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D M Jacobowitz

Publications and source records attributed to D M Jacobowitz.

At least 73 records · Page 4Linked to original sources
PubMed1994

Ca(2+)-dependent and independent interactions of calretinin with hydrophobic resins.

The ability of rat calretinin to bind to hydrophobic resins in a Ca(2+)-dependent manner was examined. Both native calretinin present in cerebellum extract and purified recombinant calretinin bound similarly to hydrophobic resins such as phenyl-, hexyl-, octyl-, and W7-agarose. Hydrophobic interactions of calretinin were partially Ca(2+)-dependent since 1/3 of bound protein was released from the resins by EGTA under varied conditions. Some calretinin tryptic fragments bound to octyl-agarose in a manner similar to uncleaved calretinin, while others bound to the resin in a Ca(2+)-independent manner. These and other results suggest that calretinin has several hydrophobic regions of varied strength and sensitivity to Ca2+. It is proposed that the local changes in hydrophobicity induced by Ca2+ binding might be relevant for calretinin functions.

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

Calretinin-immunoreactivity in trigeminal neurons innervating the nasal mucosa of the rat.

Trigeminal primary neuronal cell bodies were labeled by retrograde transport of Fluoro-gold (FG) from the nasal mucosa of rats. The trigeminal ganglion containing the labeled cell bodies were processed for double stain for calretinin- and tachykinin-immunoreactivities (CR- and TK-irs). Except for a few contralateral cells, all the cells that innervated the nasal mucosa (NM cells) were confined to the ophthalmo-maxillary division of the trigeminal ganglion ipsilateral to the FG application. In the dorsal two-thirds of the ganglion, NM cells formed a cluster in the rostromedial part of ophthalmo-maxillary division (the rostromedial cluster). In the ventral third, the number of cells in the rostromedial cluster markedly decreased. Instead, numerous NM cells were found in the caudolateral part of the ophthalmo-maxillary division (the caudoventrolateral cluster). CR- and TK-irs were detected in 18% and 54% of overall population of NM cells, respectively. Virtually all of CR-immunoreactive (-ir) NM cells coexpressed TK. Although the proportion of TK-ir cells, irrespective of CR-ir, was similar for both clusters, CR-ir cells were more frequent in the caudoventrolateral cluster than in the rostromedial cluster. In the dorsal 1/3 of the ganglion where all the NM cells belonged to the rostromedial cluster, only 8.4% exhibited CR-ir. On the other hand, as much as 30.1% of NM cells expressed CR-ir in the ventral 1/3 where most NM cells were found in the caudoventrolateral cluster. Trigeminal cell bodies innervating the cornea and conjunctivum were located in the rostromedial part of the ophthalmo-maxillary division.(ABSTRACT TRUNCATED AT 250 WORDS)

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

Calretinin-immunoreactive neurons in the trigeminal and dorsal root ganglia of the rat.

The cell body size of primary neurons were measured in the trigeminal (TG) and lumbar dorsal root ganglia (DRG) monochrome-stained for calretinin (CR)-like immunoreactivity. A trichrome stain for CR, carbonic anhydrase (CA) and tachykinin (TK) was also employed to estimate possible overlap of cellular distribution of these substances. In the DRG, the cell size spectrum of CR-positive cells was clearly bimodal; a greater proportion (84.1%) of CR-positive cells was distributed in the range > or = 800 microns2 with a mode between 1,500-1,600 microns2, while a smaller proportion (14.8%) < 700 microns2 with a mode of 400-500 microns2. They were evenly distributed throughout the DRG. Although CR-positive TG neurons were smaller than similar DRG neurons, a bimodal distribution pattern remained unchanged. 94.6% of CR-positive cells measured 100-1,400 microns2 with peak ranges of 200-300 microns2 and 400-500 microns2. Most of CR-positive cells in the ophthalmic division were 400 microns2 or larger and small CR-positive cells (< 400 microns2) were concentrated in the maxillary and mandibular divisions. Most of CR-positive DRG cells showed CA activity (76.5%), while those with TK-immunoreactivity were rare (7.2%). In the TG, 38.4% of CR-positive cells were TK-positive. They were mostly smaller than 800 microns2. On the other hand, CA was detected in 43.4% of CR-positive TG cells. Most of the TG cells co-expressing CR and CA were 400 microns2 or larger. Simultaneous co-expression of TK and CA by the CR-positive cells was negligible.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1993

Ultrastructural localization of calretinin immunoreactivity in lobule V of the rat cerebellum.

Ultrastructural localization of a calcium-binding protein, calretinin, in the cerebellar cortex of the rat was examined with an immunoperoxidase method. The cerebellar lobule V was investigated in detail. In the molecular layer, calretinin immunoreactivity was found in parallel fibers and in their varicosities that formed synapses with unlabeled Purkinje cell dendritic thorns and Golgi cell dendrites. In the granular layer, most granule cells were immunostained for calretinin. Within immunostained granule cells, the labels were found in the cytoplasmic matrix of both the perikaryon and primary dendrite, and in euchromatin patches of the nucleus. Some mossy fiber terminals also stained for calretinin. In the white matter, a few axons were labeled for calretinin and surrounded by unstained myelin sheath. It is suggested that calretinin may play a role at the presynaptic sites of the parallel and mossy fiber terminals.

Animals↗
PubMed1993

Appearance of calretinin-immunoreactive neurons in the upper layers of the rat superior colliculus after eye enucleation.

The effects of retinal deafferentation on a calcium-binding protein, calretinin, in the upper layers (superficial gray layer and optic nerve layer) of the rat superior colliculus were examined. In intact rats and on the ipsilateral side of unilaterally eye-enucleated rats, the superficial gray layer and optic nerve layer contained a few dispersed calretinin-immunoreactive cells. On the contralateral side to the enucleation, the number of immunostained cells in the superficial gray layer and optic nerve layer was increased. These findings suggest that retinal deafferentation results in an increase in contents of calretinin in some cell bodies within the upper layers of the superior colliculus.

Afferent Pathways↗
PubMed1993

Quantitative measurement of calretinin and beta-actin mRNA [correction of mRNAIN] in rat brain micropunches without prior isolation of RNA.

A microdissection technique for quantitation of neurochemicals in discrete brain nuclei has been applied to quantitative measurement of mRNA. The method permits quantitation of low abundance mRNA from submilligram amounts of tissue (10-500 micrograms protein). Discrete nuclei and other regions of the brain are solubilized in concentrated guanidine thiocyanate solution, mRNA is directly hybridized with riboprobes, and detected with a ribonuclease protection assay. This method eliminates the necessity for RNA isolation from solid tissue. No assumptions regarding RNA recovery are necessary since tissue specimens are solubilized, hybridized and treated with ribonuclease in a single tube. We have determined the mRNA levels of calretinin, a predominantly neuron-specific calcium binding protein in microdissected nuclei and other regions of rat brain. For interassay comparison, measurement of sample protein and beta-actin mRNA permits normalization and quantitation in terms of these internal controls. The quantity of calretinin mRNA ranged from 281 +/- 35 fg/micrograms protein in the thalamic paraventricular nucleus to 2.3 +/- 0.5 fg/micrograms protein for the cerebral cortex. The calretinin/beta-actin ratios ranged from 79.9 +/- 9.3% to 1.3 +/- 0.1%, respectively. The combination of microdissection techniques with a lysate RNase protection assay: (1) establishes this technique as quantitative for detection of high and low abundance mRNAs from microdissected brain specimens; (2) bypasses the inefficiencies and uncertainties associated with isolating RNA; and (3) enables large numbers of determinations from discrete brain nuclei to be analyzed in 2 to 3 days.

Actins↗
PubMed1993

Nucleotide sequence of rat calretinin cDNA.

A rat calretinin cDNA clone was selected by antibody screening of a lambda gt11 brain library. The sequence revealed remarkable nucleotide and amino acid homology with human calretinin (91.1% and 98.5%, respectively), with only four amino acid differences. A high degree of homology with chick calretinin was also observed (79.8% and 86.6%, respectively), with 36 amino acid differences. Although the role of this central nervous system protein has not been well characterized, the evolutionarily conserved calcium binding domains and connecting regions, in addition to the limited local changes observed between rat and chick primary structure, lead us to believe that calretinin interacts with other highly conserved constituents of brain cells. This calretinin cDNA clone provides a new probe for the analysis of a specific subset of neurons in the central nervous system. The probe will allow a more detailed analysis of calretinin regulation in the brain and will be useful for screening genomic libraries for the complete chromosomal gene. (GenBank accession No X66974).

Amino Acid Sequence↗
PubMed1993

Immunohistochemical localization of calretinin in the dorsal root ganglion and spinal cord of the rat.

Calretinin (CR), a recently identified calcium-binding protein, is present in nervous tissue, including sensory pathways, where it may play an important role in regulation of cellular activity. Using immunocytochemistry, we examined the cellular localization of CR in dorsal root ganglia (DRG) and spinal cord of normal rats and after multiple unilateral dorsal root ganglionectomies. In DRG, CR-immunoreactive cell bodies and axons were a small subpopulation (10%) of medium- to large-sized neurons. In the spinal cord, CR-like immunoreactivity (LI) in neurons and fibers was found in all laminae except motoneurons. Dense fiber networks were also found in Clarke's column. The densest staining of both cell bodies and fibers was in the superficial laminae, especially lamina II, and in the lateral spinal and lateral cervical nuclei. CR-immunoreactive fibers were also observed in the fasciculi cuneatus and gracilis. Fasciculus gracilis exhibited the greatest number of labeled axons at the lumbosacral levels, but few labeled axons were found at the rostral thoracic and cervical levels. In contrast, the corticospinal tract at the base of the dorsal column was devoid of CR-immunoreactive fibers. Unilateral multiple lumbar ganglionectomies resulted in a loss of CR-LI in the dorsal columns ipsilateral to the surgery. In the spinal gray matter ipsilateral to the ganglionectomies, CR-LI was reduced in Clarke's column and slightly enhanced in the medial third of lamina II. Our observations demonstrate a unique distribution pattern of CR-LI compared to other calcium-binding proteins in the spinal cord, and suggest a role for CR in nociceptive and proprioceptive pathways.

Animals↗
PubMed1992

Immunohistochemical localization of calretinin in the rat lateral geniculate nucleus and its retino-geniculate projection.

In the present study, we examined the distribution of calretinin-immunoreactive neuronal cell bodies and fibers in the lateral geniculate nucleus of the rat. In normal rats, clusters of immunoreactive cell bodies were found in: (i) the rostral portion of the ventral lateral geniculate nucleus pars medialis (VLGM), (ii) the intergeniculate leaflet (IGL), (iii) the intermediate region between the VLGM and the ventral lateral geniculate nucleus pars lateralis (VLGL), (iv) the caudomedial portion of the VLGM, and (v) the caudolateral portion of the VLGM. In the dorsal lateral geniculate nucleus (DLG), immunoreactive cell bodies were rarely observed. After uni- or bilateral eye enucleation, no significant alteration in the morphological features or distribution of immunoreactive cell bodies was detected in the lateral geniculate nucleus. In normal rats, immunoreactive fibers formed dense plexuses in: (i) the DLG, (ii) the external layer of the VLGL, (iii) the internal layer of the VLGL, (iv) the IGL, (v) the caudomedial portion of the VLGM, and (vi) the optic tract. After unilateral eye enucleation, immunoreactive fibers in the external layer of the VLG and in the optic tract almost totally disappeared on the contralateral side to the lesion. Unilateral eye enucleation caused a significant decrease of immunoreactive fibers in the DLG and in the internal layer of the VLGL, but a substantial number of immunoreactive fibers still remained there. In the IGL and the caudomedial portion of the VLGM, no observable alteration in the distribution of immunoreactive fibers was detected after uni- or bilateral eye enucleation.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

Calretinin-immunoreactivity in the oro-facial and pharyngeal regions of the rat.

Calretinin-immunoreactivity (CR-ir) was examined in the rat oro-facial and pharyngeal tissues using an immunofluorescence method. CR-ir was distributed in the entire size range of trigeminal ganglion neurons. CR-ir was also observed in nerve fibers surrounding neuronal cell bodies in autonomic ganglia, and in nerve endings in the lip, tongue, incisal papilla, soft palate, pharynx and epiglottis. CR-immunoreactive nerve endings were all in close proximity to the epithelium, and classified into 2 types; simple (free nerve ending) and taste-bud-related types. In the salivary gland, positive nerve fibers were seen around large excretory ducts. The present study indicates that viscerosensory (probably including gustatory) nerve fibers innervating the oral and pharyngeal tissues contain CR, while somotosensory nerve fibers innervating the facial skin are devoid of CR.

Animals↗
PubMed1992

Chymotrypsin-reactive antibodies in insulin-dependent diabetes mellitus.

In a previous study (Frazier et al., 1990), it was demonstrated that two patients with type 1 (insulin-dependent) diabetes mellitus had antibodies in their serum which reacted with four 29 kDa pancreas-specific proteins on two-dimensional immunoblots. This paper reports on the purification and identification of these pancreatic proteins. The protein with the pI closest to pH7 was purified through the use of ammonium sulfate fractionation and ion-exchange chromatography. Gel filtration chromatography established that the protein's molecular weight was closer to 25 kDa. Amino acid composition and sequence analyses demonstrated homology between the protein and chymotrypsin. It is suggested that an abnormal regulation of chymotrypsin activity might be related to antibodies formed in some diabetic patients.

Adolescent↗
PubMed1992

Calretinin is present in non-pyramidal cells of the rat hippocampus--I. A new type of neuron specifically associated with the mossy fibre system.

Calretinin-containing cells were visualized with immunocytochemistry in the rat dorsal hippocampal formation. Calretinin immunoreactivity was present exclusively in non-pyramidal cells in all layers of the dentate gyrus and the CA1-3 areas. Calretinin-positive neurons and processes were most abundant in the hilus of the dentate gyrus and in the stratum lucidum of the CA3 region. Several calretinin-immunoreactive cells were located within the hippocampal fissure. A distinct band of calretinin-immunoreactive fibres occupied the superficial part of the granule cell layer and the lowest part of the molecular layer. Closer examination of the calretinin-positive cells revealed that they formed two distinct cell groups. One group of cells, found exclusively in the stratum lucidum of the CA3 area and in the hilus of the dentate gyrus, was covered with numerous spines. Their somata and dendrites were restricted to stratum lucidum and to the hilus. Cells of the other group had smooth, often varicose, radially running dendrites, and were present in all areas and layers of the hippocampal formation. Two to three thick primary dendrites arose from the irregularly shaped cell body of spiny cells and emitted fine secondary branches only distally (70-100 microns) from the soma, where they formed a profuse network. The extensive dendritic tree of the cells spread horizontally within stratum lucidum and span a distance of 400-600 microns both in the septotemporal and in the transverse directions. The layer-specific location of these cells and their processes suggested that the majority of their input may derive from mossy fibres. This presumption has been confirmed by electron microscopic examination. A large number of asymmetrical synapses were found to cover the soma, the dendritic shafts and the spines (four to six synapses/spine) of the cells. A large proportion of the synapses were formed by boutons, which showed the distinctive features of mossy fibre terminals. Three to six primary dendrites arose from the multipolar, bipolar or pyramidal-shaped somata of spine-free cells, which were smaller than the somata of spiny cells. The smooth and frequently varicose dendrites branched proximally and ran primarily radially. Dendrites ascended or descended through several layers and received both asymmetrical and symmetrical synapses. In the CA1 subfield, the vertically running dendrites frequently contacted other calretinin-immunoreactive spine-free dendrites or cell bodies. Two or three calretinin-immunoreactive dendrites were often seen to be attached for over 100 or, occasionally, 200 microns and several puncta adherentia were observed between them using the electron microscope.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1992

Calretinin is present in non-pyramidal cells of the rat hippocampus--II. Co-existence with other calcium binding proteins and GABA.

The possible co-existence of calretinin with other calcium binding proteins, parvalbumin and calbindin D28k, and with GABA, was studied in non-pyramidal cells of the rat dorsal hippocampal formation, using the mirror technique. The majority of the calretinin-containing neurons (83%) were found to be immunoreactive for GABA (79% in the dentate gyrus, 84% in the CA2-3, and 88% in the CA1 subfield). Most of the GABA-negative calretinin-immunoreactive neurons were located in the hilus of the dentate gyrus and in stratum lucidum of the CA3 subfield. Detailed analysis of the calretinin-immunoreactive cells of these subfields revealed that the two morphologically distinct types of calretinin neurons, i.e. the spiny and the spine-free cells, differ in their immunoreactivity for GABA. The overwhelming majority (92%) of the spine-free neurons were GABA-positive, whereas the immunoreactivity of spiny cells was ambiguous. At the sensitivity threshold of the immunocytochemical techniques used in the present study, most of the spiny cells (89%) had to be considered as GABA-negative, although the staining intensity in their cell bodies was somewhat above background level. Colchicine treatment resulted in a degeneration of calretinin-immunoreactive neurons; therefore, its effect on the GABA content of spiny neurons could not be evaluated. Nevertheless, the observations suggest that calretinin-containing neurons are heterogeneous both morphologically and neurochemically. Examination of the co-existence of calcium binding proteins revealed that none of the hippocampal cells contained both calretinin and parvalbumin in any regions of the hippocampal formation. Some overlap was detected between the calretinin- and the calbindin D28k-containing cell populations, 5.1% of the former and 6.2% of the latter were immunoreactive for both calcium binding proteins. This may be due to a small degree of cross-reactivity of the calbindin D28k antiserum with calretinin. Thus, our results demonstrate that the majority of calretinin-immunoreactive neurons are GABAergic and represent a subpopulation of non-pyramidal cells with no or only a negligible overlap with the subpopulations containing the other calcium binding proteins, parvalbumin and calbindin.

Animals↗