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
. 2007 Jul 11;27(28):7541-52.
doi: 10.1523/JNEUROSCI.0431-07.2007.

Pilocarpine-induced seizures cause selective time-dependent changes to adult-generated hippocampal dentate granule cells

Cynthia Walter  1 , Brian L Murphy, Raymund Y K Pun, Anne L Spieles-Engemann, Steve C Danzer
Affiliations

Pilocarpine-induced seizures cause selective time-dependent changes to adult-generated hippocampal dentate granule cells

Cynthia Walter et al. J Neurosci. .

Abstract

Aberrantly interconnected granule cells are characteristic of temporal lobe epilepsy. By reducing network stability, these abnormal neurons may contribute directly to disease development. Only subsets of granule cells, however, exhibit abnormalities. Why this is the case is not known. Ongoing neurogenesis in the adult hippocampus may provide an explanation. Newly generated granule cells may be uniquely vulnerable to environmental disruptions relative to their mature neighbors. Here, we determine whether there is a critical period after neuronal birth during which neuronal integration can be disrupted by an epileptogenic insult. By bromodeoxyuridine birthdating cells in green fluorescent protein-expressing transgenic mice, we were able to noninvasively label granule cells born 8 weeks before (mature), 1 week before (immature), or 3 weeks after (newborn) pilocarpine-epileptogenesis. Neuronal morphology was examined 4 and 8 weeks after pilocarpine treatment. Strikingly, almost 50% of immature granule cells exposed to pilocarpine-epileptogenesis exhibited aberrant hilar basal dendrites. In contrast, only 9% of mature granule cells exposed to the identical insult possessed basal dendrites. Moreover, newborn cells were even more severely impacted than immature cells, with 40% exhibiting basal dendrites and an additional 20% exhibiting migration defects. In comparison, <5% of neurons from normal animals exhibited either abnormality, regardless of age. Together, these data demonstrate the existence of a critical period after the birth of adult-generated neurons during which they are vulnerable to being recruited into epileptogenic neuronal circuits. Pathological brain states therefore may pose a significant hurdle for the appropriate integration of newly born endogenous, and exogenous, neurons.

PubMed Disclaimer

Figures

Figure 1.
Figure 1.
GFP-expressing neurons in the Thy1-GFP mouse line do not colocalize with the immature neuronal marker doublecortin in control animals or animals examined 4, 8, or 11 weeks after SE. Images are confocal optical sections through the dentate gyrus showing GFP (green) and doublecortin (red) immunolabeling. No double-labeled cells were found, indicating that GFP is not expressed until neurons are ∼4 weeks old. Scale bar, 50 μm.
Figure 2.
Figure 2.
Schematic detailing the experimental design. The different colors reflect different groups of animals (groups 1–4), whereas hatched versus filled bars within a group reflect different populations of neurons within the same animals. Four groups of animals and eight different populations of neurons are depicted. A, For each group of animals, hatched bars depict the time period over which GFP-expressing cells were generated (0–12 weeks), whereas filled bars depict the period during which the subpopulation of BrdU-labeled, GFP-expressing neurons were generated (either week 4 or 11). Animals received saline or pilocarpine treatment to induce SE when they were 12 weeks old. All animals were killed when they were 16 weeks old. B, Four- to 16-week-old GFP-labeled granule cells exposed to SE when they were 0–12 weeks old (hatched blue and red bars) exhibit basal dendrites significantly more frequently than age-matched neurons from animals that did not experience SE (hatched orange and green bars). Moreover, the two epileptic groups were statistically identical, indicating that pilocarpine treatments were identical between groups receiving injections of BrdU at 4 weeks (blue bar) or 11 weeks (red bar), as expected. C, BrdU-labeled, GFP-expressing dentate granule cells exposed to SE at 1 week of age (red bar) were significantly more likely to possess hilar basal dendrites 4 weeks later relative to age-matched control cells (green bar), cells exposed to SE when they were 8 weeks old (blue bar), and 12-week-old control cells (orange bar). Granule cells exposed to status at 8 weeks of age (blue bar) did not differ from age-matched cells from control animals (orange bar). All values are means ± SEM. ***p < 0.001. P0, Postnatal day 0; DGC, dentate granule cells; HBD, hilar basal dendrites.
Figure 3.
Figure 3.
A, C, Age-matched granule cells from control animals do not have basal dendrites. B, D, Immature (D), but not mature (B), granule cells exposed to SE exhibit basal dendrites 1 month later. A–D, Digital reconstructions of BrdU-labeled, GFP-expressing granule cells (orange) superimposed on maximum projections showing BrdU-negative, GFP-expressing neighbor cells (green). BrdU labeling is shown in blue. A.1, B.1, C.1, D.1, Confocal maximum projections from confocal z-series stacks used to generate the reconstructions shown in A–D. A.2, B.2, C.2, D.2, Confocal maximum projections showing BrdU labeling. Insets, Merged 90° rotations of GFP (orange) and BrdU (blue) confocal stacks showing colocalization for each neuron. Arrowheads denote axons. Arrows denote basal dendrites. Scale bar: A–D, 20 μm; A.1–D.2, insets, 40 μm.
Figure 4.
Figure 4.
Granule cell neurogenesis persists for at least 2 months after pilocarpine-SE, as demonstrated by labeling of newborn granule cells with the immature neuronal marker doublecortin. Images are single confocal optical sections through the hippocampal dentate gyrus from a control animal and animals examined 4, 8, and 11 weeks after pilocarpine-SE. By 11 weeks, a trend toward declining doublecortin labeling was evident, although labeled cells were still present. Asterisks depict regions of the subgranular zone devoid of doublecortin labeling. Arrowheads depict doublecortin-immunoreactive hilar ectopic granule cells. Scale bar, 200 μm.
Figure 5.
Figure 5.
Schematic detailing the experimental design. At 12 weeks of age, animals received pilocarpine to induce SE (group 5). Control animals received saline (group 6). Three weeks after treatment, animals received injections of BrdU to label granule cells. Five weeks later, animals were killed, and BrdU-labeled, GFP-expressing granule cells from epileptic and control animals were analyzed. DGC, Dentate granule cell.
Figure 6.
Figure 6.
A, B, Newborn granule cells generated after SE possess basal dendrites (B), whereas age-matched granule cells from control animals do not (A). Digital reconstructions of BrdU-labeled, GFP-expressing granule cells (orange) superimposed on maximum projections showing BrdU-negative, GFP-expressing neighbor cells (green) are shown. BrdU labeling is shown in blue/green. A.1, B.1, Maximum projections from confocal z-series stacks used to generate the reconstructions shown in A and B. A.2, B.2, Maximum projections showing BrdU labeling. Insets, Merged 90° rotations of GFP (orange) and BrdU (blue/green) confocal stacks showing colocalization for each neuron. Arrowheads denote axons. Arrows denote basal dendrites. Scale bar: A, B, 20 μm; A.1–B.2, insets, 40 μm.
Figure 7.
Figure 7.
Granule cells born after SE migrate into the hilus. A, Confocal maximum projection of a control animal showing GFP-labeled granule cells with their axons in the hilus. No BrdU-labeled, GFP-expressing hilar ectopic granule cells were found in control animals. Arrows denote BrdU-positive, GFP-negative cells. B, Digital reconstructions of BrdU-labeled, GFP-expressing granule cells (orange) superimposed on the maximum of the same field showing BrdU-negative, GFP-expressing neighbor cells (green). BrdU labeling is shown in blue. B.1, Maximum projection generated from confocal z-series stacks used to create the reconstruction shown in B. B.2, Maximum projection showing BrdU labeling. Inset, Merged 90° rotation of GFP (orange) and BrdU (blue) confocal stacks showing colocalization within the two hilar ectopic granule cells. DGC, Dentate granule cell. Arrowheads denote hilar ectopic granule cells. Scale bar: A, B, 60 μm; B.1, B.2, inset, 30 μm.
Figure 8.
Figure 8.
Composite of confocal maximum projections showing increased numbers of axonal expansions (arrows) on a granule cell born 3 weeks after SE relative to an age-matched granule cell from a control animal. Scale bar, 5 μm.
Figure 9.
Figure 9.
Bright-field micoroscopy images of cresyl violet-stained sections from control (no SE) and pilocarpine-treated groups. All animals that underwent SE exhibited extensive loss of hilar neurons (asterisks) and minimal loss of granule cells. Cell loss was equivalent among pilocarpine-treated groups. Scale bar, 200 μm.
Figure 10.
Figure 10.
Proposed model for the recruitment of adult-generated granule cells into aberrant hippocampal circuits. In this model, only the youngest cells can be induced to migrate into the dentate hilus, whereas the critical period for basal dendrite formation persists for a longer period of time. With neuronal maturity, both critical periods end, and exposure to epileptogenic brain injury induces neither abnormality. DGC, Dentate granule cell.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Ackman JB, Siddiqi F, Walikonis RS, LoTurco JJ. Fusion of microglia with pyramidal neurons after retroviral infection. J Neurosci. 2006;26:11413–11422. - PMC - PubMed
    1. Acsady L, Kamondi A, Sik A, Freund T, Buzsaki G. GABAergic cells are the major postsynaptic targets of mossy fibers in the rat hippocampus. J Neurosci. 1998;18:3386–3403. - PMC - PubMed
    1. al-Hussain S, al-Ali S. A golgi study of cell types in the dentate gyrus of the adult human brain. Cell Mol Neurobiol. 1995;15:207–220. - PubMed
    1. Ambrogini P, Lattanzi D, Ciuffoli S, Agostini D, Bertini L, Stocchi V, Santi S, Cuppini R. Morpho-functional characterization of neuronal cells at different stages of maturation in granule cell layer of adult rat dentate gyrus. Brain Res. 2004;1017:21–31. - PubMed
    1. Austin JE, Buckmaster PS. Recurrent excitation of granule cells with basal dendrites and low interneuron density and inhibitory postsynaptic current frequency in the dentate gyrus of macaque monkeys. J Comp Neurol. 2004;476:205–218. - PubMed

Publication types

MeSH terms