鸡胚视顶发育过程中神经元增殖、迁移和分层形成的规律

目的:采用5-溴脱氧尿嘧啶核苷(BrdU)标记鸡胚发育过程巾视顶新生神经元,阐明鸡胚发育过程巾视顶盖神经元增殖、迁移规律.方法:采用鸡胚开窗带壳培养的方式,在培养过程巾,分别在胚胎发育的第E4天至E18.5 d滴加1 mg/ml浓度的BrdU各10μl,每组5个胚胎,滴加BrdU后密封开口,继续培养,当培养到胚胎发育的E18.5 d时,取材、甲醛固定、震荡切片,采用硫堇染色,观察视顶盖层的结构,免疫组织化学显示BrdU标记的新生细胞;显微拍照,分析不同时期表面灰质纤维层增殖细胞的数量、总面积、灰度平均值、平均光密度和平均积分光密度等参数,综合分析各阶段新生神经元的差异.结果:鸡胚视顶结构通过硫堇染色可以明显分成6层,在胚胎发育的E4 d～8 d是细胞大量增殖阶段,新生细胞数明显高于其他阶段;新生细胞在胚胎发育的E12 d后明显减少.结论:鸡胚发育过程中,视顶从胚胎发育的E4 d开始到E12 d时,6层结构已经基本形成,新生神经元E12 d后明显减少,其视顶盖6层结构的形成规律是从内向外.     

Roles of periventricular neurons in retinotectal transmission in the optic tectum

The midbrain roof is a retinorecipient region referred to as the optic tectum in lower vertebrates, and the superior colliculus in mammals. The retinal fibers projecting to the tectum transmit visual information to tectal retinorecipient neurons. Periventricular neurons are a subtype of these neurons that have their somata in the deepest layer of the teleostean tectum and apical dendrites ramifying at more superficial layers consisting of retinal fibers. The retinotectal synapses between the retinal fibers and periventricular neurons are glutamatergic, and ionotropic glutamate receptors mediate the transmission in these synapses. This transmission involves long-term potentiation, and is modulated by hormone action. Visual information processed in the periventricular neurons is transmitted to adjacent tectal cells and target nuclei of periventricular neuron axonal branches, some of which relay the visual information to other brain areas controlling behavior. We demonstrated that periventricular neurons play a principal role in visual information processing in the teleostean optic tectum; the effects of tectal output on behavior is discussed also in the present review.     

Neonatal deafferentation does not alter membrane properties of trigeminal nucleus principalis neurons

In the brain stem trigeminal complex of rats and mice, presynaptic afferent arbors and postsynaptic target cells form discrete modules ("barrelettes"), the arrangement of which duplicates the patterned distribution of whiskers and sinus hairs on the ipsilateral snout. Within the barrelette region of the nucleus principalis of the trigeminal nerve (PrV), neurons participating in barrelettes and those with dendritic spans covering multiple barrelettes (interbarrelette neurons) can be identified by their morphological and electrophysiological characteristics as early as postnatal day 1. Barrelette cells have focal dendritic processes, are characterized by a transient K(+) conductance (I(A)), whereas interbarrelette cells with larger soma and extensive dendritic fields characteristically exhibit low-threshold T-type Ca(2+) spikes (LTS). In this study, we surveyed membrane properties of barrelette and interbarrelette neurons during and after consolidation of barrelettes in the PrV and effects of peripheral deafferentation on these properties. During postnatal development (PND1-13), there were no changes in the resting potential, composition of active conductances and Na(+) spikes of both barrelette and interbarrelette cells. The only notable changes were a decline in input resistance and a slight increase in the amplitude of LTS. The infraorbital (IO) branch of the trigeminal nerve provides the sole afferent input source to the whisker pad. IO nerve transection at birth abolishes barrelette formation as well as whisker-related neuronal patterns all the way to the neocortex. Surprisingly this procedure had no effect on membrane properties of PrV neurons. The results of the present study demonstrate that distinct membrane properties of barrelette and interbarrelette cells are maintained even in the absence of input from the whiskers during the critical period of pattern formation.     

Electrophysiological properties of neurons recorded intracellularly in slices of the pigeon optic tectum

The electrical properties of pigeon's optic tectum neurons located in the non-retinorecipient region of layer II have been studied in vitro slice preparations by using intracellular recordings. As judged from the somatodendritic characteristics of cells intracellularly labeled with horseradish peroxidase recordings were obtained from pyramidal neurons, the main morphological type, as well as from ganglion cells. When stimulated with depolarizing current pulses of 300-500 ms duration, three distinct modes of firing were observed. Most neurons (Type I) responded with a continuous firing of fast action potentials whose frequency rate increased regularly when current strength was raised. Another group of cells (Type II) also exhibited sustained firing. However, in Type II cells, grouped discharges formed by 2-6 fast action potentials per group fired in rapid succession were elicited within a certain range of current intensity. Finally, another group of cells (Type III) responded at all intensities tested by a short train of fast action potentials only at the onset of the current step. At current strength close to threshold the spike undershoot of type I neurons was followed by a slow hyperpolarizing afterpotential while the spike undershoot of Type II cells was followed by a hump-like depolarization and a slow hyperpolarizing afterpotential. In Type II cells, we have also observed a pronounced increase of the hyperpolarizing afterpotential after a grouped discharge. Type III cells were characterized by a small amplitude and short duration hyperpolarizing afterpotential, barely visible in most of them. In Type I and II cells the slow hyperpolarizing afterpotential was blocked by replacing Ca2+ with Mg2+ or Cd2+ in the saline. These results support the idea that in these two types of neurons the slow hyperpolarizing afterpotential is primarily caused by a Ca2+-dependent K+ conductance. Furthermore, blocking the slow hyperpolarizing afterpotential provoked a pronounced increase of the firing frequency of Type I cells. In Type II cells blockade of the slow hyperpolarizing afterpotential had a greater effect on firing behavior: i.e. when Ca2+ was replaced with Mg2+ or Cd2+, Type II neurons exhibited repetitively fired action potentials at high frequency but were incapable of discharging repetitive grouped discharges. These observations indicate that the Ca2+-dependent K+ conductance involved in the generation of the slow hyperpolarizing afterpotential is the main modulator of the firing behavior of both types of cells.(ABSTRACT TRUNCATED AT 400 WORDS)     

Anatomy and physiology of horizontal cells in layer 5b of the chicken optic tectum

In the visual midbrain of birds, a variety of cell types has recently been characterized with both anatomical and physiological techniques to gain insight into the mechanisms of visual information processing. Here we present data from a horizontal cell type located in the retinorecipient layer 5b of the chick optic tectum. Intracellular labeling revealed that these neurons are multipolar, have no axonal structures and arborize completely within the layer 5b where they extend over considerable distances. Immunohistochemistry with an antibody against calbindin labeled a population of horizontal cells in layer 5b; however, double labeling showed that these neurons represent a subpopulation of approximately one third of the neurons in that layer. Whole-cell patch recordings with additional cell filling from horizontal cells revealed that the physiological responses to depolarization changes with maturation, from a comparatively slow oscillatory pattern reminiscent of hair cell physiology at embryonal stages to a damped series of small action potentials at posthatching. In response to electrical stimulation in the vicinity of the neurons, cells responded with either excitatory postsynaptic potentials or small action potentials. Horizontal cell types are found in the visual midbrain of both avian and mammalian species. On the basis of the data presented here and data from the literature, the functional role of these cells is discussed. As in layer 5b of the chick optic tectum specific synaptic glomeruli have been found, the horizontal cells might constitute local inhibitory circuits within the retino-tectal synapses and, in addition, contribute to mechanisms of directional selectivity in these projections.     

Spatiotemporal analysis of electrically evoked activity in the chicken optic tectum: a VSDI study

The midbrain is an important processing area for sensory information in vertebrates. The optic tectum and its mammalian counterpart, the superior colliculus, receive multimodal, topographic information and contain a sensory map that plays a role in spatial attention and orientation movements. Many studies have investigated the tectal circuitry by cytochemistry and by characterization of particular cell types. However, only a few studies have investigated network activation throughout the depth of the tectum. Our study provides the first data on spatiotemporal activity profiles in the depth and width of the avian optic tectum. We used an optical imaging approach with voltage-sensitive dyes to investigate population responses at a high temporal and spatial resolution. With the necessary caution due to cell extension across several layers, we can thus link our findings tentatively with the general layout of the avian optic tectum. Single electrical stimuli in the retinorecipient layers 1-4 evoked a complex optical response pattern with two components: a short, strong transient response and a weaker persistent response that lasted several hundred milliseconds. The response started in layer 5 and spread within this layer before it propagated into deeper layers. This is in line with neuroanatomical and earlier physiological data. Analysis of temporal sequence and pharmacological manipulations revealed that these responses were mainly driven by postsynaptic activation. Thus tectal network responses to patterned input can be studied by voltage-sensitive dye imaging.     

Progress of topographic regulation of the visual projection in the halved optic tectum of adult goldfish.

1. The patterns of re-established visual projections on to the rostral half-tectum are studied following excision of the caudal tectum at various intervals after section of either the contralateral optic nerve or the ipsilateral optic tract in adult goldfish. 2. The pattern of a newly restored retinotectal projection depends on the duration of the post-operative period given to the halved tectum before it is re-innervated by regenrating optic fibres from the retina. 3. When the duration is such that regenerating optic fibres invade the denervated rostral half-tectum at about 40 days or longer after excision of the caudal tectum, the remaining half-tectum is able to accommodate incoming optic fibres not only from the appropriate temporal hemi-retina but also from the foreign nasal hemiretina in an orderly compressed topographic pattern. 4. If the surgical operations are timed so that the halved tectum receive regenerating optic fibres earlier than 33 days after excision of the caudal tectum, the halved tectum initially accommodates only those optic fibres originating from the temporal half of the retina at this early stage. 5. This normal (uncompressed) pattern of the newly regenerated visual projection, however, eventually changes into an orderly compressed pattern at a later period. Post-operative dark-deprivation of the operated fish has no significant effect on the temporal transition. 6. The temporal transition from an initially normal pattern into an orderly compressed pattern may reflect the time course of progressive and systematic changes involved in topographic regulation of the halved tectum into a whole.     

Physiology and morphology of callosal projection neurons in mouse

In the mammalian neocortex, the corpus callosum serves as the major source of interhemispheric communication, composed of axons from callosal neurons located in supragranular (II/III) and infragranular (V/VI) layers. We sought to characterize the physiology and morphology of supragranular and infragranular callosal neurons in mice using retrograde tracers and whole-cell patch clamp recordings. Whole-cell patch clamp recordings were made from retrogradely labeled callosal neurons following unilateral injection of fluorescent latex microspheres in the contralateral sensory-motor cortex. Following recordings and biocytin dialysis, labeled neurons were reconstructed using computer-assisted camera lucida (Neurolucida) for morphological analyses. Whole-cell recordings revealed that callosal neurons in both supra- and infragranular layers display very similar intrinsic membrane properties and are characteristic regular-spiking neurons. Morphological features examined from biocytin-filled reconstructions as well as retrogradely BDA labeled cells did not reveal any differences. Analysis of spontaneous postsynaptic potentials from callosal neurons did reveal several differences including average amplitude, frequency, and decay time. These findings suggest that callosal neurons in both supra- and infragranular layers have similar phenotypes though belong to different local, intracortical networks.     

The optic tectum of the gymnotiform electric fish, Eigenmannia: labeling of physiologically identified cells

A total of 47 tectal neurons of the weakly electric fish, Eigenmannia, were studied physiologically and labelled by intracellular injection of Lucifer Yellow. With the exception of two cell types, all cells could be classified in accordance with the Golgi studies of Sas and Maler. The dominant stimulus modality of neurons was correlated with their laminar location. Neurons of the stratum opticum only responded to visual stimuli, such as modulations of the light level or the motion of an object. They showed, however, no directional preferences for motion. Neurons of the stratum griseum centrale were predominantly driven by electrosensory stimuli, most often those associated with the movement of an object, and generally were very sensitive to the direction of motion. Integration of different sensory modalities was found in neurons with dendrites invading laminae with different sensory inputs. In addition, small axons of interneurons appear to relay information across laminae. Large multipolar neurons in the deep tectum responded to the motion of objects, often preferring a particular direction of motion. Some of these large multipolar neurons of the deep tectum also discriminated the sign of the frequency difference between a mimic of a neighbor's sinusoidal electric organ discharge and the animal's own signal. These neurons are potential candidates for the control of the jamming avoidance response. These neurons were morphologically indistinguishable from large multipolar neurons of the deep tectum that either responded to moving objects or to acoustical stimuli. Individual large cells of the deep tectum project to various targets (Fig. 1) and probably contribute to the control of different behavioral responses. This suggests that the nature of such responses would then depend upon the constitution of sets of neurons recruited by a given stimulus situation, and the role of individual tectal neurons would neither be particularly specific nor very significant.     

Manipulation of inhibition in the owl's nucleus laminaris and its effects on optic tectum neurons

Differences in arrival time and intensity (or level) of sound between the ears serve as cues for localization of sound in many animals. Barn owls use interaural time difference (ITD) and interaural level difference (ILD) for localization in azimuth and elevation, respectively. The owl's brain processes these two cues in separate pathways. The nucleus laminaris is the first site that detects ITDs by methods of delay lines and coincidence detection. The nucleus ventralis lemnisci lateralis, pars posterior is the first site of processing ILDs. The two pathways merge in the inferior colliculus to give rise to sensitivity to combinations of ITD and ILD. This selectivity is relayed to the optic tectum where neurons are sensitive to both visual and auditory stimuli. The present paper reports the results of manipulating inhibition in the nucleus laminaris and its effects on the optic tectum neurons. Injection of GABA or muscimol (a GABA(A) receptor agonist) in the nucleus laminaris reduces the responses of its neurons to ITD. This finding proves that GABA(A) receptor-mediated inhibition acts on the nucleus laminaris neurons. The same treatment did not affect the neurons of the nucleus ventralis lemnisci lateralis, pars posterior, whereas it reduced the response of the optic tectum neurons to ITD-ILD pairs.We conclude that although the two pathways are independent, the process of combining ITD and ILD creates a new relationship in which the output of the neuron varies with the amplitude of either input. This conclusion is consistent with the recent finding that the combination sensitivity is due to a multiplication of ITD and ILD inputs.     

Short-term sensory learning does not alter parvalbumin neurons in the barrel cortex of adult mice: a double-labeling study

We have previously reported that a classical conditioning paradigm involving stimulation of a row of facial vibrissae produced expansion of the cortical representation of the activated vibrissae ("trained row"), this was demonstrated by labeling with 2-deoxyglucose in layer IV of the barrel cortex. We have also shown that functional reorganization of the primary somatosensory cortex is accompanied by an increase in the density of small GABAergic cells and glutamate decarboxylase 67-positive neurons in the hollows of barrels representing the "trained row." GABA neurons of the rat neocortex co-localize with calcium-binding proteins [parvalbumin, carletinin, calbindin D28k] and neuropeptides (vasoactive intestinal polypeptide, somatostatin). In the present study we have examined GABAergic parvalbumin-positive, interneurons in the cortical representation of "trained" facial vibrissae after short-term aversive training, in order to determine whether the observed changes in glutamate decarboxylase 67-positive neurons are accompanied by changes in parvalbumin-positive neurons. Using double immunofluorescent staining, it was found that (i) all parvalbumin-positive neurons in the barrel hollows were glutamate decarboxylase 67-positive, (ii) following aversive training density of glutamate decarboxylase 67-positive neurons in barrel hollows increased significantly compared with controls and (iii) density glutamate decarboxylase 67-positive/parvalbumin-positive neurons in "trained" barrel hollows did not change compared with controls. This study is the first to demonstrate that the density of double-labeled glutamate decarboxylase 67-positive/parvalbumin-positive neurons does not alter during cortical plasticity, thus suggesting that some other population (i.e. parvalbumin negative) of GABAergic interneurons is involved in learning-dependent changes in layer IV of the barrel cortex.     

Morphology of identified projection neurons in layer 5 of rat visual cortex

We studied the morphology of neurons in layer 5 of rat visual cortex (area 17) projecting to the contralateral hemisphere and the superior colliculus. Double labelling with fluorescent tracers indicated that these projections arise from different populations of cells. To reveal the morphology of the cells we stained retrogradely labelled neurons intracellularly in living brain slices. Callosal projecting pyramidal cells have 3-6 basal dendrites and an apical dendrite which never reaches higher than layer 3. Corticotectal cells have 6-8 basal dendrites and a prominent apical dendrite which always forms a large tuft in layer 1. Thus, neurons in the same cortical layer that give rise to different projections also differ in their morphology. However, each population of neurons has a rather stereotyped dendritic branching pattern, despite a large variation in soma size.     

Neurogenesis,gliogenesis and the developing chicken optic tectum: an immunohistochemical and ultrastructural analysis

We present the first comprehensive analysis of avian optic tectum development, including proliferation, migration and maturation of both neuronal and glial cells. The distribution of doublecortin, Tuj-1, vimentin and GFAP was characterized by immunohistochemistry between E3 and E20, and correlated with the electron microscopic structure in the chicken optic tectum. The immunohistological markers used in our study are known to be critical for distinct steps of neurogenesis and gliogenesis. We demonstrate that neurogenesis within the optic tectum starts at E3 with prominent doublecortin and moderate Tuj-1 expression. With the aid of electron microscopy, we also show that most of the cells are still undifferentiated at E4. Starting from E6, all postmitotic Tuj-1-positive neurons have left the ventricular zone and concurrently, with the end of proliferation around E12, doublecortin disappears from this region. Before hatching, doublecortin expression totally ceases, indicating that now all neurons have matured, this was also confirmed by ultrastructural investigations. Furthermore, vimentin expression starts around E4, prior to the appearance of the first radial glial cells at E6. Astrocytes can be detected by GFAP expression at E12. As radial glial cells (RGC) transform into astrocytes between E12 and E20, the vimentin signal is progressively replaced by the GFAP signal. We could also show that vimentin-positive RGCs do express doublecortin between E4 and E6, the time-point of prominent neurogenesis, reflecting their bipotent character.     

Dietary sodium deprivation does not alter taste sensitivity in the rat

The influence of 33 days of combined dietary sodium and water deprivation on the taste sensitivity and responsivity of the rat to a wide range of NaCl solutions and discrimination tasks was examined using double-blind procedures, high-precision gustometry, and an automated operant go/no-go task. Sodium concentration in urine and saliva but not plasma decreased significantly only in the sodium-deficient rats. The same measures remained unchanged in control animals. Nonparametric measures of taste sensitivity and responsivity to NaCl did not differ between sodium-deficient and control animals and performance on taste discrimination, reversal learning, and lick-rate tasks remained the same between the groups as well. The present results support and extend previous findings that dietary-induced sodium deficiency increases NaCl and water consumption, but alters neither the sodium-deficient rat's taste sensitivity and responsivity nor performance on simple and complex tasks in which tastants serve as cues for discriminative responding.     

An extensive projection of fish dorsolateral tegmental cells to the optic tectum revealed by intra-axonal dye marking

The cells of the dorsolateral tegmental (DLT) nucleus in the crucian carp (Carassius carassius) were physiologically identified and marked with Lucifer dye. All the identified DLT cells receive both visual and rhombencephalic inputs. These cells project their axons into the contralateral tectum via the tectal commissure as well as into the ipsilateral tectum. The most striking characteristic of the DLT cells was the wide distribution of their axonal branching in the ipsilateral tectum.     

Long-term, but not transient, threshold shifts alter the morphology and increase the excitability of cortical pyramidal neurons

Partial hearing loss often results in enlarged representations of the remaining hearing frequency range in primary auditory cortex (AI). Recent studies have implicated certain types of synaptic plasticity in AI map reorganization in response to transient and long-term hearing loss. How changes in neuronal excitability and morphology contribute to cortical map reorganization is less clear. In the present study, we exposed adult rats to a 4-kHz tone at 123 dB, which resulted in increased thresholds over their entire hearing range. The threshold shift gradually recovered in the lower-frequency, but not the higher-frequency, range. As reported previously, two distinct zones were observed 10 days after the noise exposure, an enlarged lower-characteristic frequency (CF) zone displaying normal threshold and enhanced cortical responses and a higher-CF zone showing higher threshold and a disorganized tonotopic map. Membrane excitability of layer II/III pyramidal neurons increased only in the higher-CF, but not the lower-CF, zone. In addition, dendritic morphology and spine density of the pyramidal neurons were altered in the higher-CF zone only. These results indicate that membrane excitability and neuronal morphology are altered by long-term, but not transient, threshold shift. They also suggest that these changes may contribute to tinnitus but are unlikely to be involved in map expansion in the lower-CF zone.     

Membrane properties of identified lateral and medial perforant pathway projection neurons

The physiological characteristics of neurons that project to the hippocampus and dentate gyrus via the medial perforant pathway (projection neurons) are well known, but the characteristics of neurons that project to these areas via the lateral perforant pathway (projection neurons) are less well known. We have used retrograde tracing and whole-cell recording in brain slices to compare the membrane and firing properties of medial perforant pathway and lateral perforant pathway projection neurons in layer II of the medial and lateral entorhinal cortex. The properties of medial perforant pathway projection neurons were identical to those reported previously for spiny stellate neurons in the medial entorhinal cortex. In contrast, lateral perforant pathway projection neurons were characterized by a higher input resistance, a lack of time-dependent inward (anomalous) rectification, and a lack of prominent depolarizing spike afterpotentials. Voltage-clamp recordings suggest that the absence of anomalous rectification in lateral perforant pathway projection neurons is due to smaller hyperpolarization activated cation currents in these cells, and the lack of depolarizing afterpotential may be due to smaller low-threshold calcium currents. Persistent sodium current was also smaller in lateral perforant pathway projection neurons, but the difference in persistent sodium current between medial perforant pathway and lateral perforant projection neurons was much less pronounced than the difference in low voltage activated currents. These results underscore the functional differences between the medial entorhinal cortex and lateral entorhinal cortex, and may help to explain the differing abilities of these cortical areas to participate in certain types of network activity.     

Social stress does not alter the expression of sensitization to cocaine

The effects of chronic social stress on behavioral sensitization to cocaine were investigated in the Syrian hamster. Adolescent animals received either 15 mg/kg i.p. of cocaine or saline twice per day for 7 consecutive days. Two weeks following the last injection (young adulthood), they were given a challenge dose of 5 mg/kg i.p. of cocaine and scored for locomotion. Motor activity was significantly greater in cocaine-treated animals, demonstrating sensitization to this psychostimulant. Following the results of the first study, another group of adolescent animals was exposed to either a novel clean cage (control) or an aggressive resident male hamster (social stress) for 15 min following an injection of cocaine (20 mg/kg i.p. once daily) or saline for 7 consecutive days. The groups were as follows: Social Stress/Cocaine (SSC), No Social Stress/Cocaine (NSSC), Social Stress/Saline (SSS) and No Social Stress/Saline (NSSS). Two weeks following the last injection (Day 21), all animals were given a challenge dose of cocaine (5 mg/kg i.p.) and were rescored for locomotion. At that time, the suppressive effect of stress on locomotion was no longer detectable, as the expression of sensitization was observed in the NSSC but not in the SSC group. These results suggest that chronic social stress administered during adolescence does not cross-sensitize with cocaine in young adult hamsters.