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At least 451 records · Page 25Linked to original sources

PubMed2002

Optimal colocalization of TCR and CD8 as a novel mechanism for the control of functional avidity.

The improved efficacy of high avidity CTL for clearance of virus has been well-documented. Thus, elucidation of the mechanisms that confer the increased sensitivity to peptide ligand demonstrated by high avidity CTL is critical. Using CTL lines of high and low avidity generated from a TCR transgenic mouse, we have found that functional avidity can be controlled by the expression of CD8alphaalpha vs CD8alphabeta and the ability of CTLs to colocalize the TCR and CD8 in the membrane. Colocalization of these molecules was mediated by lipid rafts and importantly, raft disruption resulted in the conversion of high avidity CTL into a lower functional avidity phenotype. These novel findings provide insights into the control of functional avidity in response to viral infection.

Animals↗

PubMed2004

Enrichment of lck in lipid rafts regulates colocalized fyn activation and the initiation of proximal signals through TCR alpha beta.

Recent results provide insight into the temporal and spatial relationship governing lck-dependent fyn activation and demonstrate TCR/CD4-induced activation and translocation of lck into lipid rafts and the ensuing activation of colocalized fyn. The prediction follows that directly targeting lck to lipid rafts will bypass the requirement for juxtaposing TCR and CD4-lck, and rescue cellular activation mediated by Ab specific for the constant region of TCRbeta chain. The present study uses a family of murine IL-2-dependent CD4(+) T cell clonal variants in which anti-TCRCbeta signaling is impaired in an lck-dependent fashion. Importantly, these variants respond to Ag- and mAb-mediated TCR-CD4 coaggregation, both of which enable the coordinated interaction of CD4-associated lck with the TCR/CD3 complex. We have previously demonstrated that anti-TCRCbeta responsiveness in this system correlates with the presence of kinase-active, membrane-associated lck and preformed hypophosphorylated TCRzeta:zeta-associated protein of 70 kDa complexes, a phenotype recapitulated in primary resting CD4(+) T cells. We show in this study that forced expression of wild-type lck achieved the same basal composition of the TCR/CD3 complex and yet did not rescue anti-TCRCbeta signaling. In contrast, forced expression of C20S/C23S-mutated lck (double-cysteine lck), unable to bind CD4, rescues anti-TCRCbeta proximal signaling and cellular growth. Double-cysteine lck targets lipid rafts, colocalizes with >98% of cellular fyn, and results in a 7-fold increase in basal fyn kinase activity. Coaggregation of CD4 and TCR achieves the same outcome. These results underscore the critical role of lipid rafts in spatially coordinating the interaction between lck and fyn that predicates proximal TCR/CD3 signaling.

Animals↗

PubMed2006

The beneficial relationship of the colocation of orthopedics and physical therapy in a deployed setting: Operation Iraqi Freedom.

In a deployment setting, orthopedic and musculoskeletal injuries represent the majority of both combat-related and noncombat-related injuries. To expeditiously and efficiently manage the large influx of patients, our experience revealed the tremendous benefits of having physical therapy colocated with an orthopedic surgeon in a level III combat support hospital. A physical therapist, working in a physician extender role, can treat the majority of nonsurgical orthopedic patients, thus allowing the orthopedic surgeon to focus his or her skills and time on surgical interventions. This physician extender role, although often overlooked during peacetime, becomes essential during wartime, when critically injured patients are abundant and physicians are in short supply. The lessons learned support the continued colocation of physical therapy and orthopedics in a deployment setting and recognize the need to have more physical therapists placed at level I and II echelons of care.

Geography↗

PubMed2002

The multiple LIM domain-containing adaptor protein Hic-5 synaptically colocalizes and interacts with the dopamine transporter.

The Na+/Cl--dependent dopamine transporter (DAT) is critical in terminating dopaminergic transmission by removing the transmitter away from the synapse. Several lines of evidence suggest that transporter-interacting proteins may play a role in DAT function and regulation. In this report, using the yeast two-hybrid system, we have identified a novel interaction between DAT and the multiple Lin-11, Isl-1, and Mec-3 (LIM) domain-containing adaptor protein Hic-5. This association involves the N-terminal portion of the intracellular tail of DAT and the LIM region of Hic-5. In human embryonic kidney 293 cells, Hic-5 colocalizes with DAT at polarized sites and reduces DAT uptake activity through a mechanism involving a decrease in the cell-surface levels of the transporter. A fragment of Hic-5 containing the LIM domains is sufficient to bind DAT but lacks the ability to inhibit transporter activity. In addition, the LIM fragment prevents the effect of the full-length Hic-5 on DAT localization and function. In the brain, Hic-5 protein is expressed in the cerebral cortex, hippocampus, hypothalamus, cerebellum, and striatum, suggesting a role for this protein in the nervous system. The association of the endogenous Hic-5 and DAT proteins was confirmed biochemically by coimmunoprecipitation from brain striatal extracts. Moreover, immunostaining of rat midbrain neurons in culture revealed a presynaptic colocalization of Hic-5 and DAT. Because Hic-5 has been shown to interact with several signaling molecules, including the nonreceptor protein tyrosine kinases focal adhesion kinase and Fyn, this raises the possibility that this adaptor protein may link DAT to intracellular signaling pathways.

Animals↗

PubMed1999

N(epsilon)(carboxymethyl)lysin and the AGE receptor RAGE colocalize in age-related macular degeneration.

PURPOSE: To investigate whether glycoxidation products and the receptor for advanced glycation end products (RAGE) are present and colocalize in subfoveal membranes of patients with age-related macular degeneration (ARMD). METHODS: Surgically removed subfoveal fibrovascular membranes from 12 patients, 11 related to ARMD and 1 to an idiopathic membrane, were analyzed for the presence of the glycoxidation product N(epsilon)-(carboxymethyl)lysin (CML), one of the receptors for advanced glycation end products, RAGE, and the activation of NFkB, using immunohistochemistry. RESULTS: CML-like immunoreactivity was found in all ARMD specimens examined adjacent or colocalized with RAGE, but not in the idiopathic membrane. RAGE immunoreactive material was found in CD68-positive cells and in the fibrous matrix. CD68-positive cells and surrounding areas stained for p50, the activated form of NFkB. CONCLUSIONS: These results indicate that glycoxidation products are present in subretinal membranes of patients with ARMD. The concomitant expression of RAGE in these membranes and the finding of activated NFkB is suggestive of an implication of glycoxidation product formation in the pathogenesis of the disease.

Adolescent↗

PubMed1999

Expression of NR2 receptor subunit in rat somatic sensory cortex: synaptic distribution and colocalization with NR1 and PSD-95.

Functional N-methyl-D-aspartate (NMDA) receptors comprise heteromeric combinations of NR1 and NR2 subunits. In the present study, we employed light and electron microscopic immunocytochemistry to study the expression of NR2A and NR2B (NR2A/B) protein in somatic sensory cortex of adult rats. To relate this distribution to that of NR1 and to the NMDA receptor anchoring protein PSD-95, we documented extensive cellular colocalization of NR2A/B with NR1 at the light microscopic level. In contrast, PSD-95 exhibited little somatic staining, being restricted mainly to dendrites and neuropil. We employed postembedding immunocytochemistry to study the ultrastructural expression of NR2A/B. Labeling in neuronal perikarya was associated with rough endoplasmic reticulum and Golgi apparatus; in dendrites, gold particles labeled microtubules. The preponderance of labeling was associated with asymmetric synapses. Double immunolabeling revealed that NR2 colocalized in many synapses with NR1 and with PSD-95. Quantitative measurements revealed that density of gold particles coding for both NR2 and PSD-95 was highest just inside the postsynaptic membrane. Tangentially along the membrane, gold particles were concentrated at the synaptic specialization. These data provide structural evidence in neocortex for heteromeric NMDA receptors anchored at the postsynaptic membrane.

Animals↗

PubMed2001

Substance P receptor expression by inhibitory interneurons of the rat hippocampus: enhanced detection using improved immunocytochemical methods for the preservation and colocalization of GABA and other neuronal markers.

Two unresolved issues regarding the identification and characterization of hippocampal interneurons were addressed in this study. One issue was the longstanding inability to detect gamma-aminobutyric acid (GABA) in the somata of several hippocampal interneuron subpopulations, which has prevented the unequivocal identification of all hippocampal interneurons as GABA neurons. The second issue was related to the identification of the hippocampal interneurons that constitutively express substance P (neurokinin-1) receptors (SPRs). The recent development of neurotoxins that specifically target SPR-expressing cells suggests that it may be possible to destroy hippocampal inhibitory interneurons selectively for experimental purposes. Although SPRs are apparently expressed in the hippocampus only by interneurons, colocalization studies have found that most interneurons of several subtypes and hippocampal subregions appear SPR-negative. Thus, the identities and locations of the inhibitory interneurons that are potential targets of an SPR-directed neurotoxin remain in doubt. Using newly developed methods designed to copreserve and colocalize GABA and polypeptide immunoreactivities with increased sensitivity, the authors report that virtually all hippocampal interneuron somata that are immunoreactive for parvalbumin (PV), calbindin, calretinin, somatostatin (SS), neuropeptide Y, cholecystokinin, and vasoactive intestinal peptide exhibited clearly detectable, somal, GABA-like immunoreactivity (LI). Hippocampal SPR-LI was detected only on the somata and dendrites of GABA-immunopositive interneurons. All glutamate receptor subunit 2-immunoreactive principal cells, including dentate granule cells, hilar mossy cells, and hippocampal pyramidal cells, were devoid of detectable SPR-LI, even after prolonged electrical stimulation of the perforant pathway that induced the expression of other neuronal proteins in principal cells. Thus, hippocampal interneurons of all subtypes and subregions were found to be SPR-immunoreactive, including the PV-positive interneurons of the dentate hilus and hippocampus, and the SS-positive cells of area CA1, both of which were previously reported to lack SPR-LI. Only minor proportions of hippocampal interneurons appeared clearly devoid of detectable SPR-LI. These results demonstrate for the first time that all identified interneuron subpopulations of the rat hippocampus are GABA-immunoreactive, and that many inhibitory interneurons of all subtypes in all subregions of the rat hippocampus express SPRs constitutively.

Animals↗

PubMed2001

Colocalization of neuropilin-1 and Flk-1 in retinal neovascularization in a mouse model of retinopathy.

PURPOSE: To investigate the mechanisms of the development of retinal neovascularization, the localizations of vascular endothelial (VEGF) receptors Flk-1 and neuropilin (NP)-1 mRNAs were examined. METHODS: The model of retinopathy of prematurity (ROP) was produced by ischemia-induced ocular neovascularization, by exposing postnatal day-7 mice to 75% oxygen for 5 days and then returning them to room air for 5 days. Retinal neovascularization was visualized by injection of fluorescein-dextran. Expression of Flk-1 and NP-1 mRNAs were examined by in situ hybridization with flatmount and serial sections of the retina. The localization of NP-1 was also confirmed by immunohistochemistry. Blood vessel patterns were characterized by immunohistochemical localization of von Willebrand factor (vWF). RESULTS: Flatmount in situ hybridization showed intense expression of NP-1 and Flk-1 mRNAs colocalized in the area of neovascularization. In situ hybridization of serial sections of the retina revealed that expression of Flk-1 and NP-1 was restricted to neovascularized vessels of the retina from ROP mice. CONCLUSIONS: The restricted expression of Flk-1 and NP-1 on neovascularized vessels suggests that these molecules may play important roles in retinal neovascularization. This is the first report of the colocalization of NP-1 and Flk-1 on neovascularized vessels of the retina from ROP mice.

Animals↗

PubMed1992

Subcellular distribution of the Rap1A protein in human neutrophils: colocalization and cotranslocation with cytochrome b559.

Rap1A, a low molecular weight guanosine triphosphate-binding protein (LMWG), has been shown previously by us to be associated with purified cytochrome b from stimulated human neutrophils. In the present studies, we show that Rap1A is also associated with affinity-purified cytochrome b from unstimulated neutrophils and use specific anti-Rap1 peptide antibodies to biochemically and immunocytochemically determine the subcellular distribution of Rap1A in resting and activated human neutrophils. Analysis of the subcellular fractionation of unstimulated cells by Western blotting of isopycnic sucrose density gradient fractions with anti-Rap1 peptide antibodies indicated that Rap1A colocalized with cytochrome b in the plasma membrane as well as in the specific granule membranes and that it was translocated, along with cytochrome b, to the plasma membrane when the cells were stimulated with phorbol myristate acetate (PMA). No evidence for a cytosolic localization of Rap1A was found in our studies; however, if the cells were disrupted by sonication, rather than N2 cavitation, a fraction of the Rap1A was released from the membrane. Electron microscopy of thin sections of cryofixed, molecular-distillation dried neutrophils labeled with anti-Rap1 antibody alone or double-labeled with anti-Rap1 and anti-cytochrome b peptide antibodies confirmed our biochemical localization, and quantitation showed that more than half of the specific granule-associated Rap1A was translocated to the plasma membrane in PMA-stimulated cells. Ultrastructural analysis of neutrophils phagocytosing Staphylococcus aureus also demonstrated the translocation of Rap1A with cytochrome b. Approximately 70% of the total Rap1A labeling was associated with the phagolysosomal membrane, the site of assembly of the superoxide-generating system. The colocalization and cotranslocation of Rap1A with cytochrome b in resting and activated neutrophils is consistent with a functional association of these two molecules in the intact cell and provides further evidence for a role of this LMWG in the structure or function of the neutrophil superoxide-generating system.

Biological Transport↗

PubMed1992

Colocalization of a large heterodimeric proteoglycan with basement membrane proteins in cultured cells.

A novel large heterodimeric dermatan sulfate proteoglycan with core proteins of 460 and 300 kDa, respectively, had been described as a secretory product of human fetal skin fibroblasts (Breuer et al., J. Biol. Chem. 266, 13224-13232 (1991)). Pulse-chase experiments showed a preferential association of the proteoglycan with the cell membrane. Immunogold labeling indicated its localization in fibrils on the cell surface as well as in fibrillar extensions from the cell body. Immunofluorescence studies yielded a fibrillar and punctate staining pattern which was also seen in cultured human and porcine endothelial cells. Dot-like structures were observed in transformed human keratinocytes. Various immunocytochemical double-labeling experiments indicated a remarkable colocalization of the proteoglycan with fibronectin, laminin, perlecan, and type IV collagen whereas only occasionally a colocalization with chondroitin-6-sulfate was found. No evidence for an enrichment of the proteoglycan in vinculin-containing structures was obtained. These results suggest that the proteoglycan is a widely distributed macromolecule which can associate with basement membrane components. Preliminary findings in rat cornea supported this conclusion.

Basement Membrane↗

PubMed1991

Substance P- and enkephalin-like immunoreactivities are colocalized in certain neurons of the substantia gelatinosa of the rat spinal cord: an ultrastructural double-labeling study.

The finding that certain cells of the substantia gelatinosa of the rat spinal cord contain both substance P (SP)- and enkephalin (ENK)-like immunoreactive material offers new insights into the mechanisms of action of these peptides in the processing of nociceptive sensory information. The simultaneous detection of these immunoreactivities was obtained in the superficial dorsal horn of the rat spinal cord at the ultrastructural level using monoclonal antibodies. An internally radiolabeled monoclonal antibody (against SP or ENK) was used to recognize one antigenic site, while the other antigenic site was identified by either a bispecific monoclonal antibody (for SP) or a monoclonal antibody (for ENK). The bispecific anti-SP antibody recognized HRP, whereas a secondary bispecific antibody recognized both the IgG of the anti-ENK monoclonal antibody and HRP. In laminae I-III, SP-like immunoreactivity (SP-LI) and ENK-like immunoreactivity (ENK-LI) were colocalized in a significant number of axonal varicosities, which contained round or pleomorphic synaptic vesicles. Such double-labeled varicosities, however, were not found to be components of synaptic glomeruli. Most of the immunostained boutons of lamina I were SP-like immunoreactive only. In rats pretreated with colchicine, SP-LI and ENK-LI were colocalized in small perikarya of lamina II and in some lamina I cells. These findings indicate that SP and ENK occur in a significant population of interneurons of the superficial dorsal horn. It is suggested that some of these neurons may correspond to stalked cells and release one or the other substance depending on physiological conditions.

Animals↗

PubMed1991

Immunohistochemical colocalization of amyloid precursor protein with cerebrovascular amyloid of Alzheimer's disease.

Molecular cloning and cDNA sequencing have indicated that the fibril-forming, amyloidogenic beta/A4 peptide of cerebrovasculature and plaque core in AD is encoded as part of a larger precursor, amyloid precursor protein (APP). A panel of antibodies directed against synthetic peptides, which correspond to distinct domains of this putative APP molecule (i.e., amino acid residues 45-62, 587-596, 597-606, 597-638 [beta/A4 peptide], 638-658 and 653-661), were used to probe immunohistochemically serial sections of formalin-fixed, paraffin-embedded Alzheimer's disease (AD) brains for the presence of APP and/or its derivatives. Histochemical staining of adjacent sections with Bielschowsky's silver impregnation and with Congo red or thioflavin S-staining techniques was also done to identify the structures with amyloid deposition. All these antibodies exhibited intense immunoreactivity with amyloidotic cerebral vessels, including meningeal and parenchymal. This observation indicates that the amyloidotic vasculature of AD brain contains, in addition to the fibril-forming beta/A4 protein, nonamyloidogenic APP and/or its derivatives. More importantly, this APP immunoreactivity colocalized with angiopathic amyloid, which is characterized by phenol-resistant, birefringent congophilia. Parallel analyses with a dual SABC/silver impregnation procedure further confirmed that APP and/or its derivatives, including the amyloidogenic beta/A4, colocalized with argentophilic amyloid in the cerebrovasculature of AD.

Alzheimer Disease↗

PubMed1991

Interaction of colocalized neuropeptides: functional significance in the circadian timing system.

The suprachiasmatic nucleus (SCN), which appears to act as a circadian clock, contains a subpopulation of local circuit neurons in which vasoactive intestinal peptide (VIP), peptide histidine isoleucine (PHI), and gastrin releasing peptide (GRP) are colocalized. To determine whether VIP, PHI, and GRP interact within the SCN to produce a signal important for circadian control, the behavioral and cellular effects of coadministration of these neuropeptides were investigated. Coadministration of VIP, PHI, and GRP within the SCN mimicked the phase-delaying effects of light on circadian control following in vivo microinjection and activated SCN single units recorded in vitro. These behavioral and cellular effects of coadministration of VIP, PHI, and GRP were significantly greater than administration of VIP, PHI, or GRP alone or coadministration of any 2 of these peptides. These data illustrate a new mechanism whereby multiple, colocalized neuropeptides interact in a functionally significant manner, and indicate that the interaction of VIP, PHI, and GRP may be involved in the regulation of circadian rhythms by the SCN.

Animals↗

PubMed1991

Identification of tubulin-binding proteins of the chicken erythrocyte plasma membrane skeleton which colocalize with the microtubular marginal band.

The plasma membrane of nucleated erythrocytes contains a microtubular marginal band which appears to be associated with the plasma membrane skeleton. In this report, we identify two families of cytoskeletal proteins which may be involved in such an association. These proteins, of molecular mass 78 kDa and 48 kDa on SDS-PAGE, are shown to bind tubulin based on a 125I-labeled tubulin binding assay. Solubilization of isolated chicken erythrocyte plasma membranes in Triton X-100 shows that these proteins centrifuge with the pellet, indicating that they are bound to the membrane skeleton. Finally, immunofluorescence studies using antisera raised against the 78 kDa and 48 kDa proteins show that they colocalize with the marginal band in intact cells. Colocalization of cytoskeletal tubulin-binding proteins with the marginal band favors a hypothesis suggesting that the 78 kDa and 48 kDa proteins are involved in the association of the two molecular superstructures.

Animals↗

PubMed1990

Immunoreactive A4 and gamma-trace peptide colocalization in amyloidotic arteriolar lesions in brains of patients with Alzheimer's disease.

Cerebral amyloid angiopathy (CAA) defines a biochemically heterogeneous entity that manifests as effacement of cerebral microvessel walls by a fibrillar material with characteristic tinctorial properties. In biochemical terms, the amyloid that infiltrates blood vessels in CAA is composed of the A4 or beta peptide of Alzheimer's disease (AD), a molecule related to gamma trace or cystatin C (seen in patients with hereditary cerebral hemorrhage with amyloidosis in Iceland, HCHWA-I), or the PrP characteristic of spongiform encephalopathy and scrapie. Using antibodies to synthetic peptides representing portions of the 4.2-kd Alzheimer A4 peptide and the gamma-trace peptide, we immunostained sections of brain from patients with AD, senile dementia of Alzheimer's type, and CAA with associated leukoencephalopathy. Immunohistochemical studies demonstrated colocalization of the A4 and gamma-trace peptides within arteriolar walls, but only rarely in A4 amyloidotic capillaries or senile plaque cores of amyloid. When gamma-tracelike reactivity was noted in capillary walls, it was sometimes noted within the cytoplasm of pericytes. Immunostaining was always more intense when the anti-A4 antibody was used as the primary antibody. Gamma-trace immunostaining was more prominent on the adventitial component of arteriolar walls, whereas A4 staining was usually seen more diffusely throughout the blood vessel wall, especially in the media. Rarely individual pericytelike cells showed prominent gamma-trace immunoreactivity. These findings suggest that A4 and gamma-tracelike molecules may colocalize within arteriolar walls within the brains of patients with AD, and highlight the fact that CAA identified with AD and HCHWA-I are not as biochemically distinct as was assumed previously. Furthermore these findings suggest that other peptidases or protease inhibitors may be found within amyloidotic microvessel walls and may contribute to senile brain change and CAA-related strokes, including hemorrhage and encephalomalacia.

Aged↗

PubMed1989

Dopamine and neurotensin storage in colocalized and noncolocalized neuronal populations.

The effects of reserpine on dopamine (DA) and neurotensin (NT) levels were studied in four different brain regions of the rat. Reserpine (0.5-5.0 mg/kg i.p., 6, 18, 48 and 72 hr) produced a dose- and time-dependent decrease in both DA and NT levels in the prefrontal cortex, a brain region innervated by a mixed DA/NT projection. The effect of reserpine was not mimicked by alpha-methylparatyrosine (200 mg/kg i.p.) pretreatment. Furthermore, the reserpine-induced decline in prefrontal cortex DA and NT levels occurred after gamma-butyrolactone (GBL)-induced inhibition of impulse flow (750 mg/kg i.p.). In contrast, in the nucleus accumbens and striatum, regions which contain colocalized (nucleus accumbens) and intrinsic (striatum and nucleus accumbens) neurotensin perikarya, reserpine produced declines in DA and increases in NT levels. alpha-Methylparatyrosine decreased striatal and nucleus accumbens DA levels without altering NT levels in these structures. GBL produced an increase in DA levels in the nucleus accumbens and striatum while decreasing nucleus accumbens and striatal NT levels. Reserpine attenuated the decline in nucleus accumbens and striatal NT levels produced by GBL. In the periaqueductal grey, a brain region densely innervated by NT which has a small population of DA perikarya, reserpine had no effect on NT levels. Because there is no known colocalization of DA and NT in the striatum, the increases in striatal NT levels after depletion of DA may indicate that striatal DA afferents control the release and/or synthesis of NT within NT cells in the striatum, thus leading to alterations in striatal tissue levels.(ABSTRACT TRUNCATED AT 250 WORDS)

4-Butyrolactone↗

PubMed1988

Calmodulin colocalization with cold-stable and nocodazole-stable microtubules in living PtK1 cells.

To investigate the association of calmodulin (CaM) with microtubules (MTs) in the mitotic apparatus (MA), the distributions of both CaM and tubulin were examined in mitotic PtK1 cells in which MT subclasses had been selectively removed or altered by treatment with cold or with the MT inhibitor, nocodazole. A fluorescent CaM conjugate with tetramethylrhodamine isothiocyanate (CaM-TRITC) was microinjected into living cells, and the CaM distribution in the living cell was compared to the distribution of MTs indicated by tubulin immunofluorescence. In cells which had been treated for 2 h at 0 to 4 degrees C or with a low (0.03 micrograms/ml) dose of nocodazole, the only MTs remaining appeared to be kinetochore MTs (kMTs). The distribution of microinjected CaM-TRITC in these cells was indistinguishable from that found in untreated cells and appeared to be colocalized with the kMTs. In cells which were treated with a high (3.0 micrograms/ml) dose of nocodazole, only short MTs remained. When CaM-TRITC was injected into these cells, it formed a somewhat punctate distribution near the chromosomes and, after tubulin immunofluorescence processing, colocalized with what appeared to be remnants of kMTs. We believe that these observations support the hypothesis that CaM exists in the MA in a structural association with kMTs.

Animals↗

PubMed1988

Colocalization of taurine- and cysteine sulfinic acid decarboxylase-like immunoreactivity in the cerebellum of the rat with monoclonal antibodies against taurine.

Two monoclonal antibodies against fixative-modified taurine, Tau1 and Tau2, were produced, characterized, and used in the present study to analyze the distribution of taurine in the cerebellum of the rat. In addition, immunohistochemical colocalization experiments were performed to determine whether cerebellar neurons contain both taurine and its synthesizing enzyme, cysteine sulfinic acid decarboxylase (CSADC). In ELISAs, both Tau1 and Tau2 displayed high affinities for taurine conjugated to various carrier proteins and possessed some cross-reactivity for other amino acids which are present in lower concentrations in the brain than taurine. Tau2 was found to recognize only taurine and hypotaurine when paraformaldehyde was used to fix the amino acids to carrier proteins. With the use of glutaraldehyde fixation, Tau1 cross-reacted with conjugates of beta-alanine and hypotaurine and Tau2 cross-reacted strongly with conjugates of cysteic acid and hypotaurine and weakly with cysteine sulfinic acid. Despite different cross-reactivities, Tau1 and Tau2 exhibited almost identical patterns of neuronal staining in bands of Purkinje cells in the cerebellum. Staining of Purkinje cell dendrites was more prominent than staining of the soma. Light immunoreactivity was present in Golgi, stellate, and basket cells. A scattered population of granule cells displayed taurine-like immunoreactivity at the electron microscopic level. Immunostaining was identified in some terminals in the Purkinje cell layer and in a limited number of mossy fibers. Tau2-like immunoreactivity was colocalized with CSADC-like immunoreactivity in the cerebellar neurons described above. These immunoreactive cells may represent a subpopulation of neurons that contain a higher concentration of taurine than neighboring cells due to their ability to synthesize taurine. The intense immunoreactive staining of Purkinje cell dendrites provides support for the hypothesis that calcium-dependent release of taurine in the cerebellum may originate primarily from dendritic rather than synaptic processes and suggests a neuromodulator role for taurine in the cerebellum.

Animals↗