The University of Maryland, Baltimore, School of Medicine, Department of Anatomy and Neurobiology, and the Program in Neuroscience, 685 West Baltimore Street, Baltimore, MD 21201, USA
* Author for correspondence (e-mail: lrich001{at}umaryland.edu)
Accepted 28 March 2003
![]() |
SUMMARY |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Key words: Midline glia, Glial sling, Glial wedge, Corpus callosum, Axon guidance, Cell migration, Cortical development, Mouse
![]() |
INTRODUCTION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Further evidence for the role of the sling in callosal axon pathfinding
came from two types of experiments; ablation of the sling in utero which
resulted in agenesis of the corpus callosum, and rescue of the sling in vivo
in surgically induced acallosal animals
(Silver et al., 1982;
Silver and Ogawa, 1983
;
Katz et al., 1983
;
Smith et al., 1986
). In the
second series of experiments nitrocellulose membranes were inserted at the
midline of acallosal mice and endogenous midline glia readily grew over the
implant. Callosal axons were able to cross the midline only in the regions of
the glial-covered implant (Silver and
Ogawa, 1983
). It was postulated that the cells that rescued the
corpus callosum were derived from the sling.
However, in the early 1980s one of the limitations to studying the sling
was the paucity of available markers for the sling. Based largely on their
cellular morphology as determined by electron microscopic (EM) analysis, the
sling was previously thought to be composed of glial cells
(Silver et al., 1982). However
since the sling did not label with GFAP and did not have the morphological
characteristics of mature glia when viewed by EM
(Silver et al., 1993
), the
cells were classified as glioblast cells rather than mature astrocytes. Given
the lack of available markers for the sling, Silver and colleagues did not
rule out the possibility that the sling may contain neurons or other cell
types (Silver et al., 1982
;
Silver et al., 1993
). In order
to determine the putative axonal guidance molecules expressed by the sling we
first sought to characterize the cellular makeup of the sling and identify
molecular markers of these cells so that they could be easily studied and
isolated.
Here we have identified a number of makers of the sling and characterized the electrophysiological properties of sling cells in whole-cell current clamp recordings. These data indicate that the sling is largely composed of neurons. We also find that the sling remains proliferative until early postnatal ages and that the disappearance of the sling by P10 is not due to widespread cell death. These findings suggest the hypothesis that the sling provides a pathway for developing neurons migrating from the subventricular zone (SVZ), possibly to populate adjacent regions of the brain.
![]() |
MATERIALS AND METHODS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Immunohistochemistry
Embryos (collected as described below) were perfused with 4%
paraformaldehyde and brains were sectioned with a vibratome into 50 µm
thick sections. Immunohistochemistry was performed as previously described
(Shu et al., 2000). Primary
antibodies used were: (1) mouse anti-neuronal nuclei (NeuN; Chemicon, CA) 1:
3,000 for nickel-DAB reaction, 1:1000 for fluorescent Cy3 detection; (2)
rabbit anti-cow glial fibrillary acidic protein (GFAP; DAKO, Denmark) 1:1000
for fluorescent Cy2 detection; (3) rabbit anti ß-tubulin III (TUJ1;
Babco, CA) 1:10,000 for Cy2 detection; (4) rabbit anti-calretinin (Swant)
1:500 for Cy2 detection; (5) mouse anti-bromodeoxyuridine (BrdU; DAKO) 1:1000
for Cy3 detection and (6) PCNA (DAKO) 1:1000 for Cy2 detection. Secondary
antibodies used were: (1) biotinylated donkey anti-mouse (Jackson
ImmunoResearch Laboratories) 1:600; (2) Cy2-conjugated donkey anti-rabbit IgG
(H+L) (Jackson ImmunoResearch Laboratories) 1:400 and (3) Cy3-conjugated
donkey anti-mouse IgG (H+L) (Jackson ImmunoResearch Laboratories) 1:400.
Controls were performed for each antibody by eliminating the primary antibody
to determine if there was non-specific staining from the secondary antibody
alone.
Preparation of slices for electrophysiological recording
C57BL/6J mice between E17 and birth were obtained by anaesthetizing their
mother with sodium phenobarbital (Nembutal; Abbott Laboratories, IL) at 0.07
mg/g body weight and then placing her on a warming pad to maintain body
temperature. Once deeply anesthetized, the mother's abdomen was opened to
expose the uterus. Pups were removed sequentially and placed on ice, if older
than E16, until deeply anesthetized. Pups were removed from the uterine horns
and then decapitated before removing the brain. Postnatal mice were first
anaesthetized on ice before decapitation and removal of the brain. Live brains
were blocked in 3% low melting point agar (Sea plaque, FMC Bioproducts) in
L-15 medium (Gibco BRL). Coronal sections were made on a vibratome (Leica) at
400 µm and the sections containing the sling were collected.
Slices were kept in a holding chamber that contained artificial cerebrospinal fluid (ACSF) at room temperature, aerated with 95% O2 and 5% CO2. ACSF was composed of (in mM): NaCl 124, NaHCO3 25, N,N-bis[2-hydroxyethyl]-2-aminoethanesulfonic acid (BES) 5, KCl 3, MgSO4 1.3, CaCl2 2.0 and glucose 15. Recordings were made at room temperature in a submersion type chamber perfused with ACSF at 2 ml/minute. Electrode placement and cell selection were performed with near-infrared differential-interference-contrast microscopy (NIR-DIC), using 40x (0.8 NA) water immersion objective in a fixed-stage, upright microscope (BX50-WI; Olympus).
Whole-cell current-clamp recordings were obtained with an Axon 1D amplifier
(Axon Instruments, Foster City, CA), from 36 sling cells from slices harvested
from E17 (n=21), E18 (n=4), P0 (n=6), P1
(n=3) or P2 (n=2) mice and 4 glial wedge cells at E17.
Recordings were digitized with an ITC-18 interface (Instrutech, Port
Washington, NY) and acquired using PULSE software (HEKA Elektronik, Germany)
on a Power PC Macintosh Computer (Apple, Inc., Cupertino, CA). The impedance
of patch electrodes was 3 to 5 M. The intracellular recording solution
contained (in mM) potassium gluconate 120, KCl 10, Hepes 10, MgCl2
1, MgATP 2.5, Tris-GTP 0.2, BAPTA 0.1 and biocytin 2.6; pH was adjusted to
7.3. Chemicals were obtained from Sigma (St. Louis, MO).
Cells were filled with biocytin through the recording pipette. We recovered
10/36 of the recorded sling cells and 3/4 of the recorded glial wedge cells
(due to fragility of the embryonic slices some cells were lost after recording
probably as the patch pipette was removed). Slices were fixed overnight in a
buffered solution containing 4% paraformaldehyde and then co-immunolabeled for
either NeuN (sling cells) or GFAP (glial wedge cells) as described above.
Visualization of the biocytin was performed as previously described
(Gottlieb and Keller, 1997).
In every case, biocytin-labeled cells stained with NeuN were localized to the
sling, whereas biocytin-labeled, GFAP-positive cells were in the wedge,
indicating that the anatomical location of these cells by infrared microscopy
was adequate to identify the cells.
BrdU and TUNEL labeling
Bromodeoxyuridine was dissolved in sterile saline and injected
intraperitoneally into pregnant dams at a concentration of 50 µg/g body
weight on E15, E16, E17 and E18 respectively. Embryos from mothers injected on
E15 and E16 were collected on E17 and those from mothers injected on E17 and
E18 were collected on P0. Postnatally, P2 pups were injected with BrdU and
collected on P3. The brains were processed for immunohistochemistry as
described above. For time-lapse experiments BrdU was injected into pregnant
dams at 9 am and embryos were collected at successive time points (separate
litters were used for time points over 4 hours).
For TUNEL labeling, E17, E18, P0, P3, P5 and P10 brains were sectioned into 10 µm thick sections with a cryostat. An ApopTag Plus Fluorescein In Situ Apoptosis Detection Kit (S7111, Intergen) was used following the manufacturer's protocol. The sections were further counterstained with ToTo-3 (Molecular Probes) to reveal brain morphology.
![]() |
RESULTS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
|
|
Cells within the sling express neuronal physiological properties
To determine if sling cells are capable of producing regenerative currents
in response to membrane depolarization we obtained whole-cell, current-clamp
recordings from sling cells in an in vitro slice preparation. Using
near-infrared video microscopy (Stuart et
al., 1993) to identify sling cells below the corpus callosum
(Fig. 2D), we obtained
recordings from 36 sling cells from slices harvested between E17 and P2.
Biocytin was included in the pipette solution to intracellularly fill the
recorded neurons (Fig. 3).
|
|
In more mature sling neurons (P0-P2), Na+ spikes dominated the
response to current injections, whereas Ca2+ spikes occurred only
in 4 of 12 cells (Fig. 2B).
When Ca2+ spikes did occur, they had a shorter duration
(12.7±4.9 mseconds at half width) and a smaller amplitude
(82.8±83.4 mV), compared with those recorded from E17-E18 sling cells.
The Na+ spikes at P0-P2 also had a shorter duration (4.4±2.3
mseconds at half-width), but an amplitude indistinguishable from that of
E17-E18 neurons (79.8±21.3 mV). TTX reversibly suppressed
Na+ spikes in 5 of 6 P0-P2 cells, and addition of Cd2+
reversibly suppressed Ca2+ spikes in 4 of 4 of these neurons. In
response to hyperpolarizing current injections, 6 of 11 neurons recorded at
P0-P2 produced a response consistent with an anomalous inward rectification,
and a rebound spike immediately following the termination of the
hyperpolarizing pulse. This response, consistent with the existence of a
hyperpolarization-activated inward current
(Solomon et al., 1993)
(Ih), was never observed in E17-E18 sling cells. Finally, 5 of 11
P0-P2 sling cells produced spontaneous action potentials at rest, whose
kinetics and suppression by TTX (5 of 5 cells), identified them as
Na+-mediated spikes (Fig.
2B).
Following the recordings cells were filled with biocytin and then processed for NeuN labeling. At E17, biocytin-labeled sling cells possessed a long leading process oriented toward the midline (Fig. 3A) and at P0 had multiple processes resembling dendrites (Fig. 3C,D). These findings, and the fact that all biocytin-filled sling cells were immunoreactive for the neuronal marker NeuN (Fig. 3D), indicate that our recordings were obtained from neurons in the sling.
As a control we compared the electrophysiological properties of the
identified sling cells with those of the glial wedge, a known population of
GFAP-positive glia present at the ventrolateral edges of the sling
(Shu and Richards, 2001). In
response to intracellular injections of depolarizing currents, all glial wedge
cells (E17, n=4) produced non-rectifying responses, and none produced
Na+ or Ca2+ spikes
(Fig. 2C). Three
biocytin-filled glial cells were recovered and processed for GFAP
immunohistochemistry. All labeled with GFAP
(Fig. 3B) and had a typical
radial glial morphology as previously described
(Shu and Richards, 2001
).
Therefore sling cells possessed the electrophysiological characteristics of
neurons and not glia, and together with the immunohistochemical data described
above provide compelling evidence that the majority of cells within the sling
are neurons.
Birthdating of the sling
It was originally reported that sling cells were generated between E15 and
E17 of mouse development (Silver et al.,
1982). We have performed a birth-dating analysis of the sling by
injecting BrdU between E12 and P2 and sacrificing the animals at E17, P0 or
P3. We found no BrdU-labeled cells within the sling when BrdU was injected at
E12-E14, but labeled sling cells were present when BrdU was injected between
E15-P2 (the oldest age examined; Fig.
4). This confirms previous results showing that sling cells are
born on E15 (Silver et al.,
1982
), but extends these results to show that sling cells continue
to be generated until at least P2.
|
|
Postnatal development of the sling
Despite the continued proliferation of the sling cells, the structure
itself does not continue to increase in size (other than to increase in
proportion to the growth of the brain), and in fact it decreases slightly in
size by P5 and disappears altogether by P10
(Fig. 6). Therefore the sling
is a transient structure. In Nissl-stained, and NeuN-labeled sections, the
sling forms a distinct structure at P0 (arrow in
Fig. 6A,B) and P5
(Fig. 6C,D) but disappears by
P10 (Fig. 6E,F).
|
|
![]() |
DISCUSSION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
The electrophysiological recordings we obtained from the sling cells
changed during development. Early in development (E17-E18) sling neurons show
long-duration Na+ and Ca2+ spikes, as do developing
neurons elsewhere in the central nervous system
(McCormick and Prince, 1987).
Postnatally the Ca2+ spikes become smaller and less frequent, and
the kinetics of Na+ spikes resemble those of more mature neurons
(Gutnick and Modi, 1995
).
However, sling neurons did not label with markers of mature neurons such as
neurofilament, GAP43 or MAP2 either pre- or postnatally, indicating they may
represent an immature population of neurons. It is therefore unclear whether
the same cells remain within the sling and become progressively more mature or
whether new cells continue to migrate through the sling. The results of our
BrdU time-lapse experiment suggest that the majority of neurons continue to
migrate through the sling even at P3. It is therefore possible that sling
cells migrating postnatally may have different cellular characteristics than
those migrating at prenatal stages. In addition, given these results, more
sling cells may be generated than previously thought. Further investigation is
required to determine both the site of origin and the final destination of
neurons migrating through the sling throughout development.
Originally the sling was described as only those cells that migrate from
the medial aspect of the SVZ to the midline
(Silver et al., 1982;
Silver and Ogawa, 1983
;
Katz et al., 1983
;
Smith et al., 1986
;
Hankin and Sliver, 1988
;
Schneider and Silver, 1990
).
We have used this definition of the sling. In a later study Silver and
colleagues (Silver et al.,
1993
) described a radial glial-like population of cells in cat
that sit at the lateral edges of the sling. They postulated these may be the
lateral extent of the sling and expanded the definition of the sling to
include these cells. These radial glial-like cells are probably the equivalent
of the glial wedge described in rodents
(Shu and Richards, 2001
).
However, here we show that the glial wedge and the sling cells derived from
the SVZ are clearly two different populations. Therefore here we have referred
to the `sling' as being only those cells originally described as the sling
that are derived from the SVZ (Silver et
al., 1982
; Hankin and Silver, 1983;
Silver and Ogawa, 1983
;
Katz et al., 1983
;
Smith et al., 1986
;
Schneider and Silver,
1990
).
The prior classification of the sling as glioblast cells, the timing of
sling formation and their location below the corpus callosum made it plausible
that a function of the sling was to guide callosal axons across the midline.
Furthermore, in vivo manipulations of the sling and rescue of the sling in
surgically acallosal mice provided compelling evidence for their role in
callosal axon guidance (Silver et al.,
1982; Silver and Ogawa,
1983
; Katz et al.,
1983
). Key to this hypothesis was the fact that the sling was
formed prior to the time callosal axons cross the midline [which was
previously thought to be at E18 (Floeter
and Jones, 1985
)]. However, recent experiments indicate that the
pioneering axons of the corpus callosum (derived from the cingulate cortex)
begin to cross the midline as early as E15.5
(Rash and Richards, 2001
),
before the sling structure has formed at E17
(Silver et al., 1982
).
In previous experiments in which the sling was severed, the corpus callosum
could be rescued by implanting a piece of cellulose membrane at the midline
over which GFAP-positive glial cells grew, followed by callosal axons
(Silver and Ogawa, 1983;
Smith et al., 1986
). We show
that the sling is largely composed of neurons indicating that the cells that
rescued the corpus callosum in these experiments may not have been sling cells
(at least not the SVZ-derived sling cells, which are not GFAP positive) but
glial cells from other midline populations (possibly from the lateral edges of
the sling). Our results do not rule out the possibility that the sling is
involved in callosal axon guidance in some capacity, particularly at E17 when
the sling is formed and the majority of callosal axons begin to cross the
midline, or that it may be involved in the maintenance of the corpus callosum.
It is still conceivable that a migratory population of neurons may be used by
callosal axons to cross the midline after the pioneering axons have crossed.
But our findings do indicate that the initial cortical axons that cross the
midline (Rash and Richards,
2001
) do not require the sling. Our data also suggest that the
sling should not be grouped with other midline glial populations involved in
guiding other midline commissures such as the optic chiasm and the anterior
commissure (Marcus et al.,
1995
; Cummings et al.,
1997
; Pires-Neto et al.,
1998
) since the sling cells derived from the SVZ are not glia.
The postnatal disappearance of the sling has been attributed to a selective
loss of the entire sling structure by an unknown mechanism of cell death
(Hankin et al., 1988).
However, our TUNEL labeling of the sling shows that very few of the cells die
at E17 and E18 and that none die at postnatal ages. Therefore, unless cell
death and clearance of the cells is occurring so rapidly that it cannot be
detected with this technique, cell death alone cannot explain either the
disappearance of the sling or the postnatal decrease in the size of the sling.
Clearance of the sling has been proposed to occur by macrophage-mediated
endocytosis, leaving the cavum septum pellucidum behind
(Hankin et al., 1988
). We have
observed macrophage-like cells at the cortical midline (T.S. and L.J.R.,
unpublished observation) but have not determined which, if any, cells are
removed by them. An alternative hypothesis for the disappearance of the sling
is that the cells continue to migrate from the SVZ of the dorsal telencephalon
to adjacent brain regions. Long distance migration of cells within the SVZ has
been shown to occur from cells that originate from the medial ganglionic
eminence (Anderson et al.,
2001
; Marin et al.,
2001
). In the rostral migratory stream, cells leave the SVZ and
migrate to the olfactory bulbs where they give rise to granule neurons
(Luskin, 1993
;
Lois and Alvaerz-Buylla,
1994
). It is possible that sling cells do not cease to migrate
when they reach the midline but continue to migrate and populate adjacent
regions of the brain.
We therefore redefine the sling as a migratory population of neurons whose
function may include the guidance of callosal axons across the midline. These
new findings open the possibility for additional hypotheses about the
developmental function of this distinct population of cells. Sling neurons
label with many of the same markers as subplate neurons of the cortical plate,
although sling neurons are born after those of the subplate. The subplate is
involved in axonal guidance, and regionalization and patterning of the
neocortex (McConnell et al.,
1989; Ghosh et al.,
1990
; De Carlos and O'Leary,
1992
; Ghosh and Shatz,
1992
; Ghosh and Shatz,
1993
; Ghosh and Shatz,
1994
; McConnell et al.,
1994
; Molnar et al.,
1998
). The developmental significance of the sling is yet to be
determined, but the possibility of novel functions for this population in
forebrain development needs to be examined in order to determine their true
potential.
![]() |
ACKNOWLEDGMENTS |
---|
![]() |
REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Anderson, S. A., Martin, O., Horn, C., Jennings, K. and
Rubenstein, J. L. R. (2001). Distinct cortical
migrations from the medial and lateral ganglionic eminences.
Development 128,353
-363.
Cummings, D. M., Malun, D. and Brunjes, P. C. (1997). Development of the anterior commissure in the opossum: midline extracellular space and glia coincide with early axon decussation. J. Neurobiol. 32,403 -414.[CrossRef][Medline]
Danbolt, N. C. (2001). Glutamate uptake. Prog. Neurobiol. 65,1 -105.[CrossRef][Medline]
De Carols, J. A. and O'Leary, D. D. (1992). Growth and targeting of subplate axons and establishment of major cortical pathways. J. Neurosci. 12,1194 -1211.[Abstract]
Feng, L., Hatten, M. E. and Heintz, N. (1994). Brain lipid-binding protein (BLBP): a novel signaling system in the developing mammalian CNS. Neuron 12,895 -908.[Medline]
Ferreira, A., Busciglio, J. and Caceres, A. (1987). An immunocytochemcical analysis of the ontogeny of the microtubule-associated proteins MAP-2 and Tau in the nervous system of the rat. Brain Res. 431,9 -31.[Medline]
Figlewicz, D. A., Gremo, F. and Innocenti, G. M. (1988). Differential expression of neurofilament subunits in the developing corpus callosum. Brain Res. 470,181 -189.[Medline]
Floeter, M. K. and Jones, E. G. (1985). The morphology and phased outgrowth of callosal axons in the fetal rat. Brain Res. 354,7 -18.[Medline]
Fonseca, M., del Rio, J. A., Martinez, A., Gomez, S. and Soriano, E. (1995). Development of calretinin immunoreactivity in the neocortex of the rat. J. Comp. Neurol. 361,177 -192.[Medline]
Ghosh, A., Antonini, A., McConnell, S. K. and Shatz, C. J. (1990). Requirement for subplate neurons in the formation of thalamocortical connections. Nature 347,179 -181.[CrossRef][Medline]
Ghosh, A. and Shatz, C. J. (1992). Pathfinding and target selection by developing geniculocortical axons. J. Neurosci. 12,39 -55.[Abstract]
Ghosh, A. and Shatz, C. J. (1993). A role for
subplate neurons in the patterning of connections from thalamus to cortex.
Development 117,1031
-1047.
Ghosh, A. and Shatz, C. J. (1994). Segregation of geniculocortical afferents during the critical period: a role for subplate neurons. J. Neurosci. 14,3862 -3880.[Abstract]
Goslin, K., Schreyer, D. J., Skene, J. H. and Banker, G. (1988). Development of neuronal polarity: GAP-43 distinguishes axonal from dendritic growth cones. Nature 336,672 -674.[CrossRef][Medline]
Gottlieb, J. P. and Keller, A. (1997). Intrinsic circuitry and physiological properties of pyramidal neurons in rat barrel cortex. Exp. Brain Res. 115, 47-60.[Medline]
Gutnick, M. J. and Crill, W. E. (1995). The cortical neuron as an electrophysiological unit. In The Cortical Neuron (ed. M. J. Gutnick and I. Modi), pp33 -51. New York: Oxford University Press.
Gutnick, M. J. and Modi, I. (1995). The Cortical Neuron. New York: Oxford University Press.
Hankin, M. H. and Silver, J. (1988). Development of intersecting CNS fiber tracts: the corpus callosum and its perforating fiber pathway. J. Comp. Neurol. 272,177 -190.[Medline]
Hankin, M. H., Schneider, B. F. and Silver, J. (1988). Death of the subcallosal glial sling is correlated with formation of the cavum septi pellucidi. J. Comp. Neurol. 272,191 -202.[Medline]
Katz, M. J., Lasek, R. J. and Silver, J. (1983). Ontophyletics of the nervous system: development of the corpus callosum and evolution of axon tracts. Proc. Natl. Acad. Sci. USA 80,5936 -5940.[Abstract]
Kurtz, A., Zimmer, A., Schnutgen, F., Bruning, G., Spener, F.
and Mullen, T. (1994). The expression pattern of a
novel gene encoding brain-fatty acid binding protein correlates with neuronal
and glial cell development. Development
120,2637
-2649.
Lois, C. and Alvarez-Buylla, A. (1994). Long-distance neuronal migration in the adult mammalian brain. Science 264,1145 -1148.[Medline]
Luskin, M. B. (1993). Restricted proliferation and migration of postnatally generated neurons derived from the forebrain subventricular zone. Neuron 11,173 -189.[Medline]
Marcus, R. C., Blazeski, R., Godement, P. and Mason, C. A. (1995). Retinal axon divergences in the optic chiasm: uncrossed axons diverge from crossed axons within a midline glial specialization. J. Neurosci. 15,3716 -3729.[Abstract]
Marin, O., Yaron, A., Bagri, A., Tessier-Lavigne, M. and
Rubenstein, J. L. (2001). Sorting of striatal and
cortical interneurons regulated by semaphorin-neruopilin interactions.
Science 293,872
-875.
McConnell, S. K., Ghosh, A. and Shatz, C. J. (1989). Subplate neurons pioneer the first axon pathway from the cerebral cortex. Science 245,978 -982.[Medline]
McConnell, S. K., Ghosh, A. and Shatz, C. J. (1994). Subplate pioneers and the formation of descending connections from cerebral cortex. J. Neurosci. 14,1892 -1907.[Abstract]
McCormick, D. A. and Prince, D. A. (1987). Post-natal development of electrophysiological properties of rat cerebral cortical pyramidal neurones. J. Physiol. 393,743 -762.[Abstract]
Meiri, K. F., Pfenninger, K. H. and Willard, M. B. (1986). Growth-associated protein, GAP-43, a polypeptide that is induced when neurons extend axons, is a component of growth cones and corresponds to pp46, a major polypeptide of a subcellular fraction enriched in growth cones. Proc. Natl. Acad. Sci. USA 83,3537 -3541.[Abstract]
Moody, S. A., Quigg, M. S. and Frankfurter, A. (1989). Development of the peripheral trigeminal system in the chick revealed by an isotope-specific anit-beta-tubulin monoclonal antibody. J. Comp. Neurol. 279,567 -580.[Medline]
Molnar, Z., Adams, R. and Blakemore, C. (1998).
Mechanisms underlying the early establishment of thalamocortical connections
in the rat. J. Neurosci.
18,5723
-5745.
Mullen, R. J., Buck, C. R. and Smith, A. M.
(1992). NeuN, a neuronal specific nuclear protein in vertebrates.
Development 116,201
-211.
Niinobe, M., Maeda, N., Ino, H. and Mikoshiba, K. (1988). Characterization of microtubule-associated protein 2 from mouse brain and its localization in the cerebellar cortex. J. Neurochem. 51,1132 -1139.[Medline]
Nowakowski, R. A., Lewin, S. B. and Miller, M. W. (1989). Bromodeoxyuridine immunohistochemical determination of the lengths of the cell cycle and the DNA-synthetic phase for an anatomically defined population. J. Neurocytol. 18,311 -318.[Medline]
Okano, H. J. and Darnell, R. B. (1997). A
hierarchy of Hu RNA binding proteins in developing and adult neurons.
J. Neurosci. 17,3024
-3037.
O'Rourke, N. A., Chenn, A. and McConnell, S. K.
(1997). Postmitotic neurons migrate tangentially in the cortical
ventricular zone. Development,
124,997
-1005.
Packard, D. S., Menzies, R. A. and Skalko, R. G. (1973). Incorporation of thymidine and its analog, bromodeoxyuridine, into embryos and maternal tissues of the mouse. Differentiation 1,397 -405.[Medline]
Pires-Neto, M., Braga-De-Souza, S. and Lent, R. (1998). Molecular tunnels and boundaries for growing axons in the anterior commissure of hamster embryos. J. Comp. Neurol. 399,176 -188.[CrossRef][Medline]
Preston, S. L., Shu, T., Corte, G., Rubenstein, J. L. R. and Richards, L. J. (2000). Emx-1 is required for the development of the glial sling and formation of the corpus callosum. Soc. Neurosci. Abst. 218.13,577 .
Rash, B.G. and Richards, L. J. (2001). A role for cingulate pioneering axons in the development of the corpus callosum. J. Comp. Neurol. 434,147 -157.[CrossRef][Medline]
Schneider, B. F. and Silver, J. (1990). Failure of the subcallosal sling to develop after embryonic X-irradiation is correlated with absence of the cavum septi. J. Comp. Neurol. 299,462 -469.[Medline]
Shu, T. and Richards, L. J. (2001). Cortical
axon guidance by the glial during development of the corpus callousm.
J. Neurosci. 21,2749
-2758.
Shu, T., Valentino, K. M., Seaman, C., Cooper, H. M. and Richards, L. J. (2000). Expression of the netrin receptor, deleted in colorectal cancer (DCC), is largely confined to projecting neurons in the developing forebrain. J. Comp. Neurol. 416,201 -212.[CrossRef][Medline]
Shu, T., Butz, K. G., Plachez, C., Gronostajski, R. M. and
Richards, L. J. (2003). Abnormal development of forebrain
midline glia and commissural projections in Nfia knock-out mice. J.
Neurosci. 23,203
-212.
Silver, J., Edwards, M. A. and Levitt, P. (1993). Immunocytochemical demonstration of early appearing astroglial structures that form boundaries and pathways along axon tracts in the fetal brain. J. Comp. Neurol. 328,415 -436.[Medline]
Silver, J., Lorenz, S. E., Wahlsten, D. and Coughlin, J. (1982). Axonal guidance during development of the great cerebral commissures: descriptive and experimental studies, in vivo, on the role of preformed glial pathways. J. Comp. Neurol. 210, 10-29.[Medline]
Silver, J. and Ogawa, M. Y. (1983). Postnatally induced formation of the corpus callosum in acallosal mice on glia-coated cellulose bridges. Science 220,1067 -1069.[Medline]
Smith, G. M., Miller, R. H. and Silver, J. (1986). Changing role of forebrain astrocytes during development, regenerative failure, and induced regeneration upon transplantation. J. Comp. Neurol. 251,23 -43.[Medline]
Solomon, J. S., Doyle, J. F., Burkhalter, A. and Nerbonne, J. M. (1993). Differential expression of hyperpolarization-activated currents reveals distinct classes of visual cortical projection neurons. J. Neurosci. 13,5082 -5091.[Abstract]
Stuart, G. J., Dodt, H. U. and Sakmann, B. (1993). Patch-clamp recordings from the soma and dendrites of neurons in brain slices using infrared video microscopy. Eur. J. Physiol. 423,511 -518.[Medline]
Wichterle, H., Garcia-Verdugo, J. M. and Alvarez-Buylla, A. (1997). Direct evidence for homotypic, glia-independent neuronal migration. Neuron 19,779 -791.