P.L.A. Gabbott

Prefrontal cortex in the rat: Projections to subcortical autonomic, motor, and limbic centers

This paper describes the quantitative areal and laminar distribution of identified neuron populations projecting from areas of prefrontal cortex (PFC) to subcortical autonomic, motor, and limbic sites in the rat. Injections of the retrograde pathway tracer wheat germ agglutinin conjugated with horseradish peroxidase (WGA‐HRP) were made into dorsal/ventral striatum (DS/VS), basolateral amygdala (BLA), mediodorsal thalamus (MD), lateral hypothalamus (LH), mediolateral septum, dorsolateral periaqueductal gray, dorsal raphe, ventral tegmental area, parabrachial nucleus, nucleus tractus solitarius, rostral/caudal ventrolateral medulla, or thoracic spinal cord (SC). High‐resolution flat‐map density distributions of retrogradely labelled neurons indicated that specific PFC regions were differentially involved in the projections studied, with medial (m)PFC divided into dorsal and ventral sectors. The percentages that WGA‐HRP retrogradely labelled neurons composed of the projection neurons in individual layers of infralimbic (IL; area 25) prelimbic (PL; area 32), and dorsal anterior cingulate (ACd; area 24b) cortices were calculated. Among layer 5 pyramidal cells, approximately 27.4% in IL/PL/ACd cortices projected to LH, 22.9% in IL/ventral PL to VS, 18.3% in ACd/dorsal PL to DS, and 8.1% in areas IL/PL to BLA; and 37% of layer 6 pyramidal cells in IL/PL/ACd projected to MD. Data for other projection pathways are given. Multiple dual retrograde fluorescent tracing studies indicated that moderate populations (<9%) of layer 5 mPFC neurons projected to LH/VS, LH/SC, or VS/BLA. The data provide new quantitative information concerning the density and distribution of neurons involved in identified projection pathways from defined areas of the rat PFC to specific subcortical targets involved in dynamic goal‐directed behavior. J. Comp. Neurol. 492:145–177, 2005.

Stress suppresses and learning induces plasticity in CA3 of rat hippocampus: a three-dimensional ultrastructural study of thorny excrescences and their postsynaptic densities

Chronic stress and spatial training have been proposed to affect hippocampal structure and function in opposite ways. Previous morphological studies that addressed structural changes after chronic restraint stress and spatial training were based on two-dimensional morphometry which does not allow a complete morphometric characterisation of synaptic features. Here, for the first time in such studies, we examined these issues by using three-dimensional (3-D) reconstructions of electron microscope images taken from thorny excrescences of hippocampal CA3 pyramidal cells. Ultrastructural alterations in postsynaptic densities (PSDs) of thorny excrescences receiving input from mossy fibre boutons were also determined, as were changes in numbers of multivesicular bodies (endosome-like structures) within thorny excrescences and dendrites. Quantitative 3-D data demonstrated retraction of thorny excrescences after chronic restraint stress which was reversed after water maze training, whilst water maze training alone increased thorny excrescence volume and number of thorns per thorny excrescence. PSD surface area was unaffected by restraint stress but water maze training increased both number and area of PSDs per thorny excrescence. In restrained rats that were water maze trained PSD volume and surface area increased significantly. The proportion of perforated PSDs almost doubled after water maze training and restraint stress. Numbers of endosome-like structures in thorny excrescences decreased after restraint stress and increased after water maze training. These findings demonstrate that circuits involving contacts between mossy fibre terminals and CA3 pyramidal cells at stratum lucidum level are affected conversely by water maze training and chronic stress, confirming the remarkable plasticity of CA3 dendrites. They provide a clear illustration of the structural modifications that occur after life experiences noted for their different impact on hippocampal function.

Remodelling of synaptic morphology but unchanged synaptic density during late phase long-term potentiation(ltp): A serial section electron micrograph study in the dentate gyrus in the anaesthetised rat

In anaesthetised rats, long-term potentiation (LTP) was induced unilaterally in the dentate gyrus by tetanic stimulation of the perforant path. Animals were killed 6 h after LTP induction and dendritic spines and synapses in tetanised and untetanised (contralateral) hippocampal tissue from the middle molecular layer (MML) were examined in the electron microscope using stereological analysis. Three-dimensional reconstructions were also used for the first time in LTP studies in vivo, with up to 130 ultrathin serial sections analysed per MML dendritic segment. A volume sampling procedure revealed no significant changes in hippocampal volume after LTP and an unbiased counting method demonstrated no significant changes in synapse density in potentiated compared with control tissue. In the potentiated hemisphere, there were changes in the proportion of different spine types and their synaptic contacts. We found an increase in the percentage of synapses on thin dendritic spines, a decrease in synapses on both stubby spines and dendritic shafts, but no change in the proportion of synapses on mushroom spines. Analysis of three-dimensional reconstructions of thin and mushroom spines following LTP induction revealed a significant increase in their volume and area. We also found an increase in volume and area of unperforated (macular) and perforated (segmented) postsynaptic densities. Our data demonstrate that whilst there is no change in synapse density 6 h after the induction of LTP in vivo, there is a considerable restructuring of pre-existing synapses, with shaft and stubby spines transforming to thin dendritic spines, and mushroom spines changing only in shape and volume.

Hemispheric asymmetry of synapses in chick medial hyperstriatum ventrale following passive avoidance training: a stereological investigation

A stereological analysis was made of synapses in the left and right hemispheres of chick medial hyperstriatum ventrale (MHV) 24h after passive avoidance training (PAL) and in water trained controls (W-control). The synaptic parameters examined were (D), the mean length of the postsynaptic thickening; (NV.syn), the number of synapses per unit volume of neuropil; (VV.syn), the volume density of the pre-synaptic bouton; (NV.ves), the number of synaptic vesicles per unit volume of neuropil and (ves.syn), the mean number of synaptic vesicles per pre-synaptic bouton. No significant differences exist in NV.syn between the left or right hemispheres of W-control and trained chicks, nor is NV.syn influenced by training. However, in W-control chicks D in the right MHV is significantly greater (12%) than in the left MHV and this difference disappears on training. There are no differences in VV.syn between left and right hemispheres of W-control chicks but following training VV.syn is 22.7% greater in the left MHV than in the right MHV. Training als influences the number of synaptic vesicles; in W-control chicks NV.ves in the right MHV is 12.25% greater than in the left MHV but following training these differences are reversed. When the data are expressed as numbers of vesicles per synapse (ves.syn), values for the left hemisphere of trained chicks exceed those in the right hemisphere by a staggering 61.38%. These results are discussed in the context of biochemical and electrophysiological studies which suggest that there is lateralization of the memory trace.

Distribution of neurons and glia in the visual cortex (area 17) of the adult albino rat: A quantitative description

The neuronal and glial cell composition of the rat visual cortex (area 17) has been determined quantitatively using stereological techniques. The volume numerical densities (number of cells per mm3 of cortex) of neurons and of the principal glial cell types (astroglia, oligodendroglia, and microglia) were calculated from tangential semithin resin sections spaced at regular intervals 50 micron apart throughout the entire depth of the visual cortex. From measurements of cortical and laminar thickness the separate volume numerical densities of neurons and glial cells were derived for each lamina in the cortex. In addition, the absolute numbers of cells in each lamina under 1 mm2 of cortical surface were calculated. The mean cortical volume numerical density of neurons was 60,020 +/- 3840/mm3 (mean +/- SEM; n = 8), and 49,040 +/- 2610/mm3 for the combined glial cell types. Astroglia, oligodendroglia, and microglia were present in a ratio of 6:3:1 respectively. It was determined from neuronal and glial somatic volume estimates that the somata of these cells occupied approximately 13.5% of unit cortical volume, with 81.3% of the unit volume being occupied by cortical neuropil. Using previously published reports that described the laminar composition of neurons in terms of the relative proportions of pyramidal and non-pyramidal cells, the laminar volume numerical densities for these neuronal categories have been derived. In addition, it has been estimated that under 1 mm2 of cortical surface there are 79,500 pyramidal and 7790 non-pyramidal neurons distributed throughout layers 1-6 of the rat visual cortex.

Calretinin neurons in human medial prefrontal cortex (areas 24a,b,c, 32′, and 25)

The calcium‐binding protein calretinin (CR) is present in a subpopulation of local‐circuit neurons in the mammalian cerebral cortex containing γ‐aminobutyric acid. This light microscopic investigation provides a detailed qualitative and quantitative morphological analysis of CR‐immunoreactive (CR+) neurons in the medial prefrontal cortex (mPFC; areas 24a,b,c, 32′, and 25) of the normal adult human.

Differences in the laminar origin of projections from the medial prefrontal cortex to the nucleus accumbens shell and core regions in the rat

The medial prefrontal cortex (mPFC) projects to the nucleus accumbens shell, core and rostral pole. In this retrograde tract-tracing study of rat mPFC to nucleus accumbens projection neurons, the advantages of Neurobiotin are utilised in order to reveal the detailed morphology of labelled projection cells, and to permit an examination of the laminar projections to shell and core compartments The retrogradely transported Neurobiotin was found in somata, proximal and distal dendrites of neurons that project from the mPFC to the nucleus accumbens. The morphology of these projection neurons was revealed in great detail and confirmed that the projection arises wholly from pyramidal cells. Interestingly, it was also found that retrogradely labelled neurons were exclusively located in prelimbic and infralimbic regions in layers V and VI, after shell injections, but also in layer II following core sites. This observation may reflect possibly different roles for cortical laminae on the nucleus accumbens.

A quantitative investigation of the neuronal composition of the rat dorsal lateral geniculate nucleus using GABA-immunocytochemistry

The proportion of neurons immunoreactive for gamma-aminobutyric acid (GABA), and their rostrocaudal distribution in the dorsal lateral geniculate nucleus of the rat, were determined quantitatively using post-embedding GABA-immunochemistry on semithin resin embedded coronal sections followed by stereological analysis. The mean total volume numerical density of neurons (total number of neurons per mm3) in the dLGN was 67,077 +/- 4412 mm-3 (mean +/- SEM; n = 5), comprising a mean volume numerical density for GABA-immunopositive neurons of 14,584 +/- 1324 mm-3, and a mean volume numerical density of GABA-immunonegative neurons of 52,493 +/- 3419 mm-3, GABA-immunopositive neurons constituted 21.7 +/- 0.5% of the total neuronal composition of the rat dorsal lateral geniculate nucleus. Although no rostrocaudal variation was detected in the total volume numerical density of neurons, the relative proportion of GABA-immunopositive neurons was significantly lower in the caudal segment (18.1 +/- 0.6%) compared with the middle (24.9 +/- 0.9%) and the rostral segments (22.1%). Furthermore, on the basis of somatic size distributions, GABA-immunonegative neurons were seen to be significantly smaller in the caudal segment than in the more anterior two segments. The somatic size of GABA-immunopositive neurons showed no rostrocaudal variation through the dorsal lateral geniculate nucleus. These data provide a morphological correlate for the structural and functional subdivision of the dorsal lateral geniculate nucleus described previously in electrophysiological and morphological studies.

A cell adhesion molecule mimetic, FGL peptide, induces alterations in synapse and dendritic spine structure in the dentate gyrus of aged rats: a three-dimensional ultrastructural study

The FGL peptide is a neural cell adhesion molecule (NCAM) mimetic comprising a 15‐amino‐acid‐long sequence of the FG loop region of the second fibronectin type III module of NCAM. It corresponds to the binding site of NCAM for the fibroblast growth factor receptor 1. FGL improves cognitive function through enhancement of synaptic function. We examined the effect of FGL on synaptic and dendritic structure in the brains of aged (22‐month‐old) rats that were injected subcutaneously (8 mg/kg) at 2‐day intervals until 19 days after the start of the experiment. Animals were perfused with fixative, brains removed and coronal sections cut at 50 µm. The hippocampal volume was measured, tissue embedded and ultrathin sections viewed in a JEOL 1010 electron microscope. Analyses were made of synaptic and dendritic parameters following three‐dimensional reconstruction via images from a series of ∼100 serial ultrathin sections. FGL affected neither hippocampal volume nor spine or synaptic density in the middle molecular layer of the dentate gyrus. However, it increased the ratio of mushroom to thin spines, number of multivesicular bodies and also increased the frequency of appearance of coated pits. Three‐dimensional analysis showed a significant decrease in both post‐synaptic density and apposition zone curvature of mushroom spines following FGL treatment, whereas for thin spines the convexity of the apposition zone increased. These data indicate that FGL induces large changes in the fine structure of synapses and dendritic spines in hippocampus of aged rats, complementing data showing its effect on cognitive processes.

Quantitative morphological effects of dark-rearing and light exposure on the synaptic connectivity of layer 4 in the rat visual cortex (area 17)

SummaryThe quantitative effects of dark-rearing and light exposure on the ultrastructural characteristics of synapses and synaptic boutons in layer 4 of the rat visual cortex (area 17) have been investigated using stereological techniques. Two experimental groups (each containing 5 animals) were investigated i) animals dark-reared upto weaning at 21 days post natum (21DPN) and then light exposed until 52DPN (Group 21/31), and ii) littermate animals totally dark-reared until 52DPN (Group 52dD). The results indicate a significantly higher mean density of synapses in the neuropil of layer 4 in group 21/31 (3.58×108 · mm-3) compared with group 52dD (2.68×108 · mm-3). Although the density per unit volume of synapses with identified asymmetrical synaptic membrane specialisations was not significantly different in group 21/31 than in group 52dD (but was significantly lower than animals reared normally), the density of synapses with identified symmetrical synaptic membrane specialisations was about 200% higher in group 21/31 versus group 52dD. However, significant differences were detected in the number of asymmetrical synapses established by single synaptic boutons in group 21/31 (1.21 ± 0.11) compared with group 52dD (1.10 ± 0.09). On the basis of the numbers of post-synaptic targets contacted by an individual synaptic bouton, a significantly higher density of synaptic boutons was found in group 21/31 (2.32×108 · mm-3) compared with group 52dD (1.82×108 · mm-3). Furthermore, planar quantitative data indicated significant inter-group differences in the ultrastructure of asymmetrical and symmetrical synaptic boutons. The results of this study provide evidence indicating marked structural alterations in the synaptic connectivity of layer 4 of the rat visual cortex following the light exposure of rats dark-reared upto weaning. Indeed visual deprivation severely affected the ‘inhibitory’ circuitry in the major thalamorecipient territory of the visual cortex.