SEARCH · PubMed Health

Results for “Colocalization”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 469 records · Page 26Linked to original sources

PubMed1993

Macromolecules that are colocalized with deposits of beta 2-microglobulin in hemodialysis-associated amyloidosis.

BACKGROUND: Common elements in many different types of amyloid may have important roles in amyloidogenesis. The proteinaceous tissue deposits have a common appearance in polarized light and other similar features. The present investigation describes for the first time the relation between beta 2-microglobulin (beta 2-M)-type amyloidosis and colocalized materials, as demonstrated using specific antibodies and hyaluronan-binding protein. EXPERIMENTAL DESIGN: Amyloid-rich carpal tunnel synovium was obtained surgically from 28 patients who were being treated by maintenance hemodialysis. Serial sections were examined using a hyaluronan (hyaluronic acid)-binding protein and antibodies against heparan sulfate-glycosaminoglycan, chondroitin sulfate-proteoglycan, dermatan sulfate-proteoglycan, alpha 1-antichymotrypsin, alpha 1-antitrypsin, inter-alpha-trypsin inhibitor, haptoglobin, and ubiquitin. RESULTS: Accumulation of hyaluronan was of three types, namely, localization around beta 2-M deposits, colocalization with deposition of beta 2-M itself and localization at a small distance from beta 2-M deposits. Immunostaining for heparan sulfate glycosaminoglycan was demonstrated at the sites of beta 2-M plaques. Chondroitin sulfate-proteoglycan did not show specific patterns of immunostaining, resembling hyaluronan rather than heparan sulfate. The other materials tested, alpha 1-antichymotrypsin, alpha 1-antitrypsin, inter-alpha-trypsin, haptoglobin and ubiquitin, were not immunostained at sites of beta 2-M plaques. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting revealed that the molecular weight of heparan sulfate-glycosaminoglycan was 16,000. CONCLUSIONS: These results suggest that HS has an important role in hemodialysis-associated amyloidosis as it does in other types of amyloidosis. Moreover, accumulation of hyaluronan may be an indication of inflammation of the carpal synovium.

Amyloidosis↗

PubMed1995

Odorant receptors and desensitization proteins colocalize in mammalian sperm.

BACKGROUND: The identification of transcripts encoding putative olfactory receptors in mammalian germ cells (1) has generated the hypothesis that olfactory receptors may serve a chemosensory role in sperm chemotaxis during fertilization. We have sought to identify and localize these receptors and their regulatory machinery in rat sperm in order to gain further insight into mammalian sperm chemotaxis and odorant receptor physiology. MATERIALS AND METHODS: We conducted reverse transcription-polymerase chain reaction (RT-PCR) using degenerate primers directed against sequences conserved across members of the known odorant receptor family to identify transcripts from testis and round spermatids. Western analysis and immunohistochemistry were performed using antibodies raised against two peptide sequences conserved among odorant receptors and using fusion protein antibodies to G-protein receptor kinase 3 (GRK3/beta ARK2) and beta-arrestin2. RESULTS: We detected transcripts encoding putative odorant receptors in both testis and round spermatids of the adult rat. Restriction digests of the PCR products demonstrated the existence of multiple gene products. Two anti-odorant receptor antibodies specifically recognized a 64 kD band in rat sperm preparations by Western blot. The proteins GRK3 and beta-arrestin2, implicated in olfactory desensitization, were detected in sperm cytosolic extracts using Western analysis. Immunohistochemistry colocalized putative odorant receptors, GRK3 and beta-arrestin2 to elongating spermatids in the testis and to the midpiece of mature sperm. CONCLUSIONS: The specific localization of odorant receptors to the respiratory center of mature sperm is consistent with a role for these proteins in transducing chemotactic signals. Based on the colocalization, it is plausible that GRK3 and beta-arrestin2 function in sperm to regulate putative chemoreceptor responses.

Animals↗

PubMed1996

Nuclear colocalization of the Ro 60 kDa autoantigen and a subset of U snRNP domains.

The Ro 60 kDa protein is an RNA binding molecule present both in the cytoplasm and in the nucleus. Cytoplasmic Ro 60 kDa is complexed to other proteins and to certain RNAs denoted hYRNAs. This RNA-protein complex is also known as the Ro/SSA antigen recognized by sera from patients with certain autoimmune disorders. Components interacting with the nuclear Ro 60 kDa protein fraction in mammalian cells have not been identified. To look for an association with previously known nuclear structures, rabbit antisera to the amino- and carboxy-terminal parts of the Ro 60 kDa protein were used in immunomorphological studies on HeLa cells. A strong speckled nuclear pattern and a weak cytoplasmic staining were detected. Double immunofluorescence staining with affinity purified anti-Ro 60 kDa antibodies and monoclonal antibodies recognizing the Sm and RNP antigens of the U snRNPs, displayed colocalization. Another U snRNP containing nuclear compartment, the coiled bodies, did not contain any Ro 60 kDa protein. Cells infected with a toga virus demonstrated redistribution of both U snRNP antigens and the Ro 60 kDa protein with retained colocalization. These results indicate a role for the nuclear fraction of the Ro 60 kDa protein in RNA processing.

Animals↗

PubMed1997

Colocalization of hyaluronan and water in rabbit corneas after photorefractive keratectomy by specific staining for hyaluronan and by quantitative microradiography.

PURPOSE: Endogenous hyaluronan (HA) has previously been shown to be a part of corneal wound healing after photorefractive keratectomy (PRK). HA is known to bind considerable amounts of water. This study aimed to show whether HA colocalized with water in the wound area after PRK. METHODS: Five New Zealand White rabbits were subjected to PRK in one eye each. After 4 weeks, the rabbits were killed and the corneas were freeze-sectioned. The freeze-dried sections were exposed to soft x-rays generated at 3 kV and the microradiographs were evaluated densitometrically. The same sections were then stained for the presence of HA using the HA-binding region of bovine nasal cartilage proteoglycans linked to biotin. RESULTS: The microradiographs showed, in a zone underlying the epithelium, that the water content was increased. In the corresponding zone, the cornea stained positive for HA. CONCLUSION: HA that is reactively formed in the corneal wound after PRK colocalized to the hydrated area of the corresponding location as revealed by quantitative microradiography. The findings suggest that HA causes local shifts in water content in the corneal wound and thereby also local shifts in transparency.

Animals↗

PubMed1997

Further similarities between astrocytes and perisinusoidal stellate cells of liver (Ito cells): colocalization of desmin and glial fibrillary acidic protein in astroglial primary cultures.

The colocalization of desmin and glial fibrillary acidic protein (GFAP) in astrocytes was inferred from previous studies demonstrating a unique antigenic composition comprising GFAP, desmin and vimentin in perisinusoidal stellate cells (PSC) of liver which share several features with astrocytes. In the present study the colocalization of GFAP and desmin was investigated by double-immunolabeling experiments in 12-day-old rat astroglial primary cultures with antiserum against GFAP and two commercial monoclonal antibodies against desmin, antibodies of clone DEU-10 and clone DEB-5. These antibodies selectively decorated the perisinusoidal stellate cells (PSC) of liver for which desmin is known to be a marker. The results obtained with astroglial cells demonstrate that both GFAP and desmin are coexpressed in morphologically different types, process-bearing and process-lacking astrocytes. The expression of desmin was apparently more pronounced in process-lacking astrocytes and was considerably lower in process-bearing ones. In process-lacking astrocytes, in contrast to filamentous cytoplasmic staining for GFAP, the immunoreactivity for desmin was non-filamentous and was irregularly spread in the perinuclear cytoplasm of the cells, while in process-bearing astrocytes the pattern of staining for desmin was similar to that of GFAP. The variability in the intensity and pattern of staining for desmin in astrocytes might be due to transitional stages of differentiation for part of the cells. This interpretation was supported by the presence of GFAP in the cells weakly expressing smooth muscle alpha-actin and the absence of GFAP in the cells enriched with microfilaments.

Actins↗

PubMed1998

Colocalization of substance P or enkephalin in serotonergic neuronal afferents to the hypoglossal nucleus in the rat.

The serotonergic innervation of the hypoglossal nucleus originates from the caudal raphe nuclei. Non-serotonergic neurons in the caudal raphe nuclei also project to the hypoglossal nucleus. We employed a triple-fluorescence technique to determine whether the substance P- or the enkephalin-containing neurons in the caudal raphe nuclei that projected to the hypoglossal nucleus also contained serotonin. Rhodamine latex microspheres were injected into the hypoglossal nucleus, and then serotonin and peptide dual-immunofluorescence was performed to colocalize perikarya containing serotonin, substance P, and rhodamine microspheres; or perikarya containing serotonin, enkephalin, and rhodamine microspheres. Our results demonstrate that most substance P-containing neuronal afferents to the hypoglossal nucleus colocalize serotonin. In contrast, few enkephalin-containing neuronal afferents to the hypoglossal nucleus also contain serotonin. These data suggest that substance P projections to the hypoglossal nucleus are a subset of serotonergic projections and that limited overlap exists between the populations of enkephalinergic and serotonergic neuronal afferents to the hypoglossal nucleus. Either substance P- or enkephalin-containing somata account for a very small proportion of non-serotonergic caudal raphe projections to the hypoglossal nucleus. Finally, these data demonstrate the medial tegmental field origins of the substance P projections and the enkephalin projections to the hypoglossal nucleus.

Afferent Pathways↗

PubMed1997

Colocalization of dopamine D1 and D2 receptor mRNAs in rat placenta.

Dopamine is present in the human placenta. The major function of dopamine is the inhibition of human placental lactogen (hPL) release from human trophoblastic cells. This effect is mediated by cAMP through dopamine D2 receptors. However, studies on the effects of cAMP in the control of hPL release have yielded conflicting results. The purpose of this study is to explore the distribution of dopamine receptors in the rat placenta. Dopamine D1 and D2 receptor mRNAs were colocalized in the rat placenta by in situ hybridization histochemistry using radiolabeled cRNA probes. Dopamine D1 and D2 receptor mRNAs were detected in large cells of the endometrium of the uterus on day 10 of gestation. On days 12-16 of gestation, hybridization signals were localized mainly in the spongiotrophoblast and giant cells of the junctional zone of the placenta. With the development of the placenta, signals were moving from the junctional zone to the labyrinth zone. Pit-1 mRNA was detected in the placental lactotrophs and was also colocalized in neighboring placental sections. Our results clearly showed that dopamine D1 and D2 receptor mRNAs were coexpressed in the placental lactotrophs that express Pit-1 mRNA.

Animals↗

PubMed1998

Discoordinate surface expression of IFN-gamma-induced HLA class II proteins in nonprofessional antigen-presenting cells with absence of DM and class II colocalization.

We compared HLA class II expression in a human melanoma line (a nonprofessional APC), induced by IFN-gamma or by stable transfection with CIITA, with constitutive class II expression in an EBV-transformed B lymphoblastoid cell line (a professional APC) from the same donor. IFN-gamma-induced and CIITA-transfected melanoma cells expressed DR, DP, and DQ at levels similar to those expressed by the professional APC; however, DP and DQ proteins and DM-dependent DR epitopes were delayed in appearing on the cell surface when induced by IFN-gamma. The delay in cell surface expression of some IFN-gamma-induced class II epitopes was observed even though Northern blots demonstrated class II and DM genes to be coordinately transcribed and their mRNA levels to be equivalent to that in B lymphoblastoid cells. Confocal microscopy suggests that discoordinate cell surface expression of class II results from different intracellular trafficking for IFN-gamma-induced class II proteins in the melanoma line compared with that in professional APCs. Specifically, although DR and DM proteins were present 2 days after IFN-gamma induction, colocalization of DR and DM proteins intracellularly was not apparent in cells at any time after induction. Failure of DR and DM proteins to colocalize suggests that IFN-gamma-induced cells lack an intracellular MIIC-like compartment. The absence of a compartment containing DR and DM to facilitate interaction between the two proteins may account for the delayed surface expression of class II epitopes whose formation requires both class II and DM.

Antigen Presentation↗

PubMed1998

CCND1 and FGFR1 coamplification results in the colocalization of 11q13 and 8p12 sequences in breast tumor nuclei.

The CCND1 gene, localized to chromosome band 11q13, is amplified in approximately 15% of human primary breast tumors. From 30 to 40% of the tumors presenting this amplification show concomitant amplification at the FGFR1 locus in 8p12. Similarly, MDA-MB-134 breast cancer cells bear CCND1 and FGFR1 coamplified, resulting in the formation of a hybrid intrachromosomal amplification assembling 11q13 and 8p12 sequences. To learn whether similar amplified structures arise in breast tumors, we used a two-color FISH approach on interphase nuclei. A cohort of 225 breast tumors was analyzed by Southern blotting and a subset of 12 tumors presenting the 11q13-8p12 coamplification was selected for further study by interphase FISH. In 6/12 tumors the FISH signals for 11q13 and 8p12 probes formed colocalizing clusters of green and red spots in the nuclei. The FISH patterns were identical to those observed on MDA-MB-134 interphase nuclei hybridized with 11q13 and 8p12. These data, suggesting the formation in these tumors of a hybrid amplification domain in which 11q13 and 8p12 sequences are joined, were reinforced by dual-color FISH on extended chromatin showing that the said were sequentially aligned in these tumors. Furthermore, 3/6 nuclei with colocalized 11q13 and 8p12 amplifications showed fusion of centromeric sequences from chromosomes 8 and 11. Our data strongly suggest the occurrence, in approximately 3% of primary breast tumors, of a recurrent rearrangement involving the proximal portions of 8p and 11q and resulting in the formation of a hybrid amplified structure composed of 11q13 and 8p12 sequences.

Base Sequence↗

PubMed1996

Glutamate and aspartate immunoreactive neurons of the rat basolateral amygdala: colocalization of excitatory amino acids and projections to the limbic circuit.

The basolateral amygdala has projections to several structures that take part in the limbic cortico-striato-pallido-thalamic circuit, including the prefrontal cortex, ventral striatum, and mediodorsal thalamic nucleus. The present investigation used a technique that combines retrograde tract tracing with immunohistochemistry for glutamate and aspartate to determine if amygdaloid neurons projecting to different targets in the limbic circuit can be distinguished on the basis of their content of excitatory amino acids. Cell counts revealed that at least 85-95% of the neurons in the basolateral nucleus projecting to the prefrontal cortex or ventral striatum were pyramidal cells that exhibited glutamate or aspartate immunoreactivity. Colocalization studies indicated that 94-100% of aspartate-immunoreactive neurons in the basolateral nucleus were also glutamate positive and that 92-94% of glutamate-immunoreactive neurons were also aspartate positive. A small number of glutamate-positive pyramidal neurons in the anterior subdivision of the cortical nucleus were found to project to the mediodorsal thalamic nucleus. However, the great majority of amygdaloid neurons with projections to the mediodorsal nucleus did not exhibit glutamate or aspartate immunoreactivity. The absence of glutamate and aspartate immunoreactivity in these cells suggests that these neurons do not use excitatory amino acids as neurotransmitters. The finding of high levels of glutamate and aspartate in basolateral amygdaloid neurons projecting to the prefrontal cortex and ventral striatum is consistent with previous reports indicating that these neurons may use excitatory amino acids as neurotransmitters, but is not a definitive criterion for this determination.

Amygdala↗

PubMed1996

Colocalization of ionotropic glutamate receptor subunits with NADPH-diaphorase-containing neurons in the rat mesopontine tegmentum.

Tegmental cholinergic neurons vary their discharge patterns across the sleep-wake cycle, and glutamate is suggested to play an important role in determining these firing patterns. Cholinergic and noncholinergic neurons in the mesopontine tegmentum have different susceptibilities to various excitotoxins, presumably because of heterogeneity in the expression of glutamate receptor subtypes in this area. By using a double-labeling procedure that combines nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-diaphorase) histochemistry and avidin-biotin-peroxidase immunocytochemistry with diaminobenzidine as the chromogen, we compared the colocalization of AMPA receptor subunits GluR1, GluR2/3, and GluR4, kainate receptor subunits GluR5/6/7, and an NMDA receptor subunit NMDAR1 on NADPH-diaphorase-positive (cholinergic) neurons in the mesopontine tegmentum. Throughout the brainstem, neurons immunoreactive for GluR2/3 and NMDAR1 were most numerous, whereas neurons labeled for GluR1, GluR4, and GluR5/6/7 were less common. Specifically within the mesopontine tegmentum, the proportion of double-labeled neurons in the diaphorase-containing cell population was highest with GluR1 (43%) and lowest with GluR5/6/7 (12%). Regardless of the receptor subunit type, the greatest numbers of double-labeled neurons were observed in the pedunculopontine tegmental nucleus pars compacta and the fewest in the dorsal aspect of the laterodorsal tegmental nucleus. In addition, there were regional differences in the relative expression of receptor subunits and diaphorase-positive neurons across the subdivisions of the tegmental cholinergic column. Because each ionotropic subunit confers distinctive properties to a receptor channel, the present results suggest that mesopontine cholinergic neurons have nonuniform responses to glutamate and are also discriminable from basal forebrain cholinergic neurons in terms of glutamate receptor configuration.

Animals↗

PubMed1999

Colocalization of androgen receptors and mating-induced FOS immunoreactivity in neurons that project to the central tegmental field in male rats.

Bilateral lesions of the central tegmental field (CTF) in male rats virtually eliminate mating behavior. This study examined if mating-induced Fos expression (a measure of neuronal activation) and androgen receptors (AR) are colocalized in brain and spinal cord neurons which project to the CTF. Animals received unilateral injections of the retrograde tracer Fluorogold (FG) in the lateral part of the CTF (CTFl), and 10 days later were killed after ejaculating with females. Brains and spinal cords were examined for FG transport, AR-immunoreactivity (AR-ir), and Fos-immunoreactivity (Fos-ir). AR-ir and Fos-ir were visualized with fluorescence microscopy using cyanine-conjugated and fluorescein-conjugated secondary antibodies. The CTFl received projections from AR-containing neurons in forebrain structures (bed nucleus of stria terminalis, medial preoptic area, lateral and ventromedial hypothalamus), in the central amygdala and various mid- and hindbrain structures (dorsolateral tegmentum, superior and inferior colliculi, pedunculopontine nucleus), and in the lumbosacral spinal cord (lamina X). Some of the AR-containing neurons in bed nucleus of stria terminalis and in the dorsal part of the medial preoptic area with projections to the CTFl were activated by mating. Most AR-containing neurons in spinal lamina X with projections to the CTFl were also activated by mating. Information from spinal cord and pontine nuclei and from outputs descending from the forebrain may be relayed in the CTFl. Thus, as part of a network of hormone-sensitive neurons linking brain and spinal cord mechanisms for mating, the CTFl could participate in the integration of visceral and somatic information relevant for sexual behavior.

Animals↗

PubMed2000

Distribution of GAD-immunoreactive neurons in the diencephalon of the african lungfish Protopterus annectens: colocalization of GAD and NPY in the preoptic area.

The distribution of GABAergic neurons was investigated in the diencephalon of the African lungfish, Protopterus annectens, by using specific antibodies directed against glutamic acid decarboxylase (GAD). A dense population of immunoreactive perikarya was observed in the periventricular preoptic nucleus, whereas the caudal hypothalamus and the dorsal thalamus contained only scattered positive cell bodies. Clusters of GAD-positive cells were found in the intermediate lobe of the pituitary. The diencephalon was richly innervated by GAD-immunoreactive fibers that were particularly abundant in the hypothalamus. In the periventricular nucleus, GAD-positive fibers exhibited a radial orientation, and a few neurons extended processes toward the third ventricle. More caudally, a dense bundle of GAD-immunoreactive fibers coursing along the ventral wall of the hypothalamus terminated into the median eminence and the neural lobe of the pituitary. Double-labeling immunocytochemistry revealed that GAD and neuropeptide tyrosine (NPY)-like immunoreactivity was colocalized in a subpopulation of perikarya in the periventricular preoptic nucleus. The proportion of neurons that coexpressed GAD and NPY was higher in the caudal region of the preoptic nucleus. The distribution of GAD-immunoreactive elements in the diencephalon and pituitary of the African lungfish indicates that GABA may act as a hypophysiotropic neurohormone in Dipnoans. The coexistence of GAD and NPY in a subset of neurons of the periventricular preoptic nucleus suggests that GABA and NPY may interact at the synaptic level.

Animals↗

PubMed1996

Novel mouse IgG-like immunoreactivity expressed by neurons in the moth Manduca sexta: developmental regulation and colocalization with crustacean cardioactive peptide.

Immunoglobulin-related molecules have been shown to play important roles in cell-cell recognition events during the development of both vertebrate and invertebrate nervous systems. In the moth, Manduca sexta, we report the presence of novel, mouse, immunoglobulin G (mIgG)-like immunoreactivity in a discrete population of identified neurosecretory neurons (the NS-Ls also known as the cell 27s) and interneurons (the IN-704s). A number of polyclonal anti-mIgG antibodies were used to immunostain these cells in wholemount. The mIgG-like-immunoreactive (IR) neurons were present during embryogenesis through the developing adult stages, but disappeared in the postemerged adult. Biochemical analysis of M. sexta ventral nerve cords revealed that the mIgG-like antigen is a membrane-associated 27-kDa protein which is likely responsible for the mIgG-like immunostaining observed. Unambiguous identification of the mIgG-like-IR neurons was based on neuronal morphology and our ability to demonstrate conclusively that these neurons expressed immunoreactivity to an antiserum against crustacean cardioactive peptide (CCAP). The NS-Ls and IN-704s were both shown to colocalize the CCAP and mIgG-like immunoreactivities. The mIgG-like and CCAP-IR neurons were identical to a subset of CCAP-IR neurons recently described by Davis et al. [(1993) J. Comp. Neurol., 338:612-627] in pupae. We found these CCAP-IR neurons, however, also to be present in larvae. The mIgG-like- and CCAP-IR neurons included the NS-L pair of the subesophageal maxillary neuromere, which projected anteriorly to the corpora cardiaca, and the NS-L of the labial neuromere whose axons projected out the dorsal nerve of the next posterior ganglion. The mIgG-like and CCAP-IR NS-Ls were also observed throughout the three thoracic ganglia, and all shared strikingly similar structural features. These cells exited out the dorsal nerve of the next posterior ganglion and eventually projected to the neurohemal release sites of the perivisceral organs. These neurons appear to be the homologues of the abdominal CCAP-IR NS-Ls, neurons that in the adult switch their neurotransmitter and release the neuropeptide bursicon. Our description of the distribution and developmental expression of this novel mIgG-like immunoreactivity may provide new insights into the regulation of neurotransmitter plasticity and/or recognition-signaling events involved in the embryonic and postembryonic assembly of the nervous system.

Animals↗

PubMed1996

Immunocytochemical colocalization of desmin and vimentin in human fetal skeletal muscle cells.

BACKGROUND: Desmin and vimentin are the major intermediate filaments in muscle. They have been extensively studied in animal experiments. This study is the first to identify the distribution and to analyse the correlation of desmin and vimentin in human fetal skeletal muscle. Vimentin might be replaced by or transformed into desmin during myogenesis in chick embryo, although the precise process remains to be elucidated. The aim of this report is to evaluate the ratio of desmin to vimentin in human fetal muscle. METHODS: By double-labeling immunoelectron microscopy, desmin and vimentin intermediate filaments were localized in developing skeletal muscles of 20-29-week-old human fetuses. The ratio of desmin and vimentin was analyzed statistically. RESULTS: Two sizes of colloidal gold particles, 5 nm (vimentin) and 10 nm (desmin), were distributed along the intermediate filaments. The commonest distance between gold particles was approximately 40-50 nm. Desmin and vimentin labeled with gold particles were arranged very close together in the same intermediate filament. The ratio of vimentin to desmin varied but the amount of vimentin decreased progressively from the undifferentiated myoblast to the differentiated myocytes. As the fetuses developed, desmin increased and vimentin decreased. Desmin and vimentin intermediate filaments were identified in the intermyofibrils of differentiated myocytes, in subsarcolemmal space, and in myoblast. CONCLUSIONS: Desmin and vimentin were colocalized in the same intermediate filaments. More vimentin existed in the less differentiated myocytes, although a small number of desmin filaments were already found in undifferentiated myoblasts. These intermediate filaments may not only connect myofibril bundles, cell organelles, and cell membrane but also provide a basis for myofibrillogenesis that is similar to relation between connective fibers and parenchymal cells.

Desmin↗

PubMed1999

Colocalization of beta-adrenergic receptors and caveolin within the plasma membrane.

The rapid amplification of beta-adrenergic receptor signaling involves the sequential activation of multiple signaling molecules ranging from the receptor to adenylyl cyclase. The prevailing view of the agonist-induced interaction between signaling molecules is based on random collisions between proteins that diffuse freely in the plasma membrane. The recent identification of G protein alpha- and betagamma-subunits in caveolae and their functional interaction with caveolin suggests that caveolae may participate in G protein-coupled signaling. We have investigated the potential interaction of beta-adrenergic receptors with caveolin under resting conditions. beta1- and beta2-adrenergic receptors were recombinantly overexpressed in COS-7 cells. Caveolae were isolated using the detergent-free sucrose gradient centrifugation method. beta1- and beta2-adrenergic receptors were localized in the same gradient fractions as caveolin, where Gsalpha- and betagamma-subunits were detected as well. Immunofluorescence microscopy demonstrated the colocalization of beta-adrenergic receptors with caveolin, indicating a nonrandom distribution of beta-adrenergic receptors in the plasma membrane. Using polyhistidine-tagged recombinant proteins, beta-adrenergic receptors were copurified with caveolin, suggesting that they were physically bound. Our results suggest that, in addition to clathrin-coated pits, caveolae may act as another plasma membrane microdomain to compartmentalize beta-adrenergic receptors.

Animals↗

PubMed1998

Corticotropin-releasing hormone (CRH)-containing neurons in the immature rat hippocampal formation: light and electron microscopic features and colocalization with glutamate decarboxylase and parvalbumin.

Corticotropin-releasing hormone (CRH) excites hippocampal neurons and induces death of selected CA3 pyramidal cells in immature rats. These actions of CRH require activation of specific receptors that are abundant in CA3 during early postnatal development. Given the dramatic effects of CRH on hippocampal neurons and the absence of CRH-containing afferents to this region, we hypothesized that a significant population of CRHergic neurons exists in developing rat hippocampus. This study defined and characterized hippocampal CRH-containing cells by using immunocytochemistry, ultrastructural examination, and colocalization with gamma-aminobutyric acid (GABA)-synthesizing enzyme and calcium-binding proteins. Numerous, large CRH-immunoreactive (ir) neurons were demonstrated in CA3 strata pyramidale and oriens, fewer were observed in the corresponding layers of CA1, and smaller CRH-ir cells were found in stratum lacunosum-moleculare of Ammon's horn. In the dentate gyrus, CRH-ir somata resided in the granule cell layer and hilus. Ultrastructurally, CRH-ir neurons had aspiny dendrites and were postsynaptic to both asymmetric and symmetric synapses. CRH-ir axon terminals formed axosomatic and axodendritic symmetric synapses with pyramidal and granule cells. Other CRH-ir terminals synapsed on axon initial segments of principal neurons. Most CRH-ir neurons were coimmunolabeled for glutamate decarboxylase (GAD)-65 and GAD-67 and the majority also contained parvalbumin, but none were labeled for calbindin. These results confirm the identity of hippocampal CRH-ir cells as GABAergic interneurons. Further, a subpopulation of neurons immunoreactive for both CRH and parvalbumin and located within and adjacent to the principal cell layers consists of basket and chandelier cells. Thus, axon terminals of CRH-ir interneurons are strategically positioned to influence the excitability of the principal hippocampal neurons via release of both CRH and GABA.

Animals↗

PubMed2000

Allatotropin-like neuropeptide in the cockroach abdominal nervous system: myotropic actions, sexually dimorphic distribution and colocalization with serotonin.

Allatotropin (AT) was isolated from the moth Manduca sexta as a peptide stimulating biosynthesis of juvenile hormone in the corpora allata, but has also been shown to be cardioactive in the same species. Here, we have investigated the presence and biological activity of AT-like peptide in the cockroaches Leucophaea maderae and Periplaneta americana with focus on abdominal ganglia and their target tissues. An antiserum to M. sexta AT was used for immunocytochemical mapping of neurons in the abdominal ganglia. A small number of interneurons and efferent neurons were found AT-like immunoreactive (AT-LI) in each of the abdominal ganglia. A prominent sexual dimorphism was detected in the terminal abdominal ganglion: in L. maderae the male ganglion there are approximately 18 AT-LI neurons with cell bodies posteriorly and efferent axons in the genital nerves; in the female ganglion 4-5 AT-LI cell bodies (with efferent axons) were found in the same region. Correlated with the extra efferents in males, the male accessory glands are richly supplied by AT-LI fibers and in females a less prominent innervation was seen in oviduct muscle. A similar dimorphism was seen in abdominal ganglia of P. americana. A sexual dimorphism was also detected in the abdominal ganglia A4-A6 of L. maderae. In each of these ganglia, approximately 8-10 large AT-LI neuronal cell bodies were found along the midline; in females these neurons have significantly larger cell bodies than in males. In both sexes, and both cockroach species, two large dorsal midline neurons were detected in A-5 and 6, which seem to send axons to the hindgut: the rectal pads of the hindgut are supplied by arborizing AT-LI axons. In males and females of both species, efferent AT-LI axons from midline neurons in A3-A6 supply the lateral heart nerves and other neurohemal release sites with arborizations. The efferent midline neurons of females contain colocalized serotonin-immunoreactivity. We tested the in vitro actions of M. sexta AT on muscle contractions in the L. maderae hindgut and the abdominal heart of both species. The frequency of contractions in the hindgut increased dose dependently when applying AT at 5 x 10(-8) to 5 x 10(-6) M (maximal response at 5 x 10(-7) M). Also the frequency of contractions of the heart increased by application of AT (threshold response at 5 x 10(-9) M). This effect was more prominent in males of both species (maximal response was a 35-40% increase in males and 10-20% in females). In conclusion, an AT-like peptide is present in neurons and neurosecretory cells of cockroach abdominal ganglia and seems to play a role in control of contractions in the hindgut and heart and also to have some function in male accessory glands and oviduct.

Animals↗