Sonia Gómez-Urquijo

Targets and Laminar Distribution of Projection Neurons with 'Inverted' Morphology in Rabbit Cortex.

This study examines the axonal projections of so‐called inverted pyramids and other neurons with their major dendritic shaft oriented in the direction of the white matter (‘inverted cells’) in the adult rabbit cortex. Single injections of horseradish peroxidase wheat germ agglutinin were made into cortical or subcortical sites. The resulting retrograde labelling in the cortex was analysed and the distribution across areas and layers of inverted cells contributing to each of these projections was estimated. In addition, the radial distribution of inverted cells was independently determined from rapid Golgi‐impregnated and Nissl‐stained material. All three procedures revealed that inverted cells lay overwhelmingly in infragranular layers, but congregated at the border between layers 5 and 6. Inverted cells, identified by retrograde labelling, seldom furnished non‐telencephalic centres; in contrast, these cells constituted a major source for the projections to the ipsi‐ or the contralateral cortex, the claustrum or the nucleus caudatus. In general, each set of inverted cells (when defined by its specific destination as a group) was located below the typically oriented cells whose axons were aimed at the same target. Thus, the inverted cells of the rabbit cortex are characterized not only by their unique morphology and their corticocortical, corticoclaustral and corticostriatal projections, but also by their distinctive radial locations. These findings suggest that inverted cells, even though possibly composed of different cell types, are a specific class of projection neurons.

Cerebral blood flow and morphological changes after hypoxic-ischaemic injury in preterm lambs

AIM To evaluate the effect of cerebral hypoxia-ischaemia induced by partial occlusion of the umbilical cord on the relationship of the regional cerebral blood flow and the cerebral cell death in near-term fetal lambs. METHODS Fifteen near-term lambs were assigned to two hypoxic-ischaemic groups with or without life support (3 h), and a healthy one. Hypoxia-ischaemia was induced by partial occlusion of the umbilical cord (60 min). Routine light and electron microscopy, and the TUNEL method for apoptosis were performed. Regional cerebral blood flow was measured by coloured microspheres. Cardiovascular, gas exchange and pH parameters were also evaluated. RESULTS Both hypoxic-ischaemic groups produced a transient acidosis and a decrease of base excess in comparison to the healthy group. Cortical and cerebellar zones, where the regional cerebral blood flow values were similar to baseline, showed an increased number of oligodendrocyte-like apoptotic cells. In contrast, in the inner zones, where regional cerebral blood flow was increased, the number of apoptotic cells did not increase. Necrotic neurons were observed in the basal nuclei, mesencephalon, pons and deep cerebellar nuclei. CONCLUSION Our results suggest that regional cerebral blood flow and the presence of apoptotic cells, 3 h after hypoxic-ischemic injury, are correlated.

Percentage incidence of γ-aminobutyric acid neurons in the claustrum of the rabbit and comparison with the cortex and putamen

We describe the incidence of gamma-aminobutyric acid (GABA)ergic neurons after post-embedding immunocytochemistry on semithin sections of the claustrum, putamen and lateral, dorsal and medial cortical areas. Twelve percent of the neurons counted in the claustrum of 11 rabbits were GABAergic. This incidence was significantly higher in the dorsal halves of both the insular and endopiriform claustra than in the ventral (13 vs. 10%). The incidence of GABAergic cells was 4% in the putamen, 14% in the insular cortex, 15% in areas 17 and 18 and 13% in area 29d. Thus, our results indicate that in contrast to the putamen the incidence of GABAergic cells was similar in the claustrum and cortical areas. We interpret this in the light of the pallial origin of the claustrum, which has recently been substantiated.

Electrophysiological characterization of substantia nigra dopaminergic neurons in partially lesioned rats: Effects of subthalamotomy and levodopa treatment

Progressive degeneration of dopaminergic neurons in the substantia nigra pars compacta is the main histopathological characteristic of Parkinsons disease. We studied the electrophysiological characteristics of the spontaneous activity of substantia nigra pars compacta dopaminergic neurons in rats with a partial, unilateral, 6-hydroxydopamine lesion of the nigrostriatal pathway. In addition, the effects of subthalamotomy and prolonged levodopa treatment on the activity of dopaminergic neurons were investigated. As a result of the lesion ( approximately 50% neuronal loss), the number of spontaneously active neurons was significantly reduced. Basal firing rate, burst firing and responsiveness to intravenously administered apomorphine remained unchanged. In contrast, the variation coefficient, a measure of interspike interval regularity, was significantly increased. Ibotenic acid (10 microg) lesion of the ipsilateral subthalamic nucleus in lesioned rats did not modify the electrophysiological parameters. However, prolonged levodopa treatment (100 mg/kg/day + benserazide 25 mg/kg/day, 14 days) reversed the irregularity observed in cells from lesioned rats, while it induced an irregular firing pattern in cells from intact rats. Our results using an experimental model of moderate Parkinsons disease indicate that surviving substantia nigra pars compacta dopaminergic neurons fire irregularly. In this model, subthalamotomy does not modify the firing pattern while levodopa treatment efficiently restores normal firing of SNpc neurons and does not appear to be toxic to them.

INVOLVEMENT OF SUBTHALAMIC NUCLEUS IN THE STIMULATORY EFFECT OF Δ9-TETRAHYDROCANNABINOL ON DOPAMINERGIC NEURONS

The cannabinoid CB1 receptor which is densely located in the basal ganglia is known to participate in the regulation of movement. The present study sought to determine the mechanisms underlying the effect of Delta(9)-tetrahydrocannabinol (Delta(9)-THC) on neurons in the substantia nigra pars compacta (SNpc) using single-unit extracellular recordings in anesthetized rats. Administration of Delta(9)-THC (0.25-2 mg/kg, i.v.) increased the firing rate of SNpc neurons (maximal effect: 33.54+/-6.90%, n=8) without modifying other firing parameters (coefficient of variation and burst firing). This effect was completely blocked by the cannabinoid receptor antagonist rimonabant (0.5 mg/kg, i.v.). In addition, the blockade of excitatory amino acids receptors by kynurenic acid (0.5 microM, i.c.v.) or a chemical lesion of the subthalamic nucleus (STN) with ibotenic acid abolished Delta(9)-THC effect. These results indicate that CB1 receptor activation modulates SNpc neuronal activity by an indirect mechanism involving excitatory amino acids, probably released from STN axon terminals in the SNpc.

Widespread Horizontal Connections Arising from Layer 5/6 Border Inverted Cells in Rabbit Visual Cortex.

Herein we describe the inverted cells [defined as those projection neurons having a major dendritic shaft abpially oriented (Bueno‐López et al., Eur. J. Neurosci., 3, 415, 1991)] originating a unique set of cortical connections characterized by extraordinarily widespread horizontal distribution. Single and multiple injections of wheatgerm agglutinin ‐ horseradish peroxidase were made in areas 17 and 18 and the resulting retrograde labelling in the cortex was analysed. The findings were assessed in independent control experiments in which Fluoro‐Gold was used as retrograde tracer. Following single injections in area 17 several separate patches of labelled cells comprising layers 2–6 were consistently found in area 18. In addition to these associational cells a number of labelled cells appeared at the layer 5/6 border but were distributed over most of the tangential extent of the visual occipital cortex. This widespread pattern was particularly striking in brains after multiple injections. In these brains a conspicuous band of labelled cells at the 5/6 border radiated from the injection sites, making up an apparently continuous horizontal sheet that intersected the striate ‐ extrastriate boundary and merged with the patches of labelled cells in area 18 and beyond. Most of the cells in the 5/6 border band were inverted cells (82%; n= 2081). Injections in area 18 failed to produce such a widespread set of labelled cells in area 17. The functional significance of these connections furnished by the 5/6 border inverted cells remains to be determined, but their distribution would allow for convergent/divergent binding interactions both intra‐areally (within area 17) and inter‐areally (from area 18 to area 17).

GABAergic neurons in the rabbit visual cortex: percentage, layer distribution and cortical projections

6250 neurons yielding either callosal or inter-areal ipsilateral projections extrinsic to area 17 was GABAergic. Comparing these findings with those reported for other mammals, it seems that the incidence and distribution of GABAergic neurons in the visual cortex is similar in rabbits and rats. In contrast to rats but akin to higher mammals, no GABAergic neuron was found to furnish cortico-cortical connections to area 17 other than intrinsic connections.

Conserved cellular distribution of the glutamate receptors GluA2/3, mGlu1a and mGlu2/3 in isolated cultures of rat cerebellum

To what extent the intrinsic glutamatergic system of the cerebellum is able to keep normal features in the absence of mossy and climbing fibres, is at present not known. To answer this question, immunocytochemistry for light and high resolution electron microscopy was used to reveal the cellular and subcellular distribution of glutamate receptors in isolated cerebellar cultures. The localization of the ionotropic glutamate receptor subunits GluA2/3 and the metabotropic glutamate (mGlu) 1a and mGlu2/3 receptor subtypes was carried out in 0 to 9-day-old rat parasagittal slices developed in vitro for 20-40 days. The typical localization of GluA2/3, mGlu1a and mGlu2/3 observed in Purkinje cells, granule cells, Golgi cells and unipolar brush cells was maintained in the organotypic cultures. Furthermore, the subcellular distribution of mGlu1a showed the characteristic in vivo perisynaptic position in Purkinje cell dendritic spines receiving parallel fibre synapses. We conclude that the cellular and subcellular localization of the studied ionotropic and metabotropic glutamate receptors is not affected by the removal of the two extrinsic cerebellar glutamatergic inputs, the mossy and climbing fibres.

Cerebral blood flow and morphological changes after hypoxic-ischaemic injury in preterm lambs: Cerebral blood flow and cell damage

Aim: To evaluate the effect of cerebral hypoxia‐ischaemia induced by partial occlusion of the umbilical cord on the relationship of the regional cerebral blood flow and the cerebral cell death in near‐term fetal lambs. Methods: Fifteen near‐term lambs were assigned to two hypoxic‐ischaemic groups with or without life support (3 h), and a healthy one. Hypoxia‐ischaemia was induced by partial occlusion of the umbilical cord (60 min). Routine light and electron microscopy, and the TUNEL method for apoptosis were performed. Regional cerebral blood flow was measured by coloured microspheres. Cardiovascular, gas exchange and pH parameters were also evaluated. Results: Both hypoxic‐ischaemic groups produced a transient acidosis and a decrease of base excess in comparison to the healthy group. Cortical and cerebellar zones, where the regional cerebral blood flow values were similar to baseline, showed an increased number of oligodendrocyte‐like apoptotic cells. In contrast, in the inner zones, where regional cerebral blood flow was increased, the number of apoptotic cells did not increase. Necrotic neurons were observed in the basal nuclei, mesencephalon, pons and deep cerebellar nuclei.