Layer Ia Research Articles

Projections from the anterodorsal and anteroveniral nucleus of the thalamus to the limbic cortex in the rat

The present study characterized the projections of the anterodorsal (AD) and the anteroventral (AV) thalamic nuclei to the limbic cortex. Both AD and AV project to the full extent of the retrosplenial granular cortex in a topographic pattern. Neurons in caudal parts of both nuclei project to rostral retrosplenial cortex, and neurons in rostral parts of both nuclei project to caudal retrosplenial cortex. Within AV, the magnocellular neurons project primarily to the retrosplenial granular a cortex, whereas the parvicellular neurons project mainly to the retrosplenial granular b cortex. AD projections to retrosplenial cortex terminate in very different patterns than do AV projections: The AD projection terminates with equal density in layers I, III, and IV of the retrosplenial granular cortex, whereas, in contrast, the AV projections terminate very densely in layer Ia and less densely in layer IV. Further, both AD and AV project densely to the postsubicular, presubicular, and parasubicular cortices and lightly to the entorhinal (only the most caudal part) cortex and to the subiculum proper (only the most septal part). Rostral parts of AD project equally to all three subicular cortices, whereas neurons in caudal AD project primarily to the postsubicular cortex. Compared to AD, neurons in AV have a less extensive projection to the subicular cortex, and this projection terminates primarily in the postsubicular and presubicular cortices. Further, the AD projection terminates in layers I, II/III, and V of postsubiculum, whereas the AV projection terminates only in layers I and V.

Selective suppression of intrinsic but not afferent fiber synaptic transmission by baclofen in the piriform (olfactory) cortex

The GABA B agonist baclofen has been shown to suppress synaptic transmission in subregions of the hippocampus and in the piriform (olfactory) cortex. Here we report a laminar selectivity of suppression of synaptic potentials in the olfactory cortex. In brain slice preparations, baclofen suppresses extracellularly recorded field potentials at the intrinsic fiber synapses proximal to the superficial pyramidal cell bodies (layer Ib) while leaving the af ferent fiber synaptic potentials recorded at the distal dendrites (layer Ia) little affected. This dose-dependent selective suppression of intrinsic fiber synaptic transmission is also correlated with an increase of paired-pulse facilitation. These results suggest that afferent and intrinsic synaptic inputs may be differentially modulated by the activation of GABA B receptors and that this selective suppression is at least partially mediated via a presynaptic mechanism.

Cortical areas are revealed by distribution patterns of proteoglycan components and parvalbumin in the Mongolian gerbil and rat

Cortical areas in rodents have been basically characterized by its cytoarchitecture, connectivity or by physiological parameters. In this study we show that they are also revealed by distribution patterns of proteoglycans and parvalbumin-immunoreactivity. Brains of young adult Mongolian gerbils ( Meriones unguiculatus) and Wistar rats were cut into series of transversal sections. Proteoglycan components were detected using the N-acetylgalactosamine binding Wisteria floribunda agglutinin (WFA) and antibodies against chondroitin sulphate proteoglycan (CSPG). Differences between cortical areas were found to exist with regard to the occurrence and the density of perineuronal nets, but were also expressed in varying staining intensities for WFA and CSPG of the neuropil. Primary neocortical areas (somatosensory, auditory, visual cortex) were characterized by an intense neuropil staining in layer IV and the upper part of layer VI. Using the same methods strong labelling was also typical of the neuropil in the retrosplenial cortex, of layer Ia in the prepiriform cortex and the hippocampal CA3 field. In tangential sections cut from gerbil cortical hemispheres, some of the heavily lectin-stained cortical areas were sharply delineated from adjacent faintly labelled regions, others showed more diffuse borders. In the rat, the area-specific staining for WFA was less clearly expressed than in the gerbil. Immunocytochemistry of the calcium-binding protein parvalbumin in alternate sections showed labelling patterns of neuropil which resembled those of WFA-binding and CSPG-immunoreactivity in the entire neocortex and hippocampus. From these results it can be concluded that functional peculiarities of cortical fields may not only be determined by neuronal network parameters but also by the spatial arrangement of extracellular matrix proteoglycans.

Laminar distribution and neuronal targets of GABAergic axon terminals in cat primary auditory cortex (AI)

The form, density, and neuronal targets of presumptive axon terminals (puncta) that were immunoreactive for gamma-aminobutyric acid (GABA) or its synthesizing enzyme, glutamic acid decarboxylase (GAD), were studied in cat primary auditory cortex (AI) in the light microscope. High-resolution, plastic-embedded material and frozen sections were used. The chief results were: 1) There was a three-tiered numerical distribution of puncta, with the highest density in layer Ia, an intermediate number in layers Ib-IVb, and the lowest concentration in layers V and VI, respectively. 2) Each layer had a particular arrangement: layer I puncta were fine and granular (less than 1 micron in diameter), endings in layers II-IV were coarser and more globular (larger than 1 micron), and layer V and VI puncta were mixed in size and predominantly small. 3) The form and density of puncta in every layer were distinctive. 4) Immunonegative neurons received, in general, many more axosomatic puncta than immunopositive cells, with the exception of the large multipolar (presumptive basket) cells, which invariably had many puncta in layers II-VI. 5) The number of puncta on the perikarya of GABAergic neurons was sometimes related to the number of puncta in the layer, and in other instances it was independent of the layer. Thus, while layer V had a proportion of GABAergic neurons similar to layer IV, it had only a fraction of the number of puncta; perhaps the intrinsic projections of supragranular GABAergic cells are directed toward layer IV, as those of infragranular GABAergic neurons may be. Since puncta are believed to be the light microscopic correlate of synaptic terminals, they can suggest how inhibitory circuits are organized. Even within an area, the laminar puncta patterns may reflect different inhibitory arrangements. Thus, in layer I the fine, granular endings could contact preferentially the distal dendrites of pyramidal cells in deeper layers. The remoteness of such terminals from the spike initiation zone contrasts with the many puncta on all pyramidal cell perikarya and the large globular endings on basket cell somata. Basket cells might receive feed-forward disinhibition, pyramidal cells feed-forward inhibition, and GABAergic non-basket cells would be the target of only sparse inhibitory axosomatic input. Such arrangements imply that the actions of GABA on AI neurons are neither singular nor simple and that the architectonic locus, laminar position, and morphological identity of a particular neuron must be integrated for a more refined view of its role in cortical circuitry.

Optical imaging of the in vitro guinea pig piriform cortex activity using a voltage-sensitive dye

The spatio-temporal patterns of signal processing in guinea pig piriform cortex (PC) slices were analyzed by optical imaging using a voltage-sensitive dye. Slices (400 μm thick) were cut in a plane parallel to the lateral olfactory tract and perpendicular to the cortical surface. In all the anterior PC and the majority of the posterior PC preparations, neural activity elicited by electrical stimulation of layer Ia propagated along the same layer, then it invaded into layers II and III and propagated along them. In addition to the above pattern, invasion of activity into the deeper area than layer III was observed in some posterior PC preparations. Real-time imaging of an active zone evoked by Ia shocks and its spatio-temporal behavior will contribute to resolving olfactory information processing.

Sylvian fissure length and angle asymmetries in normal MZ twins and in MZ twins discordant for schizophrenia

Anterior cingulate cortex (ACC) has one of the highest densities of opioid receptors in the CNS and it has been implicated in acute and chronic pain responses. Little is known, however, about which neurons express opioid receptors in their dendrites and axon terminals. The present studies employed experimental techniques to remove afferent axons or classes of projection neurons from ratACC area 24 followed by coverslip autoradiography to localize changes in binding of [3H]Tyr-d-Ala-Gly-MePhe-Gly-ol (DAMGO) to mu receptors and 2-[3H]d-penicillamine-5-d-penicillamine-en-kephalin (DPDPE) to delta receptors. Removal of all afferents to area 24 with undercut lesions did not alter DPDPE binding, but significantly reduced binding of DAMGO in layers I, III, and V. In contrast, removal of all cortical neurons with the excitotoxin ibotenic acid almost abolished DPDPE binding in all layers. The same lesions reduced DAMGO binding in most layers; however, there was a postlesion bimodal distribution in binding with high levels of binding in layer I and moderate levels in layer VI. These data suggest that delta receptors are expressed by cortical neurons, while mu receptors are expressed by both cortical neurons and afferent axons. To explore the distribution of postsynaptic receptors, immunotoxin lesions were made in area 24 by injection of OX7-saporin into the caudate and/or thalamic nuclei. Almost complete removal of projection neurons to these targets in layers Vb and VIa did not alter DPDPE binding, while the lesions reduced DAMGO binding in all but layer II. Removal of layer Vb corticostriatal projection neurons with caudate OX7-saporin injections reduced binding only in this layer. It is proposed that opioidergic circuits in area 24 are organized according to an input/output model for mu opioid regulation. In this model mu receptors regulate axon terminal activity from the thalamus in layer Ia and the locus coeruleus in layers Ic and II, whereas cortical outputs to the thalamus are modulated via postsynaptic receptors expressed in all layers by thalamocortical projection neurons with somata in layer VI. These opioidergic circuits in ACC are of particular importance because they may regulate responses to chronic nociceptive activity and associated pain perceptions.

Membrane currents evoked by afferent fiber stimulation in rat piriform cortex. I. Current source-density analysis.

1. The membrane currents evoked by afferent fiber stimulation in the piriform cortex were derived by the use of current source-density (CSD) analysis in the rat under urethan anesthesia. The primary goals were to test hypotheses concerning the sequence of synaptic events evoked by afferent fiber stimulation and to derive data required for development and testing of the model presented in the companion paper. 2. In confirmation of previous studies, it was found that afferent fiber stimulation evokes a monosynaptic excitatory postsynaptic current (EPSC) in distal segments of pyramidal cell apical dendrites (layer Ia) followed by a strong disynaptic EPSC in adjacent middle segments (superficial layer Ib). 3. Given the central importance of the strong disynaptic EPSC in models for operation of the piriform cortex, the hypothesis that it is mediated by long association fibers from the anterior piriform cortex was tested by comparing its latency in response to stimulation at anterior and posterior locations. The results confirmed the hypothesis and ruled out a significant contribution from local connections in the posterior piriform cortex. 4. Intensification of pyramidal cell activity by spatially restricted disinhibition with picrotoxin confirmed the hypothesis that associational projections from the posterior piriform cortex can mediate a long-latency disynaptic EPSC in proximal dendritic segments (mid to deep layer Ib) in the anterior piriform cortex. 5. Analysis of the time course of the monosynaptic EPSC in different areas revealed that activation of the anterior piriform cortex from afferent fiber stimulation is fast and nearly synchronous throughout its extent as a result of the relatively high conduction velocities of afferent fibers in the lateral olfactory tract (LOT). By contrast, the posterior piriform cortex is sequentially activated by this EPSC as a consequence of the slow propagation velocity of afferent fiber collaterals that course across its surface. This activation is sufficiently slow that a large phase lag is present between rostral and caudal regions. 6. The time courses of the monosynaptic and principal disynaptic EPSCs changed in characteristically different ways with increasing distance from the LOT within the posterior piriform cortex. Simulations in the companion paper indicate that initiation and propagation patterns for activity in fiber systems rather than differences in synaptic conductance waveforms are responsible for these differences. 7. Although the laminar distribution of the active inward current component of the monosynaptic EPSC remained constant over time, the peak outward current associated with this EPSC shifted from the depth of proximal apical dendrites (layer Ib) to the depth of superficial pyramidal cell somata (layer II).(ABSTRACT TRUNCATED AT 400 WORDS)

Distribution of corticotropin-releasing factor-like immunoreactivity in squirrel monkey (Saimiri sciureus) amygdala.

Previous anatomical studies of corticotropin-releasing factor (CRF)-like immunoreactivity in rat brain have reported prominent clustering of neuronal elements containing this peptide within the amygdala. The highest concentrations of both CRF-positive cells and fibers were evident in the central nucleus, an observation consistent with the putative role of this peptide in autonomic and endocrine regulation. In addition, lower densities of CRF-positive somata and processes have been noted in other amygdaloid nuclei. However, the distribution of CRF-like immunoreactivity in the amygdala has not been described for any primate species. Such a description would be of interest since substantial differences in the distribution of CRF in rodent and primate have been reported for other brain regions. The present study uses immunohistochemical methods, with a polyclonal antiserum directed against the human form of CRF, to determine the distribution of this peptide in non-colchicine-treated monkeys (Saimiri sciureus). Within the amygdaloid complex, the most numerous and concentrated collections of CRF-positive neurons were seen in the basal and lateral nuclei. The highest densities of CRF-positive fibers and terminals were seen in the lateral and central amygdaloid nuclei. Moderately dense plexuses of CRF-positive fibers also were seen in layer Ia of the periamygdaloid cortex, nucleus of the lateral olfactory tract, anterior and posterior cortical nuclei, and the medial nucleus. Thus, the distribution of CRF-like immunoreactivity differs substantially in monkey and rat amygdala. Since CRF-positive perikarya in monkey are most prominent in nuclei with pronounced interconnections with neocortex, these differences may be an integral component of the increased cortical development that characterizes the primate brain.

Cholinergic suppression specific to intrinsic not afferent fiber synapses in rat piriform (olfactory) cortex.

1. Differences in the cholinergic suppression of afferent and intrinsic fiber synaptic transmission were studied in the rat piriform cortex. Extracellular and intracellular recording techniques were applied in an in vitro transverse slice preparation. Afferent and intrinsic fiber systems were differentially stimulated with electrodes placed in layer Ia or layer Ib, respectively. Synaptic responses were monitored in the presence of cholinergic agonists and antagonists. 2. Afferent and intrinsic fiber synaptic potentials measured extracellularly showed large differences in sensitivity to micromolar concentrations of the cholinergic agonists carbachol or (+/-)-muscarine, or to acetylcholine combined with neostigmine. Intrinsic fiber synaptic responses in layer Ib were strongly reduced in the presence of cholinergic agonists, whereas afferent fiber synaptic responses in layer Ia were largely unaffected. At a concentration of 100 microM, all three agonists caused a greater than 60% decrease in the height of the intrinsic fiber synaptic potential but less than 15% reduction in the afferent fiber synaptic potential. 3. Intracellular recordings confirmed that the cholinergic agonist carbachol selectively suppresses intrinsic fiber synaptic potentials but not afferent fiber synaptic potentials recorded from the same pyramidal cell. 4. Dose-response curves to carbachol were obtained for both fiber systems using extracellular recording of evoked field potentials. Carbachol suppressed intrinsic fiber synaptic potentials with a coefficient of dissociation (KD) estimated at 2.9 microM and an inhibitory concentration for 50% response estimated at 6.6 microM. 5. Carbachol produced a proportionately greater suppression of the first pulse than the second pulse of a pulse pair. This increase in the level of facilitation accompanying suppression suggests a presynaptic mechanism.(ABSTRACT TRUNCATED AT 250 WORDS)

Localization of corticotropin-releasing factor-like immunoreactivity in monkey olfactory bulb and secondary olfactory areas.

Electrophysiological and anatomical observations suggest that terminals of olfactory bulb mitral cells ending in rat primary olfactory cortex exert certain postsynaptic effects via an excitatory amino acid neurotransmitter. Recent anatomical studies have shown that several peptides, most notably corticotropin-releasing factor (CRF) (Imaki et al., '89) Brain Res., 496: 35-44), are also localized within rat olfactory bulb projection neurons, thus raising the possibility that there is a peptide cotransmitter in this system. In contrast to the availability of data for rodents, very little is known about the distribution of peptides and other putative transmitters in the olfactory systems of primate species. In the present study, sections through the olfactory bulb and its target areas were obtained from two monkey species (Saimiri sciureus and Macaca fascicularis) and processed for immunohistochemistry with a well-characterized polyclonal antiserum directed against the human form of CRF. Virtually identical results were obtained in the two species. Within the olfactory bulb, nearly all mitral and many tufted cells contained CRF-like immunoreactivity. CRF-positive fibers were seen within the olfactory tract and olfactory stria, which contain the axons of mitral and tufted cells. Within the anterior olfactory nucleus and layer Ia of the olfactory tubercle and piriform cortex, immunoreactivity was seen within fine processes, as well as in coarse, varicose fibers and isolated puncta. CRF-positive cells were seen within layer III of the olfactory tubercle and piriform cortex. Immunoreactive fibers and varicosities were also seen within olfactory-recipient regions of the amygdala and entorhinal cortex. These observations suggest that CRF may act as a transmitter and/or neuromodulator in primate olfactory system.

Multiple heteroreceptors on limbic thalamic axons: M2 acetylcholine, serotonin1B, beta 2-adrenoceptors, mu-opioid, and neurotensin.

Ligand binding to many transmitter receptors is much higher in layer Ia of rat posterior cingulate cortex than it is in other layers, and this is where most axons from the anterior thalamus terminate. The present study explores the possibility that a number of receptors may be expressed on axons from limbic thalamic nuclei that terminate in layer Ia. Unilateral thalamic lesions were placed in rats and, 2 weeks later, five ligand binding protocols, coverslip autoradiography, and single grain counting techniques were used to quantify binding in control and ablated hemispheres. Binding to the following receptor subtypes was analyzed: M2 acetylcholine, 3H-oxotremorine-M, or 3H-AF-DX 116 with 50 nM pirenzepine; serotonin1B, 125I-(-)-cyanopindolol with 30 microM isoproterenol; beta 2-adrenoceptors, 125I-(-)-cyanopindolol with 1 microM serotonin and 10 microM atenolol; mu-opioid, 3H-T[r-D-Ala-Gly-MePhe-Gly-ol; neurotensin, 3H-neurotensin. Thalamic lesions reduced binding in two laminar patterns. In one pattern, there was a major reduction in binding in most superficial layers with that in layer Ia ranging from 50 to 70% for binding to M2 muscarinic and serotonin1B receptors. Binding to beta 2-adrenoceptors was also reduced in most superficial layers but to a lesser extent. In the second pattern, reductions were limited to layer I with losses in layer Ia of 20-30% for mu-opioid and neurotensin receptors. In no instance was layer Ia binding completely abolished (i.e., postlesion peaks remained). Since the transmitters for each of the five receptors analyzed in this study are not synthesized by anterior or laterodorsal thalamic neurons, these receptors are heteroreceptors. The greatest postlesion reduction in M2 binding was for AF-DX 116 and so most M2 heteroreceptors are of the "cardiac" subtype. Finally, the diverse population of heteroreceptors on limbic thalamic axons provides for presynaptic modulation by a wide range of transmitter systems and suggests that thalamocortical transmission may not be a simple, unmodulated event.

Early breakdown of dendritic bundles in the retrosplenial granular cortex of hypertensive rats: prevention by antihypertensive therapy.

Layer II pyramidal neurons in retrosplenial granular cortex (Rg) give rise to apical dendrites that bundle together tightly in layer Ic and Ib and spread out in layer Ia, where they arborize extensively. These dendritic bundles in Rg become disorganized in very old rats. Since hypertensive rats appear to age more rapidly than normotensive rats, we tested the hypothesis that the disorganization of the dendritic bundles in layer I of Rg is present at a much earlier age in hypertensive rats [spontaneously hypertensive rats (SHR)] compared to normotensive rats [Sprague-Dawley rats (SD)]. Further, we tested the hypothesis that long-term, antihypertensive drug treatment (captopril) prevents or attenuates the breakdown of the dendritic bundles in Rg of SHR. The retrograde tracer Fluorogold was injected into Rg to document the morphology of the apical dendrites of the layer II Rg neurons. In 4-month-old SD and SHR, most densely labeled layer II neurons have labeled apical dendrites that are confined to the dendritic bundles in layer Ib and Ic, and these dendrites arborize extensively only in layer Ia of Rg. In 14-month-old SD this pattern is preserved, but in contrast, in 14-month-old SHR most of the labeled apical dendrites lie outside the bundles, and less than half these dendrites reach layer Ia. Chronic antihypertensive therapy significantly attenuates this disorganization in 14-month-old SHR. These data indicate that in SHR, the layer II neurons in Rg are highly vulnerable to the combined effects of hypertension and aging, and the results suggest that hypertension is a primary contributor to these structural alterations.

Muscarinic receptor binding increases in anterior thalamus and cingulate cortex during discriminative avoidance learning

Training-induced neuronal activity develops in the mammalian limbic system during discriminative avoidance conditioning. This study explores behaviorally relevant changes in muscarinic ACh receptor binding in 52 rabbits that were trained to one of five stages of conditioned response acquisition. Sixteen naive and 10 animals yoked to criterion performance served as control cases. Upon reaching a particular stage of training, the brains were removed and autoradiographically assayed for 3H-oxotremorine-M binding with 50 nM pirenzepine (OXO-M/PZ) or for 3H-pirenzepine binding in nine limbic thalamic nuclei and cingulate cortex. Specific OXO-M/PZ binding increased in the parvocellular division of the anterodorsal nucleus early in training when the animals were first exposed to pairing of the conditional and unconditional stimuli. Elevated binding in this nucleus was maintained throughout subsequent training. In the parvocellular division of the anteroventral nucleus (AVp), OXO-M/PZ binding progressively increased throughout training, reached a peak at the criterion stage of performance, and returned to control values during extinction sessions. Peak OXO-M/PZ binding in AVp was significantly elevated over that for cases yoked to criterion performance. In the magnocellular division of the anteroventral nucleus (AVm), OXO-M/PZ binding was elevated only during criterion performance of the task, and it was unaltered in any other limbic thalamic nuclei. Specific OXO-M/PZ binding was also elevated in most layers in rostral area 29c when subjects first performed a significant behavioral discrimination. Training-induced alterations in OXO-M/PZ binding in AVp and layer Ia of area 29c were similar and highly correlated.(ABSTRACT TRUNCATED AT 250 WORDS)

Distribution of mRNAs coding for liver and heart gap junction proteins in the rat central nervous system

The present study examined the distributions of connexin43 mRNA and connexin32 mRNA in the central nervous system (CNS) of the rat by using in situ hybridization histochemistry. These connexins are the best studied gap junction proteins; connexin32 forms direct cell-cell channels in the liver, as does connexin43 in the heart. There was a differential distribution of cells containing connexin32 mRNA compared with the population of cells which contained connexin43 mRNA, thus implying a regional specificity in the expression of connexins in the CNS. Cells containing connexin43 mRNA were uniformly distributed throughout the gray matter of the neuraxis. Several areas had a higher concentration of cells that express connexin43, such as layer IA of the piriform cortex, supraoptic and paraventricular nuclei of the hypothalamus, anterior cortical amygdaloid nucleus, the reticular part of the substantia nigra, lateral habenula, mesencephalic trigeminal nucleus. Purkinje cell layer of the cerebellum, facial nucleus, prepositus hypoglossal nucleus, and dorsal cochlear nucleus. The pattern of connexin43 hybridization and the morphology of connexin43 mRNA containing cells suggest that this gap junction forming protein is found predominantly in astrocytes. Connexin32 mRNA was detected in discrete cell groups of the gray matter that appeared to be neurons, including cells in layer 2 of the neocortex, layer II of the piriform cortex, pyramidal cell layer of the hippocampus, granule and polymorphic cell layers of the dentate gyrus, islands of Calleja, olfactory tubercle, lateral thalamic nuclei, lateral habenula, and Purkinje cell layer of the cerebellar cortex. A large population of cells in white matter tracts that were labelled with the connexin32 riboprobe appeared to be oligodendrocytes. These studies suggest that neurons and glial cells express connexin32 mRNA, but only astrocytes express connexin43 mRNA. Many of the areas in which connexin mRNAs were demonstrated have electrically coupled cells, morphologically distinct gap junction plaques, and/or have immunocytochemically identifiable connexin proteins. These results indicate that cells with mRNAs coding for intercellular channels have a widespread distribution in the mammalian CNS.

Neuroarchitecture of the central complex in the brain of the locust Schistocerca gregaria and S. americana as revealed by serotonin immunocytochemistry

The central complex is a prominent structure in the insect brain, yet its anatomical organization and functional role is still poorly understood. To facilitate investigations on the physiology of the central complex, this study describes its anatomical organization in the brain of locusts (Schistocerca gregaria and Schistocerca americana) based on an investigation of serotonin immunocytochemistry. Most subdivisions of the central complex including the protocerebral bridge, several layers in the upper division of the central body, and the noduli of the central body are innervated by serotonin-immunoreactive neurons, while the lower division of the central body does not exhibit serotonin-like immunoreactivity. Several types of serotonin-immunoreactive neurons can be distinguished. A system of about 60 columnar neurons innervates the protocerebral bridge, layer III of the upper division of the central body, and the noduli. A group of 15-20 bilateral pairs of serotonin-immunoreactive neurons connects the posterior optic tubercles with the protocerebral bridge. About ten pairs of neurons with somata in the inferior protocerebrum innervate layer Ia of the upper division of the central body. In addition, large-field neurons arborize in layers Ia and Ib of the upper division of the central body and in the lateral accessory lobes. The detailed mapping of serotonin immunoreactivity provides further insight into the anatomical organization of the central complex and suggests that serotonin is a major neuroactive substance within this brain structure.

Dendritic bundling in layer I of granular retrosplenial cortex: Intracellular labeling and selectivity of innervation

The extrinsic projections to and from the retrosplenial cortex have been studied in detail, but the intrinsic circuitry within this region has been characterized less completely. To further define the internal connections, small injections of the retrograde, fluorescent tracer Fluorogold were made into the retrosplenial cortex of the rat. These injections label neurons in layers II-V of the contralateral homotopic cortex. In layers III-V, the labeled neurons are present over an area much larger than the injection site, but in layer II neurons are labeled in a very precise homotopic pattern. Following these injections, only the neurons in layer II display heavily labeled apical dendrites, and these labeled dendrites form tight bundles in layer Ic and Ib of the cortex and spread out in layer Ia. An examination of Golgi-stained material demonstrates that most of the neurons in layer II are small pyramidal cells with 2-3 small basal dendrites and a single, large apical dendrite that arborizes extensively in layer Ia. To verify the structure of the layer II neurons, they were intracellularly filled with Lucifer yellow. Examination of these labeled cells confirms the observations from the Golgi-stained material and demonstrates that many apical dendrites of the layer II cells angle acutely, apparently to join a bundle and/or avoid an interbundle space. Tract tracing experiments demonstrate that the anteroventral nucleus of the thalamus appears to project selectively to the region containing the dendritic bundles, whereas intracortical projections appear to terminate in layers Ib and Ic in the 30-200 microns spaces between the bundles. Furthermore, the areas containing the bundles display dense AChE staining, but the interbundle spaces are almost free of AChE staining. These findings demonstrate a form of dendritic bundling that is input and output specific and may play an important role in the regulation of thalamic inputs to the cingulate cortex.

Laminar distributions of muscarinic acetylcholine, serotonin, GABA and opioid receptors in human posterior cingulate cortex

Experimental animal studies have demonstrated a number of receptor localizations on specific cortical afferents and neurons. The present study of human posterior cingulate cortex evaluates the laminar distributions of particular receptors and their likely association with components of the neuropil. Coverslip autoradiographic and single grain counting techniques were used followed by heterogeneity analysis in which the layer of peak binding and an index of heterogeneity were determined for each ligand. The index was calculated by determining specific binding by layer as a percentage of binding in all layers. The differences from an absolutely homogeneous distribution, i.e. 11.1% for each of nine layers, were subtracted and the absolute laminar differences summed to form the index. High indices of over 15 reflected heterogeneous binding patterns in neocortex. The binding of ligands for muscarinic acetylcholine, serotonin, opioid, GABA and beta adrenoceptors was evaluated. Pirenzepine binding peaked in layer II of area 23a but was extremely homogeneous with an index of heterogeneity of 8.9. In contrast, oxotremorine-M binding had a peak in layer IIIc and an index of 16.4, while AF-DX 116 binding peaked in layer IIIa-b and had an index of 30.6. Of the ligands for serotonin uptake and receptor binding paroxetine binding was evenly distributed in layers I–III and had a low index of heterogeneity of 9.8. Ketanserin binding was also homogeneous and, since it had an index of 8.9, this pattern was virtually the same as that for paroxetine. In contrast, serotonin and 8-hydroxy-2-(di- n-propylamino)tetralin binding peaked in layer II and had very high indices of 20.8 and 50.3, respectively, suggesting only a limited association with that of the paroxetine distribution. Finally, there were three layers which contained peaks in binding for ligands for opioid, GABA and beta adrenoceptors. Firstly, layer Ia had peak dynorphin-A binding, the latter of which had an index of 22.6. Secondly, Tyr- d-Ala-Gly-MePhe-Gly-ol and 2- d-penicillamine-5- d-penicillamine-enkephalin binding peaked in layer II and had indices of 8.6 and 17.4, respectively. Thirdly, muscimol and (−)-cyanopindolol binding peaked in layer IIIa-b and had indices of 29.6 and 11.1, respectively. When viewed in the context of experimental animal studies, it is likely that heterogeneities in oxotremorine-M and paroxetine binding are associated with the termination of the thalamic and raphe nuclei, respectively. While serotonin, receptors are co-distributed with serotonin uptake sites, serotonin 1A receptors have a significant mismatch with these sites. Finally, postsynaptic receptors preferentially peak in different layers including kappa opioid receptors in layer Ia, M 1 acetylcholine, serotonin 1A, mu and delta opioid receptors in layer II, GABA A and beta adrenoceptors in layer IIIa-b and serotonin 2 receptors in layer IIIc. Thus, ligand binding analyses provide new insights into the structure and connections of human cerebral cortex.

The topographical and laminar organization of a commissural-associational entorhino-entorhinal projection in the guinea pig

A commissural-associational entorhino-entorhinal projection in the guinea pig was analyzed using anterograde and retrograde axonal tracing techniques. The projection originated in layer II and terminated in layer Ia of both the medial entorhinal area (MEA) and the lateral entorhinal area (LEA). A few cells in other layers, especially layer III, also contributed to the commissural system. The protection was largely homotopic with the exception of the most medial MEA, which projected ventrally like previously described projections from the para- and presubiculum to the superficial layers of the entorhinal area. The commissural fibers crossed the midline in the dorsal hippocampal commissure. The antero-posterior position of the fibers within the commissure reflected the ventrodorsal position of their origin in the entorhinal area.

Widespread expression of corticotropin-releasing factor messenger RNA and immunoreactivity in the rat olfactory bulb

Immunohistochemical, in situ hybridization histochemical, and Northern blot methods were used to demonstrate and charaterize the distribution of corticoptropin-releasing immunoreactivity (CRF-IR) and mRNA in the rat olfactory system. Northern analysis demonstrated the presence of an mRNA species in the olfactory bulb indistinguishable from, and in greater abundance than, CRF and mRNA isolated from whole hypothalamus. Results from hybridization histochemical and immunohistochemical studies converged to indicate tha CRF is expressed in a majority of mitral and tufted in the main accessory bulbs, and in subsets of granule and periglo3erular cells. Consistent with cellular localization in primary output neurons, a dense network of fine CRF-immunoreactive varicosities was demonstrated in the external plexiform layer of the olfactory bulb and in layer Ia of piriform cortex. Other acknowledged terminal fields of the projection neurons of the main and accessory bulbs also displayed CRF-IR. The results suggest that CRF is the most broadly distributed neuroactive agent yet charted in olfactory bulb somata. This peptide may serve as a modulator or co-transmitter of importance in several cell types in the main and accessory olfactory bulbs, including the principal output neurons.

Deep neurons in piriform cortex. I. Morphology and synaptically evoked responses including a unique high-amplitude paired shock facilitation.

1. Synaptic responses of cells in layer III of the piriform cortex and the subjacent endopiriform nucleus (layer IV) were analyzed with intracellular recording techniques in a slice preparation from the rat, cut perpendicular to the pial surface. 2. Micropipettes containing Lucifer yellow (LY) were used to correlate response properties with morphology. An antiserum to LY was used to intensify staining and to prevent fading during detailed morphological study. Response properties were also examined with potassium acetate-containing electrodes. 3. Morphologically, two cell types were identified: pyramidal cells that were confined to layer III of the piriform cortex and multipolar cells that were in layer III and the endopiriform nucleus. 4. In morphology, deep pyramidal cells in layer III closely resembled superficial pyramidal cells in layer II, with the exception that primary apical dendritic trunks were longer and basal dendritic arborizations were more extensive than apical. Like superficial pyramidal cells, apical dendrites of all deep pyramidal cells stained extended through the afferent fiber termination zone in layer Ia and gave rise to local axonal arbors that were concentrated in layer III and the endopiriform nucleus. 5. Multipolar cells were morphologically indistinguishable in layer III and the endopiriform nucleus. All gave rise to nonvaricose spiny dendrites that never extended into layer II and local axonal arbors. 6. Response properties of deep pyramidal and multipolar cells were similar; responses of both of these populations were very different from those of superficial pyramidal cells. The primary difference between responses of deep pyramidal and multipolar cells was a shorter latency of postsynaptic potentials evoked in deep pyramidal cells by stimulation of afferent fibers, consistent with the extension of their dendrites into layer Ia. 7. Responses of most deep cells to stimulation of afferent and association fibers at sufficiently high strength consisted of an initial excitatory postsynaptic potential (EPSP), followed by a fast Cl- -mediated and a slow K+-mediated inhibitory postsynaptic potential (IPSP). 8. A characteristic feature of deep cells, which was only rarely observed in superficial pyramidal cells, was the presence of variable EPSPs evoked at long latencies (greater than 100 ms) by stimulation of afferent or association fibers. 9. A striking finding for deep pyramidal and multipolar cells, when studied with LY-containing pipettes, was a variable slowly rising depolarizing potential triggered at depolarized membrane potentials by stimulation of afferent or association fibers.(ABSTRACT TRUNCATED AT 400 WORDS)