Diego Minciacchi

The thalamo-caudate versus thalamo-cortical projections as studied in the cat with fluorescent retrograde double labeling

SummaryThe distribution of thalamic cells projecting to the head of the caudate and their interrelations with thalamo-cortical cells were studied in the cat with different combinations of fluorescent tracers. Injections in the head of the caudate were combined with the injections in the pericruciate, proreal, suprasylvian, anterior cingulate, occipital and ectosylvian cortices. The following results were obtained: (i) Injections in the head of the caudate resulted in retrograde labeling of thalamic cells medially and laterally to the anteromedial (AM) nucleus, and in the medioventral part of the ventral anterior (VA) nucleus. Further, labeled cells were distributed throughout the anterior intralaminar central medial (CeM), paracentral (Pc) and central lateral (CL) nuclei, and the posterior intralaminar center median-parafascicular complex (CM-Pf). Labeled cells were mainly grouped in the mediodorsal parts of the anterior intralaminar nuclei; they were also found in the more dorsal part of the mediodorsal (MD) nucleus, ventral to the thalamic paraventricular (Pv) nucleus and to the habenular complex, (ii) Thalamo-cortical and thalamo-caudate cells overlapped in the medial part of the VA; in the anterior intralaminar nuclei they were either intermingled or were distributed in separate clusters or longitudinal bands. The two cell populations also overlapped in the posterior intralaminar complex. The greatest overlap occurred with the thalamic cell population projecting to the pericruciate cortex. (iii) Thalamic cells bifurcating to the head of the caudate and to the pericruciate cortex were found lateral to the AM, within the VA, and throughout the anterior intralaminar nuclei, especially in the CeM and in the posterior part of the CL; a few branched cells were also found in the CM. Thalamic cells bifurcating to caudate and anterior suprasylvian cortex were also found in the VA. Very few cells (scattered in the anterior thalamus lateral to the AM, as well as in the CeM, Pc and CL) were found to bifurcate to the head of the caudate and the other cortical fields here examined.

The organization of the ipsi- and contralateral claustrocortical system in rat with notes on the bilateral claustrocortical projections in cat

The organization of the claustrocortical system was investigated in rat by means of cortical injections of either lectin-conjugated horseradish peroxidase or retrograde fluorescent tracers. The latter were also employed in cat. Evans Blue, Fast Blue, True Blue, Nuclear Yellow and Diamidino Yellow were used in different combinations and were injected, uni- or bilaterally, in different cortical fields. Cells retrogradely labeled from each cortical injection were observed in the ipsi- and contralateral claustrum. Anterogradely labeled terminals were also seen in the claustra of both sides in the horseradish peroxidase experiments. The topographic and quantitative study of the distribution of labeled neurons showed a topographic organization of the rats claustrocortical system, although a certain degree of overlap of the cell populations projecting to frontal and occipital fields was also evident. Four types of branched claustrocortical neurons were observed in the double labeling experiments: neurons branching ipsilaterally (A) or contralaterally (B) to anterior and posterior cortical fields; neurons branching bilaterally to homotopic (C) or heterotopic (D) cortical fields. Each population of branched neurons was equivalent to a different percent value of the total labeled cell populations; the percent value decreased from type A to type D. Type C branched neurons were also identified in the claustrofrontal system of the cat. The intricate organization of the claustral-ascending projections suggests that the nucleus is involved in different cortical activities and that its efferents may also provide the substrate of a powerful subcortical mechanism of interhemispheric communication.

Efferent fibers from the motor cortex terminate bilaterally in the thalamus of rats and cats

SummaryThe anterograde transport of lectin-conjugated horseradish peroxidase (WGA-HRP) was here employed in order to visualize crossed cortico-thalamic efferents of the motor cortex in rats and cats. After WGA-HRP cortical injections in the rat retrogradely labeled cells were observed in the ipsilateral thalamus, and heavy anterograde labeling was observed both in the ipsi-and contralateral thalamus. The contralateral anterograde labeling was less intense than the ipsilateral one and it was distributed in the anterior intralaminar structures, in the parafascicular nucleus, in the ventromedial, ventrolateral and ventrobasal nuclei and in the posterior complex, symmetrically to the labeling observed on the ipsilateral side. Further experiments were made in the rat in order to ascertain that the bilateral anterograde labeling in the thalamus derived unilaterally from the cortex. To this purpose, kainic acid was injected unilaterally either into the frontal cortex or into the thalamus, and WGA-HRP was later injected on the same side in the frontal cortex. Moreover, WGA-HRP was injected into the frontal cortex after splitting of the corpus callosum. The results obtained in these experiments confirmed that cortical neurons projected bilaterally upon the thalamus. Further, these experiments indicated that at least the majority of the contralateral fronto-thalamic fibers crossed the midline in the thalamic massa intermedia. WGA-HRP injections into the pericruciate cortex in the cat confirmed the presence of anterogradely labeled terminals in the contralateral anterior and posterior intralaminar, ventral anterior, ventromedial and ventrolateral nuclei. The labeling was in all cases heavier in the intralaminar nuclei than in the other structures, but it was less intense than that observed in the rat. The labeling in the contralateral principal nuclei was heavier in one case in which the injection area diffused into the proreal cortex than in the other cats. Thus, the present results indicate that frontal efferents terminate more densely in the rats than in the cats contralateral thalamus. In the latter species frontal fibers are mainly distributed upon the contralateral intralaminar nuclei, whereas in the rat frontal efferents are also heavily distributed upon the ventral nuclear complex.

GABAergic interneurons and neuropil of the intralaminar thalamus: an immunohistochemical study in the rat and the cat, with notes in the monkey

SummaryImmunohistochemistry using antibodies to glutamic acid decarboxylase (GAD) was used to investigate the intralaminar nuclei of the thalamus in rat, cat and monkey. Antibodies to gamma aminobutyric acid (GABA) were also used in the cat. Intralaminar immunoreactive cell bodies were not detected in the rat, but were clearly present in cat and monkey. In the latter species, GABA- or GAD-immunopositive perikarya were distributed throughout the anterior intralaminar nuclei, whereas in the posterior intralaminar complex they prevailed in the lateral part of the centre median nucleus and around the fasciculus retrofiexus. Measurements of the area of immunostained intralaminar cell bodies in cat and monkey indicated that they are represented by small neurons. Experiments in the cat, based on retrograde tracers injections involving large sectors of the frontal and parietal cortices and the head of the caudate nucleus, revealed that the GABA- or GAD-immunoreactive cells and the retrogradely labeled projection neurons represented two separate intralaminar cell populations, although the latter also included small cells. Considerable differences were observed in the immunoreactive GABAergic neuropil of the anterior and posterior intralaminar nuclei. Clusters of densely packed bouton-like immunoreactive elements were detected in the former structures in the rat, cat and monkey, and were especially evident in the central lateral nucleus; immunopositive varicose fibers and puncta were diffusely distributed in the posterior intralaminar structures. Taken together with data from the literature, the present findings indicate that in cat and monkey local circuit inhibitory cells regulate not only the activity of principal thalamic nuclei which project densely upon restricted cortical fields, but also of the intralaminar structures which are widely connected with the cerebral cortex and the striatum. Regional variations in the distribution of GABAergic fibers and terminals suggest major differences in the organization of inhibitory circuits and synaptic arrangements of the anterior and posterior intralaminar thalamus.

Organization of claustro-cortical projections to the primary somatosensory area of primates.

Different fluorescent dyes were injected in the face (S1fa) and hand (S1hn) representations of the primary somatosensory cortex, involving both areas 3b and 1. Claustral neurons labeled by either S1fa or S1hn were divided in two populations. One population was located in the dorsal part of the nucleus: neurons labeled from S1fa were placed laterally to those labeled from S1hn. The second population was located more ventrally, with a rostro-caudal distribution of S1fa vs S1hn neurons. These findings demonstrate the existence of ordered and possibly multiple somatosensory representations in the monkey claustrum.

Spatial analysis reveals alterations of parvalbumin- and calbindin-positive local circuit neurons in the cerebral cortex of mutant mdx mice

The aim of the present study was to investigate the spatial organization of selected populations of local circuit neurons in the cerebral cortex of the mutant mdx mouse, an acknowledged model of Duchenne Muscular Dystrophy. To this purpose, we quantified and compared the distribution of parvalbumin- and calbindin-positive neurons in the motor, somatosensory, visual, and anterior cingulate cortices of wild-type and mdx mice. The methodological approach was based on generation of two-dimensional Voronoi polygons from digital charts of the cell populations visualized immunohistochemically. Polygon areas were then analyzed and the derived coefficients of variation were statistically compared. Using this strategy, we were able to reveal, in mdx mice, changes involving both the above populations of interneurons. These changes were evident in the motor and anterior cingulate cortices but not in the somatosensory and visual cortices. In addition, the changes of coefficients of variation were area-specific in the cortex of mdx mice. The values increased in the motor cortex and decreased in the anterior cingulate cortex with respect to the corresponding values of wild-type animals. The present findings point out widespread alterations in the mdx cortex involving also areas not primarily related to sensorimotor integration. In addition, we demonstrate that cortical alterations of the local circuit machinery are characterized in mdx mice by individual regional differences.

Multifaceted alterations of the thalamo-cortico-thalamic loop in adult rats prenatally exposed to ethanol

The thalamo-cortico-thalamic loop was investigated in adult rats exposed to ethanol during the last week of fetal life. Animals underwent either cortical or thalamic injections of lectin-conjugated horseradish peroxidase. Results demonstrate that prenatal exposure to ethanol causes permanent changes in the thalamocortical circuits. Alterations of thalamo-cortical and cortico-thalamic projections are concentrated at the level of axon terminal fields. The most severe thalamic damage is observed in the anterior intralaminar and midline nuclei; crossed cortico-thalamic projections also appear to be severely impaired. In the cortex, the damage to thalamic terminals displays a medio-lateral gradient of increasing severity through sensori-motor areas, with the lateral fields more impaired. Cells of origin of thalamo-cortical and cortico-thalamic projections are less affected by prenatal ethanol exposure: in the thalamus and layer 5 of sensori-motor cortex labeled cells exhibit normal values of areal numeric density. Conversely, cortico-thalamic neurons of layer 6, especially in the lateral agranular sensori-motor field, display smaller values of areal density than those of normal animals. Possible mechanisms underlying the establishment of these abnormalities are discussed.

Spinal afferents and cortical efferents of the anterior intralaminar nuclei: an anterograde-retrograde tracing study

The topographical relations among the terminal field of spinothalamic fibers and the cells projecting upon areas 4 and 5 were studied in the anterior intralaminar nuclei of the cat. Terminals anterogradely labeled from the spinal cord and cell populations retrogradely labeled from the lateral pericruciate and anterior suprasylvian cortex were simultaneously observed by means of a multiple fluorescent tracing strategy. The present findings confirm that spinal afferents in the central lateral and paracentral nuclei overlap with the cells projecting to area 4. Further, the present data demonstrate that spinal terminals are largely segregated from the intralaminar cell population projecting to area 5.

GABAergic miniature spontaneous activity is increased in the CA1 hippocampal region of dystrophic mdx mice

Duchenne muscular dystrophy (DMD), a genetic disease due to dystrophin gene mutation and characterised by skeletal muscle failure, is associated with non-progressive cognitive deficits. In human and mouse brain, full-length dystrophin is localised postsynaptically in neocortical, hippocampal and cerebellar neurons. Evidence obtained in the CNS of dystrophic mice (mdx) suggested alterations of the GABAergic system. However, a direct functional evaluation of GABAergic synaptic transmission in mdx mice has not been conducted in the hippocampus, which is involved in cognitive processes and is rich in full-length dystrophin. Here, we investigated evoked and miniature inhibitory postsynaptic currents (IPSCs) in CA1 pyramidal neurons of mdx mice with patch clamp recording techniques. Results showed an increased frequency of miniature spontaneous IPSCs in mdx mice compared with controls, whereas evoked IPSCs did not show significant variations. Paired-pulse facilitation (PPF) analysis showed lack of facilitation at short intervals in mdx mice compared with that in wild-type mice. Analysis of density of synapses that innervate CA1 pyramidal cell bodies did not indicate significant differences between mdx mice and controls. Therefore, we suggest that increased miniature spontaneous IPSC frequency is due to altered pre-synaptic release probability. The present findings are discussed in the light of the accrued evidence for alterations of inhibitory synaptic transmission in the brain of dystrophic mice.

Cortical and brainstem neurons containing calcium‐binding proteins in a murine model of Duchenne's muscular dystrophy: Selective changes in the sensorimotor cortex

In the muscular dystrophic (mdx) mouse, which is characterized by deficient dystrophin expression and provides a model of Duchennes muscular dystrophy, we previously demonstrated marked central nervous system alterations and in particular a quantitative reduction of corticospinal and rubrospinal neurons and pathologic changes of these cells. Prompted by these findings and in view of the relations between calcium ions and dystrophin, we analyzed with immunohistochemistry the neurons containing the calcium‐binding proteins parvalbumin, calbindin D28k, and calretinin in cortical areas and brainstem nuclei of mdx mice. In the sensorimotor cortex, parvalbumin‐positive and calbindin‐positive neurons, which represent a subset of cortical interneurons, were significantly more numerous in mdx mice than in wild‐type ones. In addition, the laminar distribution of parvalbumin‐positive neurons in the motor and somatosensory cortical areas of mdx mice was altered with respect to wild‐type animals. No alterations in the number and distribution were found in the parvalbumin‐ or calbindin‐expressing cell populations of the visual and anterior cingulate cortices of mdx mice. The pattern of calretinin immunoreactivity was normal in all investigated cortical areas. The cell populations containing either calcium‐binding protein were similar in brainstem nuclei of mdx and wild‐type mice. The present findings demonstrated selective changes of subsets of interneurons in the motor and somatosensory cortical areas of mdx mice. Therefore, the data showed that, in the cortices of these mutant animals, the previously demonstrated pathologic changes of corticospinal cell populations are accompanied by marked alterations in the local circuitry. J. Comp. Neurol. 456:48–59, 2003.