Glutamatergic axons from the lateral habenula mainly terminate on GABAergic neurons of the ventral midbrain

The concept of cortical-subcortical loops emphasizes the importance of the basal ganglia for motor, psychomotor, and emotional cortical functions. These loops are bidirectionally controlled by the midbrain dopaminergic system, predominantly but not exclusively at the level of the striatum including the accumbens nucleus. Successful behaviors increase the activities of the mesostriatal (arising in the complex part of the substantia nigra) and mesolimbic (arising in the ventral tegmental area, VTA) neurons, thereby reinforcing the corresponding actions. In contrast, unsuccessful behaviors result in an increased activation of the lateral habenular complex (LHb), thereby decreasing the activities of mesolimbic neurons. Correspondingly, electrical stimulation of the LHb effectively blocks neuronal activity in the VTA. Whether this block is due to an inhibitory projection from the LHb to the VTA, or whether axons from excitatory LHb neurons target inhibitory neurons within the VTA, is presently not known. Here we show, using in situ hybridization and immunocytochemical double labeling at the light and electron microscopic level, that GABAergic neurons are scarce in the LHb and that glutamatergic axons from the LHb mostly target GABAergic neurons in the VTA and the mesopontine rostromedial tegmental nucleus (RMTg), also known as tail of the VTA (tVTA). These data explain the inhibitory effect of LHb activation on the VTA. In addition, however, a small number of LHb terminals in the VTA actually contacts dopaminergic neurons. The biological importance of these terminals requires further investigation.     

Differential expression of vesicular glutamate transporters 1 and 2 may identify distinct modes of glutamatergic transmission in the macaque visual system

Glutamate is the primary neurotransmitter utilized by the mammalian visual system for excitatory neurotransmission. The sequestration of glutamate into synaptic vesicles, and the subsequent transport of filled vesicles to the presynaptic terminal membrane, is regulated by a family of proteins known as vesicular glutamate transporters (VGLUTs). Two VGLUT proteins, VGLUT1 and VGLUT2, characterize distinct sets of glutamatergic projections between visual structures in rodents and prosimian primates, yet little is known about their distributions in the visual system of anthropoid primates. We have examined the mRNA and protein expression patterns of VGLUT1 and VGLUT2 in the visual system of macaque monkeys, an Old World anthropoid primate, in order to determine their relative distributions in the superior colliculus, lateral geniculate nucleus, pulvinar complex, V1 and V2. Distinct expression patterns for both VGLUT1 and VGLUT2 identified architectonic boundaries in all structures, as well as anatomical subdivisions of the superior colliculus, pulvinar complex, and V1. These results suggest that VGLUT1 and VGLUT2 clearly identify regions of glutamatergic input in visual structures, and may identify common architectonic features of visual areas and nuclei across the primate radiation. Additionally, we find that VGLUT1 and VGLUT2 characterize distinct subsets of glutamatergic projections in the macaque visual system; VGLUT2 predominates in driving or feedforward projections from lower order to higher order visual structures while VGLUT1 predominates in modulatory or feedback projections from higher order to lower order visual structures. The distribution of these two proteins suggests that VGLUT1 and VGLUT2 may identify class 1 and class 2 type glutamatergic projections within the primate visual system (Sherman and Guillery, 2006).     

Low accumulation of drebrin at glutamatergic postsynaptic sites on GABAergic neurons

Glutamatergic synapses form onto both glutamatergic and GABAergic neurons. These two types of glutamatergic synapses differ in their electrical responses to high-frequency stimulation and postsynaptic density protein composition. However, it is not known whether they differ in the actin cytoskeleton composition. In the present study, we used hippocampal neuronal cultures prepared from glutamate decarboxylase 67 (GAD67)-GFP knock-in mice and analyzed the differences in the actin cytoskeleton at glutamatergic synapses contacting GABAergic and glutamatergic neurons. Drebrin-binding actin filaments enriched in dendritic spines are known to play a pivotal role in spine formation. Immunocytochemical analyses demonstrated that drebrin accumulated at glutamatergic synapses on GABAergic neurons as well as at those on glutamatergic neurons. However, the density of drebrin clusters along dendrites in GABAergic neurons was significantly lower than those of glutamatergic neurons. Furthermore, the level of drebrin accumulating at glutamatergic synapses was lower on GABAergic neurons than on glutamatergic neurons. In neurons overexpressing drebrin, drebrin cluster density and accumulation levels in GABAergic and glutamatergic neurons were similar, suggesting that the low drebrin levels in the glutamatergic postsynaptic sites on GABAergic neurons may be because GABAergic neurons express low levels of drebrin. On the other hand, pharmacological analysis demonstrated that the postsynaptic localization of drebrin depended on actin cytoskeleton organization in both GABAergic and glutamatergic neurons. Together these results indicated that, although GABAergic and glutamatergic neurons share common regulatory systems affecting drebrin localization, the density of drebrin-positive glutamatergic synapses formed on GABAergic neurons is lower than those on glutamatergic neurons. This is probably due to the low expression of drebrin in GABAergic neurons.     

Brain-wide mapping of efferent projections of glutamatergic (Onecut3+) neurons in the lateral mouse hypothalamus

Aim

This study mapped the spatiotemporal positions and connectivity of Onecut3⁺ neuronal populations in the developing and adult mouse brain.

Methods

We generated fluorescent reporter mice to chart Onecut3⁺ neurons for brain-wide analysis. Moreover, we crossed Onecut3-iCre and Mapt-mGFP (Tau-mGFP) mice to visualize axonal projections. A dual Cre/Flp-dependent AAV construct in Onecut3-iCre cross-bred with Slc17a6-FLPo mice was used in an intersectional strategy to map the connectivity of glutamatergic lateral hypothalamic neurons in the adult mouse.

Results

We first found that Onecut3 marks a hitherto undescribed Slc17a6⁺/Vglut2⁺ neuronal cohort in the lateral hypothalamus, with the majority expressing thyrotropin-releasing hormone. In the adult, Onecut3⁺/Vglut2⁺ neurons of the lateral hypothalamus had both intra- and extrahypothalamic efferents, particularly to the septal complex and habenula, where they targeted other cohorts of Onecut3⁺ neurons and additionally to the neocortex and hippocampus. This arrangement suggests that intrinsic reinforcement loops could exist for Onecut3⁺ neurons to coordinate their activity along the brain's midline axis.

Conclusion

We present both a toolbox to manipulate novel subtypes of hypothalamic neurons and an anatomical arrangement by which extrahypothalamic targets can be simultaneously entrained.     

Stress-induced,glucocorticoid-dependent strengthening of glutamatergic synaptic transmission in midbrain dopamine neurons

Stress facilitates development of addictive behaviors in part by stress-induced increase in the strength of glutamatergic synapses at dopamine (DA) neurons within the ventral tegmental area (VTA). Here, we further demonstrate that this stress-induced synaptic adaptation is glucocorticoid-dependent and is progressively developed. Activation of glucocorticoid receptors (GRs) either by in vivo injection of dexamethasone (Dex) or incubation of the VTA slice with Dex potentiate the synaptic strength of glutamatergic synapses at VTA DA neurons, whereas preventing the activation of GRs by Ru486 abolishes this effect. These results suggest that the VTA GRs play a critical role in stress-induced cellular adaptations.     

APPswe转基因小鼠海马结构中谷氨酸和γ-氨基丁酸能神经元的数量变化

目的:探讨谷氨酸和γ-氨基丁酸在阿尔茨海默病(AD)发生、发展中的作用.方法:利用硫磺素S染色技术检测APPswe转基因小鼠脑内沉积的老年斑;免疫荧光技术标记正常及APPswe转基因小鼠海马发育过程中谷氨酸、γ-氨基丁酸(GABA)阳性细胞. 结果: 模型组小鼠海马CA1、CA3区以及齿状回谷氨酸阳性细胞的数量明显少于正常对照组,而海马各区的GABA阳性细胞在两者之间变化不明显.结论: 谷氨酸与GABA能神经元数量的改变与Aβ诱导的神经元凋亡以及AD的病理发生有关.     

Effects of medial olivocochlear efferent stimulation on the activity of neurons in the auditory midbrain

Medial olivocochlear (MOC) efferents are known to suppress spontaneous activity and sound-evoked responses of primary afferents by their actions on outer hair cells in the cochlea. This study investigated the effects of MOC activation on the responses of single neurons in the central nucleus of the inferior colliculus (CNIC) of anaesthetized guinea pigs. Extracellular responses of CNIC neurons to contralateral tones were recorded with and without MOC stimulation in normal animals and in animals acutely treated with gentamicin to eliminate peripheral effects of MOC activation. In normal animals, input–output functions of CNIC neurons showed a variety of changes. Some effects resembled qualitatively those reported for primary afferents. However, other effects were also observed, including an increase of firing rates at medium- to high-tone levels and in a small number of neurons (10%), an increase in spontaneous activity. In addition, larger threshold shifts and larger reductions of spontaneous firing rates were observed as compared to effects seen in the periphery. In gentamicin-treated animals, activation of MOC efferents did not produce any changes in the input–output functions or spontaneous activity of CNIC neurons. This observation is consistent with the majority of MOC-induced changes in monaural responses in the CNIC being mediated by the actions of MOC terminals in the cochlea and resulting from the interplay between altered afferent input and central circuitry.     

The effect of intraocular injection of tetrodotoxin on fast axonal transport of [3H]proline- and [3H]fucose-labeled materials in the developing rat optic nerve

The fast axonal transport of [3H]proline-labeled proteins and [3H]fucose-labeled glycoproteins delivered to the dorsal lateral geniculate nucleus in the developing rat optic nerve was investigated during tetrodotoxin-induced monocular impulse blockade. Repeated intraocular injections of various dosages of tetrodotoxin or citrate buffer vehicle were made every two days in rats aged 5-21 days postnatal, and the accumulation of rapidly transported radioactivity in the lateral geniculate nucleus measured between three and twelve hours post-injection at each age. The effectiveness of prolonged tetrodotoxin treatment was monitored by loss of the pupillary light reflex and the level of cytochrome oxidase activity in the contralateral superior colliculus and dorsal lateral geniculate nucleus. Numbers of optic axons proximal to the chiasm and the frequency of retinal ganglion cells per unit distance from the optic disc were examined for signs of tetrodotoxin-induced degeneration of the retinofugal pathway. Tetrodotoxin-treatment reduced the amount of fucosyl glycoproteins, but not proline-labeled proteins, axonally transported to the lateral geniculate nucleus during the first three weeks of postnatal development. Other studies indicated that tetrodotoxin significantly reduced the incorporation of [3H]fucose into retinal proteins indicating that the reduction in transport was probably due to a decrease in precursor incorporation into retinal ganglion cells. Electron microscopy of ganglion cells at 21 days revealed dilated and vacuolated Golgi cisternae associated with tetrodotoxin treatment, suggesting that tetrodotoxin may alter fucose metabolism by secondarily disrupting Golgi organization. Other protein synthetic machinery in these cells, including ribosomes and rough endoplasmic reticulum, appeared normal throughout tetrodotoxin treatment. These data indicate that Na+-dependent optic impulse activity may be indirectly related to the axonal transport of glycoproteins during early postnatal development by mediating the incorporation of precursor into glycoproteins at the Golgi apparatus and their subsequent entrance into the fast transport system.     

Distribution of the tecto-thalamic projection neurons in the hereditary microphthalmic rat

Summary Tecto-thalamic projections in the hereditary bilaterally microphthalmic rat were studied by means of WGA-HRP injection into the dorsal lateral geniculate nucleus (LGNd) and the lateroposterior thalamic nucleus (LP). Histological study in the mutant rats showed that whereas LGNd and superficial layers of the superior colliculus (SC) suffered from a remarkable reduction in size, LP had no histological changes as compared to the normal animals. Unilateral injection of the tracer into the microphthalmic LGNd showed that WGA-HRP positive neurons were present mostly in the ipsilateral str. griseum superficiale (SGS) of the SC. However, the number of labeled SGS neurons of the microphthalmic animals was about 3% of the normal. Although cell bodies of the normal tecto-LGNd neurons in the SGS were spindle-form in shape and issued one or two proximal dendrites from each pole, the microphthalmic tecto-LGNd neurons showed an irregular contour and their dendrites were not so intensively labeled. Unilateral injections of WGA-HRP into the LP revealed that the tecto-LP neurons were mainly distributed in the ipsilateral str. opticum of the colliculus. (SO) in both normal and microphthalmic animals. However, the number of labeled SO cells in the microphthalmic rat was about one-half of the normal. Furthermore, the size of labeled tecto-LP neurons was smaller than that of the normal ones, and they showed irregular round to oval cell bodies with equivocally labeled dendrites, in contrast to the normal tecto-LP neurons with polygonal cell bodies extending three or more dendrites in a radial fashion. These results indicate that there exist the tecto-LGNd and -LP projection neurons in the microphthalmic rat and that their laminally segregated projection is fundamentally preserved. However, the number of the tecto-thalamic projection neurons, especially of the tecto-LGNd cells, was markedly diminished in the mutant tectum compared to normals.Abbreviations CST cortico-spinal tract - DRN dorsal raphe nucleus - DTN dorsal tegmental nucleus - LGNd pars dorsalis of the lateral geniculate nucleus - LLN nucleus of the lateral lemniscus - LM medial lemniscus - LP lateroposterior thalamic nucleus - MGN medial geniculate nucleus - MRF midbrain reticular formation - OT optic tract - P pretectal area - PAG periaqueductal gray - PB parabigeminal nucleus - PN pontine nuclei - PCS superior cerebellar peduncle - SGS superficial gray layer of the superior colliculus - SO stratum opticum of the superior colliculus - SN substantia nigra - Vm motor nucleus of the trigeminar nerve - Vs sensory nucleus of the trigeminar nerve     

Presynaptic modulation by GABAB receptors of glutamatergic excitation and GABAergic inhibition of neostriatal neurons.

1. The present experiments investigated the effects of gamma-amino-butyric acidB (GABAB) receptor stimulation on the excitatory and inhibitory responses of neostriatal neurons evoked by stimulation of the subcortical white matter in a rat neostriatal slice preparation. 2. Intracellular recordings showed that single-impulse stimulation of the corpus callosum evoked monosynaptic, 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX)-sensitive excitatory postsynaptic potentials (EPSPs) that were attenuated by the GABAB receptor agonist, p-chlorophenyl-GABA (baclofen, 0.5-10 microM) in a concentration-dependent manner. Baclofen also blocked the GABAA-mediated inhibition of neostriatal cell responses, which were revealed by paired-impulse stimulation of the subcortical white matter. Both of these effects persisted in slices in which the anterior cortex was removed, indicating that the site of action for baclofen was intrinsic to the neostriatum. The GABAB antagonist 3-amino-2-hydroxy-2-(4-chlorophenyl)-propanesulfonic acid (saclofen, 250-500 microM) reversed the depressant actions of baclofen on both the excitatory and inhibitory responses of neostriatal cells. 3. Concentrations of baclofen as high as 100 microM, which markedly attenuated EPSP amplitude, did not exert direct effects on resting membrane potential, current-voltage relationship, input resistance, or spike threshold and thus appeared to have no postsynaptic effect on the population of neurons recorded. 4. These results indicate that, in contrast to other regions of the CNS, the depressant effects of baclofen on glutamate-dependent EPSPs are mediated exclusively through GABAB receptors located presynaptically on the terminals of glutamatergic afferents.(ABSTRACT TRUNCATED AT 250 WORDS)     

The connections between the parabigeminal nucleus and the superior colliculus in the golden hamster

The present study in hamsters shows that the horseradish peroxidase conjugated with wheat germ agglutinin (WGA-HRP) injected into the eye can be transported transneuronally in both anterograde and retrograde directions to the parabigeminal nuclei (PBN). Moreover, the patterns of labelling in the PBN are similar to those observed after introduction of horseradish peroxidase (HRP) into the superior colliculus. These results suggest that the WGA-HRP is a reliable pathway tracer for studying second-order connections at least in the visual system in mammals.     

Plasticity of tonotopic maps in auditory midbrain following partial cochlear damage in the developing chinchilla

 There is substantial reorganization of the midbrain (inferior colliculus) tonotopic map following neonatally induced partial cochlear lesions in the chinchilla. The most obvious feature of this remapping is a large ”iso-frequency” region in the ventral sector of the central nucleus of inferior colliculus (ICC). Neurons in this region exhibit similar threshold and tuning properties, with a common characteristic frequency which corresponds to the high-frequency audiometric cutoff. This overrepresented frequency range also corresponds to the high-frequency border of the cochlear lesion. Alterations to the tonotopic map corresponding to lower frequencies, in more dorsal regions of ICC, depend on the extent and degree of the cochlear lesion. When there is minimal damage to apical (low-frequency) cochlear areas, the dorsal ICC has relatively normal frequency representations. With more extensive apical cochlear lesions there is a corresponding disruption of ICC tonotopic representation of the low frequencies. We conclude that the tonotopic map within the ICC can become (re)organized postnatally according to the abnormal pattern of neural activity from the auditory periphery. Similar reorganization can be expected to occur in human infants with a partial cochlear hearing loss from birth. Received: 2 February 1998 / Accepted: 3 July 1998     

The reciprocal connections of the suprageniculate nucleus and the superior colliculus in the rat

Reciprocally bilateral connections between the superior colliculus and the suprageniculate nucleus have been studied in the rat, using the anterograde and retrograde transport techniques of HRP. In those cases where the HRP deposit was restricted to the superficial layers of the colliculus, anterogradely labeled fibers and retrogradely labeled neurons were observed in the ipsilateral suprageniculate nucleus. However, upon HRP injection extended into the intermediate layers of the colliculus, the number of labeled fibers and neurons was not only increased ipsilaterally but were also observed in the contralateral suprageniculate nucleus. The density of the labeled fibers and the number of labeled neurons was always greater in the ipsilateral side. These results show that the suprageniculate nucleus and the superior colliculus are connected reciprocally and bilaterally, with an ipsilateral dominance.     

Effect of Bcl-2 overexpression on establishment of ipsilateral retinocollicular projection in mice

During perinatal development in rodents, ipsilateral retinofugal projection spreading over the superior colliculus is eventually restricted to the rostromedial region. Since this restriction is accompanied by the apoptotic death of more than half of the retinal ganglion cells (RGCs), cell death is believed to play a major role in the restriction of transient ipsilateral projection from the retina to the superior colliculus. To determine the role of RGC death in the establishment of ipsilateral retinofugal projection, we examined the projection pattern in the superior colliculus and the dorsal lateral geniculate nucleus of transgenic mice overexpressing the human bcl-2 gene, which protects against cell death in the CNS. Retrograde labeling of RGCs showed that the number of ipsilaterally projecting RGCs in adult transgenic mice was approximately twice that in adult wild-type mice, indicating that the naturally occurring death of RGCs was prevented in these mutant mice. However, anterograde labeling of ipsilateral retinofugal pathways revealed that the innervation of retinogeniculate and retinocollicular projections was as restricted in transgenic mice as in wild-type mice.

From these results we suggest that restriction of ipsilateral retinofugal projection during development is due to retraction or elimination of excessive terminals rather than to naturally occurring RGC death.     

Functional and pharmacological differences between two types of GABA receptor on embryonic chick sensory neurons

Embryonic chick sensory neurons grown in dissociated cell culture exhibit two functional responses to GABA: an increase in resting membrane permeability to chloride (Cl) ions (resulting in membrane depolarization) and a decrease in voltage-dependent calcium (Ca) channel current (resulting in a decreased action potential duration). These two functional effects differ in a number of ways. (1) The increase in resting membrane permeability desensitizes in the maintained presence of GABA, while the decrease in action potential duration does not. (2) Muscimol is a selective agonist for the increase in resting conductance, while baclofen is a selective agonist for the decrease in action potential duration. (3) Bicuculline inhibits the GABA- or muscimol-induced increase in Cl permeability, but it does not block the GABA- or baclofen-induced decrease in action potential duration. These functional and pharmacological differences between the two effects of GABA suggest that two separate receptors are involved.     

Absolute number of parvicellular and magnocellular neurons in the red nucleus of the rat midbrain: a stereological study

The absolute number of parvicellular and magnocellular neurons in the red nucleus was estimated using design‐based stereological counting methods and systematic random sampling techniques. Six young adult male rats, and a complete set of serial 40‐μm glycolmethacrylate sections for each rat, were used to quantify neuronal numbers. After a random start, a systematic subset (i.e. every third) of the serial sections was used to estimate the total volume of the red nucleus using Cavalieri's method. The same set of sampled sections was used to estimate the number of neurons in a known subvolume (i.e. the numerical density N_v) by the optical disector method. Multiplication of the total volume by N_v yielded the absolute number of neurons. It was found that the right red nucleus consisted, on average, of 8400 parvicellular neurons (with a coefficient of variation of 0.16) and 7000 magnocellular neurons (0.12). These total neuronal numbers provide important data for the transfer of information through these nuclei and for species comparisons.     

Enhanced glutamatergic phenotype of mesencephalic dopamine neurons after neonatal 6-hydroxydopamine lesion

There is increasing evidence that a subset of midbrain dopamine (DA) neurons uses glutamate as a co-transmitter and expresses vesicular glutamate transporter (VGLUT) 2, one of the three vesicular glutamate transporters. In the present study, double in situ hybridization was used to examine tyrosine hydroxylase (TH) and VGLUT2 mRNA expression during the embryonic development of these neurons, and postnatally, in normal rats and rats injected with 6-hydroxydopamine (6-OHDA) at P4 to destroy partially DA neurons. At embryonic days 15 and 16, there was a regional overlap in the labeling of TH and VGLUT2 mRNA in the ventral mesencephalon, which was no longer found at late embryonic stages (E18-E21) and postnatally. In normal pups from P5 to P15, only 1-2% of neurons containing TH mRNA in the ventral tegmental area (VTA) and substantia nigra, pars compacta, also displayed VGLUT2 mRNA. In contrast, after the cerebroventricular administration of 6-OHDA at P4, 26% of surviving DA neurons in the VTA of P15 rats expressed VGLUT2. To search for a colocalization of TH and VGLUT2 protein in axon terminals of these neurons, the nucleus accumbens of normal and 6-OHDA-lesioned P15 rats was examined by electron microscopy after dual immunocytochemical labeling. In normal rats, VGLUT2 protein was found in 28% of TH positive axon terminals in the core of nucleus accumbens. In 6-OHDA-lesioned rats, the total number of TH positive terminals was considerably reduced, and yet the proportion also displaying VGLUT2 immunoreactivity was modestly but significantly increased (37%). These results lead to the suggestion that the glutamatergic phenotype of a VTA DA neurons is highly plastic, repressed toward the end of normal embryonic development, and derepressed postnatally following injury. They also support the hypothesis of co-release of glutamate and DA by mesencephalic neurons in vivo, at least in the developing brain.