Parvalbumin- and vasoactive intestinal polypeptide-expressing neocortical interneurons impose differential inhibition on Martinotti cells.

Disinhibition of cortical excitatory cell gate information flow through and between cortical columns. The major contribution of Martinotti cells (MC) is providing dendritic inhibition to excitatory neurons and therefore they are a main component of disinhibitory connections. Here we show by means of optogenetics that MC in layers II/III of the mouse primary somatosensory cortex are inhibited by both parvalbumin (PV)- and vasoactive intestinal polypeptide (VIP)-expressing cells. Paired recordings revealed stronger synaptic input onto MC from PV cells than from VIP cells. Moreover, PV cell input showed frequency-independent depression, whereas VIP cell input facilitated at high frequencies. These differences in the properties of the two unitary connections enable disinhibition with distinct temporal features.

Electrophysiological and morphological properties of Cajal-Retzius cells with different ontogenetic origins.

The different origins of Cajal-Retzius cells (CRc) as well as their diverse molecular profile suggest that this cell type may represent different neuronal subpopulations. In order to investigate whether CRc from different origins show distinct functional or morphological characteristics we used transgenic Dbx1(cre);ROSA26(YFP) mice in which two subpopulations of CRc, originating from the septum and ventral pallium (VP) at the pallial-subpallial border (PSB), were permanently labeled by yellow fluorescent protein (YFP) expression. Electrophysiological properties of YFP(+) and YFP(-) CRc were investigated by whole-cell patch-clamp recordings, while a thorough somatodendritic and axonal reconstruction of the biocytin labeled CRc was subsequently performed using a Neurolucida system. Our experiments revealed that no significant differences in resting membrane potential, input resistance or capacitance, hyperpolarization activated currents and most action potentials properties could be observed between YFP(+) and YFP(-) CRc. Both YFP(+) and YFP(-) CRc displayed spontaneous and carbachol-induced GABAergic postsynaptic currents with similar properties and comparable NMDA-receptor mediated glutamatergic inward currents that were equally affected by the NR2B specific antagonist ifenprodil. Morphological reconstructions revealed that dendritic and axonal parameters are similar between YFP(+) and YFP(-) CRc, while the dendritic compartment of YFP(+) CRc was slightly larger. In summary, no considerable differences in functional and morphological properties between YFP(+) and YFP(-) CRc could be observed in this study. These observations suggest that CRc of different ontogenic origins display comparable functional properties in the early postnatal cortex and therefore perform similar functions within the transient neuronal networks of the developing cortex.

Morphology, electrophysiology and functional input connectivity of pyramidal neurons characterizes a genuine layer va in the primary somatosensory cortex.

Cortical layer V classically has been subdivided into sublayers Va and Vb on cytoarchitectonic grounds. In the analysis of cortical microcircuits, however, layer Va has largely been ignored. The purpose of this study was to investigate pyramidal neurons of layer Va in view of their potential role in integrating information from lemniscal and paralemniscal sources. For this we combined detailed electrophysiological and morphological characterization with mapping of intracortical functional connectivity by caged glutamate photolysis in layer Va of rat barrel cortex in vitro. Electrophysiological characterization revealed pyramidal cells of the regular spiking as well as the intrinsically burst firing type. However, all layer Va pyramidal neurons displayed uniform morphological properties and comparable functional input connectivity patterns. They received most of their excitatory and inhibitory inputs from intracolumnar sources, especially from layer Va itself, but also from layer IV. Those two layers were also the main origin for transcolumnar excitatory inputs. Layer Va pyramidal neurons thus may predominantly integrate information intralaminarly as well as from layer IV. The functional connectivity maps clearly distinguish layer Va from layer Vb pyramidal cells, and suggest that layer Va plays a unique role in intracortical processing of sensory information.

Excitatory and inhibitory neurons express c-Fos in barrel-related columns after exploration of a novel environment.

Recent work has shown that behaviorally meaningful sensory information processing is accompanied by the induction of several transcription factors in the barrel cortex of rodents. It is now generally accepted that stimulus-transcription coupling is an important step in the sequence of events leading to long-term plastic changes in neuronal structure and function. Nevertheless, so far few data are available as to what types of neurons are involved in such a genomic response. Here, we determined the morphological and neurochemical identity of neurons in rat barrel cortex showing a c-Fos-immunoreactive nucleus after exploration of an enriched environment. Double stainings of c-Fos and glial fibrillary acidic protein excluded astrocytes as a possible cell type expressing this transcription factor. By morphological phenotyping with intracellular Lucifer Yellow injections, it was found that a large majority were probably excitatory pyramidal cells, but inhibitory interneurons were also found to contain c-Fos-immunoreactive nuclei. By neurochemical phenotyping of GABAergic interneurons with specific antibodies, a significant induction was found, in a layer-dependent manner, for the populations of glutamic acid decarboxylase-, parvalbumin-, calbindin- and vasoactive intestinal polypeptide-immunoreactive neurons but not for calretinin-immunoreactive cells in experimental compared to control columns. From these data we conclude that thalamic afferents effectively drive cortical excitatory as well as inhibitory intracortical circuits. Thus, the adaptations of receptive field properties of cortical neurons after different manipulations of the sensory periphery are likely to be caused by plastic changes in excitatory and inhibitory networks.

Layer-specific intracolumnar and transcolumnar functional connectivity of layer V pyramidal cells in rat barrel cortex.

Layer V pyramidal cells in rat barrel cortex are considered to play an important role in intracolumnar and transcolumnar signal processing. However, the precise circuitry mediating this processing is still incompletely understood. Here we obtained detailed maps of excitatory and inhibitory synaptic inputs onto the two major layer V pyramidal cell subtypes, intrinsically burst spiking (IB) and regular spiking (RS) cells, using a combination of caged glutamate photolysis, whole-cell patch-clamp recording, and three-dimensional reconstruction of biocytin-labeled cells. To excite presynaptic neurons with laminar specificity, the release of caged glutamate was calibrated and restricted to small areas of 50 x 50 microm in all cortical layers and in at least two neighboring barrel-related columns. IB cells received intracolumnar excitatory input from all layers, with the largest EPSP amplitudes originating from neurons in layers IV and VI. Prominent transcolumnar excitatory inputs were provided by presynaptic neurons also located in layers IV, V, and VI of neighboring columns. Inhibitory inputs were rare. In contrast, RS cells received distinct intracolumnar inhibitory inputs, especially from layers II/III and V. Intracolumnar excitatory inputs to RS cells were prominent from layers II-V, but relatively weak from layer VI. Conspicuous transcolumnar excitatory inputs could be evoked solely in layers IV and V. Our results show that layer V pyramidal cells are synaptically driven by presynaptic neurons located in every layer of the barrel cortex. RS cells seem to be preferentially involved in intracolumnar signal processing, whereas IB cells effectively integrate excitatory inputs across several columns.

Kainate receptors regulate unitary IPSCs elicited in pyramidal cells by fast-spiking interneurons in the neocortex.

Unitary IPSCs elicited by fast-spiking (FS) interneurons in layer V pyramidal cells of the neocortex were studied by means of dual whole-cell recordings in acute slices. FS to pyramidal cell unitary IPSCs were depressed by (RS)-S-amino-3-(3-hydroxy-5-tert-butylisoxazol-4-yl) (ATPA), a kainate (KA) receptor agonist, and by the endogenous agonist l-glutamate in the presence of AMPA, NMDA, mGluR, and GABA(B) receptor antagonists. This effect was accompanied by an increase in failure rate of synaptic transmission, in the coefficient of variation, and in the paired pulse ratio, indicating a presynaptic origin of the IPSC depression. Pairing the activation of the presynaptic neuron with a depolarization of the postsynaptic cell mimicked the decrease of unitary IPSCs, and this effect persisted when postsynaptic sodium action potentials were blocked with the local anesthetic QX314. The effects of ATPA, glutamate, and of the pairing protocol were almost totally blocked by CNQX. These data suggest that KA receptors located on presynaptic FS cell terminals decrease the release of GABA and can be activated by glutamate released from the somatodendritic compartment of the postsynaptic pyramidal cells.

Exploration of a novel environment leads to the expression of inducible transcription factors in barrel-related columns.

Tactile information acquired through the vibrissae is of high behavioral relevance for rodents. Numerous physiological studies have shown adaptive plasticity of cortical receptive field properties due to stimulation and/or manipulation of the whiskers. However, the cellular mechanisms leading to these plastic processes remain largely unknown. Although genomic responses are anticipated to take place in this sequel, virtually no data so far exist for freely behaving animals concerning this issue. Thus, adult rats were placed overnight in an enriched environment and most of them were also subjected to clipping of different sets of whiskers. This type of stimulation led to a specific and statistically significant increase in the expression of the protein products of the inducible transcription factors c-Fos, JunB, inducible cyclic-AMP early repressor and Krox-24 (also frequently named Zif268 or Egr-1), but not c-Jun. The response was found in columns of the barrel cortex corresponding to the stimulated vibrissae; it displayed a layer-specific pattern. However, no induction of transcription factors was observed in the subcortical relay stations of the whisker-to-barrel pathway, i.e. the trigeminal nuclei and the ventrobasal complex. These results strongly suggest that a coordinated transcriptional response is initiated in the barrel cortex as a consequence of processing of novel environmental stimuli.

Laminar characteristics of functional connectivity in rat barrel cortex revealed by stimulation with caged-glutamate.

In rodent somatosensory (barrel) cortex input is processed by whisker-related columns before the integrated output is fed into behaviorally-relevant circuits. The layer-specific activation patterns of the rat barrel cortex were examined with a set-up for scanning functional connectivity in brain slices. Flash-induced release of caged-glutamate at a large number of stimulation sites was used in combination with simultaneous field potential recordings from layers II to VI with five electrodes. The field potentials revealed striking differences between the cortical layers. Glutamate-release in layer IV and lower layer III was most effective in evoking excitation in all other cortical layers, whereas field potentials recorded from layer IV itself were caused by stimulation of a very restricted columnar zone only. Field potentials in layers II and III were strongly driven by stimulation in layer IV and less consistently and much weaker by layer V. Layer V was the only lamina capable of responding to stimulation of all other cortical layers, thus displaying the largest input maps. Layer VI possessed functional connectivity intrinsically and with layer V. These data lead us to suggest that thalamic input may be boosted by its main target layer IV to start a sequence of excitation in layer IV, passing to the supragranular layers and finally reaching the infragranular layers. This sequence is likely to be backed-up by other simultaneous steps of transmission including a layer IV-to-V interaction. We proposed that the increasing size of the receptive fields when sampling granular, supragranular and infragranular layers in vivo, might have its functional basis in the laminar interactions described here in an in vitro preparation.

Neurons immunoreactive for vasoactive intestinal polypeptide in the rat primary somatosensory cortex: morphology and spatial relationship to barrel-related columns.

Vasoactive intestinal polypeptide (VIP) in neocortex affects neuronal excitability as well as cortical blood flow and metabolism. Interneurons immunoreactive for VIP (VIP-IR neurons) are characterized by their predominantly bipolar appearance and the radial orientation of their main dendrites. In order to determine whether the morphology of VIP-IR neurons is related to the functional organization of the cortex into vertical columns, we combined both immunostaining of neurons containing VIP and cytochrome oxidase histochemistry for visualizing barrels, morphological layer IV correlates of functional columns, in the primary somatosensory (barrel) cortex of rats. VIP-IR neurons were localized in supragranular (48%), granular (16%), and infragranular layers (36%) as well as in the white matter. In the granular layer, a clear trend that more neurons were located in interbarrel septa rather than in barrels could be observed, resulting in a neuronal density which was about one-third higher in the septal area. VIP-IR neurons from the different cortical layers were three-dimensionally reconstructed from serial sections by using a computer microscope system. The neurons were mostly bipolar. Striking morphological differences in both axonal and dendritic trees were found between neurons whose cell bodies were located in supragranular, granular, and the upper part of infragranular layers, and those whose cell bodies were located in the area below. The former had dendrites which often reached layer I, where they bifurcated several times, and axonal trees which were particularly oriented vertically, with a tangential extent smaller than the width of barrels. Therefore, these neurons were mostly confined to either a barrel- or septum-related column. By contrast, the dendrites of neurons of the latter group did not reach the granular layer. Furthermore, these neurons had axons with sometimes very long horizontal collaterals, which often spanned two, in one case three, barrel columns. It is proposed that the differential morphology of neurons with different locations as stated above parallels to some extent the divergence of input streaming into the corresponding layer-defined areas. As a possible consequence of this, VIP-IR neurons may be capable of adapting the excitability and metabolism of cortical compartments either in a spatially limited or more extensive way.

Cellular morphology and physiology of the perinatal rat cerebral cortex.

The cellular morphology and electrophysiology of the rat neocortex between embryonic day (E) 18 and postnatal day (P) 3 was studied in vitro by extracellular biocytin injections and whole-cell recordings, respectively. Most neurons were characterized by a small number of short-range dendrites and a main axon that was directed towards the white matter. Biocytin injections into the marginal zone and the cortical plate labeled far-reaching connections extending up to 2 mm in horizontal direction, indicating the existence of a dense network of long-range intrinsic projections in the neonatal cortex. Action potentials could be elicited as early as E18 and repetitive firing could first be observed at P0. Electrical stimulation of the immature cortex at various positions elicited polyphasic and long-lasting (up to 1 s) excitatory postsynaptic potentials and currents, which were significantly reduced in amplitude by a selective N-methyl-D-aspartate receptor antagonist. Our data indicate that the perinatal cortex manifests the structural and functional conditions for powerful excitatory interactions, which increase the likelihood for the generation of epileptiform activity during this developmental period.

Connectivity in the somatosensory cortex of the adolescent rat: an in vitro biocytin study.

A promising way to elucidate neuronal information processing is to establish detailed structure-function relationships of identified single neurons or populations of nerve cells, especially their synaptic connectivity. This has been greatly improved by the development of acute brain slice preparations. The cellular physiology of the rodent primary somatosensory (barrel) cortex has been extensively studied. However, for a meaningful interpretation of physiological experiments the degree and pattern of connectivity has to be known for the particular preparation. Since such studies are not available for rat (P15-25) barrel cortex in vitro, we have traced the cortico-cortical and thalamo-cortical connections in 400-microm-thick slices with biocytin. In coronal slices, a wealth of axonal connections in retrograde and anterograde directions were heavily labeled, resembling the full pattern of cortico-cortical projections described in vivo. The most striking connections were vertical and horizontal connections within the primary somatosensory cortex, as well as a columnar projection to the secondary somatosensory cortex and beyond (mainly the parietal ventral area). Electron microscopic extensions of the study indicated that the full possible set of synaptic contacts with an adult-like appearance was already established in these connections. In thalamo-cortical slices, strong reciprocal connections with the ventrobasal (and to a much lesser extent also the posterior) thalamic nucleus were always observed, together with an intensive ramification of fibers in the reticular nucleus. A striatal terminal field was also consistently found. We conclude that all major intracortical and thalamo-cortical connection are richly preserved in the in vitro slice preparations of rats. Thus, these preparations are suitable for elucidation of the functional interaction of the most crucial brain structures involved in somatosensory information processing combining an in vivo-like anatomical structure with the controlled environment of an in vitro slice.

Enlargement of cortical vibrissa representation in the surround of an ischemic cortical lesion.

It has been shown that cortical lesions are associated with an increase of excitability in surrounding brain regions, and with a downregulation of GABA(A) receptors. In the present study we investigated whether this increased excitability affects the cortical map of inputs represented in areas surrounding the lesioned brain area. Focal lesions with a diameter of 2-2.5 mm were induced photochemically in the hindlimb area at the border of the primary somatosensory cortex of the rat. One week after lesioning, the cortical representation of the B3 vibrissa was studied using 14C-deoxyglucose (DG) autoradiography. In all animals mechanical stimulation of the B3 vibrissa produced a column-shaped DG-labeling in the somatosensory cortex, corresponding to the B3-barrel with a maximum of the glucose uptake in layer IV. In control animals without cortical lesions (n=6), stimulation increased the glucose uptake rate by 50.8+/-10.5% in layer IV. In lesioned animals (n=6) maximum DG-uptake in layer IV (54.8+/-8.6%) did not differ significantly from that in controls. However, as compared to control animals, lesioned animals showed also increased glucose uptake within the activated column in layers II/II (51.+/-11.1%, lesioned animals; 31.8+/-11.2%, controls; P<0.05, lesioned vs. control) and V (47.5+/-5.8%, lesioned animals, 28.8+/-10.5%, controls; P<0.05, lesioned vs. control). The diameter of the metabolically activated B3-barrel area of layer IV was expanded from 461.8+/-77.6 microm in control animals to 785.5+/-103.6 microm; P<0.01) in lesioned animals. Lesioned animals also showed expansion of the activated area in layers II/III (890.4+/-134.8 microm, lesioned animals; 430.6+/-95.1 microm, controls; P<0.01) and layer V (1117.5+/-163.6 microm, lesioned animals; 648.7+/-114.1 microm, controls; P<0.01). The depth profile of the activation columns showed a maximum in layer IV in control animals, which was expanded towards layers II/III and layer V in lesioned animals. It is concluded that cortical lesions alter the representational area of neighboring afferent inputs through disinhibition or 'unmasking' of pre-existing silent or ineffectual intracortical synapses. The present observations raise the possibility that the brain supports recovery from lesions by decreasing GABAergic inhibition, thereby facilitating a remapping of the cortical representation in neighboring brain areas.

Developmental synaptic changes increase the range of integrative capabilities of an identified excitatory neocortical connection.

Excitatory synaptic transmission between pyramidal cells and fast-spiking (FS) interneurons of layer V of the motor cortex was investigated in acute slices by using paired recordings at 30 degrees C combined with morphological analysis. The presynaptic and postsynaptic properties at these identified central synapses were compared between 3- and 5-week-old rats. At these two postnatal developmental stages, unitary EPSCs were mediated by the activation of AMPA receptors with fast kinetics at a holding potential of -72 mV. The amplitude distribution analysis of the EPSCs indicates that, at both stages, pyramidal-FS connections consisted of multiple functional release sites. The apparent quantal size obtained by decreasing the external calcium ([Ca2+]e) varied from 11 to 29 pA near resting membrane potential. In young rats, pairs of presynaptic action potentials elicited unitary synaptic responses that displayed paired-pulse depression at all tested frequencies. In older animals, inputs from different pyramidal cells onto the same FS interneuron had different paired-pulse response characteristics and, at most of these connections, a switch from depression to facilitation occurred when decreasing the rate of presynaptic stimulation. The balance between facilitation and depression endows pyramidal-FS connections from 5-week-old animals with wide integrative capabilities and confers unique functional properties to each synapse.

Immunohistochemical evidence for dysregulation of the GABAergic system ipsilateral to photochemically induced cortical infarcts in rats.

Deficits of GABAergic transmission have been reported to occur in tissue surrounding ischemic cortical lesions between a few days and several weeks after the insult. In the present experiments, we used immunohistochemistry with antibodies against parvalbumin and two major subunits of the GABA(A) receptor (alpha1, alpha2) to characterize the events that underlie these changes at different levels of circuit organization. Neocortical infarcts (2 mm diameter) consistently affecting medial parts of the primary somatosensory cortex were induced photochemically in adult male Wistar rats; animals were allowed to recover for one week before perfusion-fixation. When compared to controls the pattern of immunoreactivity had changed for the al subunit of the GABA(A) receptor seven days after the insult. Ipsilateral to the ischemic lesions, we found a decrease in staining intensity reaching up to 4 mm laterally, resulting in a partial or complete absence of the normal laminar staining pattern. No consistent changes were observed for the alpha2 subunit. Parvalbumin staining revealed pathological alterations in a rim of tissue surrounding the infarct, measuring up to 1 mm from the border of the infarcts. Parvalbumin-positive interneurons in this region showed signs of degeneration; both a reduction of the number of dendrites and, to a lesser extent and only immediately adjacent to the ischemic lesions, a reduction of the number of parvalbumin-positive neurons was readily apparent. The results provide evidence for both a differential regulation of two GABA(A) receptor subunits and degenerative changes of parvalbumin-containing interneurons ipsilateral to cortical infarcts. The relevance of these findings for mechanisms underlying long-term recovery, transient functional deficits and postinfarct seizures warrants further investigation.

Analysing functional connectivity in brain slices by a combination of infrared video microscopy, flash photolysis of caged compounds and scanning methods.

We evaluate a novel set-up for scanning functional connectivity in brain slices from the somatosensory cortex of the rat. Upright infrared video microscopy for targeted placement of electrodes is combined with rapid photolysis of bath-applied caged neurotransmitter induced by a xenon flash lamp. Flash photolysis of caged glutamate and electrical stimulation produce comparable field potential responses and demonstrate that the viability of the submerged slices exceeds several hours. Glutamate release leads to field potential responses whose two phases are differentially affected by selective blockade of N-methyl-D-aspartate- and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate-type glutamate receptors with DL-2-amino-5-phosphonovaleric acid and 1,2,3,4-tetrahydro-6-nitro-2,3-dioxo-benzo[f]quinoxaline-7-sulphonamide, respectively. Rapid computer-controlled scanning of hundreds of distinct stimulation sites with simultaneous recordings at a fixed reference site allows construction of functional input maps from peak amplitudes and delays to peak of field potential responses. Selective laminar expansion of the functional input maps after bicuculline application demonstrates that the combination of this conveniently assembled set-up with pharmacological and physical manipulations can provide insights into the determinants of functional connectivity in brain slices.

Unsolved problems in comparing brain sizes in Homo sapiens.

When brain size is compared across taxonomic levels, there is a clear relation between body parameters and brain size. It is generally stated that the correlation between brain size and body parameters becomes very small at the species level (Aboitiz, 1996), but this is not the case for Homo sapiens where there is a strong correlation between brain size and body size across racial groups that differ in body size. The control for body size across racial groups (and sexes) is rendered difficult because bodies do not just differ only in height and weight. Within groups different studies show weak and inconsistent brain size/body height correlations. A better understanding of brain size/body height relations must await better quality data and a better understanding of how exactly body parameters should be scaled between groups and sexes. We attribute the clear between-group and weak within-group correlations to the large variety of body sizes and body types in our species, a variety which is only equalled in selectively bred species of animals. At present, there is no meaningful basis for the comparison of brain sizes within and between racial groups and sexes.

Disturbance of the cortical cholinergic innervation in Borna disease prior to encephalitis.

Rats experimentally infected with the highly neurotropic Borna disease virus (BDV) display a wide variety of dysfunction such as learning deficiencies and behavioral abnormalities. Prior to the onset of encephalitis alterations of one of the major cortical neurotransmitters, acetylcholine, were monitored immunohistochemically by light and electron microscopy of its synthesizing enzyme choline acetyltransferase (ChAT). We found a progressing decrease in the number of ChAT-positive fibers, starting with discrete changes at day 6 post infection (p.i.) and ending with a nearly complete loss of cholinergic fibers, especially in the hippocampus and neocortex, suggesting a massive disturbance of the cholinergic innervation by day 15 p.i.. The fiber pathways (e.g., fimbria-fornix) connecting the basal forebrain with these target areas in the cortex displayed axon spheroids which are often linked to axonal transport dysfunction. No evidence for significant cellular destruction was seen in the brain, including the cells of origin of these axons in the basal forebrain. We conclude that the motor, mood, learning and memory disabilities in BDV-infected rats are likely to result, in part, from cortical cholinergic denervation. The present study gives new insights into the pathogenesis of neurological disease caused by a noncytopathogenic virus.

Properties of bipolar VIPergic interneurons and their excitation by pyramidal neurons in the rat neocortex.

In the rat neocortex, a subset of GABAergic interneurons express the neuropeptide vasoactive intestinal peptide (VIP). Previously, we demonstrated that a population of VIPergic interneurons could be accurately identified by their irregular spiking (IS) pattern and their bipolar morphology. IS interneurons were studied in neocortical slices from 16-22-day-old rats using whole-cell recordings, intracellular labelling and single-cell RT-PCR. In response to a depolarizing pulse, IS interneurons typically discharged a burst of action potentials followed by spikes emitted at an irregular frequency. Several seconds of depolarization, micromolar concentrations of 4-aminopyridine, and nanomolar concentrations of either dendrotoxin I or K converted this irregular pattern to a sustained discharge, suggesting the involvement of an ID-like K+ current. The main glutamate receptor subunits detected in IS cells were GluR1 flop and GluR2 flop, GluR5 and GluR6, and NR2B and NR2D for the alpha-amino-3-hydroxyl-5-methyl-4-isoxazolepropionic acid (AMPA), kainate and N-methyl-D-aspartic acid (NMDA) subtypes, respectively. Paired whole-cell patch-clamp recordings indicated that pyramidal neurons provide intracortical glutamatergic inputs onto IS interneurons. Most connections had high probabilities of response and exhibited frequency-dependent paired pulse depression. Comparison of the amplitude distribution of paired responses suggested that most of these connections consisted of multiple functional release sites. Finally, two discrete subpopulations of IS cells could be identified based on the duration of the initial burst of action potentials and the differential expression of calretinin and choline acetyltransferase.

Geniculo-cortical afferents form synaptic contacts with vasoactive intestinal polypeptide (VIP) immunoreactive neurons of the rat visual cortex.

The lateral geniculate nucleus of the rat was injected with the anterograde tracer Phaseolus vulgaris leucoagglutinin (PHA-L) to see if geniculo-cortical axons terminate on vasoactive intestinal polypeptide immunoreactive (VIP-IR) neurons of the primary visual cortex. PHA-L-labelled boutons attached to VIP-IR perikarya and dendrites were identified as presynaptic parts of asymmetrical synapses. This geniculo-cortical projection to VIP-IR cells in the visual cortex and comparable findings in the somatosensory cortex suggest that sensory input from specific thalamic nuclei may influence local circuit inhibition and the metabolic state within the cortical domain via VIP-IR neurons.

Co-localization of vasoactive intestinal polypeptide, gamma-aminobutyric acid and choline acetyltransferase in neocortical interneurons of the adult rat.

Interneurons immunoreactive for vasoactive intestinal polypeptide (VIP) are integral elements of columnar organization patterns in the rat cerebral cortex. By application of the sensitive mirror technique, the co-localization of VIP with the classical inhibitory neurotransmitter gamma-aminobutyric acid (GABA) and the acetylcholine-synthesizing enzyme, choline acetyltransferase (ChAT), was investigated in neocortical neurons. Furthermore, the frequency of co-localization of ChAT with GABA was determined. In a sample of 118 VIP-immunoreactive neurons, mostly from the primary somatosensory cortex, it was demonstrated that virtually all of them reveal immunoreactivity for GABA and, therefore, are to be GABAergic. Moreover, 34% of mostly bipolar, VIP-positive neurons contained ChAT and are, thus, supposedly cholinergic as well. Co-localization of VIP and ChAT varied according to cortical laminae. Finally, 88% of a total of 60 ChAT-immunoreactive neurons were also immunostained for GABA. It is concluded that almost all VIP-immunoreactive neurons and most of the cholinergic neurons in rat neocortex represent partly overlapping subpopulations of inhibitory interneurons utilizing GABA.