Journal of Neurocytology最新文献

Human natural killer antigen-1 (HNK-1) is a carbohydrate epitope associated with sulfoglucuronylglycolipids and glycoproteins. Biochemical analyses have demonstrated associations between the HNK-1 epitope and isoforms of the neural cell adhesion molecule (N-CAM) family. In the cerebellum, HNK-1 is prominently expressed in Purkinje cell dendrites and Golgi cells. Purkinje cell expression of HNK-1 reveals an array of parasagittal stripes and transverse zones. Interestingly, the parasagittal expression pattern of HNK-1 is different from those reported with several other markers such as zebrin II/aldolase C and the small heat shock protein HSP25. N-CAM null knockout mice were used to explore the possible role of the HNK-1/N-CAM interaction during the topographical organization of the cerebellar cortex. N-CAM null mice have no N-CAM immunoreactivity but otherwise the cerebellum appears morphologically normal. Further, in the N-CAM null HNK-1 immunoreactivity is abolished from Purkinje cell dendrites but is retained on Golgi cells and neurons of the cerebellar nuclei. Despite the absence of N-CAM/HNK-1, parasagittal stripes and transverse zones in the cerebellum as revealed by using zebrin II immunocytochemistry appear normal.

Combined confocal microscopy and freeze-fracture replica immunogold labeling (FRIL) were used to examine the connexin identity at electrical synapses in goldfish brain and rat retina, and to test for "co-localization" vs. "close proximity" of connexins to other functionally interacting proteins in synapses of goldfish and mouse brain and rat retina. In goldfish brain, confocal microscopy revealed immunofluorescence for connexin35 (Cx35) and NMDA-R1 (NR1) glutamate receptor protein in Mauthner Cell/Club Ending synapses. By FRIL double labeling, NR1 glutamate receptors were found in clusters of intramembrane particles in the postsynaptic membrane extraplasmic leaflets, and these distinctive postsynaptic densities were in close proximity (0.1-0.3 microm) to neuronal gap junctions labeled for Cx35, which is the fish ortholog of connexin36 (Cx36) found at neuronal gap junctions in mammals. Immunogold labeling for Cx36 in adult rat retina revealed abundant gap junctions, including several previously unrecognized morphological types. As in goldfish hindbrain, immunogold double labeling revealed NR1-containing postsynaptic densities localized near Cx36-labeled gap junction in rat inferior olive. Confocal immunofluorescence microscopy revealed widespread co-localization of Cx36 and ZO-1, particularly in the reticular thalamic nucleus and amygdala of mouse brain. By FRIL, ZO-1 immunoreactivity was co-localized with Cx36 at individual gap junction plaques in rat retinal neurons. As cytoplasmic accessory proteins, ZO-1 and possibly related members of the membrane-associated guanylate kinase (MAGUK) family represent scaffolding proteins that may bind to and regulate the activity of many neuronal gap junctions. These data document the power of combining immunofluorescence confocal microscopy with FRIL ultrastructural imaging and immunogold labeling to determine the relative proximities of proteins that are involved in short- vs. intermediate-range molecular interactions in the complex membrane appositions at synapses between neurons.

During development radial glia (RG) are neurogenic, provide a substrate for migration, and transform into astrocytes. Cells in the RG lineage are functionally and biochemically heterogeneous in subregions of the brain. In the subventricular zone (SVZ) of the adult, astrocyte-like cells exhibit stem cell properties. During examination of the response of SVZ astrocytes to brain injury in adult mice, we serendipitously found a population of cells in the walls of the ventral lateral ventricle (LV) that were morphologically similar to RG. The cells expressed vimentin, glial fibrillary acidic protein (GFAP), intermediate filament proteins expressed by neural progenitor cells, RG and astrocytes. These RG-like cells had long processes extending ventrally into the nucleus accumbens, ventromedial striatum, ventrolateral septum, and the bed nucleus of the stria terminalis. The RG-like cell processes were associated with a high density of doublecortin-positive cells. Lesioning the cerebral cortex did not change the expression of vimentin and GFAP in RG-like cells, nor did it alter their morphology. To study the ontogeny of these cells, we examined the expression of molecules associated with RG during development: vimentin, astrocyte-specific glutamate transporter (GLAST), and brain lipid-binding protein (BLBP). As expected, vimentin was expressed in RG in the ventral LV embryonically (E16, E19) and during the first postnatal week (P0, P7). At P14, P21, P28 as well as in the adult (8-12 weeks), the ventral portion of the LV retained vimentin immunopositive RG-like cells, whereas RG largely disappeared in the dorsal two-thirds of the LV. GLAST and BLBP were expressed in RG of the ventral LV embryonically and through P7. In contrast to vimentin, at later stages BLBP and GLAST were found in RG-like cell somata but not in their processes. Our results show that cells expressing vimentin and GFAP (in the radial glia-astrocyte lineage) are heterogeneous dorsoventrally in the walls of the LV. The results suggest that not all RG in the ventral LV complete the transformation into astrocytes and that the ventral SVZ may be functionally dissimilar from the rest of the SVZ.

The relationship between efferents of the hypothalamic suprachiasmatic nucleus (SCN) and neurons of the thalamic paraventricular nucleus (PVT) projecting to the amygdala was investigated in the rat using tract tracing in light and electron microscopy. Biotinylated dextran amine was used to label anterogradely SCN efferents. These fibers were found to reach the thalamic midline, terminating in PVT, through three pathways: anterodorsally through the preoptic region, dorsally through the periventricular hypothalamus, and through the contralateral medial hypothalamic and preoptic areas after crossing the midline in the optic chiasm. Preterminal and terminal-like elements labeled from the SCN were distributed throughout the rostrocaudal extent of PVT, with an anteroposterior gradient of density. Labeled terminal elements were densest in the dorsal portion of PVT beneath the ependymal lining and some of them entered the ependyma. Anterograde tracing of SCN fibers was combined with injections of retrograde tracers in the amygdala. Numerous retrogradely labeled cell bodies were seen throughout PVT, with a prevalence in its anterodorsal portion. Overlap was detected between puncta labeled from the SCN and retrogradely labeled neurons, especially in the anterodorsal sector of PVT, where numerous puncta were in close apposition to thalamo-amygdaloid cells. Electron microscopy revealed that boutons labeled from the SCN established synaptic contacts with dendritic profiles of PVT neurons labeled from the amygdala. The findings demonstrate that information processed in the biological clock is conveyed to the amygdala through PVT, indicating that this nucleus plays a role in the transfer of circadian timing information to the limbic system.

The zonal organization of the corticonuclear and the olivocerebellar climbing fiber projections to the vermis of the cerebellum of the rat was compared to the pattern of zebrin-positive and zebrin-negative bands in material double-stained for zebrin II and for different anterograde tracers injected in subnuclei of the inferior olive, or retrograde tracers injected in the cerebellar and vestibular target nuclei of the Purkinje cells of the vermis. Projection zones A(1), A(X), X, B, C(X) in the vermis and A(2) (accessory A zone) and C(2) in the hemisphere were defined by their efferent corticonuclear and their afferent climbing fiber connections, and were found to share the same topographical framework with the zebrin pattern.