Generated Granule Cells Research Articles

Distribution of glutamate receptors GluR 2/3 and NR1 in the developing rat cerebellum

The distribution of glutamate receptors GluR2/3 and NR1 was analysed immunohistochemically during development of the rat cerebellum. GluR2/3 immunoreactivity appeared by postnatal day P0 in somata of Purkinje cells. Throughout P7, P15, P20 and adulthood, GluR2/3 immunoreactivity was found in the entire Purkinje cell dendritic arbor reaching to the external granular layer and, by P15, the surface of the cerebellum. By P7, the granular layer revealed scattered, mildly reactive, cells. NR-1 immunoreactivity first gained prominence about P7 in the region of the multi-layered Purkinje cell somata. By P15, NR1 was prominent in Purkinje cell somata and Golgi cells. The reaction product extended into the primary main dendrite of Purkinje cells. By P21, stellate and basket cells had intense reactivity throughout the molecular layer and reactive large-diameter dendrites of Golgi cells projected toward the molecular layer. Granule cells remained very weak among strongly reactive Golgi cell somata and dendrites. Ultrastructural immunohistochemistry revealed NR1 reaction product in Purkinje cell somata, in stellate cell somata and dendrites and on postsynaptic membranes of scattered spines throughout the molecular layer. The later appearance and restricted location of NR1 in somata and proximal dendrites of Purkinje cells contrasted markedly with GluR2/3 which appeared before birth and remained prominent throughout Purkinje cell dendritic arbors of adults. The time of NR1 expression correlated with the generation of granule cells, their synaptogenesis on Purkinje cells, the formation of stellate/basket cells and the shift of climbing fibre synapses from distal to proximal dendrites. The developmental appearance of stellate/basket cells and Golgi cells as well as their high reactivity remaining into adulthood suggests that these inhibitory molecular and granular layer interneurons are the principal targets of glutamate axons serving NR1 synaptic properties while Purkinje cells and brush type granule cells are targets for glutamate connections with GluR2/3 characteristics.

Effect of prenatal protein deprivation on postnatal granule cell generation in the hippocampal dentate gyrus.

The effect of prenatal malnutrition, produced by protein deprivation, on postnatal neurogenesis of granule cells in the fascia dentata of the rat hippocampal formation was examined by injecting tritiated thymidine on P8 and P15 and sacrificing the pups on P30, or by injecting on P30 and sacrificing on P90. The number of labeled granule cells was significantly decreased in prenatally malnourished rats injected on P8, and unaffected in those injected on P15. In contrast, the number of labeled granule cells in prenatally malnourished rats was significantly increased in animals injected in P30. The study shows that prenatal malnutrition significantly alters the postnatal pattern of granule cell neurogenesis in rat hippocampal formation and that the effect persists despite nutritional rehabilitation at birth.

Local Control of Granule Cell Generation by Cerebellar Purkinje Cells

Cerebellar Purkinji cells were ablated by the specific expression of diphtheria toxin in these cells in transgenic mice. Purkinje cell degeneration during early postnatal development shows a zonally restricted pattern which has been exploited in order to look for local secondary effects. The most obvious early effect is the alignment of gaps in the Purkinje cell layer with dramatically thinned zones in the overlying EGL, the germinal layer from which granule cells are generated. Within these EGL zones in the transgenic mutant, markers that distinguish matrix from mantie cells demonstrate a preferential loss of the proliferative cells. Comparison of BrdU incorporation in the mutant vs wild-type confirms the reduction in proliferation. In the mutant, in situ labeling of DNA fragmentation associated with apoptotic cell death shows abundant labeling of granule cells that have exited the EGL, but not of progenitor cells In the EGL. Thus, although a trophic role for Purkinje cells has been well documented, these observations further suggest a mitogenic role which can be exerted locally.

Prenatal malnutrition effect on pyramidal and granule cell generation in the hippocampal formation

The effects of prenatal malnutrition produced by protein deprivation on the neurogenesis of granule and pyramidal cells in the rat hippocampal formation was investigated by injecting pregnant rats with tritiated thymidine on E12, E16, or E20 and sacrificing the pups on P30. Granule cell neurogenesis was significantly decreased in the pups injected on E20, but not in E12 or E16 groups. There was no effect on the generation of pyramidal cells at the times noted, indicating a differential effect of prenatal malnutrition on the generation of these two different neuronal types in the hippocampal formation.

Possible involvement of radial glia in the development of the newly generated granule cells of the adult rat dentate gyrus

Possible involvement of radial glia in the development of the newly generated granule cells of the adult rat dentate gyrus

Highly polysialylated neural cell adhesion molecule (NCAM-H) is expressed by newly generated granule cells in the dentate gyrus of the adult rat

We have found in the adult rat that the persistent expression of a highly polysialylated neural cell adhesion molecule (NCAM-H) that is generally specific to developing tissues, remains restrictively in the cells of the deepest portion of the dentate granular layer. Since the granule cells are known to continue to be generated in this region during the adult period, we have tried to determine whether NCAM-H is expressed by newly generated granule cells. Immunoelectron microscopic observation revealed that about half of the NCAM-H-expressing cells had the features of dentate granule cells, and that the rest of these cells appeared to be immature cells. Double immunostaining for NCAM-H and glial fibrillary acidic protein (GFAP) revealed that the NCAM-H-expressing cells differed from GFAP-positive glial cells. In rats injected with 5-bromo-2'-deoxyuridine (BrdU) at post-natal day 35, double immunostaining for NCAM-H and BrdU demonstrated that the BrdU-labeled cells expressed NCAM-H at 12 d after the injection but not at 80 d. These results provide the first direct evidence that NCAM-H is expressed transiently by newly generated granule cells that may add new neuronal circuits to the adult hippocampal formation.

1408 New neuronal circuits are age-dependently added to adult hippocampal formation by newly generated granule cells

1408 New neuronal circuits are age-dependently added to adult hippocampal formation by newly generated granule cells

The persistent expression of a highly polysialylated NCAM in the dentate gyrus of the adult rat

The expression of a highly polysialylated form of the neural cell adhesion molecule (NCAM-H), often termed 'embryonic NCAM', has been investigated in the hippocampal formation of developing and adult rats. To determine the immunohistochemical localization of NCAM-H, a monoclonal antibody that reacts with the polysialic acid portion of NCAM-H was used. In the late embryonic and early postnatal periods, immunoreactivity for NCAM-H was found throughout the hippocampal formation, except for the ventricular layer. Thereafter, the immunoreactivity gradually decreased and almost vanished in most parts in the adult. However, a strong immunoreactivity remained on a number of cells in the dentate gyrus of adult rats, particularly in the deepest part of the granular layer. The immunoreactive arborized dendrites, mostly arising from the primary apical pole of the granule cells, were found to enter the molecular layer. The mossy fibers also were positive. Electron-microscopic examination of the hilus portion showed that the immunoreactivity was detected on the plasma membrane of some axons in the mossy fiber bundles. Since postnatal neurogenesis is known to continue into adulthood in the deepest part of the granule cell layer of the dentate gyrus, these results suggest that, in the adult dentate gyrus, NCAM-H is expressed by newly generated granule cells, and that the NCAM-H-expressing new cells may participate in the formation of new neural circuits.

Highly polysialylated NCAM is expressed in newly generated granule cells of the adult dentate gyrus

Highly polysialylated NCAM is expressed in newly generated granule cells of the adult dentate gyrus

Migration and distribution of two populations of hippocampal granule cell precursors during the perinatal and postnatal periods

Methacrylate-embedded sections and short-survival thymidine radiograms of the hippocampal dentate gyrus were examined in perinatal and postnatal rats in order to trace the site of origin and migration of the precursors of granule cells and study the morphogenesis of the granular layer. The densely packed, spindle-shaped cells of the secondary dentate matrix (a derivative of the primary dentate neuroepithelium) stream in a subpial position towards the granular layer of the internal dentate limb during the perinatal and early postnatal periods. By an accretionary process, the crest of the granular layer forms on day E21 and on the subsequent days the granular layer of the internal dentate limb expands progressively in a lateral direction. Granule cells differentiation, as judged by the transformation of polymorph, darkly staining small cells into rounder, lightly staining larger granule cells, follows the same gradient from the external dentate limb to the internal dentate limb. The secondary dentate matrix is in a process of dissolution by day P5. This matrix is the source of what will later become the outer shell of the granular layer composed of early generated granule cells. The thicker inner shell of the granular layer, formed during the infantile and juvenile periods, derives from an intrinsic, tertiary germinal matrix. On day E22, the dentate migration of the secondary dentate matrix becomes partitioned into two components: a) the subpial component of extradentate origin, referred to in this context as the first dentate migration, and b) the second dentate migration. The latter is distributed in the basal polymorph layer throughout the entire dentate gyrus and is henceforth recognized as the tertiary dentate matrix. The tertiary dentate matrix is prominent between days P3 and P10. It is postulated that the great increase in granule cell population during the infantile period is principally due to cells derived from this intrinsic matrix of the dentate gyrus. Between days P20 and P30 the tertiary dentate matrix disappears in the basal polymorph layer and henceforth proliferative cells become largely confined to the subgranular zone at the base of the granular layer. The subgranular zone is the source of granule cells produced during the juvenile and adult periods.

Neuromorphological changes in mouse olfactory bulb after neonatal exposure to phenobarbital

Heterogeneous Sabra mice pups were injected daily with 50 mg/kg phenobarbital (B 50) or 40 mg/kg (B 40) from postnatal day 2 (P 2) to P 21. Control litter mates (C) received vehicle injections. All animals were killed on P 50 and their brains removed for histological analysis. The volumetric growth of the main olfactory bulb (MOB) was assessed in hematoxylin and eosin-stained serial sections cut in the sagittal plane. The volume of MOB was reduced 20 to 25% in barbiturate-treated mice. The greatest reductions in volume were seen in the external layers of the bulb (20 to 28%). There were moderate reductions in the internal granular layer (IGL, 6 to 20%) and no reductions in the subependymal layer that contains the migrating and immature cells. The number of prenatally-formed mitral cell neurons was unaffected by the treatment while the number of the pre- and postnatally generated granule cells was significantly reduced in the B 40 group. There were no treatment effects on the sizes or packing densities of the mitral or granule cells. Like the hippocampus and the cerebellum, the developing MOB is vulnerable to barbiturate-exposure.

Numerical matching in the mammalian CNS: lack of a competitive advantage of early over late-generated cerebellar granule cells.

In this report we use postnatal 3H-thymidine injections to test whether granule cells that are generated early in postnatal cerebellar development and whose axons have access to their Purkinje cell target beginning in the first postnatal week have an advantage over granule cells generated 9 days later in the competition for target-related stabilization. In the wild-type mouse, 3-5% of the adult granule cell population is labeled by injection of 3H-thymidine at either postnatal day 4 (P4) or P13. In the lurcher mutant, however, over 40% of the surviving granule cells are labeled by P4 injection while less than 1% are labeled after a P13 injection. Together, these results suggest that time of target contact is a critical factor in the competition for neuronal survival. The results from the lurcher chimeras, however, reveal that the situation is likely to be more complicated. In all chimeras examined, with target sizes ranging from 3 to 108% of wild type, equivalent numbers of granule cells were labeled at P4 and P13. These data lead to the contradictory conclusion that, in this experimental situation, early generated granule cells do not have a competitive advantage over later-generated granule cells. The results are discussed in terms of various models of target stabilization. We propose that, of the various hypotheses, our results are best explained by postulating two distinct mechanisms for developmental cell death. Supporting evidence for this hypothesis from other neuronal systems is also briefly reviewed.

Stability of synaptic density and spine volume in dentate gyrus of aged rats

The number of synapses per unit volume and per granule cell and the size of dendritic spines were studied in the dentate gyrus of Sprague-Dawley rats 6, 24, and 30 months of age. Neither synaptic density nor mean spine volume showed any age-related trends. An increase in granule cell packing density at 24 months and concomitant stability of the height of the granule cell layer is consistent with the idea that postnatal generation of granule cells may continue late into life. Possible explanations for the discrepancies in the literature regarding synaptic loss in this area include differences in morphometric techniques, age of animals used, regional differences within dentate gyrus, and sampling variability. Generalized synapse loss in the senescent rodent brain remains to be established.

Infrapyramidal Mossy Fibers in the Hippocampus of the Hyperthyroid Rat

Hyperthyroidism, initiated at birth, produces an infrapyramidal (IP) bundle of mossy fibers in stratum oriens of field CA3_a-b of Ammon''s horn. The distribution of these axons, their mode of passage to the infrapyramidal region, and their synaptic relationship to the basal pyramidal cell dendrites were studied using the Timm-Haug silver sulfide method, rapid Golgi, and electron microscopy. Terminals of the hyperthyroid IP mossy fibers were consistently found to synapse with ''excrescences'' on the basal pyramidal cell dendrites in field CA3_a-b at all levels of the hippocampus. The excrescences, normally characteristic of synapses between the suprapyramidal mossy fibers and apical pyramidal cell dendrites are uncommon on the basal pyramidal cell dendrites of CA3_a-b in untreated animals except at septodorsal levels where a few are found in association with a small number of IP mossy fibers. The apparently exclusive association of excrescences with mossy fiber boutons even when they are in an ectopic position suggests that the mossy fibers may have the ability to induce formation of these postsynaptic specializations oh their pyramidal cell targets during development. Topographic differences with respect to the magnitude of the hyperthyroid effect at different anatomical levels were also observed. For example, although IP mossy fibers were numerous in CA3_a-b at the midseptotemporal level, this effect was even more pronounced at septodorsal levels. These differences may reflect the interaction of the hyperthyroid state with certain developmental relationships such as the temporoseptal gradient for granule cell generation and the septotemporal gradient for development of cholinergic afferents to stratum oriens. Since thyroid hormone requires immature cells to exert its differentiation related effects, the more immature IP mossy fibers at septodorsal levels might react more strongly to the growth stimulating effect of hyperthyroidism and thus be able to successfully compete for postsynaptic sites on the basal pyramidal cell dendrites. In untreated animals, the presence of a small number of IP mossy fibers in CA3_a-b at septodorsal levels might reflect these same spatio-temporal relationships during phases of hippocampal development normally modulated by thyroid hormone. Formation of the IP mossy fiber bundle in hyperthyroidism is <i>dose dependent, </i>raising the possibility that it results from a direct effect of excess thyroxine on axonal growth. Furthermore, it is <i>long-lasting </i>indicating that the mossy-fiber pyramidal cell circuitry is permanently altered. Finally, the <i>''critical period'' </i>for its production occurs within the first 3 weeks after birth which coincides with the main period of granule cell generation and mossy fiber development in the rat. This experimentally produced aberration in the development of intrinsic connections emphasizes the importance of temporal factors and their hormonal modulation in postnatal hippocampal ontogeny. Future studies of circuitry construction in this discrete hodologic system may benefit from the use of hyperthyroidism as an analytical tool.