Histogenesis of ferret somatosensory cortex

The ferret has emerged as an important animal model for the study of neocortical development. Although detailed studies of the birthdates of neurons populating the ferret visual cortex are available, the birthdates of neurons that reside in somatosensory cortex have not been determined. The current study used bromodeoxyuridine to establish when neurons inhabiting the somatosensory cortex are generated in the ferret; some animals also received injections of [³H]thymidine. In contrast to reports of neurogenesis in ferret visual cortex, most neurons populating the somatosensory cortex have been generated by birth. Although components of all somatosensory cortical layers have been produced at postnatal day 0, the layers are not distinctly formed but develop over a period of several weeks. A small number of neurons continue to be produced for a few days postnatally. The majority of cells belonging to a given layer are born over a period of approximately 3 days, although the subplate and last (layer 2) generated layer take somewhat longer. Although neurogenesis of the neocortex begins along a similar time line for visual and somatosensory cortex, the neurons populating the visual cortex lag substantially during the generation of layer 4, which takes more than 1 week for ferret visual cortex. Layer formation in ferret somatosensory cortex follows many established principles of cortical neurogenesis, such as the well-known inside-out development of cortical layers and the rostro-to-caudal progression of cell birth. In comparison with the development of ferret visual cortex, however, the generation of the somatosensory cortex occurs remarkably early and may reflect distinct differences in mechanisms of development between the two sensory areas. J. Comp. Neurol. 387:179–193, 1997. © 1997 Wiley-Liss, Inc.     

Genesis of GABA-immunoreactive neurons in the ferret visual cortex

The pattern of neurogenesis of GABA-immunoreactive neurons in the ferret primary visual cortex was determined using immunohistochemical and 3H-thymidine autoradiographic techniques. Neurons in the visual cortex of the ferret undergo their final cell division during a period extending from embryonic day 20 (E20) to postnatal day 14 (P14) and follow an inside-out pattern of neuronal production (Jackson et al., 1984) similar to that observed in other mammals. Earlier-generated neurons are found at deeper cortical positions in the adult than are those generated later. Layer I is an exception to this rule, since neurons destined for this layer are produced at both the beginning and end of neurogenesis. In this study, the pattern of neurogenesis of GABA-immunoreactive neurons is compared to the pattern observed for nonimmunoreactive neurons. The overall pattern of cortical neurogenesis (inside-out pattern) is similar for GABA-immunoreactive neurons and neurons that are not GABA-immunoreactive. However, the GABA-immunoreactive neurons born on a given day of development are more broadly distributed across the radial axis of the adult cortex than are nonimmunoreactive neurons generated on the same day. GABA-immunoreactive neurons generated later in neurogenesis are, on average, slightly smaller than those generated early. If GABA-immunoreactive neurons in the visual cortex are interneurons, then these findings suggest that interneurons follow the same pattern of neurogenesis as do projecting neurons in the visual cortex.     

Neurogenesis of glutamic acid decarboxylase immunoreactive cells in the hippocampus of the mouse. I: Regio superior and regio inferior

The neurogenetic gradients of neurons showing glutamic acid decarboxylase (GAD) immunoreactivity were determined in the regio superior and in the regio inferior of the mouse hippocampus. Pregnant C57Bl mice received pulse injections of (3H)thymidine from E11 through E17 (E0 being the day of mating). Distributions of (3H)thymidine-labeled, GAD-positive neurons in the different strata of the hippocampus proper were recorded in adult animals. GAD-positive neurons in this region are generated prenatally. Radial gradients of neurogenesis of GAD-positive cells are characterized by two main features: 1) with the exception of the stratum lacunosum-moleculare and its interface with the stratum radiatum, GAD-positive neurons of the plexiform strata are generated before those destined for the pyramidal layer; 2) within the pyramidal layer, GAD-positive cells are positioned according to an inside-out sequence. In the transverse axis, neurogenesis of GAD-positive cells follows a regio inferior to regio superior gradient. This gradient is due to prolonged neurogenesis of GAD-positive cells for the pyramidal layer in the regio superior. Given the selective laminar disposition of the GABAergic interneurons in the hippocampus, the present authors explored whether or not the diverse types of these interneurons could have specific birth dates and concluded that no relationship exists between birth dates and adult phenotypes of GAD-immunoreactive cells in the mouse hippocampus proper.     

Neurogenesis of the cat's primary visual cortex

The 3H-thymidine method of birth-dating was used to determine when the cells belonging to each of the principal cellular layers of the cat's primary visual cortex are generated. In order to detect systematic differences in the position of radioactively labeled cells following 3H-thymidine administration at different prenatal ages, a geometric method was devised to represent the distribution of labeled cells in the form of depth histograms. Results show that visual cortical neurogenesis occurs largely during the second half of gestation between embryonic day 31 (E31) and E57. Cells of layer 6 are generated early, between E31 and E38, whereas cells destined for successively more superficial layers are generated at progressively later times. Layer 4 cells, the principal targets of geniculocortical afferents, are generated between E37 and E44. In addition, a special population of cells embedded in the white matter below layer 6 was found to be produced throughout the week-long period immediately prior to the onset of layer 6 neurogenesis. Overall, this radial pattern of cortical neurogenesis closely resembles the inside-first, outside-last, spatiotemporal sequence of development described for the monkey's primary visual cortex (Rakic, '74). In addition to finding this pronounced gradient in the radial dimension, we were also able to detect a less pronounced gradient along the tangential dimension: neurons destined for any given layer in the anterior part of the cortex (inferior visual field representation) are generated slightly in advance of neurons destined for more posterior regions (superior visual field). However even our more quantitative histogram analysis failed to reveal a mediolateral (central to peripheral visual field) gradient within area 17. In the cat, layers 6, 5, and 4 each take about a week to be generated, although their total cell numbers and packing densities differ in the adult. About 2 weeks are required to produce the cells of layers 2 and 3 combined. Furthermore, we found that neurons belonging to different layers and different morphological classes can be generated simultaneously. This suggests that the identity of a cortical neuron is not solely a function of the time of neurogenesis.     

Times of generation of glutamic acid decarboxylase immunoreactive neurons in mouse somatosensory cortex

The birth dates of neurons showing glutamic acid decarboxylase (GAD) immunoreactivity have been determined in mouse somatosensory cortex. Pregnant C57Bl mice received pulse injections of (3H)thymidine from E10 through E17 (E0 being the day of mating). The distributions of thymidine-labeled, GAD-positive and nonimmunoreactive (non-GAD) cells as a function of depth under the pial surface were recorded in adult animals. The maximum rate of generation of GAD-positive neurons occurred at E14, whereas the generation of non-GAD neurons reached its maximum rate at E13. Except for those in layer I, GAD-positive neurons followed an inside-out sequence of positioning. GAD-positive neurons born at E12 and E13 were located in layers VI-IV. GAD-positive neurons born at E14 were found throughout the cortical thickness, with a maximum in layer IV. The GAD-positive neurons labeled after pulses at E15 or E16 or E17 were limited to the superficial strata, forming a band that became narrower as it moved toward the pial surface with increase in age of pulse labeling. GAD-positive neurons in layer I were generated at a constant rate during the whole embryonic period analyzed. Non-GAD neurons also followed an inside-out spatiotemporal gradient. Two partially overlapping phases were distinguished in non-GAD neurogenesis. During the first phase (from E12 to E14) neurons populating adult layers VI and V originated, while neurons located in layers IV through I were generated during the second phase (from E13 to E17). Since GAD-immunoreactive neurons form a heterogeneous population, we envisage further studies in order to test whether differences exist in birth dates among the classes.     

Neurogenesis in a marsupial: the brush-tailed possum (Trichosurus vulpecula). I. Visual and auditory pathways

The times of origin of neurons in the visual and auditory systems were studied in a marsupial, the brush-tailed possum, using tritiated thymidine autoradiography. Within the subcortical visual pathways, most neurons are generated between postnatal days 5 and 21, and the neurons of the primary visual cortex up to postnatal day 68. In the subcortical auditory pathways, most neurons are generated between postnatal days 5 and 28, and all auditory cortex neurons have appeared by postnatal day 46. Neurons in a single layer of cerebral cortex are generated during a period of about 2 weeks. Thus cortical neurogenesis in marsupials extends over a period similar to that seen in primates.     

Time of origin of cortico-collicular projection neurons in the rat visual cortex.

The time of origin and the radial gradient of neurogenesis of cortico-collicular neurons have been studied in the rat visual area 17. We used a combined technique for the histochemical detection of the retrogradely transported horseradish peroxidase from the superior colliculus and the autoradiographic detection of the [3H]-thymidine administered during the gestational period. The cortico-collicular neurons of visual area 17 are located in layer V and are generated on gestational day (GD) 15 (59.78%), GD 16 (36.21%), and GD 17 (4.01%). This finding reveals that, for the cortico-collicular neuronal population, the birth date is well-correlated with the laminar position in the adult animal. To see whether the cortico-collicular neurons located at various radial levels of layer V are generated concurrently, or whether they follow an "inside-out" pattern of positioning, we divided layer V into three (upper, middle and lower) sublaminae. Most cortico-collicular neurons located in the lower two-thirds of layer V are generated on GD 15 (65%), whereas the neurons located in the upper third of the layer are generated both on GD 15 and GD 16 in almost equal proportions (52.53% and 44.39%, respectively).     

Laminar fate of cortical GABAergic interneurons is dependent on both birthdate and phenotype

Pioneering work indicates that the final position of neurons in specific layers of the mammalian cerebral cortex is determined primarily by birthdate. Glutamatergic projection neurons are born in the cortical proliferative zones of the dorsal telencephalon, and follow an "inside-out" neurogenesis gradient: later-born cohorts migrate radially past earlier-born neurons to populate more superficial layers. GABAergic interneurons, the major source of cortical inhibition, comprise a heterogeneous population and are produced in proliferative zones of the ventral telencephalon. Mechanisms by which interneuron subclasses find appropriate layer-specific cortical addresses remain largely unexplored. Major cortical interneuron subclasses can be identified based on expression of distinct calcium-binding proteins including parvalbumin, calretinin, or calbindin. We determined whether cortical layer-patterning of interneurons is dependent on phenotype. Parvalbumin-positive interneurons populate cortical layers with an inside-out gradient, and birthdate is isochronous to projection neurons in the same layers. In contrast, another major GABAergic subtype, labeled using calretinin, populates the cerebral cortex using an opposite "outside-in" gradient, heterochronous to neighboring neurons. In addition to birthdate, phenotype is also a determinant of cortical patterning. Discovery of a cortical subpopulation that does not follow the well-established inside-out gradient has important implications for mechanisms of layer formation in the cerebral cortex.     

Neurogenesis of the amygdaloid nuclear complex in the rhesus monkey.

The time course of neurogenesis for neurons which comprise the amygdaloid complex in Rhesus monkeys was determined using tritiated thymidine autoradiography. Fourteen pregnant monkeys received injections of tritiated thymidine between embryonic days 27 (E27) and 56 of their 165 day gestation and offspring were sacrificed during the early postnatal period. The first neurons destined for the amygdaloid complex were generated at E33 making them among the earliest postmitotic neurons in the telencephalon. Neurogenesis peaked within all nuclei of the amygdaloid complex between E38 and E48 and had ceased between E50 and E56. While amygdaloid neurogenesis in postnatally sacrificed monkeys displayed a dorsal-to-ventral gradient of radiolabeled neurons, the considerable rotation of the temporal lobe during the latter stages of primate development indicates that neurogenesis in the embryo, during the first third of gestation, actually occurs across a medial-to-lateral gradient. This medial-to-lateral gradient occurs as a smooth wave across the amygdaloid nuclei and does not respect neuroanatomical subdivisions or patterns of connectivity of the amygdaloid nuclei in the Rhesus monkey.     

Maturation of rat visual cortex: IV. The generation, migration, morphogenesis, and connectivity of atypically oriented pyramidal neurons

The generation, migration, and morphogenesis of atypically oriented pyramidal neurons in the rat visual cortex were examined. In the mature cortex, these neurons were distributed through layers II-VI. Moreover, the atypically oriented pyramidal neurons in a particular layer tended to be oriented in a specific way; atypically oriented pyramidal neurons in layer II, layers III-VIa, and layer VIb were obliquely, radially, and obliquely oriented, respectively. Ultrastructurally, the somata of atypically oriented pyramidal neurons contained large euchromatic ovoid nuclei and cytoplasm that was replete with rough endoplasmic reticulum and Golgi apparatus. These somata formed only symmetric axosomatic synapses. Many atypically oriented pyramidal neurons projected axons into the white matter as demonstrated by a Golgi method and by a retrograde tract-tracing technique; however, some of these pyramidal neurons in layers III-V had axons that ascended to layer I. By using a technique which combined retrograde tract tracing with [3H]thymidine autoradiography, it was determined that most atypically oriented pyramidal neurons in layers V and VIa, layer IV, and layer II were generated on gestational days (GD) 15-17, GD 17-19, and GD 20-21, respectively. Atypically oreinted pyramidal neurons were identified during the period from postnatal day 0 (day of birth) to day 30. On day 0, obliquely oriented pyramidal neurons were distributed in the deep cortical plate, i.e., immature layer VI. On day 3, the youngest atypically oriented pyramidal neurons were radially oriented and were located in layer IV. Some obliquely oriented pyramidal neurons were present in layer II on day 6, but the greatest number and the most severely canted pyramidal neurons in layer II were evident on day 9. The orientations of the cell body and the apical dendrite did not change appreciably after migration was complete, except for those in layers V and VI with obliquely oriented cell bodies and radially oriented apical dendrites. The second and third postnatal weeks were marked by substantial morphological differentiation of all pyramidal neurons as noted by the lengthening and branching of dendrites and by the appearance of dendritic spines. By the fourth postnatal week, atypically oriented pyramidal neurons achieved their mature morphology. The generation, migration, and morphogenesis of atypically oriented pyramidal neurons proceed by an inside-to-outside sequence. This development is similar and concurrent with that of typically oriented pyramidal neurons.     

Development of GABA-immunoreactivity in the neocortex of the mouse.

The prenatal and postnatal development of GABAergic elements in the neocortex of the mouse was analyzed by GABA-immunocytochemistry. Radial distribution of cells and laminar numerical densities were calculated at each developmental stage to substantiate qualitative observations. The first immunoreactive neurons were observed in the cortical anlage at embryonic day 12-embryonic day 13 (E12-E13) in the primitive plexiform layer. At following prenatal stages (E14-E19), most GABA-positive neurons were present in the marginal zone, subplate, and subventricular zone. GABA-immunoreactivity in the cortical plate appeared early (E14), although the complete maturation of its derivatives was achieved postnatally. At prenatal stages we noted a well-developed system of immunopositive fibers in the subplate. As indicated by the direction of growth cones, most of these fibers had an extracortical origin and invaded the cortex laterally through the internal capsule and striatum. In rostral and middle telencephalic levels, fibers originating in the septal region contributed to the cingulate bundle. Presumably corticofugal fibers and callosal axons were also noticed. At postnatal stages the maturation of GABA-immunoreactivity appeared to be a complex, long-lasting process, in which the adult pattern was produced at the same time as the appearance of certain regressive phenomena. Thus, between postnatal day 0 and postnatal day 8 (P0-P8), GABA-positive populations disappeared from the subventricular zone, marginal zone and to a lesser extent from the subplate. At the same ages we noticed the presence of morphologically abnormal, GABA-immunoreactive neurons in the subventricular zone and subplate which are interpreted as correlates of neuronal degeneration. Most GABA-positive subplate fibers also disappeared whereas GABA-immunoreactive axons were seen in the cingulate bundle until the adult stage. In the derivatives of the cortical plate, the maturation of GABA-immunoreactive elements progressed according to the "inside-out" gradient of cortical development, with the important exception of layer IV, which was the last layer to exhibit an adult-like appearance. Within each layer deriving from the cortical plate (layers VIa to II-III), GABA-immunoreactivity showed a protracted maturation in which the first GABA-positive cells were detected a few days after cell birth but substantial numbers of neurons began to express GABA considerably later. The later phase occurred concurrently with the maturation of GABA-positive axonal plexuses. These results suggest that different GABA-positive populations show different developmental regulation of GABA expression during cortical ontogenesis.     

The development of Ammon's horn and the fascia dentata in the cat: a [3H]thymidine analysis

The present [3H]thymidine autoradiographic analysis of neurogenesis demonstrates that the neurons which populate the adult cat hippocampus are born between embryonic day (E)22 and E42. In contrast, although neuronal production in the fascia dentata begins on the same day, granule cells in this area continue to be produced throughout prenatal life and into early postnatal life, and probably continues at an extremely low rate well into adulthood. Three major sets of spatiotemporal gradients characterize the production of neurons in Ammon's horn and the fascia dentata. The first set involves the radial axis. Within the hippocampus there exists an inside-out gradient. The reverse gradient is present in the fascia dentata, i.e. outside-in. The second set of gradients involves the transverse or rhinodentate axis. In general the CA3 neurons are born earlier than the CA1 neurons. Within both neuronal layers of the fascia dentata, the hidden blade cells tend to be born earlier than those of the exposed blade. Again, the pattern in the fascia is the reverse of that in the hippocampus proper. A temporal to septal gradient is also present, but this is the weakest of the gradients.     

An analysis of the time of origin of neurons in the entorhinal and subicular cortices of the cat

The [³H]thymidine autoradiographic method was used to determine the birth dates of neurons in the cat parahippocampal gyrus. Cat fetuses were exposed to a single pulse of the radioactive marker between the 20th and 55th embryonic days. All animals were delivered normally and allowed to survive for 2–6 months postnatal. The resulting autoradiographs demonstrate three spatiotemporal gradients of cell birth in the entorhinal and subicular cortices. First, an inside-out gradient is apparent; i.e., neurons in the deeper layers are born earlier than those in the more superficial layers. Second, a rhino to dentate gradient exists. Accordingly, cells closer to the lateral entorhinal region tend to be generated earlier than those further away. Third, a temporal to septal gradient is present. Neurons close to the anterior pole of the temporal lobe are born earlier than those more caudally located. Whereas the first two gradients have been observed in other species, the latter gradient has not been reported consistently. Three exceptions to these overall gradients exist. First, neurons near the layer I/II border are born earlier than the majority of the layer II neurons. Second, neurons near the transition zone between two adjacent regions are born earlier than neurons located in the middle of each region. Third, the prosubiculum and subiculum do not exhibit a clear inside-out or temporal to septal gradient.     

Neurogenesis in the visual system of the rat. An autoradiographic investigation.

Rats of the BD III strain were injected with a single dose of 3H-thymidine on either the twelfth, fourteenth, sixteenth, eighteenth or twentieth day of gestation (ED 12. . . . .ED 20) or on the postnatal day one, three, or seven. Animals were killed at age 22 to 24 days. DNA synthesis, as an indicator of cell division, was studied in matrix precursors of nerve and glial cells in the visula centers, including the lateral geniculate body (LGB), the superior colliculus (SC) and the visual cortex (VC). It was found that proliferation of matrix precursors of nerve cells destined for all the regions studied was in progress on ED 12. In the subcortical regions (LGB, SC) this process was substantially more advanced than in the VC. The first neuroblasts appeared in the SC (ED 12) and only later (ED 14) in the LGB and VC. In comparison with the LGB, VC neuroblasts were quite rare on ED 14 and were present only in layer VI. They appeared more frequently in this region only after injection of isotope on ED 16. Matrix cell proliferation and nerve cell formation ceased in the LGB between ED 16 and ED 18. The number of labeled cells arising after injection of the isotope on ED 16 indicates that neurogenesis ceased somewhat earlier in the dorsal nucleus of the LGB than in the ventral. In the SC the last neurons arose between ED 18 and ED 20, and in the VC, with the possible exception of a few granular neurons (which may continue division into the first few days postnatally), proliferation continued until the end of gestation. The origin of neuroblasts initially followed a caudo-rostral gradient. Later, the times of neurogenesis in the regions studied overlapped significantly. This is clear, for example. on ED 16, when neurogenesis in the mesencephalic SC continued for about two days longer than in the more postral LGB, and coincided with that in the VC, especially in the deep layers. The end of neurogenesis in the LGB, especially in the ventral nucleus, coincided with the time of neurogenesis in the deep cortical layers. In the VC, and partly also in the SC, an inside-out pattern of proliferation and neuron formation was confirmed. The times of proliferation of precursor cells, with the exception of the very end of neurogenesis, substantially overlapped within both these regions. The degree of this overlapping, described in terms of Labeling Index values, decreased towards the end of the neurogenetic period. Division of neuroglial cell precursors, started as early as on ED 14 in/for subcortical centers (LGB, SC), but not until ED 18 in/for the VC. A few labeled endothelial-like cells were observed in all regions studied after isotope injection on ED 12.     

Neurogenesis of the magnocellular basal forebrain nuclei in the rhesus monkey

The time of origin of the neurons that comprise the magnocellular basal forebrain nuclei in rhesus monkeys was determined by using [3H]thymidine autoradiography. Thirteen pregnant animals received an injection of [3H]thymidine between embryonic days 27 (E27) and E50 of their 165 day gestation, and their offspring were sacrificed during the early postnatal period. Neurons within this region were generated in a biphasic pattern. An initial burst of [3H]thymidine-labeled magnocellular neurons was first observed throughout short quiescent period, cells of the remaining anterior basal forebrain (inclusive of magnocellular neurons comprising the vertical limb of the diagonal band and the anteromedial and anterolateral regions of the nucleus basalis) were generated between E36 and E45 with a peak of neurogenesis seen on E40-E43. The intermediate division of the nucleus basalis was generated about the same time, but the peak period of neurogenesis in this region occurred slightly earlier (E36 and E40) and was completed by E43. During the second phase of neurogenesis, neurons within the posterior division of the basal forebrain were generated first, with their genesis virtually completed between E33 and E36. The genesis of all neurons comprising the magnocellular basal forebrain nuclei was completed by E48 of gestation. A general caudal to rostral gradient of neurogenesis was observed within this telencephalic region. In contrast, a neurogenic gradient was not discerned in the radial direction. The present data demonstrate that neurons comprising the basal forebrain magnocellular nuclei in monkeys are generated early in gestation with two peak times of neuronal genesis. These nuclei are among the earliest to be generated in the entire telencephalon, which, like neurons of the thalamus and cortical neurons giving rise to cortical-cortical connections, places them in a strategic position to potentially influence their target neurons within the cortical mantle that are generated later in gestation.     

Development of layer I and the subplate in the rat neocortex

Development of layer I and the subplate of the rat neocortex was examined with [3H]thymidine autoradiography. The experimental animals used for neurogenesis were the offspring of pregnant females injected with [3H]thymidine on 2 consecutive days: Embryonic Day (E) 13-E14, E14-E15, . . . E21-E22, respectively. On Postnatal Day 5, the proportion of layer I and subplate cells originating during 24-h periods were quantified at three anteroposterior levels. Presumptive Cajal-Retzius cells (large horizontal cells) are generated mainly on E14 and subplate cells on E14 and E15 ("outside-in" gradient). Both populations are generated earlier than cells in the cortical plate, which has an "inside-out" gradient. The subplate also has a ventrolateral/older to dorsomedial/younger neurogenetic gradient. The small- to medium-sized horizontal cells in layer I have an extensive period of neurogenesis with an "outside-in" gradient. To study morphogenesis, pregnant females were given single injections of [3H]-thymidine during gestation and embryos were removed in successive 24-h intervals (sequential-survival). On E15 and E16, cells accumulate outside the neuroepithelium in the primordial plexiform layer with older presumptive Cajal-Retzius cells superficial and younger presumptive subplate cells deep. The Cajal-Retzius cells permanently settle superficially among a first system of extracellular channels that appears on E14. Before reaching their final settling sites, subplate cells form the incipient cortical plate in the ventrolateral neocortex on E16. On E17, a seocnd system of extracellular channels appears below the cortical plate. On E18 and E19, subplate cells leave the cortical plate and permanently settle among the deep extracellular channels in a separate layer.     

Neurogenetic patterns in the medial limbic cortex of the rat related to anatomical connections with the thalamus and striatum

[3H]Thymidine autoradiography was used to investigate neurogenesis in all areas of the limbic cortex in the medial wall of the hemisphere. The experimental animals were the offspring of pregnant females injected with [3H]thymidine on 2 consecutive days: Embryonic Day (E)13-E14, E14-E15, ...E21-E22, respectively. On Postnatal Day (P)60, the proportion of neurons originating during 24-h periods were quantified at nine anteroposterior levels. Three types of neurogenetic gradients are found. (i) Deep cells are older than superficial cells: layer VI is generated mainly on E15-E16, layer V on E16-E18, and layers IV-II on E18-E20. (ii) There is a ventral/older to dorsal/younger gradient between the dorsal peduncular, infralimbic, and anterior cingulate areas rostral to the genu of the corpus callosum. A ventral/older to dorsal/younger gradient is also found between superficial cells (layers II-IV) in anterior cingulate (CG3/CG1), posterior cingulate (CG2/CG1), and retrosplenial areas (RSG/RSA). (iii) An anterior/older to posterior/younger gradient is found between areas throughout the medial limbic cortex. Some of these neurogenetic patterns correlate with anatomical interconnections between the supracallosal medial limbic cortex (posterior cingulate and retrosplenial areas) and the anteroventral/anteromedial thalamic nuclei: older thalamic cells have longer axons that terminate in cortical areas containing younger cells, while younger thalamic cells have shorter axons that terminate in cortical areas containing older cells. Projections from the medial limbic cortex to the striatum also correlate with neurogenetic gradients: older cortical source cells in the infralimbic area project to the older striatal cells in the enkephalin-rich patches, while younger cortical source cells in the cingulate areas project to younger striatal cells in the surrounding matrix.     

Development of the brain stem in the rat. III. Thymidine-radiographic study of the time of origin of neurons of the vestibular and auditory nuclei of the upper medulla

Groups of pregnant rats were injected with two successive daily doses of ³H-thymidine form gestational days 12 and 13 (E12 + 13) until the day before parturition (E21 + 22). In adult progeny of the injected rats the proportion of neurons generated on specific embryonic days was determined quantitatively in the vestibular and auditory nuclei of the upper medulla. In the vestibular nuclei, neurons are generated between days E11 and E15 in an overlapping sequential order, yielding a lateral-to-medial and a rostral-to-caudal internuclear gradient. In the lateral vestibular nucleus peak production time is day E12; in the superior nucleus, E13; in the inferior nucleus, E13 and E14; and in the medial nucleus, E14. The early generation of neurons of the lateral vestibular nucleus may reflect the early differentiation of the circuit from the gravity receptors (utricle) to neurons of the spinal cord controlling postural balance. The later production of neurons of the superior vestibular nucleus may reflect the subsequent differentiation of the circuit from the rotational receptors (semicircular canals) to the neurons of the brain stem controlling eye movements. The generation time of neurons of the nucleus prepositus hypoglossi overlaps with that of the medial vestibular nucleus. The neurons of the anteroventral and posteroventral cochlear nuclei are produced form days E13 to E17, with no temporal differences between the two nuclei. The neurons of the dorsal cochlear nucleus are generated over a very long time span, beginning on day E12 and extending into the postnatal period. There is a sequence in the production of neurons forming the different layers of the dorsal cochlear nucleus in the following order: pyramidal cells, cells of the inner layer, cells of the outer layer and, finally, cells of the granular layer. There is also a sequential production of neurons in four nuclei of the superior olivary complex. In the lateral trapezoid nucleus peak production time is day E12; in the medial superior olivary nucleus, day E13; in the medial trapezoid nucleus, day E15; and in the lateral superior olivary nucleus, day E16. This order yields a medial-to-lateral gradient in the dorsal aspect of the superior olivary complex, and a lateral-to-medial gradient ventrally. These mirror-image gradients were also seen intranuclearly in the lateral superior olivary nucleus and the medial trapezoid nucleus. The cytogenetic gradients could not be related to tonotopic representation; however, they could be related to the lateral location of ipsilateral cochlear nucleus input to the lateral superior olivary nucleus and the medial location of the contralateral cochlear nucleus input to the medial trapezoid nucleus.     

Neurogenesis of basal forebrain cholinergic neurons in rat

The basal forebrain cholinergic system embodies a heterogeneous group of neurons distributed in the basal telencephalon that project topographically to the cortical mantle. We sought to examine the generation of these neurons to determine whether basal forebrain neurons have unique patterns of neurogenesis or, if, in contrast, they are born along general neurogenic gradients. The techniques of tritiated thymidine autoradiography and choline acetyltransferase (ChAT) immunocytochemistry were combined to determine the birthdays and neurogenic gradients of cholinergic cells in this region of rat brain. Cholinergic neurogenesis throughout the basal forebrain ranged from embryonic days 12 to 17 (E12-17). Neurogenesis in the nucleus basalis magnocellularis occurred over E12-16, with a peak day of generation on E13. The horizontal limb nucleus of the diagonal band which is located rostral to the nucleus basalis was generated over E12-17, with the majority of cells arising on E14-15. The rostral-most nuclei of the basal forebrain cholinergic system, the vertical limb of the diagonal band and the medial septum, were generated between E13-17, with peak days of neurogenesis on E15 and E15-16, respectively. These results were evaluated quantitatively and demonstrated that the basal forebrain cholinergic neurons were generated along the general caudal-to-rostral gradient previously described for all neurons in this brain region. The results of this study, in combination with those of similar investigations, emphasize that position-dependent epigenetic factors appear to be more potent determinants of the time of neuronal origin than factors which influence a cell's transmitter phenotype.