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PubMed1990

Relative sparing in Parkinson's disease of substantia nigra dopamine neurons containing calbindin-D28K.

The distribution of calbindin-D28K (CaBP)-positive neurons was investigated by immunohistochemistry in 4 controls, 5 cases of Parkinson's disease and a single case of strionigral degeneration. CaBP-positive neurons were preferentially localized to the mediodorsal portion of the substantia nigra pars compacta (SNC) in the beta layer, while CaBP-negative, melanin-positive neurons were concentrated in the ventrolateral SNC in the alpha layer. In Parkinson's disease and the case of strionigral degeneration, there was a relative sparing of the CaBP-positive neurons compared with CaBP-negative, pigmented neurons. These data imply that CaBP may confer some protection to SNC dopaminergic neurons against the pathological process which is responsible for Parkinson's disease and strionigral degeneration.

Aged↗

PubMed1990

Calbindin D28k-containing splanchnic and cutaneous dorsal root ganglion neurons of the rat.

Calbindin D28k (CaBP)-containing splanchnic and cutaneous sensory neurons in the rat dorsal root ganglia (DRGs) were investigated immunocytochemically in combination with a fluorescent dye tracer (Fluoro gold). About 15% of the DRG neurons at Th9-10 levels showed CaBP-like immunoreactivity. Eighty-four % of the splanchnic sensory neurons were immunoreactive to CaBP, while only 3% of the cutaneous sensory neurons were. The diameters of the splanchnic and cutaneous sensory neurons containing CaBP were 23.4 +/- 6.3 microns and 38.4 +/- 8.8 microns, respectively. Splanchnic sensory neurons containing CaBP were sensitive to capsaicin while cutaneous ones were not. These findings suggest that CaBP-containing splanchnic and cutaneous sensory neurons constitute different subgroups among the DRG neurons at the lower thoracic level.

Animals↗

PubMed1990

Long-term survival of GABA-, enkephalin-, NADPH-diaphorase- and calbindin-d28k-containing neurons in fetal striatal grafts.

Neurons in long-term striatal grafts were examined to determine if they retain the neurotransmitter characteristics of cells in younger grafts. In addition, calbindin-d28k, a ubiquitous marker of medium spiny neurons, was used to examine the overall frequency and ultrastructural characteristics of spiny neurons in the older grafts. Grafts from 17-day fetal striata were injected into the quinolinic acid-lesioned caudate nucleus in 5 adult rats. After 16 months, the neostriatum was processed for the localization of immunoreactive GABA, calbindin, enkephalin and NADPH-diaphorase (-d) activity. The proportions of GABA-, enkephalin- and NADPH-d-labeled neurons to total Nissl-stained neurons in the 16-month-old grafts (25 +/- 6, 13 +/- 4, and 3 +/- 3, respectively) were similar to findings in 2-month-old grafts. Calbindin-positive cells formed the highest proportion (36.3 +/- 3) of labeled neurons in the older grafts. Nuclear and spine morphology of immunoreactive calbindin cells varied more in the grafts than in host caudate. Results show that there is long-term survival and stability of GABA, enkephalin and NADPH-d cell populations in the grafts and that some grafted spiny neurons may exhibit altered phenotype from those of host striatum.

Animals↗

PubMed1991

Neurofilament M and calbindin D28k are present in mutually exclusive subpopulations of enteric neurons in the rat submucous plexus.

Immunocytochemical methods have been used to examine the localisation of 3 neurofilament proteins and the calcium binding protein, calbindin D28k, in whole mount preparations of the submucous plexus in the Wistar rat. Neurofilament-M (160 kDA protein) was present in 40% of the submucosal neurons, staining fine filaments in the soma and the axonal processes. Calbindin D28k was present in 40% of the submucosal neurons staining both the soma and nerves within the plexus. The neurofilament proteins and calbindin D28k were never observed within the same neurons. Neurofilament-M was co-localised with substance P and calcitonin gene-related peptide but not somatostatin or the other neuropeptides investigated. Calbindin D28k was co-localised with vasoactive intestinal polypeptide and neuropeptide Y. Galanin- and somatostatin-immunoreactive neurons did not contain either the neurofilament proteins or calbindin D28k. The results demonstrate the presence of subsets of submucosal neurons that can be distinguished by the presence of neurofilament-M or calbindin D28k.

Animals↗

PubMed1992

Nerve growth factor increases calcium binding protein (calbindin-D28K) in rat olfactory bulb.

Calbindin-D28K (CaBP28K) is a soluble intracellular protein capable of sequestering micromolar concentrations of calcium. The in vivo regulation of CaBP28K by recombinant human nerve growth factor (rhNGF) was studied in adult, male rats. Via Alzet 2002 pumps, each rat received, for 14 days, a lateral ventricle infusion (i.c.v.; n = 5-6/group) of 12 microliters PBS/day containing 1.0 microgram cytochrome C (control) or an equal amount of rhNGF. Six other animals received a vehicle or rhNGF infusion into the central neostriatum. CaBP28K was elevated by 75% (P less than 0.01) in the olfactory bulb following i.c.v. rhNGF in each of two experiments and was not altered in the temporal cortex, hippocampus, olfactory tubercle, cerebellum, or neostriatum. Direct striatal injections of rhNGF did not alter CaBP28K in the neostriatum or other regions (including the olfactory bulb). The increases in olfactory bulb CaBP28K protein levels were verified via Western blot analysis. CaBP28K immunocytochemistry revealed that 33% of olfactory bulb neurons are immunoreactive for CaBP28K and that the number or proportion of immunoreactive neurons did not change with i.c.v. infusions of rhNGF, suggesting that exogenously delivered rhNGF augments the content of CaBP28K in olfactory bulb neurons that normally express the protein. Endogenous NGF may function as a neuroprotective factor by enhancing the ability of these cells to sequester cytoplasmic calcium and retard calcium-mediated neurodegeneration.

Animals↗

PubMed1992

Melanin, tyrosine hydroxylase, calbindin and substance P in the human midbrain and substantia nigra in relation to nigrostriatal projections and differential neuronal susceptibility in Parkinson's disease.

The anatomy of melanin-containing neurons and other midbrain structures was examined by tyrosine hydroxylase (TH), calbindin D28k, and substance P immunostaining. Greater than 95% of cells in the substantia nigra pars compacta contained melanin, but densely packed cells in a ventral tier had a low content of melanin and loosely packed cells in a dorsal tier had a high content of melanin. Approximately 60% in the gamma group and 40% in the retrorubral nucleus had a low content of melanin. TH immunostaining was moderate in both the ventral and dorsal tiers, but more intense in the gamma group and retrorubral nucleus. Calbindin D28k was absent from the ventral and dorsal tiers, but present in the gamma group and retrorubral nucleus. In the light of primate tracing studies these findings suggest that the ventral tier of the pars compacta projects to striosomes of the striatum and the dorsal tier, gamma group and retrorubral nucleus to the matrix compartment. The ventral tier is more vulnerable than the dorsal tier in Parkinson's disease, but the cells contain less melanin. Neither tier contains calbindin D28k. This differential vulnerability between the ventral and dorsal tiers cannot be explained by melanin or calbindin D28k.

Aged↗

PubMed1992

Immunohistochemical markers in rat brain: colocalization of calretinin and calbindin-D28k with tyrosine hydroxylase.

Many dopaminergic cells of the substantia nigra are known to contain the calcium-binding proteins calretinin and calbindin-D28k. Catecholaminergic cell groups throughout the rat brain were therefore examined by two-colour immunofluorescence to determine whether they too contained these calcium-binding proteins as well as tyrosine hydroxylase (TH). Some TH+ cell groups are mostly positive for both calretinin and calbindin, notably in the ventral tegmental area, the interfascicular nucleus, and parts of the substantia nigra. Other TH+ cell groups in the midbrain, hindbrain and hypothalamus are very diverse; different cell groups are positive for calretinin, or calbindin, or both, or neither. In the olfactory bulb, entirely separate sets of periglomerular cells are positive for TH, calretinin and calbindin. However, there is considerable heterogeneity in calcium-binding protein expression within most cell groups, even in the substantia nigra. This could be a sign that calcium-binding proteins are regulated according to aspects of neuronal activity.

Animals↗

PubMed1992

Convergence and segregation of septal and median raphe inputs onto different subsets of hippocampal inhibitory interneurons.

The convergence and segregation of medial septal and median raphe afferents in the innervation of different subpopulations of GABAergic interneurons was investigated in the rat hippocampal formation. Following local injections of 5,7-dihydroxytryptamine into the median raphe nucleus destroying all serotonergic neurons, iontophoretic injections of Phaseolus vulgaris leucoagglutinin (PHAL) into the medial septum resulted in anterograde labelling of axons in the hippocampus. The labelled varicose fibres made multiple contacts with calbindin D28K-, parvalbumin-, and cholecystokinin-immunoreactive interneurons. These results disproved the possibility that PHAL-labelled afferents innervating hippocampal interneurons following septal PHAL injections would have been raphe axons passing through the injection site. In the second set of experiments a double anterograde tracing technique (PHAL from the septum and biotinylated-PHAL from the median raphe) and a triple or double immunostaining procedure was used to determine the types of interneurons (calbindin D28K-, parvalbumin-, or cholecystokinin-immunoreactive) innervated by one, or the other, or both pathways. The results showed that parvalbumin-containing neurons were innervated by septal afferents but avoided by raphe axons, whereas calbindin D28K-containing cells, and to a smaller extent cholecystokinin-containing cells served as targets for both pathways. In some cases the same individual calbindin D28K- or cholecystokinin-containing neurons received multiple contacts from afferents of both septal and raphe origin. Thus, our results indicate that different subcortical nuclei modulate largely different inhibitory circuits in the hippocampal formation. However, considering the occasional convergence of the two subcortical nuclei not only onto the same type, but even onto the same individual calbindin D28K-containing interneurons, we propose that a particular inhibitory function, most probably feed-forward inhibition in the distal dendritic region, is under the control of both pathways.

Afferent Pathways↗

PubMed1992

Immunohistochemical markers in rat cortex: co-localization of calretinin and calbindin-D28k with neuropeptides and GABA.

Calretinin and calbindin-D28k are two calcium-binding proteins which are present in separate populations of interneurons in cerebral cortex and hippocampus. To identify these cells with the populations expressing different transmitters, two-colour immunofluorescence was done with antibodies against the calcium-binding proteins plus antibodies against vasoactive intestinal peptide (VIP), somatostatin (SRIF), or gamma-aminobutyric acid (GABA). In neocortex, calretinin is partially co-localized with VIP (especially in the deeper layers) and is not co-localized with SRIF. Calbindin is largely co-localized with SRIF, and not with VIP. Both calretinin and calbindin are partially co-localized with GABA. In piriform and entorhinal cortex, the patterns resemble those in neocortex. In hippocampus, preliminary data indicate greater heterogeneity, especially in the ventral part; at least a few double-positive cells are present for every combination of calcium-binding protein and neuropeptide. These results expand the known diversity of local-circuit neurons in cortical regions.

Animals↗

PubMed1993

Calbindin-D28k immunoreactivity and selective vulnerability to ischemia in the dentate gyrus of the developing rat.

Hippocampal dentate granule cells normally express the calcium-binding protein calbindin-D28k and, in the adult, are the hippocampal neurons least vulnerable to an ischemic insult. We evaluated hippocampal structure 2-3 days after hypoxic/ischemic insult at postnatal day 7-10, and discovered that, unlike adult granule cells, developing granule cells were irreversibly injured. Localization of calbindin-D28k-like immunoreactivity (LI) revealed that the vulnerable cells were the immature granule cells at the base of the cell layer that were not yet calbindin-immunoreactive. Adjacent granule cells that did not die in response to the hypoxic/ischemic insult were calbindin-immunoreactive. Whether the lack of calbindin-LI in immature granule cells is causally related to their vulnerability, or is a coincidental reflection of cellular immaturity, remains to be determined.

Animals↗

PubMed1993

Colocalization of calbindin-D28k with vasopressin in hypothalamic cells of the rat: a double-labeling immunofluorescence study.

By use of a double-labeling immunofluorescence method, we examined whether vasopressin-containing cells possess a calcium-binding protein, calbindin-D28k, in the hypothalamus of the rat. Subpopulations of vasopressin-containing cells varied in their ability to possess calbindin-D28k immunoreactivity in different regions. In the supraoptic nucleus, most vasopressin-immunoreactive cells were also stained for calbindin-D28k. By contrast, in the magnocellular part of the hypothalamic paraventricular nucleus, all vasopressin-labeled cells lacked calbindin-D28k. In the suprachiasmatic nucleus, no calbindin-D28k was found in vasopressin-stained cells. This study shows a further characterization of vasopressin-containing cells of the rat hypothalamus.

Animals↗

PubMed1993

Immunohistochemical localization of calretinin-, calbindin-D28k- and parvalbumin-containing cells in the hypothalamic paraventricular and supraoptic nuclei of the rat.

The localization of three calcium-binding proteins, calretinin, calbindin-D28k and parvalbumin, in the hypothalamic paraventricular and supraoptic nuclei of the rat was immunohistochemically examined on adjacent sections and their distribution patterns were compared. Overlap between the distribution of calretinin-immunoreactive cells and that of calbindin-D28k-immunostained cells was found in the rostrodorsal part of the supraoptic nucleus, and the caudoventral part of this nucleus contained predominantly calbindin-D28k-stained cells. Cells of the medial and lateral magnocellular subdivisions of the paraventricular nucleus were almost devoid of the calcium-binding proteins examined. No parvalbumin-immunostained cells were observed in either nucleus. This study provides a further characterization of cell bodies in the hypothalamic paraventricular and supraoptic nuclei.

Animals↗

PubMed1993

Two populations of calbindin D28k-immunoreactive neurones in the striatum of the rat.

In order to examine the localisation of calbindin D28k in the normal striatum of the rat, immunocytochemistry using monoclonal antibodies was carried out at both the light and electron microscopic levels. As has previously been shown, many striatal spiny neurones were immunopositive, however, a small population of neurones with smooth dendrites and indented nuclei were also identified. At least some of these cells also displayed NADPH-diaphorase activity. These findings indicate that, in common with the primate striatum and grafted striatal tissue in the rat, the normal rat striatum contains at least two populations of neurones that express calbindin D28k.

Animals↗

PubMed1993

Calretinin, calbindin-D28k and parvalbumin-like immunoreactivity in mouse chemoreceptor neurons.

Calretinin immunoreactivity was demonstrated in adult mouse olfactory receptor neurons and in the vomeronasal and septal chemoreceptor neurons, whereas parvalbumin expression was restricted to the vomeronasal receptor neurons. Calbindin-D28k-like immunoreactivity was primarily localized in the vomeronasal and septal chemoreceptor neurons although an occasional neuronal staining with calbindin-D28k was also found in restricted areas of the main olfactory epithelium.

Animals↗

PubMed1994

Calbindin-like immunoreactivity in epithelial cells of the newborn and adult human vomeronasal organ.

The vomeronasal organs (VNOs) of two humans, a male neonate and a female adult, were examined for immunolocalization of calbindin-D28k (calbindin) which has been immunolocalized to VNO receptor cells in other mammals. The present study demonstrates that epithelial cells within the VNOs of both subjects expressed calbindin-like immunoreactivity. These results suggest that human VNO epithelial cells of both genders express calbindin during development and in the adult.

Adult↗

PubMed1994

Infrequent co-localization of nitric oxide synthase and calcium binding proteins immunoreactivity in rat neocortical neurons.

Immunoreactivity to nitric oxide synthase (NOS-IR) was detected in a small population of rat neocortical neurons scattered throughout the frontal, parietal, temporal and occipital cortices. Two-color double-labeling studies revealed that the number of NOS-IR neocortical neurons expressing immunoreactivity to the calcium binding proteins parvalbumin (PV-IR), calbindin-D28K (CB-IR) or calretinin (CR-IR), was low. The absence or low level of calcium binding proteins may imply a reduced Ca2+ buffering capacity in NOS-containing neurons, thereby contributing to their vulnerability to Ca2+ influx through the activated AMPA/kainate receptors reported by others.

Amino Acid Oxidoreductases↗

PubMed1994

Calbindin-D28k immunoreactive neurons form two populations in the rat nucleus accumbens: a compartmental study.

This study shows localisation of a second class of calbindin-D28k containing neuron other than the medium-sized spiny neuron in the rat nucleus accumbens, using immunohistochemistry with monoclonal antibodies. Ultrastructural characteristics were used to identify this second population of accumbens neurons within its compartments. Staining with NADPH-diaphorase and calbindin-D28k immunoreactivity revealed that some neurons contained both markers.

Animals↗

PubMed1994

Combination of alkaline phosphatase in situ hybridization with immunohistochemistry: colocalization of calretinin-mRNA with calbindin and tyrosine hydroxylase immunoreactivity in rat substantia nigra neurons.

We describe a method to combine non-radioactive in situ hybridization using alkaline phosphatase (AP) labelled oligonucleotide-probes with immunohistochemistry on the same thin paraffin section. The simultaneous detection of calretinin-mRNA and calbindin- or tyrosine hydroxylase-like immunoreactivity in neurons of rat substantia nigra, pars compacta, was used as a test system to develop the method. Brains were fixed by perfusion with 4% paraformaldehyde and embedded in paraffin. Five-microns-thick sections were processed for non-radioactive in situ hybridization with a 33-base alkaline phosphatase conjugated synthetic oligonucleotide complementary to calretinin mRNA. After hybridization and colour reaction to visualize calretinin mRNA, sections were incubated with antibodies against calbindin D28K or tyrosine hydroxylase. Immunoreaction was visualized using the avidin-biotin-complex-technique and diaminobenzidine. As the colour of both reaction products differ markedly, the distribution of calretinin mRNA-containing neurons (purple-blue, alkaline phosphatase product) and calbindin/tyrosine hydroxylase immunopositive cells (brown peroxidase product) could be differentiated easily on the same section. Calbindin- and tyrosine hydroxylase-like immunoreactivity was found in the majority of calretinin mRNA-containing cells within the substantia nigra, pars compacta, indicating that in this nucleus a proportion of the dopaminergic neurons contain both calcium binding proteins calbindin and calretinin. In conclusion, non-radioactive in situ hybridization using alkaline phosphatase labelled oligonucleotide probes can be readily combined with immunohistochemistry.

Alkaline Phosphatase↗