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International audience; Several studies suggest that neurons from the lateral region of the SuM (SuML) innervating the dorsal dentate gyrus (DG) display a dual GABAergic and glutamatergic transmission and are specifically activated during paradoxical (REM) sleep (PS). The objective of the present study is to characterize the anatomical, neurochemical and electrophysiological properties of the SuML-DG projection neurons and to determine how they control DG oscillations and neuronal activation during PS and other vigilance states. For this purpose, we combine structural connectivity techniques using neurotropic viral vectors (rabies virus, AAV), neurochemical anatomy (immunohistochemistry, in situ hybridization) and imaging (light, electron and confocal microscopy) with in vitro (patch clamp) and in vivo (LFP, EEG) optogenetic and electrophysiological recordings performed in transgenic VGLUT2-cre male mice. At the cellular level, we show that the SuML-DG neurons co-release GABA and glutamate on dentate granule cells and increase the activity of a subset of DG granule cells. At the network level, we show that activation of the SuML-DG pathway increases theta power and frequency during PS as well as gamma power during PS and waking in the DG. At the behavioral level, we show that the activation of this pathway does not change animal behavior during PS, induces awakening during slow wave sleep and increases motor activity during waking. These results suggest that the SuML-DG pathway is capable of supporting the increase of theta and gamma power in the DG observed during PS and plays an important modulatory role of DG network activity during this state.

International audience; The striatum, the main input structure of the basal ganglia, is critical for action selection and adaptive motor control. To understand the neuronal mechanisms underlying these functions, an analysis of microcircuits that compose the striatum is necessary. Among these, cholinergic interneurons (ChIs) provide intrinsic striatal innervation whose dysfunction is implicated in neuropsychiatric diseases, such as Parkinson's disease and Tourette syndrome. The ability to experimentally manipulate the activity of ChIs is critical to gain insights into their contribution to the normal function of the striatum and the emergence of behavioral abnormalities in pathological states. In this study, we generated and tested CAV-pChAT-GFP, a replication-defective canine adenovirus type 2 (CAV-2) vector carrying the green fluorescent protein (GFP) sequence under the control of the human choline acetyltransferase (ChAT) promoter. We first tested the potential specificity of CAV-pChAT-GFP to label striatal ChIs in a rat before performing experiments on two macaque monkeys. In the vector-injected rat and monkey striatum, we found that GFP expression preferentially colocalized with ChAT-immunoreactivity throughout the striatum, including those from local circuit interneurons. CAV-2 vectors containing transgene driven by the ChAT promoter provide a powerful tool for investigating ChI contributions to circuit function and behavior in nonhuman primates.

INTRODUCTION: It has been shown that total neuronal pool in the primate associative cortex contains up to 12% of calretinin neurons whereas in rodents their proportion does not exceed 3%. Conjointly it is unclear if such an increase is reflected in laminar distribution and present in entire cortex. In this study, we have compared phylogenetically and functionally different areas of the macaque monkey prefrontal cortex (Brodmann areas 24, 32 and 9). MATERIALS AND METHODS: Stereological analysis of serially cut sections, immunohistochemically stained for the neuronal nuclear marker (NeuN) and calretinin (CR), revealed that calretinin neurons are mostly located (80%) in the upper cortical layers. Across all analysed areas, upper cortico cortical layers have a higher proportion of calretinin neurons: in layer III 20%, in layer II 30% and in layer I almost 50% of neurons express calretinin. The increased number of calretinin neurons in the lower part of the layer I of the primate prefrontal cortex might be the reason for the increased cellularity (3% of total neuronal number) of this layer. RESULTS: Our data showed that in the primate prefrontal areas, GABAergic neurons are overrepresented in upper layers, implying more complex network organization and information processing. Absence of differences between prefrontal areas in macaque monkeys confirms that principal neocortical input and output cell types are a conserved feature of the dorsal telencephalon. CONCLUSION: Comparison of orbito-medial prefrontal cortex (Džaja et al. 2014, Society for neuroscience 446.19) with lower densities in dorso-medial parts suggests that during primate evolution increase in neuron complexity and connections precedes in dorsal areas of prefrontal cortex.

International audience; HIGHLIGHTS Blockade of dopamine D1 receptors in ACC suppressed instrumental learning when overt responding was required.Covert learning through observation was not impaired.After treatment with a dopamine antagonist, instrumental learning recovered but not the rat's pretreatment level of effort tolerance.ACC dopamine is not necessary for acquisition of task-relevant cues during learning, but regulates energy expenditure and effort based decision. Dopamine activity in anterior cingulate cortex (ACC) is essential for various aspects of instrumental behavior, including learning and effort based decision making. To dissociate learning from physical effort, we studied both observational (covert) learning, and trial-and-error (overt) learning. If ACC dopamine activity is required for task acquisition, its blockade should impair both overt and covert learning. If dopamine is not required for task acquisition, but solely for regulating the willingness to expend effort for reward, i.e., effort tolerance, blockade should impair overt learning but spare covert learning. Rats learned to push a lever for food rewards either with or without prior observation of an expert conspecific performing the same task. Before daily testing sessions, the rats received bilateral ACC microinfusions of SCH23390, a dopamine D1 receptor antagonist, or saline-control infusions. We found that dopamine blockade suppressed overt responding selectively, leaving covert task acquisition through observational learning intact. In subsequent testing sessions without dopamine blockade, rats recovered their overt-learning capacity but not their pre-treatment level of effort tolerance. These results suggest that ACC dopamine is not required for the acquisition of conditioned behaviors and that apparent learning impairments could instead reflect a reduced level of willingness to expend effort due to cortical dopamine blockade.

Although the basic principles of cortical development are conserved across mammals, evolutionary increased complexity of cortical GABAergic network leads to significant variability in origin and migration of GABAergic neurons when primates are compared to rodents. GABAergic cortical network in primates is particularly distinguished by tremendous increase in the proportion of calretinin neurons which contribute to half of GABAergic neuron population and are two times more numerous than other two major subpopulations, parvalbumin and calbindin/somatostatin. Therefore, it could be expected that the evolution is changing developmental rules to enable substantial changes that appear inside primate GABAergic network. To establish place of origin and migratory routes of different GABAergic neuron subpopulations in primates, we examined postmortem human and monkey fetal brain tissue during early fetal period (8-12 postconceptional weeks in human and embryonic day 47-56 in cynomologus monkey) processed immunohistochemicaly for markers of GABAergic neurons (GAD65 and GAD67) as well as for molecular markers of three major subpopulations. Data confirmed that, as in all mammals described so far, the most important source of GABAergic neurons during the early fetal period is the ganglionic eminence. The vast majority of progenitors in the ganglionic eminence express GAD65 (but not GAD67). After leaving the ganglionic eminence tangentially migrating cells continue to express GAD65, emerge laterally on the cortico-striatal border and form main tangential migratory stream in the subventricular/intermediate zone of the lateral telencephalic wall. Intensive GAD67 expression is observed by progenitors in the proliferative zones in rostro- dorsal and ventro-caudal part of the medial telencephalic wall. These so far undescribed proliferative regions are a source of GABAergic neurons which accumulate in the basal telencephalon and continue to migrate through the marginal zone and the layer below the cortical plate (primordial subplate). Part of the GAD67+ migrating cells in this superficial stream originates also from the preoptic area and hypothalamic proliferative zone which are in continuation with GAD67+ proliferative zones of medial telencephalon. Data also showed that already during the early fetal period the majority of GABAergic cells that originate from the ganglionic eminence and migrate via deeper migratory stream express calretinin, while GAD67+ proliferative zones of the medial telencephalic wall are the main source of somatostatin cells that migrate via the superficial stream. Our results suggest that primate specific features of GABAergic neuronal development are present already during the early fetal period. Two main subpopulations of GABAergic cortical neurons, somatostatin and calretinin, have clearly distinguishable origin and migratory routes, and significant production of calretinin subpopulation occurs earlier than in rodents. In addition to their dorsal production later on, this significant early production also accounts for tremendous increase in the proportion of calretinin neurons in primates.

Previous studies in primate associative cortex have shown that calretinin neurons represent 12% of the total neuronal pool whereas in rodents their proportion does not exceed 3%. However it is still unclear whether such high proportion of calretinin neurons is a general feature of all primate cortical areas, whether it is layer specific and whether all these calretinin neurons are GABAergic. To address these questions we performed a quantitative study of calretinin neurons within three phylogenetically and functionally different areas of the macaque monkey prefrontal cortex (Brodmann areas 24, 32 and 9). Our results showed that calretinin labelled neurons represent around 15% of the total number of neurons revealed by NeuN immunolabelling in all examined cortical areas. Calretinin neurons were mostly located (75%) in the upper cortical layers. They account for 50% of total number of the neurons in layer I ; 30% in layer II and 20% in layer III. Simultaneous immunohistochemical detection of calretinin with parvalbumin, or calbindin, showed that the population of calretinin neurons does not overlap with the other two major subpopulations of GABAergic neurons. It further demonstrated that calretinin neurons are at two times more numerous than the population of parvalbumin neurons and of calbindin neurons. These three subpopulations of GABAergic neurons account for 25% of all cortical neurons in the primate prefrontal cortex. Our data indicated that the high proportion of calretinin neurons in the upper cortical layers is a general feature of the primate prefrontal cortex. It suggests that this subpopulation of GABAergic neurons is instrumental to control information processing of a more complex cortico-cortical neuronal network.

Previous studies in primate associative cortex have shown that calretinin neurons represent 12% of the total neuronal pool whereas in rodents their proportion does not exceed 3%. It is still unclear if such an increase is reflected in laminar distribution and present in entire cortex. In this study, we have compared phylogenetically and functionally different areas of the macaque monkey prefrontal cortex (Brodmann areas 24, 32 and 9). Stereological analysis of serially cut sections, immunohistochemically stained for the neuronal nuclear marker (NeuN) and calretinin (CR), revealed that calretinin neurons are mostly located (80%) in the upper cortical layers. Across all analysed areas, upper cortico cortical layers have a higher proportion of calretinin neurons: in layer III 20%, in layer II 30% and in layer I almost 50% of neurons express calretinin. The increased number of calretinin neurons in the lower part of the layer I of the primate prefrontal cortex might be the reason for the increased cellularity (3% of total neuronal number) of this layer. Our data showed that in the primate prefrontal areas, GABAergic neurons are overrepresented in upper layers, implying more complex network organization and information processing. Absence of differences between prefrontal areas in macaque monkeys confirms that principal neocortical input and output cell types are a conserved feature of the dorsal telencephalon. Comparison of orbito- medial prefontal cortex (Džaja et al. 2014, Society for neuroscience 446.19) with lower densities in dorso-medial parts suggests that during primate evolution increase in neuron complexity and connections precedes in dorsal areas of prefrontal cortex.

Cortical column may have structural differences between species of different cognitive abilities. Several studies illustrate that the ratio of excitatory VS inhibitory neurons, in cortex of different species, is not constant. This variability could result from evolutionary increased number of GABAergic cells, especially those containing calretinin (CR). In order to test this hypothesis, a quantitative study of the CR neuron density was performed in the orbitofrontal cortex (OFC) of Wistar rat and Rhesus monkey. The analyzed region corresponds to agranular or dysgranular cortex, with strong hippocampal projections, i.e. medial orbital (MO) area in the rat and Brodmann area 14 caudal (14c) in monkey. Cresyl violet staining and imunohistochemical labeling for calretinin and NeuN were performed on adjacent sections throughout the entire region. Unbiased stereological method was used to estimate the total number of neurons (NeuN) and CR cells in each cortical layer of the region of interest. The rat MO region has a volume of 2.6 mm3 with ~ 115 000 NeuN cells, giving density of ~44 000/mm3. Number of CR cells in MO is ~3 000, with the density of ~ 1100/mm3. CR neurons represent 2.6% of the total neuronal population. Monkey region 14c displays a volume of 13.8 mm3 with ~ 1 050 000 NeuN cells, giving density of ~ 75 800/mm3. Number of CR cells is ~ 130 000, with density of ~9400/mm3. CR neurons represent ~12% of the entire neuronal population. This study establishes a significant difference between primates and rodents regarding CR neuron proportion.

Cortical hem, a putative signaling centre at the interface of the prospective hippocampus and the choroid plexus, is a significant source of Cajal-Retzius cell population that invade cortical marginal zone. Analyzing Nissl stained sections of human fetuses during the period from 8 to 14 postconceptional weeks (pcw), we observed, in the medial telencephalic wall, a stream of cells extending from the proliferative zones of cortical hem to the marginal zone (MZ) of the prospective archicortex. Golgi staining showed that this stream is composed of small unipolar migratory like cells. At corresponding ages in the macaque monkey fetuses (embryonic day/E 47-55), many cells in the proliferative zones of cortical hem were labeled for calbindin (CB). From this zone, a stream of unipolar migratory like CB-containing cells extended to the subpial position. In addition, at E47 few CB-containing neurons were observed in the thick MZ of the prospective archicortex. They were in apparent continuity with the stream of migratory like CB-containing cells extending from proliferative zone of cortical hem. At E55, CB-containing cells in the prospective hippocampus were organized in clusters. The clusters of CB-containing cells close to the proliferative zones were formed of immature cells, while more distant clusters included cells with typical pyramidal neuron body shape. During this period, GAD-containing migrating cells were mostly observed in the lateral wall, only very few were present in the medial telencephalic wall. Therefore, our results suggest that in primates the cortical hem gives rise to populations of non-GABA-ergic calbindin neurons, some of them destined to the hippocampus.

Cortical hem, a putative signaling centre at the interface of the prospective hippocampus and the choroid plexus, is a significant source of Cajal-Retzius cell population that invade cortical marginal zone. Analyzing Nissl stained sections of human fetuses during the period from 8 to 14 postconceptional weeks (pcw), we observed, in the medial telencephalic wall, a stream of cells extending from the proliferative zones of cortical hem to the marginal zone (MZ) of the prospective archicortex. Golgi staining showed that this stream is composed of small unipolar migratory like cells. At corresponding ages in the macaque monkey fetuses (embryonic day/E 47-55), many cells in the proliferative zones of cortical hem were labeled for calbindin (CB). From this zone, a stream of unipolar migratory like CB-containing cells extended to the subpial position. In addition, at E47 few CB-containing neurons were observed in the thick MZ of the prospective archicortex. They were in apparent continuity with the stream of migratory like CB-containing cells extending from proliferative zone of cortical hem. At E55, CB-containing cells in the prospective hippocampus were organized in clusters. The clusters of CB-containing cells close to the proliferative zones were formed of immature cells, while more distant clusters included cells with typical pyramidal neuron body shape. During this period, GAD-containing migrating cells were mostly observed in the lateral wall, only very few were present in the medial telencephalic wall. Therefore, our results suggest that in primates the cortical hem gives rise to populations of non-GABA-ergic calbindin neurons, some of them destined to the hippocampus.

Quantification of calretinin interneurons in the medial orbital region of the rat orbitofrontal cortex

Enduring, abnormal expression and function of the ion channel hyperpolarization-activated cyclic adenosine monophosphate gated channel type 1 (HCN1) occurs in temporal lobe epilepsy (TLE). We examined the underlying mechanisms, and investigated whether interfering with these mechanisms could modify disease course.Experimental TLE was provoked by kainic acid-induced status epilepticus (SE). HCN1 channel repression was examined at mRNA, protein, and functional levels. Chromatin immunoprecipitation was employed to identify the transcriptional mechanism of repressed HCN1 expression, and the basis for their endurance. Physical interaction of the repressor, NRSF, was abolished using decoy oligodeoxynucleotides (ODNs). Video/electroencephalographic recordings were performed to assess the onset and initial pattern of spontaneous seizures.Levels of NRSF and its physical binding to the Hcn1 gene were augmented after SE, resulting in repression of HCN1 expression and HCN1-mediated currents (I(h) ), and reduced I(h) -dependent resonance in hippocampal CA1 pyramidal cell dendrites. Chromatin changes typical of enduring, epigenetic gene repression were apparent at the Hcn1 gene within a week after SE. Administration of decoy ODNs comprising the NRSF DNA-binding sequence (neuron restrictive silencer element [NRSE]), in vitro and in vivo, reduced NRSF binding to Hcn1, prevented its repression, and restored I(h) function. In vivo, decoy NRSE ODN treatment restored theta rhythm and altered the initial pattern of spontaneous seizures.Acquired HCN1 channelopathy derives from NRSF-mediated transcriptional repression that endures via chromatin modification and may provide insight into the mechanisms of a number of channelopathies that coexist with, and may contribute to, the conversion of a normal brain into an epileptic one.

GABA development

Gamma-Aminobutyric acidergic (GABAergic) neurons that correspond mainly to local circuit neurons play a crucial role in regulating the activity of cortical neuronal networks and the complex interactions among principal neurons. These functions depend on a very heterogeneous population of neurons, whose number and diversity may increase during mammalian evolution. Interestingly dysfunction and cell death of several specific types of GABAergic neurons is a hallmark of various psychiatric and neurological disorders such as schizophrenia, autism and epilepsies to which defect in genesis or migration of these neurons can be a predisposing factor. Understanding primate specific developmental mechanisms that regulate generation, migration and maturation of cortical GABA neurons, is a crucial step in elucidating such pathologies. However, there are few such studies in monkeys and humans, whereas they are numerous in lower mammals. In rodents, cortical GABAergic neurons are generated exclusively in the subcortical proliferative zone of the ventral telencephalon, the ganglionic eminence (GE), and migrate tangentially to the dorsal telencephalon to reach their target cortical layer. We reported (Petanjek et al. 2009, Cerebral Cortex 19:249-62) that in contrast to rodents, in cynomolgus monkey cortical GABAergic neurons are generated massively in the proliferative zones of the dorsal telencephalon, in addition to the GE. This neurogenesis occurs very early during brain development in both dorsal and ventral telencephalon but with distinct temporal profiles. GABAergic neuron progenitors labeled for transcription factor Mash1 and GABA synthesizing enzyme GAD65 were present mainly in the GE at embryonic days (E) 47-55, and in the entire dorsal telencephalon at E64-75. These progenitors within the dorsal telencephalon are generated locally rather than in the GE. In the human embryonic forebrain (Letinic et al. 2002, Nature 417:645-9) retroviral labeling in organotypic slice cultures also demonstrated the existence of two distinct lineages of neocortical GABAergic neurons. One lineage expresses Dlx1/2 and Mash1 transcription factors, represents 65% of neocortical GABAergic neurons in humans, and originates from Mash1-expressing progenitors of the neocortical proliferative zones of the dorsal telencephalon. The second lineage, characterized by the expression of Dlx1/2 but not Mash1, forms in human brain only around 35% of the GABAergic neurons and originates from the GE of the ventral forebrain, in comparison to rodent where all GABAergic neurons originate in GE. The dorsal telencephalic origin of cortical GABA-ergic neurons is a unique feature of human and non-human primates and appears to be, in the primate lineage, an early evolutionary modification which provided increased number and variety of GABA-ergic neurons to an expanding neocortex.

Gamma-Aminobutyric acidergic (GABAergic) neurons that correspond mainly to local circuit neurons play a crucial role in regulating the activity of cortical neuronal networks and the complex interactions among principal neurons. These functions depend on a very heterogeneous population of neurons, whose number and diversity may increase during mammalian evolution. Interestingly dysfunction and cell death of several specific types of GABAergic neurons is a hallmark of various psychiatric and neurological disorders such as schizophrenia, autism and epilepsies to which defect in genesis or migration of these neurons can be a predisposing factor. Understanding primate specific developmental mechanisms that regulate generation, migration and maturation of cortical GABA neurons, is a crucial step in elucidating such pathologies. However, there are few such studies in monkeys and humans, whereas they are numerous in lower mammals. In rodents, cortical GABAergic neurons are generated exclusively in the subcortical proliferative zone of the ventral telencephalon, the ganglionic eminence (GE), and migrate tangentially to the dorsal telencephalon to reach their target cortical layer. We reported (Petanjek et al. 2009, Cerebral Cortex 19:249-62) that in contrast to rodents, in cynomolgus monkey cortical GABAergic neurons are generated massively in the proliferative zones of the dorsal telencephalon, in addition to the GE. This neurogenesis occurs very early during brain development in both dorsal and ventral telencephalon but with distinct temporal profiles. GABAergic neuron progenitors labeled for transcription factor Mash1 and GABA synthesizing enzyme GAD65 were present mainly in the GE at embryonic days (E) 47-55, and in the entire dorsal telencephalon at E64-75. These progenitors within the dorsal telencephalon are generated locally rather than in the GE. In the human embryonic forebrain (Letinic et al. 2002, Nature 417:645-9) retroviral labeling in organotypic slice cultures also demonstrated the existence of two distinct lineages of neocortical GABAergic neurons. One lineage expresses Dlx1/2 and Mash1 transcription factors, represents 65% of neocortical GABAergic neurons in humans, and originates from Mash1-expressing progenitors of the neocortical proliferative zones of the dorsal telencephalon. The second lineage, characterized by the expression of Dlx1/2 but not Mash1, forms in human brain only around 35% of the GABAergic neurons and originates from the GE of the ventral forebrain, in comparison to rodent where all GABAergic neurons originate in GE. The dorsal telencephalic origin of cortical GABA-ergic neurons is a unique feature of human and non-human primates and appears to be, in the primate lineage, an early evolutionary modification which provided increased number and variety of GABA-ergic neurons to an expanding neocortex.

Dysfunction and cell death of several specific types of GABAergic neurons is a hallmark of various psychiatric and neurological diseases such as schizophrenia and epilepsies to which defect in neurogenesis or migration of these neurons can be a predisposing factor. Understanding primate specific developmental mechanisms that regulate the generation, migration, and maturation of cortical GABA neurons is a crucial step to understand such pathologies. In this study, we demonstrate that in monkey as in human but in contrast to rodents, cortical GABA neurons are generated in the proliferative zones of the dorsal telencephalon, in addition to the proliferative region of the ventral telencephalon, the ganglionic eminence (GE). This neurogenesis occurs very early during brain development in both ventral and dorsal telencephalon but with distinct temporal profiles. It starts at the beginning of the main neurogenesis period in the ventral telencephalon and only 2 weeks later in the dorsal telencephalon. Strong arguments favor a direct induction of this later event in the prospective neocortex rather than to result from migration of progenitors from the GE. The ventral and dorsal lineages of cortical GABA neurons display different laminar distribution. Early generated GABA neurons from the GE mostly populate the marginal zone and subplate whereas cortical plate GABA neurons originate from both ventral and dorsal telencephalon. The dorsal telencephalic origin of cortical GABA neurons is a unique feature of human and non-human primates and appears as a former event in the ascent of evolution that could provide GABAergic neurons to an expending neocortex.

Telencephalic GABA-ergic interneurons are very heterogeneous population that differs by their morphology, neurochemical contents, specificity of their connections and their physiological properties. In rodent all hippocampal and almost all cortical GABA-ergic interneurons are generated in the ventral (basal, subcortical) proliferative zones, the ganglionic eminence, and migrate tangentially to their target regions. Evolution of the cortex increased extremely the distance that interneurons have to pass, so in human fetuses migration of interneurons becomes extremely complex and more vulnerable event then in any other species. This might explain why disturbances of the GABA-ergic network are found in majority of neurological and psychiatric disorders. To study migrational routes of prospective GABA-ergic interneurons we describe spatial and temporal patterns of tangentionaly and nonradialy migrating like cells on frontal sectioned slices of the human fetal brain in period from 13.5 to 22 weeks of gestation using Stensaas modification of the Golgi-Del Rio Hortega method. This material was obtained from the Zagreb Neuroembryological Collection. Tangentional and nonradial migrating cells have oval, up to 10 microns in length, cell body with one process. In the ganglionic eminence many cells in the stream lack, or have shorter process, as the cell body was smaller. Through all stages examined, we found densely packed tangentionally oriented (in parallel with ventricular surface) migrating like cells that form a continuous stream through whole subventricular zone of the ganglionic eminence. This stream continues on the ventral and dorsal border of ganglionic eminence to upper subventricular and lower intermediate zone of the telencephalic anlage. This stream could be followed to the most medial located telencephalic parts, but their density decreased. There were individual cells leaving it, more on some specific position: (1) many individual cells leave ganglionic eminence and enter directly to striatum ; (2) on the dorsal emerge from ganglionic eminence, group of cells change their direction in the way to cortex ; (3) on the level of olfactory tubercule a ventral stream is divided on two parts: one continuing to medial located septal region, and another continuing to olfactory tract. However, many tangentionaly and nonradialy oriented migratory like cells were observed in all other fetal layers, with respect to ventricular zone in the early stages analyzed. The density of tangetionaly migrating cells was highest in the oldest stage analyzed and the thickness of the stream has also increased. However, in opposite to earlier stages density of nonradialy migrating neurons in other cortical layers, principally subplate, has decreased. This might be explained by dilution regarding huge increase in volume of the subplate through this period. Increase in number of tangentionaly migrating like cells up to second half of gestation becomes more important in the view that radial migration has decreased substantionaly. This suggests that intensive production of interneurons continued into the second half of gestation, while the production of projection neurons is decreasing rapidly. So, the final establishment of cortical circuitry seems mostly to be connected with formation of inhibitory elements.

Fundamental differences regarding the generation and migration of cortical GABAergic neurons exist between rodents and human. In rodents, these interneurons are generated in the subcortical proliferative zone of the ventral telencephalon, the ganglionic eminence (GE), and migrate tangentially into the developing neocortex, whereas in human most of these neurons are produced in the proliferate region of the dorsal epencephalon. To clarify whether this developmental mechanism is human- or primate- specific, we studied sources and migratory pathways of cortical GABAergic neurons in the cynomolgus monkey. We performed immunohistochemical experiments using markers of GABAergic neuron progenitors (MASH1) and interneurons (GAD65 and GAD67) in fetuses from embryonic day 47 (E47) to E90 (term 165 days). From E47 to E55 precursors of GABAergic neurons expressing MASH1 and GAD65 were observed almost exclusively in the proliferative zones of the ventral telencephalon including the ganglionic eminence (GE) and adjacent septal region. Many post-mitotic migrating like GAD65-containing neurons that did not express MASH1 were present in the intermediate zone and at a lower extend in the marginal zone and subplate of the dorsal and medial wall of the telencephalon. From E67 to E75, numerous cells labeled for MASH1 were now observed in the proliferative ventricular (VZ) and sub-ventricular (SVZ) zones of the entire dorso-medial wall of the telencephalon and these cells did co-express GAD65. In these regions, numerous tangentially and non-tangential oriented GAD65-labeled migrating neurons were present. In the monkey as well as in human, cortical GABAergic neurons are massively generated in the proliferative cortical regions: the VZ and SVZ of the dorsal telencephalon, in addition to the proliferative sub-cortical regions and follow different migratory pathways that those described in rodents.

interneurons