Postnatal development of somatostatin-containing neurons in the visual cortex of normal and dark-reared rats

Summary The distribution of somatostatin (SRIF)-immunoreactive neurons in the visual cortical areas 17, 18 and 18a of Wistar rats from birth to adulthood was followed in both normal and dark-reared animals. The SRIF neurons show difference in distribution amongst the three cortical areas studied as early as the first postnatal week. Area 17 was distinguished by fewer SRIF cells in the upper layers (I–III), which results in a lower overall density. The SRIF neurons in all areas appeared to increase in numbers up to about 3 weeks and then decline dramatically to adult levels, which were 14–19% of the peak levels. Although this decline was still obvious, it moderated to 25–31% in dark-reared animals. The greatest effect was seen in area 18 where, at 60 days of age, there were twice as many SRIF cells in darkreared as in normal controls. It is suggested that, under conditions of dark rearing, the overall pattern of development of SRIF neurons, being uninfluenced by extrinsic factors, reveals the cells' genetic potential.     

The effects of binocular suture and dark rearing on the induction of c-fos protein in the rat visual cortex during and after the critical period

It has been demonstrated in kittens that binocular lid suture has more deleterious and irreversible effects on plasticity of the developing visual system than rearing in complete darkness. The present study using immunocytochemistry focuses on the effects of the two types of visual deprivation on the inducibility of c-fos protein in visual cortical neurons of rats. Rats were subjected to binocular suture or dark rearing for 1 week during (postnatal days 14-21; P14-P21) and after (P50-P57) the critical period for activity-dependent modifiability of cortical ocular dominance. In rats of both age groups reared in the normal light-dark condition, only a small number of Fos-immunoreactive neurons was obtained in the visual cortex. By contrast, in dark-reared pups and adult rats, numerous c-fos neurons were detected in the layers II-IV and VI of the visual cortex following a brief light exposure (1 h). In rats of both ages subjected to binocular suture, Fos neurons were detected in the same layers as in the dark-reared rats, but significantly less in number. We speculate that the reduced plasticity of the visual cortex in the rats subjected to binocular suture may be due partly to the repressed AP-1 activity in visual cortical neurons. No significant difference was detected in c-fos expression in the visual cortex between visually manipulated pups and adult rats.     

The postnatal development of visual callosal connections in the absence of visual experience or of the eyes

Summary Counts of callosal neurons retrogradely labeled by horseradish peroxidase (visualized using multiple substrates) were obtained in areas 17 and 18 of five kittens reared with their eyelids bilaterally sutured and of three kittens which had undergone bilateral enucleation on postnatal days 1–4. These counts were compared with those obtained in normal adult cats.The normal adult distribution of the callosal neurons results from the gradual postnatal reduction of a more widespread juvenile population. Binocular visual deprivation by lid suturing dramatically decreases the final number of callosal neurons and narrows their region of distribution (callosal zone) in areas 17 and 18. A less severe reduction in the final number of callosal neurons is caused by bilateral enucleation, which also increases the width of the callosal zone compared to that of normal cats. Thus, visual experience is necessary for the normal stabilization of juvenile callosal connections. However, since some callosal neurons form connections in the absence of vision, other influences capable of stabilizing juvenile callosal neurons also exist. These influences are probably antagonized by destabilizing influences or inhibited, when the eyes are intact.This work was supported by Swiss National Science Foundation grant 2.219.0.78 to Dr. G.M. Innocenti; Dr. D.O. Frost received a fellowship from the American-Swiss Foundation for Scientific Exchange     

The quantitative effects of dark-rearing and light exposure on the laminar composition and depth distribution of neurons and glia in the visual cortex (area 17) of the rat

Summary The effects of dark-rearing and light-exposure on the distribution of neurons and glial cells types in the rat visual cortex (area 17) have been investigated. Three groups of animals were studied: i) rats reared in the dark until weaning at 21 days post natum (21 DPN) and subsequently light-exposed for 31 days (Group 21/31); ii) rats darkreared until 52 DPN and then exposed to light for 3 days (Group 3 dL); and iii) rats totally dark-reared until 52 DPN (Group 52 dD). Semithin sections tangential to the pial surface were obtained at sampling intervals 50 m apart throughout the depth of the left visual cortex. The volume numerical densities of neurons, astroglia, oligodendroglia, and microglia, at each sampling strata in the cortex were calculated using stereological techniques. The laminer density and distribution of neurons was not significantly different between the three groups. In comparison with group 21/31 there was a marked reduction in the densities of astroglia, oligodendroglia, and microglia in lower layer 5 of groups 3 dL and 52 dD. Additionally, the density of microglia in thalamorecipeint layer 4 was greatly increased in group 3 dL compared with groups 21/31 and 52 dD. These results indicate specific alterations in the glial cell composition of the rat visual cortex following periods of dark-rearing and light-exposure. Furthermore, changes in the density of glial cells in layer 5 may reflect functional modifications in neurons projecting to the superior colliculus.     

Effects of alternating monocular occlusion on the development of visual callosal connections

Summary During normal postnatal development there is a partial elimination of the callosal projections of cortical areas 17 and 18 in the cat. Visual experience early in life can modulate this process. In the present study, we investigated how restricting visual experience to alternating monocular occlusion affects the development of the callosal connections of cortical areas 17 and 18. Alternating monocular occlusion exaggerates the normally occurring partial elimination of immature callosal projections: it causes a significant reduction in the total number of neurons in the supragranular layers that send an axon through the corpus callosum and marginally increases the distribution of these neurons across areas 17 and 18. Examination of these data in the context of the effects of other types of abnormal early visual experience on the corpus callosum and on the anatomy and physiology of areas 17 and 18 indicates that the postnatal development of the corpus callosum is under the control of multiple, interacting influences which differ in the magnitude and quality of their effects. The data also support the conclusion, drawn from our results in prior studies, that normal visual stimulation is necessary for the stabilization of the normal complement of callosal projections.     

Morphology of visual callosal neurons with different locations,contralateral targets or patterns of development

In kittens, callosally projecting neurons were labeled by retrograde transport of FITC- (fluorescein isothiocyanate)- and TRITC- (tetramethylrhodamine isothiocyanate)-conjugated latex microspheres injected in two different visual areas (17, 17/18, 19, or postero-medial lateral suprasylvian; PMLS) at postnatal day 3. At postnatal day 57 more than 1200 labeled neurons in visual cortical areas were intracellularly injected with 3% lucifer yellow (LY) in perfusion-fixed slices of the contralateral hemisphere. The distribution of labeled neurons was charted, and LY-filled neurons were classified on the basis of their area and layer of location, and dendritic pattern. The dendritic arbors of 120 neurons were computer reconstructed. For the basal dendrites of supragranular pyramidal neurons a statistical analysis of number of nodes, internodal and terminal segment lengths, and total dendritic length was run relative to the area of location and axonal projection. Connections were stronger between homotopic than between heterotopic areas. Overall tangential and laminar distributions depended on the area injected. Qualitative morphological differences were found among callosally projecting neurons, related to the area of location, not to that of projection. In all projections from areas 17 and 18, pyramidal and spinous stellate neurons were found in supragranular layers. In contrast, spinous stellate neurons lacked in projections from area 19, 21a, PMLS and postero-lateral lateral suprasylvian (PLLS). In all areas, the infragranular neurons showed heterogeneous typology, but in PMLS no fusiform cells were found. Quantitative analysis of basal dendrites did not reveal significant differences in total dendritic length, terminal, or intermediate segment length among neurons in area 17 or 18, and this was related to whether they projected to contralateral areas 17–18 or PMLS. All injections produced exuberant labeling in area 17. No differences could be found between neurons in area 17 (with transient axons through the corpus callosum) and neurons near the 17/18 border (which maintain projections to the corpus callosum). In conclusion, morphology of callosally projecting neurons seems to relate more to intrinsic specificities in the cellular composition of each area than to the area of contralateral axonal projection or the fate of callosal axons.     

Zinc histochemistry reveals circuit refinement and distinguishes visual areas in the developing ferret cerebral cortex

A critical question in brain development is whether different brain circuits mature concurrently or with different timescales. To characterize the anatomical and functional development of different visual cortical areas, one must be able to distinguish these areas. Here, we show that zinc histochemistry, which reveals a subset of glutamatergic processes, can be used to reliably distinguish visual areas in juvenile and adult ferret cerebral cortex, and that the postnatal decline in levels of synaptic zinc follows a broadly similar developmental trajectory in multiple areas of ferret visual cortex. Zinc staining in all areas examined (17, 18, 19, 21, and Suprasylvian) is greater in the 5-week-old than in the adult. Furthermore, there is less laminar variation in zinc staining in the 5-week-old visual cortex than in the adult. Despite differences in staining intensity, areal boundaries can be discerned in the juvenile as in the adult. By 6 weeks of age, we observe a significant decline in visual cortical synaptic zinc; this decline was most pronounced in layer IV of areas 17 and 18, with much less change in higher-order extrastriate areas during the important period in visual cortical development following eye opening. By 10 weeks of age, the laminar pattern of zinc staining in all visual areas is essentially adultlike. The decline in synaptic zinc in the supra- and infragranular layers in all areas proceeds at the same rate, though the decline in layer IV does not. These results suggest that the timecourse of synaptic zinc decline is lamina specific, and further confirm and extend the notion that at least some aspects of cortical maturation follow a similar developmental timecourse in multiple areas. The postnatal decline in synaptic zinc we observe during the second postnatal month begins after eye opening, consistent with evidence that synaptic zinc is regulated by sensory experience.     

Alteration of rat hippocampal neurogenesis and neuronal nitric oxide synthase expression upon prenatal exposure to tamoxifen

The present study delineates the effect of tamoxifen on neuronal density and expression of neuronal nitric oxide synthase (nNOS) in hippocampal nerve cells during prenatal and postnatal periods in rats. Pregnant rats were administered with tamoxifen one day prior to labor (E21) and on the childbirth day (E22). Hippocampi of embryos at E22 and newborns at postnatal days of 1, 7, and 21 (P1, P7, and P21) were investigated. Density of the neurons in areas of the developing hippocampus including cornu ammonis (CA1, CA3), dentate gyrus, and subiculum were studied. Our findings show that the number of pyramidal neurons was significantly decreased in CA1 and subiculum of tamoxifen-treated rats in E22, P1, and P7. We found that cellular density was lower in early stages of development, however, cellular density and thickness gradually increased during the development particularly in the third week. We found that nNOS expression was decreased in E22, P1, and P7 in animals treated with tamoxifen. The present study shows that tamoxifen affects development and differentiation of postnatal rat hippocampus, CA1 neurons, and nNOS expression.     

Effects of dark rearing on the development of visual callosal connections

Summary It is now well established that during normal postnatal development there is a partial elimination of the callosal projections of cortical areas 17 and 18 in the cat and that visual experience early in life can modulate this process. In the present experiments, we quantitatively studied the influence of light, per se, by rearing cats in total darkness. Dark rearing exaggerates the normally occurring partial elimination of immature callosal projections: it causes a significant reduction in the total number of neurons in both the supra-and infragranular layers that send an axon through the corpus callosum and slightly narrows the distribution of these neurons across areas 17 and 18. These data demonstrate that visual stimulation is not necessary either to initiate the partial elimination of immature callosal projections or to stabilize a large fraction of the callosal projections present at birth. However, normal visual stimulation is necessary for the stabilization of the normal complement of callosal projections.     

Chronic NMDA exposure accelerates development of GABAergic inhibition in the superior colliculus

Maturation of excitatory synaptic connections depends on the amount and pattern of their activity, and activity can affect development of inhibitory synapses as well. In the superficial visual layers of the superior colliculus (sSC), developmental increases in the effectiveness of gamma-aminobutyric acid (GABA(A)) receptor-mediated inhibition may be driven by the maturation of visual inputs. In the rat sSC, GABA(A) receptor currents significantly jump in amplitude between postnatal days 17 and 18 (P17 and P18), approximately when the effects of cortical inputs are first detected in collicular neurons. We manipulated the development of these currents in vivo by implanting a drug-infused slice of the ethylene-vinyl acetate copolymer Elvax over the superior colliculus of P8 rats to chronically release from this plastic low levels of N-methyl-D-aspartate (NMDA). Sham-treated control animals received a similar implant containing only the solvent for NMDA. To examine the effects of this treatment on the development of GABA-mediated neurotransmission, we used whole cell voltage-clamp recording of spontaneous synaptic currents (sPSCs) from sSC neurons in untreated, NMDA-treated, and sham-treated superior colliculus slices ranging in age from 10 to 20 days postnatal. Both amplitude and frequency of sPSCs were studied at holding potentials of +50 mV in the presence and absence of the GABA(A) receptor antagonist, bicuculline methiodide (BMI). The normal developmental increase in GABA(A) receptor currents occurred on schedule (P18) in sham-treated sSC, but NMDA treatment caused premature up-regulation (P12). The average sPSCs in early NMDA-treated neurons were significantly larger than in age-matched sham controls or in age-matched, untreated neurons. No differences in average sPSC amplitudes across treatments or ages were present in BMI-insensitive, predominantly glutamatergic synaptic currents of the same neurons. NMDA treatment also significantly increased levels of glutamate decarboxylase (GAD), measured by quantitative western blotting with staining at P13 and P19. Cell counting using the dissector method for MAP 2 and GAD(67) at P13 and P19 indicated that the differences in GABAergic transmission were not due to increases in the proportion of inhibitory to excitatory neurons after NMDA treatment. However, chronic treatments begun at P8 with Elvax containing both NMDA and BMI significantly decreased total neuron density at P19 ( approximately 15%), suggesting that the NMDA-induced increase in GABA(A) receptor currents may protect against excitotoxicity.     

肾血管性高血压大鼠尾壳核NPY阳性神经元的变化

目的 :研究易卒中型肾血管性高血压大鼠尾壳核 NPY阳性神经元的变化。方法 :雄性 SD大鼠 6 0只 ,随机分为两组 (n=30 )。手术组用单肾单夹法复制高血压模型 ,假手术组手术过程同手术组 ,但不上银夹。于术后 7d,30 d,90 d处死动物 ,取脑 ,用 ABC免疫组化方法对大鼠尾壳核 NPY阳性神经元进行定量分析。结果 :假手术组尾壳核 NPY阳性神经元密度随年龄增加而减少 ,而手术组 NPY阳性神经元密度先减少 ,再增加 ,后保持不变。手术组和假手术组 NPY神经元密度相比 ,早期两组间无显著差异 ;中期手术组小于假手术组 (P<0 .0 5 ) ;晚期 ,手术组大于假手术组 (P<0 .0 5 )。结论 :NPY阳性神经元在高血压的发生、发展过程中起一定调控作用。     

大鼠视觉中枢生后发育过程中的一氧化氮合酶阳性神经元的形态及分布变化

本文用NADPH-黄递酶组织化学技术研究了生后不同年龄段（0、7、14、21、28、35、60、120d）Wistar大鼠视皮层（17区）和上丘表层中一氧化氮合酶阳性神经元的形态及分布的变化。结果表明：视皮层17区中的一氧化氮合酶阳性神经元在生后7d时开始出现，但胞体小，树突分枝少而短，以双极细胞为主，占68.0％;14d时数量达到高峰;且14～21d中，该神经元胞体截面积明显增大，树突分枝复杂化，长度增加，多极细胞占64％;28d后一氧化氮合酶阳性神经元胞体面积，树突分枝长度明显减小，分枝简单化;35d后接近成年动物水平（120d）.生后7d时，一氧化氛合酶阳性神经元主要位于视区的白质和皮层的5、6层中，7d后该神经元及神经纤维在第2／3及4层中明显增多，这种分布状态在35d后趋于稳定。上丘表层中的一氧化氮合酶阳性细胞在生后第2周逐渐出现，第3周迅速增加，第4周达最大值，以后下降并趋于稳定水平。结果提示：大鼠生后发育过程中视觉中枢的一氧化氮合酶阳性神经元数量及形态变化与视觉发育的可塑性有关。     

Morphology and dendritic maturation of developing principal neurons in the rat basolateral amygdala

The basolateral nucleus of the amygdala (BLA) assigns emotional valence to sensory stimuli, and many amygdala-dependent behaviors undergo marked development during postnatal life. We recently showed principal neurons in the rat BLA undergo dramatic changes to their electrophysiological properties during the first postnatal month, but no study to date has thoroughly characterized changes to morphology or gene expression that may underlie the functional development of this neuronal population. We addressed this knowledge gap with reconstructions of biocytin-filled principal neurons in the rat BLA at postnatal days 7 (P7), 14, 21, 28, and 60. BLA principal neurons underwent a number of morphological changes, including a twofold increase in soma volume from P7 to P21. Dendritic arbors expanded significantly during the first postnatal month and achieved a mature distribution around P28, in terms of total dendritic length and distance from soma. The number of primary dendrites and branch points were consistent with age, but branch points were found farther from the soma in older animals. Dendrites of BLA principal neurons at P7 had few spines, and spine density increased nearly fivefold by P21. Given the concurrent increase in dendritic material, P60 neurons had approximately 17 times as many total spines as P7 neurons. Together, these developmental transitions in BLA principal neuron morphology help explain a number of concomitant electrophysiological changes during a critical period in amygdala development.     

Developmental changes in NMDA receptor-mediated visual activity in the rat superior colliculus, and the effect of dark rearing

N-methyl-D-aspartate (NMDA) receptor-mediated activity is considered important for experience-dependent plasticity in the developing visual system. We investigated the influence of age and experience on the role of NMDA receptors in the visual transmission in the superficial grey layer of the superior colliculus (SGS) of the superior colliculus, where, in the adult, NMDA receptors mediate a substantial part of the visual response. In normally reared (postnatal day 14, P14, to adult) rats, visual responses were challenged with NMDA receptor-selective iontophoretic applications of the antagonist D-2-amino-5-phosphonovalerate (AP5). After eye opening (at P14), there was a significant increase in the number of neurones whose visual responses were reduced during AP5 ejection, which peaked at P22 (85%; n = 21), and then declined to adult levels (66%; n = 47) at P25. The mean reduction of the response (from control levels) by AP5 was similar at all ages (approximately 40%). Dark rearing had striking effects on the role of NMDA receptors in visual transmission, especially when comparisons were made between age-matched subjects greater than P25. In these subjects, AP5 ejection reduced the visual responses of all neurones studied. In addition, AP5 ejection caused a significantly larger reduction of visual responses in dark-reared rats (mean reduction 62 ± 4; n = 29) compared with age-matched controls (mean reduction 44 ± 8; n = 23). The D,L-alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) reduced the visual responses of every neurone studied and there were no age- or experience-dependent effects. We conclude that NMDA receptors, but not AMPA receptors, assume greater importance for visual transmission in the SGS of dark-reared rats. Received: 2 May 1997 / Accepted: 19 November 1997     

N-methyl-d-aspartate receptor activation exerts a dual control on postnatal development of nucleus tractus solitarii neurons in vivo

We have used a morphological approach to evaluate the role of NMDA receptors (NMDAR) in postnatal development of brainstem neurons in awake rats. Chronic NMDAR blockade was performed by placing drug-impregnated Elvax implants over the brainstem at the fifth postnatal day (P5). Compared with control, NMDAR blockade led to a transient increase in dendritic arbor area and filopodium density until P12 followed by a rapid decline in both parameters. Electron microscopy observations showed that these changes correlated with an increase in synapse density at P14 followed by a decrease in synapse density at P28 if chronic NMDAR blockade was maintained until P21. These results support the hypothesis that synapse formation does not require NMDAR activation. In addition, our data suggest a dual role for NMDAR in controlling the synapse number. Early in development NMDARs may be involved in controlling the rate of synapse elimination. Later on, they may subserve synapse stabilization. The physiological significance of these results is discussed.     

A morphological analysis of the cluster organization of the cortico-cortical neurons in area 17 of the cat visual cortex

Spatial organisation and quantitative features of cortico-cortical neurons in area 17 that project to area 21a of cat visual cortex were investigated. We used the HRP technique and two-dimensional reconstruction from serial sections of brain. Pattern of distribution of labelled cells in some regions of areas 17 and 18, on the lateral and medial surface of the brain were analysed. The data about cluster organisation of cortico-cortical neurones, localisation, layer distribution and density of neurons were obtained.     

A comparison of the postnatal development of muscle-spindle and periodontal-ligament neurons in the mesencephalic trigeminal nucleus of the rat

The trigeminal mesencephalic nucleus (Vmes) is known to include primary afferent neurons of jaw muscle spindles (MS neurons) and periodontal ligament receptors (PL neurons). The aim of this study was to clarify the postnatal development of Vmes neurons by comparing MS neurons with PL neurons using horseradish peroxidase labeling. We measured somal diameter and somal shape of MS and PL neurons in rats from postnatal day (P)7 to P70. No significant changes were seen between postnatal day P7 and P70 in somal diameter or somal shape of MS neurons. Conversely, PL neurons showed a larger somal diameter at P7 than at P14, and in terms of somal profile, multipolar neurons comprised 0% at P7, but 4.8% at P14 and 16.9% at P70. These findings suggest that PL neurons develop with the eruption of teeth, taking into account the fact that tooth eruption occurs from around P14 in rats. Conversely, the lack of postnatal changes in MS neurons is due to the fact that these neurons have been active since the embryonic period, as swallowing starts in utero.     

Transgeniculate signal transmission to middle suprasylvian cortex in intact cats and following early removal of areas 17 and 18: a morphological study

 Removal of cat areas 17 and 18 early, but not late, in postnatal development results in the sparing of certain reflexive and nonreflexive visually guided behaviors. These spared behaviors are accompanied by an expansion of geniculocortical projections to middle suprasylvian (MS) cortex. However, little is known about the types of visual signals relayed along these pathways. The purpose of our study was to reveal the morphologies of the neurons participating in the rewired circuits and, by relating them to the morphologies of functionally characterized neurons described by others, infer the types of visual signals transmitted via the lateral geniculate nucleus (LGN) to MS cortex. To do this, we retrogradely labeled LGN neurons from MS cortex with fluorescent microspheres, and subsequently intracellularly filled them with Lucifer Yellow. We then classified well-filled neurons according to a battery of morphological parameters (such as soma size and shape, and dendritic field-form and specializations), and compared them with already defined structure/function relationships. By doing this, we found that the large majority of visual thalamic relay neurons to MS cortex of both normal cats and cats that incurred removal of areas 17 and 18 were types I and IV. These results indicate that visual Y and W signals, respectively, are relayed directly from LGN to MS cortex in both types of cats. Following the early lesions, some of the MS-projecting type I neurons were found in layers A and A1, where they are never found in intact cats. Thus, some layer A and A1 type I neurons redirect axons to MS cortex following early removal of areas 17 and 18. For the type IV MS-projecting neurons in early lesioned cats, the somas were hypertrophied and they had more profuse and broader dendritic arbors than equivalent neurons in intact cats. These results suggest that dynamic interactions take place between inputs and outputs of LGN neurons during development that influence final LGN neuron morphology. Moreover, they suggest that signals transferred to MS cortex by type IV neurons may be modified by early lesions of areas 17 and 18. Overall, these results contribute to our understanding of the types of behaviors that may be spared by early lesions of areas 17 and 18. Received: 22 May 1996 / Accepted: 3 September 1996     

Strabismus modifies intrinsic and inter-areal connections in cat area 18

The development of long-range horizontal connections depends on visual experience. Previous experiments have shown that in area 17 of strabismic but not in normal cats, horizontal fibers preferentially connect cell groups driven by the same eye indicating that fibers between coactive neurons are selectively stabilized. To test whether this is a general organizing principle of intracortical long-range circuitry we extended our analyses to both intrinsic horizontal connections within area 18 and to inter-areal connections between areas 17 and 18. To this end, we visualized the functional architecture of area 18 by intrinsic signal imaging. Horizontal circuitry was labeled by injecting fluorescent latex microspheres into functionally identified domains. Additionally, domains sharing the same ocular dominance as the neurons at the injection sites were visualized by 2-deoxyglucose autoradiography to allow comprehensive labeling of functional domains in regions far from the injection sites. Quantitative analyses revealed that in strabismic cats, 72% of the retrogradely labeled neurons in area 18 and 68% of the neurons in area 17 were located in the same ocular dominance domains as the injection sites. In contrast, these numbers were 52% and 54% in normal animals. These data show that experience modifies both intrinsic connections within area 18 and inter-areal projections from area 17 to area 18 as has been previously described for intrinsic and callosal connections in area 17. This provides further evidence for the hypothesis that the correlation of activity is a major selection criterion for the stabilization of neuronal circuits during postnatal development.