Association. Dr. Matsumoto was a postdoctoral fellow on leave from the Department of Fine Morphology,
University of Tokyo.
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