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

---

Query: EC:3.1.3.1 (alkaline phosphatase)
47,916 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In situ hybridization experiments, using oligodeoxyribonucleotides specific for the two major expressed human tyrosine hydroxylase mRNAs, were performed on human brain sections at the level of the mesencephalon. The specificity of the probes was ascertained by Northern blot experiments carried out with independently in vitro synthesized human tyrosine hydroxylase mRNAs. For in situ hybridization experiments, oligodeoxyribonucleotides were labelled with nucleotides tagged with digoxigenin or biotin molecules. The hybridized oligonucleotides were detected by antibodies coupled with peroxidase and alkaline phosphatase enzymes, which yield, with appropriate substrates, brown and purple products, respectively. The simultaneous detection of the two mRNAs with digoxigeninated and biotinylated probes revealed that these two mRNAs are co-expressed in single cells. The purple product obtained with alkaline phosphatase exhibits a discrete distribution within the dopaminergic cells suggesting these mRNAs are associated with sub-cellular structures. Finally, a heterogeneity in the intensity of the labelling of reactive cells with both probes was visualized as well as the expression of the two mRNA species in neurites.
...
PMID:Co-expression of tyrosine hydroxylase messenger RNA 1 and 2 in human ventral mesencephalon revealed by digoxigenin- and biotin-labelled oligodeoxyribonucleotides. 135 96

Progesterone (P)-induced PRL secretion in estradiol (E)-primed monkeys is not due to direct pituitary stimulation, because lactotropes do not express progestin receptors (PR). However, the hypothalamus, particularly the tuberoinfundibular dopaminergic system (TIDA), plays a major role in the regulation of PRL secretion. To determine whether hypothalamic dopamine neurons are progestin target cells, the colocalization of PR and tyrosine hydroxylase (TH), a phenotypic marker of dopaminergic neurons, was examined with double immunocytochemistry. Two methods for visualizing the antigens were applied; the first was a dual peroxidase method, and the second was a peroxidase-alkaline phosphatase method. In addition, the question of whether E induces PR in dopamine neurons was explored. Spayed female monkeys were treated with empty Silastic capsules, E-filled capsules for a period of 28 days, or E capsules supplemented with P capsules for the last 14 days of E treatment. Only the E- plus P-treated monkeys exhibited an increase in serum PRL during the P treatment period. Frontal sections at the level of the optic chiasm and arcuate nucleus were examined for the colocalization of TH and PR. After E treatment, hypothalamic PR-positive cells increased in both intensity and number. Neurons expressing both TH and PR were detected in the rostral hypothalamus, lateral to the third ventricle (A11-rostral) and in a discrete subventricular population (A11-subvent). The lateral population continued caudally (A11-caudal). The A11-subvent population exhibited little steroid regulation. Of the remaining A11 TH neurons, approximately 20% exhibited PR in the spayed and E-treated groups. Addition of P doubled the percentage of PR-containing TH neurons in this group. Although very few TH-positive neurons in the ventral arcuate nucleus contained PR (A12-ventral), many double labeled neurons were observed in the dorsal arcuate region (A12-dorsal). Ventral arcuate TIDA neurons were not regulated by steroids, but E plus P increased PR expression in A12-dorsal. Double labeled cells were rarely seen in the zona incerta (A13) or the emerging ventral tegmental area (A10). In summary, P probably does not act directly on ventral arcuate TIDA neurons to stimulate PRL secretion. However, the frequency of PR-positive dopamine neurons in the A11-rostral, A11-caudal, and A12-dorsal groups increased with E and P treatment. Therefore, the contribution of the PR-positive periventricular dopamine neurons to progestin-stimulated PRL secretion may be important.
...
PMID:Immunocytochemical colocalization of hypothalamic progestin receptors and tyrosine hydroxylase in steroid-treated monkeys. 135 39

An alkaline phosphatase-labelled anti-sense oligonucleotide probe specific for tyrosine hydroxylase (TOH) mRNA has been used for visualisation of TOH mRNA in the rat brain and adrenal gland. Both ribonuclease pre-treatment and the use of excess non-labelled probe abolished the specific hybridization signal. Furthermore the TOH mRNA-positive signal was only found in cells known from earlier studies to react with anti-TOH antibodies. To determine if the alkaline phosphatase-labelled probe could be used in a semiquantitative manner for measurement of the density of TOH mRNA signal, we used reserpine pre-treatment which induces TOH mRNA expression. The results revealed a significant increase in TOH mRNA signal in locus coeruleus and substantia nigra neurons, and in adrenal medulla chromaffin cells. The increased signal in these areas agreed with the increase in TOH mRNA signal previously observed by Northern analysis and suggests that this type of alkaline phosphatase-labelled probe allows sensitive detection of changes in TOH gene expression.
...
PMID:Sensitive non-radioisotopic in situ hybridization histochemistry: demonstration of tyrosine hydroxylase gene expression in rat brain and adrenal. 197 Aug 45

Tyrosine hydroxylase (TH) in freshly prepared 45,000 g supernatant from rat striatum was fractionated by DEAE-cellulose chromatography. The elution was made with 2 vols. of buffer (50 mM Tris, pH 7.4; 2 mM dithiothreitol) followed by 4 vols. of a linear NaCl gradient (0 0.3 M) in the same buffer. TH activity was eluted in two distinct peaks: one at about 0.1 M salt (I), and the other at 0.2 M salt (II). The relationship between the two enzymes peaks was examined as follows. (1) Incubation of the supernatant in the presence of cAMP-dependent protein kinase, 1 mM ATP, 10 mM Mg2+, and 0.1 mM cAMP resulted in the elimination of peak I, with a concomitant increase of peak II. This shift of TH peaks was prevented when the protein kinase was blocked by the addition of its inhibitory modulator. (2) Incubation of the supernatant with alkaline phosphatase, an enzyme known to dephosphorylate a variety of phosphoproteins, resulted in the elimination of peak II, with a concomitant increase of peak I. (3) Only freshly prepared supernatants showed two distinct TH peaks from DEAE-cellulose. From supernatants held at 0 degrees C for 24 h. peak II was markedly reduced and peak I concomitantly increased. Since peak II appears to be readily convertible to peak I, no further fractionation was attempted. From the data obtained here, we believe that peaks I and II are respectively the nonphosphorylated and phosphorylated forms of TH. Furthermore, the endogenous distribution of the two TH forms in striatum was altered by the administration of haloperidol (2 mg/kg. i.p.), a neuroleptic drug known to activate the enzyme via a cAMP-dependent mechanism. At 90 min after the treatment, there was a marked increase of peak II, with a concomitant decrease of peak I. Thus, this procedure provides a simple means for estimating the degree of phosphorylation of TH in vivo in catecholaminergic neurons under various physiological and pharmacological conditions.
...
PMID:Two forms of striatal tyrosine hydroxylase from DEAE-cellulose chromatography. 613 70

In the developing and adult human paraventricular (PVN) and supraoptic (SON) nucleus, a large proportion of neurons contains the catecholamine-synthesizing enzyme tyrosine hydroxylase (TH). In the present study we investigated the possible colocalization of TH with oxytocin (OXT) or vasopressin (VP) in the adult and neonatal PVN and SON. Adjacent paraffin sections were incubated simultaneously with two antibodies: a polyclonal against TH and a monoclonal against OXT or VP and stained with a double peroxidase-antiperoxidase/alkaline phosphatase method. We observed that TH-immunoreactive(IR) perikarya in the human PVN and SON were also positive for OXT or VP. A clear difference between the neonates and adult cases of our sample was observed in the proportion of TH-IR neurons that colocalize OXT or VP. In the neonates the majority of the TH-IR perikarya was also stained for VP, while only few TH-IR neurons were also positive for OXT. The opposite was observed in the adults, where the majority of the double-stained TH-IR neurons colocalizes OXT while only few TH-IR perikarya appear to contain VP. Our study establishes the colocalization of TH with OXT or VP in the adult and neonatal PVN and SON and indicates that antemortem factors such as perinatal hypoxia might increase TH-immunoreactivity of the VP neurons in man.
...
PMID:Colocalization of tyrosine hydroxylase with oxytocin or vasopressin in neurons of the human paraventricular and supraoptic nucleus. 769 71

Radioactive in situ hybridization was used to map the cellular localization of dopamine (DA) transporter mRNA-containing cells in the adult rat central nervous system. The distribution of DA transporter mRNA-containing cells was compared to adjacent sections processed to visualize tyrosine hydroxylase (TH) mRNA, a marker of catecholamine containing neurones. TH mRNA-containing cells, visualized using an alkaline phosphatase labelled probe, were detected in the hypothalamus, midbrain and pons; the strongest hybridization signals being detected in the substantia nigra, ventral tegmental area and locus coeruleus. The distribution of DA transporter mRNA-containing cells was more restricted; a strong signal being detected in the substantia nigra pars compacta and ventral tegmental area only. No hybridization signal was detected in the locus coeruleus. By simultaneously hybridizing mesencephalic tissue with both the alkaline phosphatase-labelled TH probe and the 35S-labelled DA transporter probe we were able to demonstrate that both DA transporter and TH mRNAs are expressed by the same cells in the substantia nigra and ventral tegmental area. The restricted anatomical localization of DA transporter mRNA-containing cells and the lack of expression in the locus coeruleus and other adrenergic and noradrenergic cell groups confirms the DA transporter as a presynaptic marker of DA containing nerve cells in the rat brain.
...
PMID:Co-expression of dopamine transporter mRNA and tyrosine hydroxylase mRNA in ventral mesencephalic neurones. 790 51

We describe a method to combine non-radioactive in situ hybridization using alkaline phosphatase (AP) labelled oligonucleotide-probes with immunohistochemistry on the same thin paraffin section. The simultaneous detection of calretinin-mRNA and calbindin- or tyrosine hydroxylase-like immunoreactivity in neurons of rat substantia nigra, pars compacta, was used as a test system to develop the method. Brains were fixed by perfusion with 4% paraformaldehyde and embedded in paraffin. Five-microns-thick sections were processed for non-radioactive in situ hybridization with a 33-base alkaline phosphatase conjugated synthetic oligonucleotide complementary to calretinin mRNA. After hybridization and colour reaction to visualize calretinin mRNA, sections were incubated with antibodies against calbindin D28K or tyrosine hydroxylase. Immunoreaction was visualized using the avidin-biotin-complex-technique and diaminobenzidine. As the colour of both reaction products differ markedly, the distribution of calretinin mRNA-containing neurons (purple-blue, alkaline phosphatase product) and calbindin/tyrosine hydroxylase immunopositive cells (brown peroxidase product) could be differentiated easily on the same section. Calbindin- and tyrosine hydroxylase-like immunoreactivity was found in the majority of calretinin mRNA-containing cells within the substantia nigra, pars compacta, indicating that in this nucleus a proportion of the dopaminergic neurons contain both calcium binding proteins calbindin and calretinin. In conclusion, non-radioactive in situ hybridization using alkaline phosphatase labelled oligonucleotide probes can be readily combined with immunohistochemistry.
...
PMID:Combination of alkaline phosphatase in situ hybridization with immunohistochemistry: colocalization of calretinin-mRNA with calbindin and tyrosine hydroxylase immunoreactivity in rat substantia nigra neurons. 790 17

We describe here a simple method for combining non-radioactive and radioactive in situ hybridization and immunohistochemistry on the same brain tissue section. This approach was first developed on the well-characterized hypothalamo-neurohypophyseal system, facilitating the optimization of the triple-labeling procedure and the verification of labeling specificity. We report the simultaneous detection of vasopressin (VP) mRNA with a digoxigenin-labeled oligonucleotide, oxytocin (OT) mRNA with a 35S-labeled oligonucleotide, and OT peptide in the same 12-microns cryostat section. This was performed on floating sections as follows: first, the two probes were hybridized simultaneously; second, the peptide was detected with an immunoperoxidase-DAB procedure; third, the digoxigenin-labeled probe was detected with an alkaline phosphatase-NBT/BCIP technique; and finally, the 35S-labeled probe was detected by histological autoradiography. We also demonstrate that this approach is suitable for the simultaneous detection of tyrosine hydroxylase and two less abundant mRNAs, vasoactive intestinal peptide and vasopressin mRNAs, in the suprachiasmatic nucleus. The combination of the three techniques did not significantly diminish their specificity or sensitivity. In conclusion, this new method, permitting the simultaneous detection of three different products of gene expression in the same section, could be useful for further analysis of the phenotypic organization and its plasticity in endocrine or neural tissues.
...
PMID:Combination of non-radioactive and radioactive in situ hybridization with immunohistochemistry: a new method allowing the simultaneous detection of two mRNAs and one antigen in the same brain tissue section. 809 8

Neurotrophic factors that improve the survival of specific neuronal types during development and after exposure to various neuronal insults hold potential for treatment of neurodegenerative diseases. In particular, brain-derived neurotrophic factor (BDNF) has been shown to exert trophic and protective effects on dopaminergic neurons, the cell type known to degenerate in Parkinson's disease. To determine whether increased levels of biologically produced BDNF affect the function or regeneration of damaged dopaminergic neurons, the effects of grafting astrocytes transduced with the human BDNF gene into the striatum of the partially lesioned hemiparkinsonian rat were examined. Replication deficient retroviruses carrying either human prepro-BDNF or human alkaline phosphatase (AP) cDNA were used to transduce primary type 1 astrocytes purified from neonatal rat cortex. In vitro, BDNF mRNA was expressed by BDNF transduced astrocytes (BDNF astrocytes), but not control AP transduced astrocytes (AP astrocytes), as determined by reverse transcription polymerase chain reaction (RT-PCR). The modified astrocytes were injected into the right striatum 15 days after partial lesioning of the right substantia nigra with 6-hydroxydopamine. Transplantation of BDNF astrocytes, but not AP astrocytes, significantly attenuated amphetamine-induced rotation by 45% 32 days after grafting. Apomorphine-induced rotation increased over time in both groups, but was not significantly different in the BDNF-treated group. The modified BDNF astrocytes survived well with non-invasive growth in the brain for up to 42 days. Although BDNF mRNA positive cells were not detected within the graft site using in situ hybridization, alkaline phosphatase immunoreactive (IR) cells were present in control graft sites suggesting that the retroviral construct continued to be expressed at 42 days. Analysis of the density of tyrosine hydroxylase (TH)-IR fibers showed no effect of BDNF on TH-IR fiber density in the striatum on the lesioned side. These findings suggest that ex vivo gene therapy with BDNF ameliorates parkinsonian symptoms through a mechanism(s) other than one involving an effect of BDNF on regeneration or sprouting from dopaminergic neurons.
...
PMID:Astrocytes retrovirally transduced with BDNF elicit behavioral improvement in a rat model of Parkinson's disease. 859 62

We have developed enhanced immunohistochemical protocols for detecting autonomic nerve fibers and splenocyte-associated proteins in rat spleen. This includes norepinephrine-synthesizing enzymes (dopamine-beta hydroxylase (DBH) and tyrosine hydroxylase (TH)), neuropeptide Y (NPY), tumor necrosis factor-alpha (TNF-alpha), interferon-gamma (IFN-gamma), c-fos protein, inducible nitric oxide synthase (iNOS), and the macrophage cell marker ED1. Animals were divided into sham-operated and splenic nerve-sectioned groups for detection of DBH, TH, and NPY. For immunodetection of TNF-alpha, iNOS, IFN-gamma and c-fos, animals were injected IV with saline or 100 microg of lipopolysaccharide (LPS) and were sacrificed at various time intervals post injection. Rats were perfused with 4% paraformaldehyde, spleens removed and cryoprotected, and 50-microm floating sections were cut on a freezing microtome. Immunodetection was performed with various detection systems and substrate/chromogen solutions, and in some cases using pretreatment with proteinase K (PK) for antigen unmasking. PK pretreatment increased immunostaining for DBH, TH, NPY, IFN-gamma, iNOS, and ED1, and the improvement was concentration-dependent. Using NPY immunostaining to index the signal-to-noise ratio for various substrates and detection systems, we found that an alkaline phosphatase detection system with NBT/BCIP as a substrate was the best procedure for light microscopy, whereas the CY3-labeled secondary antibody technique proved optimal for fluorescent microscopy. Surgical transection of the splenic nerve eliminated all nerve fiber staining for DBH, TH, and NPY. TNF-alpha, IFN-gamma, c-fos, and iNOS proteins were observed in the spleen in a time-dependent manner after LPS stimulation. Fluorescent double labeling, visualized with fluorescent confocal scanning laser microscopy, revealed many NPY fibers distributed among the ED1-labeled macrophages. These results demonstrate that immunohistochemistry can be used to index the activational effects of an immune challenge on splenocytes in situ and verifies that splenic immune cells are innervated by the sympathetic nervous system.
...
PMID:Enhanced immunohistochemical detection of autonomic nerve fibers, cytokines and inducible nitric oxide synthase by light and fluorescent microscopy in rat spleen. 911 Dec 38


1 2 3 Next >>

---