Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:1.14.16.2 (tyrosine hydroxylase)
14,760 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Radiolabelled meta-iodobenzylguanidine (MIBG) has been widely used in scintigraphy and targeted radiotherapy in patients with neuroblastoma. Recently, it has been demonstrated that MIBG is incorporated into neuroblastoma cells by the noradrenaline transporter. In vitro experiments on SK-N-SH human neuroblastoma cells performed in the present study showed that uptake of MIBG is inhibited by noradrenaline, more so by dopamine and to a lesser extent, by serotonin, indicating that the respective transporters may also contribute to MIBG uptake. However, neither dopamine nor serotonin transporter gene expression was detected. Noradrenaline transporter gene expression was found in 4 of 6 investigated cell lines, which correlated with specific MIBG uptake. Furthermore, an inverse correlation of noradrenaline transporter and tyrosine hydroxylase gene expression, the key regulatory enzyme of catecholamine synthesis, was observed. These data show that MIBG is specifically incorporated only in neuroblastoma cells in which there is noradrenaline transporter gene expression. Furthermore, the catecholamine status in neuroblastoma cells is regulated by a coordinate expression of the key elements of catecholamine synthesis and reuptake systems.
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PMID:Reverse transcriptase-polymerase chain reaction (RT-PCR) analysis of monoamine transporters in neuroblastoma cell lines: correlations to meta-iodobenzylguanidine (MIBG) uptake and tyrosine hydroxylase gene expression. 757 74

A subpopulation of nerve fibers in the rat pituitary intermediate lobe (IL) have been shown to exhibit colocalization of serotonin (5-HT-IR) and tyrosine hydroxylase immunoreactivities and they are sensitive to neurotoxins specific to catecholamine neurons. This study was set out to examine the uptake and synthesis mechanisms of serotonin in these fibers. We developed an in vitro technique in which the neurointermediate lobe explants were incubated (14 and 48 h) in the presence of various drugs and serotonin was subsequently visualized by immunohistochemistry. Control incubation in the presence of serotonin (10(-6) M) resulted in a rich plexus of 5-HT-IR fibers in both posterior and intermediate lobes. Fluoxetine and citalopram (10(-6) M and 10(-5) M), inhibitors of 5-HT transporter, did not affect 5-HT-IR in the IL fibers, unless they were used in concentrations high enough (10(-4) M and 10(-3) M) to block unspecifically a number of monoamine transporters. The same applied for desipramine (10(-5)-10(-7) M), an inhibitor of the noradrenaline transporter. However, cocaine (10(-5)-10(-6) M) blocked serotonin uptake into these terminals, suggesting that serotonin uptake occurs through a dopamine transporter. Incubation of the IL in presence of L-tryptophan (10(-4) M) did not result in 5-HT-IR in the IL fibers showing colocalization of 5-HT-IR and tyrosine hydroxylase, which suggests that these fibers do not synthesize serotonin. The present results suggest that serotonin is taken up into the IL terminals by a dopamine transporter and is not synthesized in them, at least in vitro.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Pharmacological characterization of serotonin synthesis and uptake suggest a false transmitter role for serotonin in the pituitary intermediate lobe. 772 65

The noradrenaline transporter (NAT) is present in noradrenergic neurons and a few other specialized cells such as adrenal medullary chromaffin cells and the rat pheochromocytoma (PC12) cell line. We have raised antibodies to a 49-residue segment (NATM2) of the extracellular region (residues 184-232) of bovine NAT. Affinity-purified NATM2 antibodies specifically recognized an 80-kDa band in PC12 cell membranes by western blotting. Bands of a similar size were also detected in membranes from human neuroblastoma (SK-N-SH) cells expressing endogenous NAT and human embryonic kidney (HEK293) cells stably expressing bovine NAT. Immunocytochemistry of rat adrenal tissue showed that NAT staining was colocalized with tyrosine hydroxylase in medullary chromaffin cells. Most NAT immunoreactivity in rat adrenal chromaffin and PC12 cells was present in the cytoplasm and had a punctate appearance. Cell surface biotinylation experiments in PC12 cells confirmed that only a minor fraction of the NAT was present at the cell surface. Subcellular fractionation of PC12 cells showed that relatively little NAT colocalized with plasma membrane, synaptic-like microvesicles, recycling endosomes, or trans-Golgi vesicles. Most of the NAT was associated with [3H]noradrenaline-containing secretory granules. Following nerve growth factor treatment, NAT was localized to the growing tip of neurites. This distribution was similar to the secretory granule marker secretogranin I. We conclude that the majority of NAT is present intracellularly in secretory granules and suggest that NAT may undergo regulated trafficking in PC12 cells.
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PMID:Localization of the noradrenaline transporter in rat adrenal medulla and PC12 cells: evidence for its association with secretory granules in PC12 cells. 1046 91

The rostral ventrolateral medulla (RVL) contains neurons which are critically involved in the tonic and reflex control of blood pressure. Some of these neurons project to the intermediolateral cell column of the thoracolumbar spinal cord and excite preganglionic sympathetic neurons. In order to gain a better understanding of the properties of the RVL neurons at the cellular and molecular level, a protocol was developed utilizing acute dissociation and the reverse transcription-polymerase chain reaction (RT-PCR) to study the expression of several genes in single RVL neurons. Neurons were dissociated from the RVL region of young rats, and classified as spinally projecting or non-spinal by the presence or absence of retrogradely transported fluorescent beads injected into the upper thoracic segments of the spinal cord. Individual neurons were collected by aspiration into a glass micropipette and analysed by RT-PCR. The presence of either glyceraldehyde 3-phosphate dehydrogenase (GAPDH) or neuron-specific enolase (NSE) mRNA was used as the criterion for selecting cells for further analysis. A subpopulation (50%) of spinally projecting, GAPDH- or NSE-positive neurons expressed mRNA for tyrosine hydroxylase (TH) or phenylethanolamine N-methyltransferase (PNMT), indicative of catecholaminergic or C1 adrenergic neurons, respectively. Some bulbospinal RVL neurons, including those that were TH- or PNMT-positive, were also found to express mRNA for the mineralocorticoid receptor (MR), the glucocorticoid receptor (GR), noradrenaline transporter (NET), and neuronal glutamate transporter (EAAC1). The glial glutamate transporter (GLT), glycine transporter (GLYT2), glutamic acid decarboxylase (GAD67) and gamma-amino butyric acid (GABA) transporter (GAT-1) were not expressed. The single-cell RT-PCR protocol is a powerful, yet simple and relatively rapid method for analysis of mRNA expression in a defined neuronal population. It can be combined with whole-cell patch-clamp recording prior to RT-PCR analysis, allowing linkage of the molecular analysis of mRNA expression to the electrophysiological and pharmacological properties of single neurons. The method is very sensitive, enabling mRNA transcripts in low abundance to be detected, and its application in our recent studies provided novel information about neurons involved in blood-pressure regulation at the molecular and cellular level.
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PMID:Detection of mRNA species in bulbospinal neurons isolated from the rostral ventrolateral medulla using single-cell RT-PCR. 1059 47

Expression of the noradrenaline transporter (NAT) was identified in various cell and fibre populations of the rat adrenal medulla, examined with immunohistochemistry and confocal microscopy. Immunoreactivity for the catecholamine biosynthetic enzymes tyrosine hydroxylase (TH), aromatic-L-amino-acid decarboxylase (AADC) and dopamine beta-hydroxylase (DBH) was present in all chromaffin cells, while phenylethanolamine N-methyltransferase (PNMT) was used to determine adrenergic chromaffin cell groups. Labelling with NAT antibody was predominantly cytoplasmic and colocalised with PNMT immunoreactivity. Noradrenergic chromaffin cells were not NAT immunoreactive. Additionally, NAT antibody labelling demonstrated clusters of ganglion cells (presumably Type I) and nerve fibres. Expression of TH, AADC, DBH, PNMT and NAT mRNA was examined using reverse transcription-polymerase chain reaction (RT-PCR) from adrenal medulla punches and single chromaffin cells, and results were consistent with those obtained with immunocytochemistry. Chromaffin cells and fibres labelled with antibodies against growth associated protein-43 (GAP-43) were not NAT immunoreactive, while ganglion cells were doubled labelled with the two antibodies. The presence of NAT in adrenergic chromaffin cells, and its absence from noradrenergic cells, suggests that the adrenergic cell type is primarily responsible for uptake of catecholamines in the adrenal medulla.
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PMID:Differential expression of the noradrenaline transporter in adrenergic chromaffin cells, ganglion cells and nerve fibres of the rat adrenal medulla. 1117 23

The analysis of colocalization of multiple catecholamine biosynthetic enzymes within the ventrolateral part of the medulla oblongata of the rat revealed distinct subpopulations of neurons within the C1 region (Phillips et al., J Comp Neurol 2001, 432:20-34). In extending this study to include the caudal pons, it was shown for the first time that the A5 cell group could be distinguished by the presence of immunoreactivity to tyrosine hydroxylase (TH), aromatic l-amino acid decarboxylase (AADC), and dopamine beta hydroxylase (DBH). A novel cell group was also identified. The cells within this new group were immunoreactive to DBH but not TH, AADC, or phenylethanolamine N-methyltransferase (PNMT) and will be referred to as the TH-, DBH+ cell group. The TH-, DBH+ neurons were not immunoreactive for either the dopamine or noradrenaline transporters, suggesting that these neurons do not take up these transmitters. A5 neurons were immunoreactive for the noradrenaline transporter but not the dopamine transporter (as previously shown). Retrograde tracing with cholera toxin B revealed that the TH-, DBH+ neurons do not project to the thoracic spinal cord or to the rostral ventrolateral medulla, but A5 neurons do. A calbindin immunoreactive cell group is located in a region overlapping TH-, DBH+ cell group. However, only a few neurons were immunoreactive for both markers. The physiological role of the TH-, DBH+ cell group remains to be determined.
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PMID:Differential expression of catecholamine synthetic enzymes in the caudal ventral pons. 1155 1

The noradrenergic neurons of the locus coeruleus (LC) are damaged in Parkinson's disease (PD). Neurotoxin ablation of the LC noradrenergic neurons has been shown to exacerbate the dopaminergic toxicity of MPTP, suggesting that the noradrenergic system protects dopamine neurons. We utilized mice that exhibit elevated synaptic noradrenaline (NA) by genetically deleting the noradrenaline transporter (NET), a key regulator of the noradrenergic system (NET KO mice). NET KO and wild-type littermates were administered MPTP and striatal dopamine terminal integrity was assessed by HPLC of monoamines, immmunoblotting for dopaminergic markers and tyrosine hydroxylase (TH) immunohistochemistry. MPTP significantly reduced striatal dopamine in wild-type mice, but not in the NET KO mice. To confirm that the protection observed in the NET KO mice was due to the lack of NET, we treated wild-type mice with the specific NET inhibitor, nisoxetine, and then challenged them with MPTP. Nisoxetine conferred protection to the dopaminergic system. These data indicate that NA can modulate MPTP toxicity and suggest that manipulation of the noradrenergic system may have therapeutic value in PD.
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PMID:Reduced MPTP toxicity in noradrenaline transporter knockout mice. 1556 55

Expression of the noradrenaline transporter (NAT) was examined in normal human adrenal medulla and phaeochromocytoma by using immunohistochemistry and confocal microscopy. The enzymes tyrosine hydroxylase (TH) and phenylethanolamine N-methyltransferase (PNMT) were used as catecholamine biosynthetic markers and chromogranin A (CGA) as a marker for secretory granules. Catecholamine content was measured by using high performance liquid chromatography (HPLC). In normal human adrenal medulla (n=5), all chromaffin cells demonstrated strong TH, PNMT and NAT immunoreactivity. NAT was co-localized with PNMT and was located within the cytoplasm with a punctate appearance. Human phaeochromocytomas demonstrated strong TH expression (n=20 samples tested) but variable NAT and PNMT expression (n=24). NAT immunoreactivity ranged from absent (n=3) to weak (n=10) and strong (n=11) and, in some cases, occupied an apparent nuclear location. Unlike the expression seen in normal human adrenal medullary tissue, NAT expression was not consistently co-localized with PNMT. PNMT also showed highly variable expression that was poorly correlated with tumour adrenaline content. Immunoreactivity for CGA was colocalized with NAT within the cytoplasm of normal human chromaffin cells (n=4). This co-localization was not consistent in phaeochromocytoma tumour cells (n=7). The altered pattern of expression for both NAT and PNMT in phaeochromocytoma indicates a significant disruption in the regulation and possibly in the function of these proteins in adrenal medullary tumours.
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PMID:Expression of the noradrenaline transporter and phenylethanolamine N-methyltransferase in normal human adrenal gland and phaeochromocytoma. 1604 63

We have compared PC12 cell lines derived from different laboratories and the newly developed mouse pheochromocytoma (MPC) cell line. Morphologically, there were distinct differences in size, shape, adherence, and clumping behavior, which varied in response to different culture media, growth substrates, and nerve growth factor. Quantitative messenger ribonucleic acid (mRNA) analysis showed significant variability in the expression of the catecholaminergic biosynthetic enzymes tyrosine hydroxylase (TH), phenylethanolamine N-methyltransferase (PNMT), the noradrenaline transporter (NAT), and neuron-specific enolase (NSE) between all lines examined. Of most significance were the increased levels of PNMT mRNA in the MPC cells, which were to 15-fold greater than in the PC12 cell lines grown under the same conditions in Dulbecco modified Eagle medium (P < or = 0.05). Growth of MPC cells in Roswell Park Memorial Institute media induced a further significant increase in PNMT gene expression (P < or = 0.05). Immunohistochemistry for TH, PNMT, and NAT was generally consistent with mRNA analysis, with the MPC cells demonstrating strong immunoreactivity for PNMT. The MPC cells showed the highest levels of desipramine-sensitive [(3)H] noradrenaline uptake activity (threefold > than PC12 American Type Culture Center line, P < or = 0.05), despite relatively low levels of NAT mRNA. These results indicate that PC12 cell lines should be carefully chosen for optimal utility in the study of chromaffin cell or sympathetic neuron biology and that cell features will be influenced by type of media and substrate chosen. Furthermore, they confirm that the new MPC cell line is likely a useful model for the study of adrenergic mechanisms or studies involving NAT.
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PMID:Comparative studies of PC12 and mouse pheochromocytoma-derived rodent cell lines as models for the study of neuroendocrine systems. 1622 34

Although generally considered a prototypical movement disorder, Parkinson's disease is commonly associated with a broad-spectrum of non-motor symptoms, including autonomic dysfunctions caused by significant alterations in catecholaminergic neurons of the peripheral sympathetic nervous system. Here we present evidence that alpha-synuclein is highly expressed by sympathetic ganglion neurons throughout embryonic and postnatal life and that it is found in tyrosine hydroxylase-positive sympathetic fibers innervating the heart of adult mice. However, mice deficient in alpha-synuclein do not exhibit any apparent alterations in sympathetic development. Sympathetic neurons isolated from mouse embryos and early postnatal mice are sensitive to the parkinsonian drug MPTP/MPP(+) and intoxication requires entry of the neurotoxin through the noradrenaline transporter. Furthermore, recovery of noradrenaline from cardiac sympathetic fibers is reduced in adult mice treated with MPTP systemically. However, MPP(+)-induced sympathetic neuron loss in vitro or MPTP-induced cardiac noradrenaline depletion in vivo is not modified in mice lacking alpha-synuclein. This is in clear contrast with the observation that dopaminergic neurons of the central nervous system are significantly less vulnerable to MPTP/MPP(+) in the absence of alpha-synuclein, suggesting different actions of this molecule in central and peripheral catecholaminergic neurons.
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PMID:Vulnerability of peripheral catecholaminergic neurons to MPTP is not regulated by alpha-synuclein. 2007 41


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