Wnt5a promotes differentiation and development of adult-born neurons in the hippocampus by noncanonical Wnt signaling.

In the adult hippocampus, new neurons are generated in the dentate gyrus. The Wnt signaling pathway regulates this process, but little is known about the endogenous Wnt ligands involved. We investigated the role of Wnt5a on adult hippocampal neurogenesis. Wnt5a regulates neuronal morphogenesis during embryonic development, and maintains dendritic architecture of pyramidal neurons in the adult hippocampus. Here, we determined that Wnt5a knockdown in the mouse dentate gyrus by lentivirus-mediated shRNA impaired neuronal differentiation of progenitor cells, and reduced dendritic development of adult-born neurons. In cultured adult hippocampal progenitors (AHPs), Wnt5a knockdown reduced neuronal differentiation and morphological development of AHP-derived neurons, whereas treatment with Wnt5a had the opposite effect. Interestingly, no changes in astrocytic differentiation were observed in vivo or in vitro, suggesting that Wnt5a does not affect fate-commitment. By using specific inhibitors, we determined that Wnt5a signals through CaMKII to induce neurogenesis, and promotes dendritic development of newborn neurons through activating Wnt/JNK and Wnt/CaMKII signaling. Our results indicate Wnt5a as a niche factor in the adult hippocampus that promotes neuronal differentiation and development through activation of noncanonical Wnt signaling pathways.

ALS-linked protein disulfide isomerase variants cause motor dysfunction.

Disturbance of endoplasmic reticulum (ER) proteostasis is a common feature of amyotrophic lateral sclerosis (ALS). Protein disulfide isomerases (PDIs) areERfoldases identified as possibleALSbiomarkers, as well as neuroprotective factors. However, no functional studies have addressed their impact on the disease process. Here, we functionally characterized fourALS-linked mutations recently identified in two majorPDIgenes,PDIA1 andPDIA3/ERp57. Phenotypic screening in zebrafish revealed that the expression of thesePDIvariants induce motor defects associated with a disruption of motoneuron connectivity. Similarly, the expression of mutantPDIs impaired dendritic outgrowth in motoneuron cell culture models. Cellular and biochemical studies identified distinct molecular defects underlying the pathogenicity of thesePDImutants. Finally, targetingERp57 in the nervous system led to severe motor dysfunction in mice associated with a loss of neuromuscular synapses. This study identifiesERproteostasis imbalance as a risk factor forALS, driving initial stages of the disease.

Role of the Axon Initial Segment in the Control of Spontaneous Frequency of Nigral Dopaminergic Neurons In Vivo.

The spontaneous tonic discharge activity of nigral dopamine neurons plays a fundamental role in dopaminergic signaling. To investigate the role of neuronal morphology and architecture with respect to spontaneous activity in this population, we visualized the 3D structure of the axon initial segment (AIS) along with the entire somatodendritic domain of adult male mouse dopaminergic neurons, previously recorded in vivo We observed a positive correlation of the firing rate with both proximity and size of the AIS. Computational modeling showed that the size of the AIS, but not its position within the somatodendritic domain, is the major causal determinant of the tonic firing rate in the intact model, by virtue of the higher intrinsic frequency of the isolated AIS. Further mechanistic analysis of the relationship between neuronal morphology and firing rate showed that dopaminergic neurons function as a coupled oscillator whose frequency of discharge results from a compromise between AIS and somatodendritic oscillators. Thus, morphology plays a critical role in setting the basal tonic firing rate, which in turn could control striatal dopaminergic signaling that mediates motivation and movement.SIGNIFICANCE STATEMENT The frequency at which nigral dopamine neurons discharge action potentials sets baseline dopamine levels in the brain, which enables activity in motor, cognitive, and motivational systems. Here, we demonstrate that the size of the axon initial segment, a subcellular compartment responsible for initiating action potentials, is a key determinant of the firing rate in these neurons. The axon initial segment and all the molecular components that underlie its critical function may provide a novel target for the regulation of dopamine levels in the brain.

Morphological and Biophysical Determinants of the Intracellular and Extracellular Waveforms in Nigral Dopaminergic Neurons: A Computational Study.

Action potentials (APs) in nigral dopaminergic neurons often exhibit two separate components: the first reflecting spike initiation in the dendritically located axon initial segment (AIS) and the second the subsequent dendro-somatic spike. These components are separated by a notch in the ascending phase of the somatic extracellular waveform and in the temporal derivative of the somatic intracellular waveform. Still, considerable variability exists in the presence and magnitude of the notch across neurons. To systematically address the contribution of AIS, dendritic and somatic compartments to shaping the two-component APs, we modeled APs of previously in vivo electrophysiologically characterized and 3D-reconstructed male mouse and rat dopaminergic neurons. A parsimonious two-domain model, with high (AIS) and lower (dendro-somatic) Na+ conductance, reproduced the notch in the temporal derivatives, but not in the extracellular APs, regardless of morphology. The notch was only revealed when somatic active currents were reduced, constraining the model to three domains. Thus, an initial AIS spike is followed by an actively generated spike by the axon-bearing dendrite (ABD), in turn followed mostly passively by the soma. The transition from being a source compartment for the AIS spike to a source compartment for the ABD spike satisfactorily explains the extracellular somatic notch. Larger AISs and thinner ABD (but not soma-to-AIS distance) accentuate the AIS component. We conclude that variability in AIS size and ABD caliber explains variability in AP extracellular waveform and separation of AIS and dendro-somatic components, given the presence of at least three functional domains with distinct excitability characteristics.SIGNIFICANCE STATEMENT Midbrain dopamine neurons make an important contribution to circuits mediating motivation and movement. Understanding the basic rules that govern the electrical activity of single dopaminergic neurons is therefore essential to reveal how they ultimately contribute to movement and motivation as well as what goes wrong in associated disorders. Our computational study focuses on the generation and propagation of action potentials and shows that different morphologies and excitability characteristics of the cell body, dendrites and proximal axon can explain the diversity of action potentials shapes in this population. These compartments likely make differential contributions both to normal dopaminergic signaling and could potentially underlie pathological dopaminergic signaling implicated in addiction, schizophrenia, Parkinson's disease, and other disorders.

Differential Somatic Ca2+ Channel Profile in Midbrain Dopaminergic Neurons.

UNLABELLED: Dopaminergic (DA) neurons located in the ventral midbrain continuously generate a slow endogenous pacemaker activity, the mechanism of which is still debated. It has been suggested that, in the substantia nigra pars compacta (SNc), the pacemaking relies more on Ca(2+) channels and that the density of L-type Ca(2+) channels is higher in these DA neurons than in those located in the ventral tegmental area (VTA). This might lead to a higher Ca(2+) load in SNc DA neurons and explain their higher susceptibility to degeneration. However, direct evidence for this hypothesis is lacking. We found that the L-type current and channel density are indeed higher in the somata of rat SNc DA neurons and that this current undergoes less inactivation in this region. Nonstationary fluctuation analysis measurements showed a much higher number of L-type channels in the soma of SNc DA neurons, as well as a smaller single-channel conductance, pointing to a possible different molecular identity of L-type channels in DA neurons from the two areas. A major consequence of this is that pacemaking and, even more so, bursting are associated with a larger Ca(2+) entry through L-type channels in SNc DA neurons than in their VTA counterparts. Our results establish a molecular and functional difference between two populations of midbrain DA neurons that may contribute to their differential sensitivity to neurodegeneration. SIGNIFICANCE STATEMENT: Dopamine neurons from the substantia nigra pars compacta (SNc) and ventral tegmental area (VTA) are involved in various brain functions, such as movement initiation and goal directed behavior, respectively. This work shows that, although both neurons fire in a similar regular and slow pacemaker mode, this firing activity is supported by different calcium channel landscapes. Indeed, the L-type calcium current is larger in the soma of dopamine neurons of the SNc, leading to a higher charge transfer through L-type channels during pacemaking and bursting. Therefore, these neurons may be physiologically exposed to a larger stress than their neighbors from the VTA.

Mechanisms of Kappa Opioid Receptor Potentiation of Dopamine D2 Receptor Function in Quinpirole-Induced Locomotor Sensitization in Rats.

Background: Increased locomotor activity in response to the same stimulus is an index of behavioral sensitization observed in preclinical models of drug addiction and compulsive behaviors. Repeated administration of quinpirole, a D2/D3 dopamine agonist, induces locomotor sensitization. This effect is potentiated and accelerated by co-administration of U69593, a kappa opioid receptor agonist. The mechanism underlying kappa opioid receptor potentiation of quinpirole-induced locomotor sensitization remains to be elucidated. Methods: Immunofluorescence anatomical studies were undertaken in mice brain slices and rat presynaptic synaptosomes to reveal kappa opioid receptor and D2R pre- and postsynaptic colocalization in the nucleus accumbens. Tonic and phasic dopamine release in the nucleus accumbens of rats repeatedly treated with U69593 and quinpirole was assessed by microdialysis and fast scan cyclic voltammetry. Results: Anatomical data show that kappa opioid receptor and D2R colocalize postsynaptically in medium spiny neurons of the nucleus accumbens and the highest presynaptic colocalization occurs on the same dopamine terminals. Significantly reduced dopamine levels were observed in quinpirole, and U69593-quinpirole treated rats, explaining sensitization of D2R. Presynaptic inhibition induced by kappa opioid receptor and D2R of electrically evoked dopamine release was faster in U69593-quinpirole compared with quinpirole-repeatedly treated rats. Conclusions: Pre- and postsynaptic colocalization of kappa opioid receptor and D2R supports a role for kappa opioid receptor potentiating both the D2R inhibitory autoreceptor function and the inhibitory action of D2R on efferent medium spiny neurons. Kappa opioid receptor co-activation accelerates D2R sensitization by contributing to decrease dopamine release in the nucleus accumbens.

Applying the area fraction fractionator (AFF) probe for total volume estimations of somatic, dendritic and axonal domains of the nigrostriatal dopaminergic system in a murine model.

BACKGROUND: The Cavalieri estimator is used for volume measurement of brain and brain regions. Derived from this estimator is the Area Fraction Fractionator (AFF), used for efficient area and number estimations of small 2D elements, such as axons in cross-sectioned nerves. However, to our knowledge, the AFF has not been combined with serial sectioning analysis to measure the volume of small-size nervous structures. NEW METHOD: Using the nigrostriatal dopaminergic system as an illustrative case, we describe a protocol based on Cavalieri's principle and AFF to estimate the volume of its somatic, nuclear, dendritic, axonal and axon terminal cellular compartments in the adult mouse. The protocol consists of (1) systematic random sampling of sites within and across sections in regions of interest (substantia nigra, the nigrostriatal tract, caudate-putamen), (2) confocal image acquisition of sites, (3) marking of cellular domains using Cavalieri's 2D point-counting grids, and 4) determination of compartments' total volume using the estimated area of each compartment, and between-sections distance. RESULTS: The volume of the nigrostriatal system per hemisphere is ∼0.38 mm3, with ∼5 % corresponding to perikarya and cell nuclei, ∼10 % to neuropil/dendrites, and ∼85 % to axons and varicosities. COMPARISON WITH EXISTING METHODS: In contrast to other methods to measure volume of discrete objects, such as the optical nucleator or 3D reconstructions, it stands out for its versatility and ease of use. CONCLUSIONS: The use of a simple quantitative, unbiased approach to assess the global state of a system may allow quantification of compartment-specific changes that may accompany neurodegenerative processes.

Differences in vocal brain areas and astrocytes between the house wren and the rufous-tailed hummingbird.

The house wren shows complex song, and the rufous-tailed hummingbird has a simple song. The location of vocal brain areas supports the song's complexity; however, these still need to be studied. The astrocytic population in songbirds appears to be associated with change in vocal control nuclei; however, astrocytic distribution and morphology have not been described in these species. Consequently, we compared the distribution and volume of the vocal brain areas: HVC, RA, Area X, and LMAN, cell density, and the morphology of astrocytes in the house wren and the rufous-tailed hummingbird. Individuals of the two species were collected, and their brains were analyzed using serial Nissl- NeuN- and MAP2-stained tissue scanner imaging, followed by 3D reconstructions of the vocal areas; and GFAP and S100ß astrocytes were analyzed in both species. We found that vocal areas were located close to the cerebral midline in the house wren and a more lateralized position in the rufous-tailed hummingbird. The LMAN occupied a larger volume in the rufous-tailed hummingbird, while the RA and HVC were larger in the house wren. While Area X showed higher cell density in the house wren than the rufous-tailed hummingbird, the LMAN showed a higher density in the rufous-tailed hummingbird. In the house wren, GFAP astrocytes in the same bregma where the vocal areas were located were observed at the laminar edge of the pallium (LEP) and in the vascular region, as well as in vocal motor relay regions in the pallidum and mesencephalon. In contrast, GFAP astrocytes were found in LEP, but not in the pallidum and mesencephalon in hummingbirds. Finally, when comparing GFAP astrocytes in the LEP region of both species, house wren astrocytes exhibited significantly more complex morphology than those of the rufous-tailed hummingbird. These findings suggest a difference in the location and cellular density of vocal circuits, as well as morphology of GFAP astrocytes between the house wren and the rufous-tailed hummingbird.

Susceptibility to express amphetamine locomotor sensitization correlates with dorsolateral striatum bursting activity and GABAergic synapses in the globus pallidus.

Repeated psychostimulant administration results in behavioral sensitization, a process that is relevant in the early phases of drug addiction. Critically, behavioral sensitization is not observed in all subjects. Evidence shows that differential neuronal activity in the dorsolateral striatum (DLS) accompanies the expression of amphetamine (AMPH) locomotor sensitization. However, whether individual differences in DLS activity previous to AMPH administration can predict the expression of locomotor sensitization has not been assessed. Here, we examined DLS neuronal activity before and after repeated AMPH administration and related it to the susceptibility of rats to sensitize. For that, single-unit recordings on DLS medium spiny neurons (MSNs) were carried out in freely moving male Sprague Dawley rats during repeated AMPH administration. We also examined differences in neurostructure that could accompany sensitization. We quantified the density of the inhibitory postsynaptic marker gephyrin (Geph) in the entopeduncular nucleus (EP) and globus pallidus (GP). A higher burst firing and a lower percentage of correlation between MSNs post-Saline firing rate vs. locomotion predicted the expression of locomotor sensitization. Moreover, during the AMPH challenge, we observed that burst firing decreased in sensitized rats, in contrast to non-sensitized rats in which burst firing was maintained. Finally, a higher Geph density on GP but not EP was observed in non-sensitized rats after AMPH challenge. These results indicate that initial differences in DLS burst firing might underlie the susceptibility to express locomotor sensitization and suggest that the potentiation of dorsal striatum indirect pathway could be considered a protective mechanism to locomotor sensitization.

Primary germinoma of the medulla oblongata: illustrative case.

BACKGROUND: Primary central nervous system germinomas of the medulla oblongata are extremely rare and usually have been found in young female Asian patients. The authors present an illustrative case of a patient who presented with severe medullary and posterior cord syndrome, the first South American case published to date, to the authors' knowledge. OBSERVATIONS: Initially, the radiological differential diagnosis did not include this entity. The lesion was located at the obex and exhibited a well-delineated contrast enhancement without hydrocephalus. An emergency decompressive partial resection following functional limits was performed. After histological confirmation, radiotherapy was indicated, with complete remission achieved at a 6-month follow-up. The patient, however, continued to have a severe proprioceptive disorder. The literature review identified 21 other such patients. The mean age for this location was 23 years, with a strong female and Asian origin predilection. All tumors exhibited contrast enhancement, and only one presented with hydrocephalus. LESSONS: In the absence of elevated tumor markers, radiological clues such as a well-delineated, contrast-enhanced lesion arising from the obex, without hydrocephalus, associated with demographic features such as young age, female sex, and Asian heritage, should evoke a high level of suspicion for this diagnosis. Gross total resection must not be attempted, because this tumor is potentially curable with high-dose radiotherapy.

Immunolocalization of kappa opioid receptors in the axon initial segment of a group of embryonic mesencephalic dopamine neurons.

The dopamine mesolimbic system is a major circuit involved in controlling goal-directed behaviors. Dopamine D2 receptors (D2R) and kappa opioid receptors (KOR) are abundant Gi protein-coupled receptors in the mesolimbic system. D2R and KOR share several functions in dopamine mesencephalic neurons, such as regulation of dopamine release and uptake, and firing of dopamine neurons. In addition, KOR and D2R modulate each other functioning. This evidence indicates that both receptors functionally interact, however, their colocalization in the mesostriatal system has not been addressed. Immunofluorescent assays were performed in cultured dopamine neurons and adult mice's brain tissue to answer this question. We observed that KOR and D2R are present in similar density in dendrites and soma of cultured dopamine neurons, but in a segregated manner. Interestingly, KOR immunolabelling was observed in the first part of the axon, colocalizing with Ankyrin in 20% of cultured dopamine neurons, indicative that KOR is present in the axon initial segment (AIS) of a group of dopaminergic neurons. In the adult brain, KOR and D2R are also segregated in striatal tissue. While the KOR label is in fiber tracts such as the striatal streaks, corpus callosum, and anterior commissure, D2R is located mainly within the striatum and nucleus accumbens, surrounding fiber tracts. D2R is also localized in some fibers that are mostly different from those positives for KOR. In conclusion, KOR and D2R are present in the soma and dendrites of mesencephalic dopaminergic neurons, but KOR is also found in the AIS of a subpopulation of these neurons.

Dendritic Architecture Predicts in vivo Firing Pattern in Mouse Ventral Tegmental Area and Substantia Nigra Dopaminergic Neurons.

The firing activity of ventral tegmental area (VTA) and substantia nigra pars compacta (SNc) dopaminergic (DA) neurons is an important factor in shaping DA release and its role in motivated behavior. Dendrites in DA neurons are the main postsynaptic compartment and, along with cell body and axon initial segment, contribute to action potential generation and firing pattern. In this study, the organization of the dendritic domain in individual VTA and SNc DA neurons of adult male mice, and their relationship to in vivo spontaneous firing, are described. In comparison with dorsal VTA DA neurons, ventrally located VTA neurons (as measured by cell body location) possess a shorter total dendritic length and simpler dendritic architecture, and exhibit the most irregular in vivo firing patterns among DA neurons. In contrast, for DA neurons in the SNc, the higher irregularity of firing was related to a smaller dendritic domain, as measured by convex hull volumes. However, firing properties were also related to the specific regional distribution of the dendritic tree. Thus, VTA DA neurons with a larger extension of their dendritic tree within the parabrachial pigmented (PBP) nucleus fired more regularly compared with those with relatively more dendrites extending outside the PBP. For DA neurons in the SNc, enhanced firing irregularity was associated with a smaller proportion of dendrites penetrating the substantia nigra pars reticulata. These results suggest that differences in dendritic morphology contribute to the in vivo firing properties of individual DA neurons, and that the existence of region-specific synaptic connectivity rules that shape firing diversity.

Organization of the Catecholaminergic System in the Short-Lived Fish Nothobranchius furzeri.

The catecholaminergic system has received much attention based on its regulatory role in a wide range of brain functions and its relevance in aging and neurodegenerative diseases. In the present study, we analyzed the neuroanatomical distribution of catecholaminergic neurons based on tyrosine hydroxylase (TH) immunoreactivity in the brain of adult Nothobranchius furzeri. In the telencephalon, numerous TH+ neurons were observed in the olfactory bulbs and the ventral telencephalic area, arranged as strips extending through the rostrocaudal axis. We found the largest TH+ groups in the diencephalon at the preoptic region level, the ventral thalamus, the pretectal region, the posterior tuberculum, and the caudal hypothalamus. In the dorsal mesencephalic tegmentum, we identified a particular catecholaminergic group. The rostral rhombencephalon housed TH+ cells in the locus coeruleus and the medulla oblongata, distributing in a region dorsal to the inferior reticular formation, the vagal lobe, and the area postrema. Finally, scattered TH+ neurons were present in the ventral spinal cord and the retina. From a comparative perspective, the overall organization of catecholaminergic neurons is consistent with the general pattern reported for other teleosts. However, N. furzeri shows some particular features, including the presence of catecholaminergic cells in the midbrain. This work provides a detailed neuroanatomical map of the catecholaminergic system of N. furzeri, a powerful aging model, also contributing to the phylogenetic understanding of one of the most ancient neurochemical systems.

Discharge profiles of identified GABAergic in comparison to cholinergic and putative glutamatergic basal forebrain neurons across the sleep-wake cycle.

Whereas basal forebrain (BF) cholinergic neurons are known to participate in processes of cortical activation during wake (W) and paradoxical sleep (PS or P, also called REM sleep), codistributed GABAergic neurons have been thought to participate in processes of cortical deactivation and slow-wave sleep (SWS or S). To learn the roles the GABAergic neurons might play, in relation to cholinergic and glutamatergic neurons, we juxtacellularly recorded and labeled neurons during natural sleep-wake states in head-fixed rats. Neurobiotin (Nb)-labeled cells were identified immunohistochemically as choline acetyltransferase (ChAT)+, glutamic acid decarboxylase (GAD)+, or ChAT-/GAD-. Of the latter, some were identified as glutamatergic by immunostaining of their terminals with the vesicular glutamate transporter (VGluT2). In contrast to ChAT+ neurons, which all discharged maximally during W and PS, GAD+ neurons comprised multiple sleep-wake subgroups. Some GABAergic neurons discharged maximally during W and PS, as WP-max active cells (36%), and in positive correlation with gamma electroencephalographic (EEG) activity. Some discharged maximally during SWS, as S-max active cells (28%), and in positive correlation with delta EEG activity. Others increased their discharge progressively during sleep to discharge maximally during PS, as P-max active cells (36%), and in negative association with electromyographic (EMG) activity. ChAT-/GAD- cells comprised WP-max (46%), S-max (17%), P-max (17%), and W-max active cells (14%), whose discharge was positively correlated with EMG activity. GABAergic neurons would thus play similar or reciprocal roles to other cholinergic and glutamatergic BF neurons in regulating cortical activity and muscle tone along with behavior across sleep-wake states.

Activity of neurochemically heterogeneous dopaminergic neurons in the substantia nigra during spontaneous and driven changes in brain state.

Dopaminergic neurons of the substantia nigra (SN) and ventral tegmental area (VTA) are collectively implicated in motor- and reward-related behaviors. However, dopaminergic SN and VTA neurons differ on several functional levels, and dopaminergic SN neurons themselves vary in their intrinsic electrical properties, neurochemical characteristics and connections. This heterogeneity is not only important for normal function; calbindin (CB) expression by some dopaminergic SN neurons has been linked with their increased survival in Parkinson's disease. To test whether the activity of CB-negative and CB-positive dopaminergic SN neurons differs during distinct spontaneous and driven brain states, we recorded single units in anesthetized rats before, during and after aversive somatosensory stimuli. Recorded neurons were juxtacellularly labeled, confirmed to be dopaminergic, and tested for CB immunoreactivity. During cortical slow-wave activity, the firing of most dopaminergic neurons was slow and regular/irregular and unrelated to cortical slow oscillations. During spontaneous cortical activation, dopaminergic SN neurons fired in a more regular manner, with fewer bursts, but did not change their firing rate. Regardless of brain state, CB-negative dopaminergic neurons fired significantly faster than CB-positive dopaminergic neurons. This difference in firing rate was not mirrored by different firing patterns. Most CB-negative and CB-positive dopaminergic neurons did not respond to the aversive stimuli; of those that did respond, most were inhibited. We conclude that CB-negative and CB-positive dopaminergic neurons exhibit different activities in vivo. Furthermore, the firing of dopaminergic SN neurons is brain state-dependent, and, unlike dopaminergic VTA neurons, they are not commonly recruited or inhibited by aversive stimuli.

GABAergic neurons intermingled with orexin and MCH neurons in the lateral hypothalamus discharge maximally during sleep.

The lateral hypothalamus (LH), where wake-active orexin (Orx)-containing neurons are located, has been considered a waking center. Yet, melanin-concentrating hormone (MCH)-containing neurons are codistributed therein with Orx neurons and, in contrast to them, are active during sleep, not waking. In the present study employing juxtacellular recording and labeling of neurons with Neurobiotin (Nb) in naturally sleeping-waking head-fixed rats, we identified another population of intermingled sleep-active cells, which do not contain MCH (or Orx), but utilize gamma-aminobutyric acid (GABA) as a neurotransmitter. The 'sleep-max' active neurons represented 53% of Nb-labeled MCH-(and Orx) immunonegative (-) cells recorded in the LH. For identification of their neurotransmitter, Nb-labeled varicosities of the Nb-labeled/MCH- neurons were sought within sections adjacent to the Nb-labeled soma and immunostained for the vesicular transporter for GABA (VGAT) or for glutamate. A small proportion of sleep-max Nb+/MCH- neurons (19%) discharged maximally during slow-wave sleep (called 'S-max') in positive correlation with delta electroencephalogram activity, and from VGAT staining of Nb-labeled varicosities appeared to be GABAergic. The vast proportion of sleep-max Nb+/MCH- neurons (81%) discharged maximally during paradoxical sleep (PS, called 'P-max') in negative correlation with electromyogram amplitude, and from Nb-labeled varicosities also appeared to be predominantly GABAergic. Given their discharge profiles across the sleep-wake cycle, P-max together with S-max GABAergic neurons could thus serve to inhibit other neurons of the arousal systems, including local Orx neurons in the LH. They could accordingly dampen arousal with muscle tone and promote sleep, including PS with muscle atonia.

Projections from basal forebrain to prefrontal cortex comprise cholinergic, GABAergic and glutamatergic inputs to pyramidal cells or interneurons.

The present study was undertaken to characterize the pre- and postsynaptic constituents of the basal forebrain (BF) projection to the prefrontal cortex in the rat, and determine whether it includes glutamatergic in addition to established gamma-aminobutyric acid (GABA)ergic and cholinergic elements. BF fibres were labelled by anterograde transport using biotin dextran amine (BDA) and dual-stained for the vesicular transporter proteins (VTPs) for glutamate (VGluT), GABA (VGAT) or acetylcholine (VAChT). Viewed by fluorescence microscopy and estimated by stereology, proportions of BDA-labelled varicosities were found to be stained for VGluT2 (and not VGluT1 or 3), VGAT or VAChT (representing, respectively, approximately 15%, approximately 52% and approximately 19% within the infralimbic cortex). Each type was present in all, though commonly most densely in deep, cortical layers. Material was triple-stained for postsynaptic proteins to examine whether BDA+VTP+ varicosities might form excitatory or inhibitory synapses, respectively, labelled by postsynaptic density-95 kDA (PSD-95) or gephyrin (Geph). Viewed by confocal microscopy, a majority of BDA+/VGluT2+ varicosities were found to be apposed to PSD-95+ elements, and a majority of BDA+/VGAT+ varicosities to be apposed to Geph+ elements. Other series were triple-stained for cell marker proteins to assess whether the varicosities contacted interneurons or pyramidal cells. Viewed by confocal microscopy, BDA-labelled VGluT2+, VGAT+ and VAChT+ BF terminals were all found in contact with calbindin+ interneurons, whereas VGAT+ BF terminals were also seen in contact with parvalbumin+ interneurons and non-phosphorylated neurofilament+ pyramidal cells. Through distinct glutamatergic, GABAergic and cholinergic projections, the BF can thus influence cortical activity in a diverse manner.

Estudio dimensional de los puntos craneométricos mediosagitales en cráneos de población chilena / Dimensional study of midsagital craneometric points in skulls of Chilean population

RESUMEN: La utilidad de los puntos craneométricos en neurocirugía radica en la estudiada relación que poseen con las estructuras encefálicas y vasculares que subyacen, siendo relevantes al momento de planificar y realizar diversos abordajes y disminuir la morbilidad asociada. A pesar de esto, hasta el momento no se disponen de datos publicados de las distancias de estos puntos craneométricos en población chilena. Se utilizaron 45 cráneos secos de cadáveres adultos. Se realizaron mediciones de las distancias superficiales entre diversos puntos craneométricos mediosagitales. La distancia superficial promedio entre nasion y bregma fue de 12,71 cm; entre bregma y lambda, 12,34 cm; entre lambda e inion, 6,64 cm; entre inion y opistocranio, 5,16 cm; entre lambda y opistocráneo, 3,88 cm y entre inion y opistion, 5,16 cm. Se encontraron diferencias estadísticamente significativas (p<0,05) entre el grupo estudiado con datos obtenidos de la literatura en las distancias nasion-bregma, bregma-lambda, lambda-opistocranio. Las distancias entre los distintos puntos craneométricos en cráneos de población chilena fueron caracterizadas en el presente estudio existiendo algunas diferencias con los datos de la literatura que deben ser considerados en el contexto de la práctica quirúrgica relacionada.

Characterization of the axon initial segment of mice substantia nigra dopaminergic neurons.

The axon initial segment (AIS) is the site of initiation of action potentials and influences action potential waveform, firing pattern, and rate. In view of the fundamental aspects of motor function and behavior that depend on the firing of substantia nigra pars compacta (SNc) dopaminergic neurons, we identified and characterized their AIS in the mouse. Immunostaining for tyrosine hydroxylase (TH), sodium channels (Nav ) and ankyrin-G (Ank-G) was used to visualize the AIS of dopaminergic neurons. Reconstructions of sampled AIS of dopaminergic neurons revealed variable lengths (12-60 µm) and diameters (0.2-0.8 µm), and an average of 50% reduction in diameter between their widest and thinnest parts. Ultrastructural analysis revealed submembranous localization of Ank-G at nodes of Ranvier and AIS. Serial ultrathin section analysis and 3D reconstructions revealed that Ank-G colocalized with TH only at the AIS. Few cases of synaptic innervation of the AIS of dopaminergic neurons were observed. mRNA in situ hybridization of brain-specific Nav subunits revealed the expression of Nav 1.2 by most SNc neurons and a small proportion expressing Nav 1.6. The presence of sodium channels, along with the submembranous location of Ank-G is consistent with the role of AIS in action potential generation. Differences in the size of the AIS likely underlie differences in firing pattern, while the tapering diameter of AIS may define a trigger zone for action potentials. Finally, the conspicuous expression of Nav 1.2 by the majority of dopaminergic neurons may explain their high threshold for firing and their low discharge rate.

On the Role of DT-Diaphorase Inhibition in Aminochrome-Induced Neurotoxicity In Vivo.

Dopamine oxidation in the pathway leading to neuromelanin formation generates the ortho-quinone aminochrome, which is potentially neurotoxic but normally rapidly converted by DT-diaphorase to nontoxic leukoaminochrome. However, when administered exogenously into rat striatum, aminochrome is able to produce damage to dopaminergic neurons. Because of a recent report that substantia nigra pars compacta (SNpc) tyrosine hydroxylase (T-OH) levels were unaltered by aminochrome when there was cell shrinkage of dopaminergic neurons along with a reduction in striatal dopamine release, the following study was conducted to more accurately determine the role of DT-diaphorase in aminochrome neurotoxicity. In this study, a low dose of aminochrome (0.8 nmol) with or without the DT-diaphorase inhibitor dicoumarol (0.2 nmol) was injected into the left striatum of rats. Intrastriatal 6-hydroxydopamine (6-OHDA, 32 nmol) was used as a positive neurotoxin control in other rats. Two weeks later, there was significant loss in numbers of T-OH immunoreactive fibers in SNpc, also a loss in cell density in SNpc, and prominent apomorphine (0.5 mg/kg sc)-induced contralateral rotations in rats that had been treated with aminochrome, with aminochrome/dicoumarol, or with 6-OHDA. Findings demonstrate that neurotoxic aminochrome is able to exert neurotoxicity only when DT-diaphorase is suppressed-implying that DT-diaphorase is vital in normally suppressing toxicity of in vivo aminochrome, generated in the pathway towards neuromelanin formation.