Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 1996 Sep 15;16(18):5762-76.
doi: 10.1523/JNEUROSCI.16-18-05762.1996.

Interkinetic and migratory behavior of a cohort of neocortical neurons arising in the early embryonic murine cerebral wall

T Takahashi  1 , R S Nowakowski, V S Caviness Jr
Collaborators, Affiliations

Interkinetic and migratory behavior of a cohort of neocortical neurons arising in the early embryonic murine cerebral wall

T Takahashi et al. J Neurosci. .

Abstract

Neocortical neuronogenesis occurs in the pseudostratified ventricular epithelium (PVE) where nuclei of proliferative cells undergo interkinetic nuclear movement. A fraction of daughter cells exits the cell cycle as neurons (the quiescent, or Q, fraction), whereas a complementary fraction remains in the cell cycle (the proliferative, or P, fraction). By means of sequential thymidine and bromodeoxyuridine injections in mouse on embryonic day 14, we have monitored the proliferative and post-mitotic migratory behaviors of 1 and 2 hr cohorts of PVE cells defined by the injection protocols. Soon after mitosis, the Q fraction partitions into a rapidly exiting (up to 50 microns/hr) subpopulation (Qr) and a more slowly exiting (6 microns/hr) subpopulation (Qs). Qr and Qs are separated as two distributions on exit from the ventricular zone with an interpeak distance of approximately 40 microns. Cells in Qr and Qs migrate through the intermediate zone with no significant change in the interpeak distance, suggesting that they migrate at approximately the same velocities. The rate of migration increases with ascent through the intermediate zone (average 2-6.4 microns/hr) slowing only transiently on entry into the developing cortex. Within the cortex, Qr and Qs merge to form a single distribution most concentrated over layer V.

PubMed Disclaimer

Figures

Fig. 1.
Fig. 1.
Neocortical histogenesis: the sequence of cellular events. Histogenesis is initiated with cell proliferation in the pseudostratified ventricular epithelium (PVE), which is approximately coextensive with the ventricular zone (VZ) lying at the ventricular margin. P fraction cells of the PVE (PPVE) undergo interkinetic nuclear migration as they progress through the G1-to-S-to-G2-to-M phases of the cell cycle. Q fraction cells of the PVE (QPVE) exit the VZ and migrate through the subventricular zone (SVZ) and intermediate zone (IZ) to the developing cortex (CTX). The earliest formed neurons take up positions in the deepest cortical layers, whereas cells formed progressively later are distributed to progressively more superficial cortical layers (Redistribution in CTX). The secondary proliferative population (SPP), thought to be the progenitor population for glial cells, arises from the PVE. Its cells become distributed through a broad zone spanning the outer VZ, SVZ, and IZ. Cells of the SPP do not undergo interkinetic nuclear migration.
Fig. 2.
Fig. 2.
Protocols for defining 1 and 2 hr cell cohorts. Proliferating cells of the PVE are first exposed to [3H]TdR (arrowhead with dots) either at 7:00 A.M. (2 hr cohort) or at 8:00 A.M. (1 hr cohort) on E14. All (and only) proliferating cells that are in S phase incorporate the marker (white nuclei with dots). At 9:00 A.M., which is either 1 hr (1 hr cohort) or 2 hr (2 hr cohort) after exposure to [3H]TdR, the PVE is exposed to BUdR (solid arrowhead). This second marker will be incorporated by cells that already contain [3H]TdR and are still in S phase (gray-filled nuclei with dots) and also by cells that have entered S phase subsequent to the [3H]TdR exposure (gray-filled nuclei without dots). Thus, the cells that leave S phase between the [3H]TdR and BUdR exposures (i.e., between 8:00 A.M. and 9:00 A.M. for the 1 hr cohort or between 7:00 A.M. and 9:00 A.M. for the 2 hr cohort) will be marked only by [3H]TdR (asterisks), and it is these [3H]TdR-only labeled cells that are referred to as the 1 and 2 hr cohorts of cells. For the 1 hr cohort where the experimental interval is ≤8.0 hr, there is only the single exposure to BUdR. For the 1 hr cohort where the experimental interval is 12.5 hr and for all time points for the 2 hr cohort, the exposure to BUDR is repeated every 3 hr (open arrowheads) through an interval corresponding to the length of the cell cycle minus the length of S phase (TC − TS) to double label the P fraction cells of the cohort after they re-enter S phase. The leading edge of the 1 and 2 hr cohorts corresponds to the cells that leave S phase immediately after exposure to [3H]TdR, whereas the trailing edge corresponds to the last cells to leave S phase before BUdR exposure. The 1 and 2 hr cohorts of cells advance through G2 and M phases as a coherent wave. After mitosis, the daughter cells partition into Q (exiting cell cycle) and P (re-entering S phase) subsets of the original cohort. The progression of the Q fraction is followed as it exits the VZ (Exit from VZ) and migrates across the IZ (Migration through IZ) at 12.5–60 hr to reach the cortex (CTX). The Q fraction is followed further at P4 and P22 within the cortex, where it will be positioned above cells that migrated before and below cells that migrated afterward (Redistribution in CTX).
Fig. 3.
Fig. 3.
Strata of the murine dorsomedial cerebral wall during the experimental interval (modified from Takahashi et al., 1995a). One and two hour cohorts of cells were established by sequential [3H]TdR- and BUdR-labeling protocols (Fig. 2) on E14, 3 d after the onset of the neuronogenetic interval that extends in mouse from early E11 through early E17 (gray bar at the base of the graph) (Takahashi et al., 1995a). The movements of the cohorts were tracked from E14 through E16 in the course of their migrations (shaded area) and also postnatally at P4 and P22 (day of birth, P0). The heights of the cerebral strata were determined by direct measurements in histological sections (Takahashi et al., 1995a). From E11 through early E14, the cerebral wall is principally the VZ with narrow overlying primitive plexiform zone (PPZ). On E14, the SVZ, IZ, and cortical strata [subplate (SP), cortical plate (CP), and molecular layer (ML)] replace the PPZ. The ventricular surface corresponds to 0 on they-axis. The pial surface is the outer limit of the ML. The contours tracing progressive growth of strata were made initially by a least-squares fit to a fourth order curve and then smoothed by eye. Three photomicrographic inserts depict the principal histological features of the dramatic transitions that occur in the cerebral strata over the neuronogenetic interval. Each is taken from a 4-μm-thick coronal section stained immunocytochemically for BUdR and counterstained with basic fuchsin. The embryos had been exposed to BUdR only 30 min before killing so that the distribution of black, BUdR-positive nuclei corresponds to the zone of S phase at each illustrated age. Insert E13 is representative of the cerebral wall when the strata include only VZ and PPZ. Astar marks the pia. Insert E15 represents the cerebral wall relatively late in the course of neuronogenesis when the full stratification plan is established, and insert E18 represents the cerebral wall after the neuronogenetic interval is completed. The S-phase zone at E13 and E15 corresponds principally to the outer half of the VZ, although with some S-phase activity in cells of the SPP in the SVZ and IZ at E15. The increase in the width of the cerebral wall between E15 and E18 largely represents increase in width of the cortical strata. By E18, the VZ has become reduced to a simple cuboidal epithelium, and S-phase activity is limited to cuboidal ependymal cells and the SPP. Scale bar, 50 μm.
Fig. 4.
Fig. 4.
Distribution of 1 hr cohort of [3H]TdR-only labeled cells in relation to the distribution of cells in S phase at E14. The photomicrograph (left) illustrates an example of cells of the 1 hr cohort ([3H]TdR-only labeled cells) located both in the VZ (small arrows) and above the VZ (arrowhead). Also illustrated are S-phase cells labeled with BUdR (darkly stained nuclei) in the S-phase zone.Right, The number of cells of the cohort per 10 μm bin (see Materials and Methods) is plotted as histograms. Those within the VZ concentrated in bins 1–2 (filled bars, inner subset) belong to the PVE. A smaller number of cells of the cohort above the VZ (unfilled bars, outer subset) belongs to the SPP. Superimposed on the histogram is abroken line reflecting the BUdR-labeling index, as obtained from the sections that were labeled with BUdR 30 min before killing. BUdR-labeled cells in S phase are most concentrated in the outer half of the VZ (S-phase zone) (see also Takahashi et al., 1993).
Fig. 5.
Fig. 5.
The 1 hr cohort at the 6.5 hr time point as illustrated in a photomicrograph. The boundary between VZ and SVZ is indicated by a dashed line. [3H]TdR-only labeled cells, representing both rapidly exiting cells of the PVE Q fraction (Qr) and G1 phase cells of the SPP, are found in the SVZ and IZ (arrowheads). Cells of the 1 hr cohort located within the VZ (arrows) must be either cells of the PVE in G1 phase (i.e., the P fraction of the cohort) or slowly ascending cells of the Q fraction of the cohort (Qs) that have not yet exited the VZ. Scale bar, 20 μm.
Fig. 6.
Fig. 6.
The 1 hr cohort distribution in the 3.5–8.0 hr interval mapped as histograms. [3H]TdR-only labeled cells are scored with respect to 10 μm bin position and plotted as histograms at the 3.5, 5.0, 6.5, and 8.0 hr time points after exposure to BUdR. Through the first 8.0 hr, the two subsets of cells (the inner and outer subsets) remain well separated and distinct from each other. Because the distributions of the inner and outer subsets of cells are well approximated by two normally distributed populations, the mean positions and relative sizes of the two distributions can be calculated from a least-squares fit to the frequency histograms (dashed lines) (Table 1). The sum of the two distributions is shown with solid lines.
Fig. 7.
Fig. 7.
Changing positions of the two subpopulations of the 1 hr cohort over the 3.5–8.0 hr interval. This interval corresponds approximately to the G1 phase of the P cells of the cohort. The locations of the inner subset within the VZ and the outer subset above the VZ are plotted with respect to average bin position. Approximately 15% of the cells of the cohort move from the inner subset to the outer subset as early as 5 hr after BUdR injection, which corresponds to Qr of the PVE. This movement is indicated by an arrow with dashed line(for details, see Results).
Fig. 8.
Fig. 8.
Comparative distributions of the 1 and 2 hr cohorts at the 12.5 hr time point. The distributions of cells of the 1 and 2 hr cohorts are plotted with respect to bin position. At the 12.5 hr time point, which is at a time > TC − TS, the two cohorts have been reduced to their Q fractions only and already have exited the VZ) The Q fractions of both cohorts are represented by two distributions corresponding to the slowly exiting (Qs) subpopulation, located at the VZ–SVZ border, and the rapidly exiting (Qr) subpopulation, located more superficially in the IZ. The number of cells in the 2 hr cohort is approximately twice the number in the 1 hr cohort (Table 2). Reflecting the 1 hr longer interval between [3H]TdR and BUdR injections, theQs and Qrdistributions of the 2 hr cohort have shifted slightly outward with respect to those of the 1 hr cohort.
Fig. 9.
Fig. 9.
The 2 hr cohort distributions in the 12.5–60 hr interval mapped as histograms. [3H]TdR-only labeled cells are scored with respect to 10 μm bin position and plotted as histograms at the 12.5, 24, 36, 48, and 60 hr time points after exposure to BUdR. The Qr (rapidly exiting) and Qs (slowly exiting) subpopulations of the cohort are well separated through the 36 hr time point, that is, until the Qr subpopulation begins to enter the cortex. At the 48 and 60 hr time points, after bothQr and Qs have entered the cortex, the two populations are completely overlapping. If for the 12.5–36 hr time points, we assume that theQr and Qs subsets of cells approximate two normally distributed populations, the mean positions and relative sizes of the two distributions can be calculated from a least-squares fit to the frequency histograms (dashed lines) (Table 3). The trace shown with asolid line at each time point is a best fit to the total population of the 2 hr cohort.
Fig. 10.
Fig. 10.
The 2 hr cohort at 24 hr (A) and 60 hr (B) and at P22 (C) as illustrated in photomicrographs. [3H]TdR-only labeled cells representing both rapidly (Qr) (arrows in A) and slowly (Qs) (arrowheads inA) exiting cells of the PVE Q fraction are distinctly separated from each other in the IZ at 24 hr, but are not distinguishable as two separate distributions within the cortex (CTX) at 60 hr and P22 (arrowheads inB, C). The ML and layers II–V are indicated for P22 (C). (An atypically large number of [3H]TdR-only labeled cells is evident in the section selected for illustration at P22.) Scale bars: A(equivalent also for B), 30 μm; C, 50 μm. A star marks the pia.
Fig. 11.
Fig. 11.
Comparative patterns of migration ofQr and Qs of the 2 hr cohort. The mean positions of cells of the rapidly exiting (Qr, closed circles) and slowly exiting (Qr, open circles) subpopulations of the 2 hr cohort are plotted after they have left the VZ with respect to micrometers from the ventricular surface at the 12.5–60 hr time points. Also superimposed is the stratification pattern of the cerebral wall (taken from Fig. 3). Migrations are initiated deep in the SVZ–IZ border at 12.5 hr, at which timeQr and Qs are well separated from each other. There is no significant change in the distance between the two subpopulations in the course of migrations through the 36 hr time point when Qr enters the cortex (CTX). At 48 and 60 hr, after bothQr and Qs have entered the cortex, the two subpopulations are indistinguishable. The time elapsed after exit of the cohort from S phase is indicated in hours and in embryonic dates on the abscissa.
Fig. 12.
Fig. 12.
Comparative distributions of the 2 hr cohort within the cortex at P4 and P22. [3H]TdR-only labeled cells are scored with respect to 10 μm bin position (referenced to the pial surface) and plotted as histograms at P4 and P22. Thetrace shown with a solid line at the two ages is a best fit to the total population of the 2 hr cohort. At P4, the cohort distribution spans the full width of the cortex below the ML with the peak of the distribution located over layer V. At P22, the distribution extends only through layers VI and V, with cells most concentrated in layer V. The size of the cohort has become reduced by 45% in this 3 week interval (see Results). WM indicates cerebral white matter.
Fig. 13.
Fig. 13.
Histogenetic life cycle of a strictly defined cohort of cells. A cohort of neurons arising over an interval of 1–2 hr on E14 in the dorsomedial murine neocortical PVE separates after mitosis into a Q fraction that has left the cell cycle and a P fraction that will continue to cycle (Neuronogenesis). The Q fraction, in turn, separates into rapidly exiting (Qr, light spheres) and slowly exiting (Qs, dark spheres) subpopulations. Once above the VZ in the SVZ and IZ, the two subpopulations migrate (Migration) at indistinguishable rates until the leading, or Qr, subpopulation of the cohort is slowed on entry into the cortex (CTX). Once in the cortex and in the course of ascent to the outer margin of the CP, the two subpopulations totally overlap and become indistinguishable. Later, they come to lie deep to neurons arising at later dates and are moderately reduced in numbers by histogenetic cell death (Redistribution & Cell Death). After neuronal migrations are completed, the IZ and SVZ are replaced by the cerebral central white matter (WM) and subependyma (SE), and the PVE within the VZ is replaced by a cuboidal ependymal ventricular lining (Ep). Provided at the base of the diagram is a temporal profile of these events with the nuclear velocities for cells of the proliferative cycle in S, G2, M, and G1 phases, and VZ exit and migration velocities ofQr and Qsregistered on the ordinate. The diagram is schematic both with respect to scaling of time and distances.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Bayer SA, Altman J. Raven; New York: 1991. Neocortical development. .
    1. Berry M, Rogers AW. The migration of neuroblasts in the developing cerebral cortex. J Anat. 1965;99:691–701. - PMC - PubMed
    1. Bisconte J-C, Marty R. Etude quantitative du marquage radioautographique dans le systeme nerveux du rat. II. Caracteristiques finales dans le cerveau de l’animal adulte. Exp Brain Res. 1975;22:37–56. - PubMed
    1. Blaschke A, Staley K, Chun J. Widespread programmed cell death in proliferative and postmitotic regions of the fetal cerebral cortex. Development. 1996;122:1165–1174. - PubMed
    1. Cai L, Hayes NL, Nowakowski RS. Comparison of the cumulative labeling S-phase method and the percent labeled mitoses method in the developing cerebral cortex. Soc Neurosci Abstr. 1993;19:30.

Publication types

LinkOut - more resources