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Neurons possessing enzymes of dopamine synthesis in the mediobasal hypothalamus of rats. Topographic relations and axonal projections to the median eminence in ontogenesis.

We evaluated the topographic relations between tyrosine hydroxylase (TH)- and/or aromatic L-amino acid decarboxylase (AADC)-immunoreactive neurons in the arcuate nucleus (AN), as well as between TH- and/or AADC-immunoreactive axons in the median eminence (ME) in rats at the 21st embryonic day, 9th postnatal day, and in adulthood. The double-immunofluorescent technique in combination with confocal microscopy was used. Occasional bienzymatic neurons but numerous monoenzymatic TH- or AADC-immunoreactive neurons were observed in fetuses. There was almost no overlap in the distribution of monoenzymatic neurons, and therefore few appositions were observed in between. In postnatal animals, numerous bienzymatic neurons appeared in addition to monoenzymatic neurons. They were distributed throughout the AN resulting in the increased frequency of appositions. Furthermore, specialized-like contacts between monoenzymatic TH- and AADC-immunoreactive neurons appeared. The quantification of the fibers in the ME showed that there were large specific areas of the monoenzymatic TH-immunoreactive fibers and bienzymatic fibers in fetuses, followed by the gradual reduction of the former and the increase of the latter to adulthood. The specific area of the monoenzymatic AADC-immunoreactive fibers in fetuses was rather low, and thereafter increased progressively to adulthood. The fibers of all the types were in apposition in the ME at each studied age. Close topographic relations between the neurons containing individual complementary enzymes of dopamine synthesis at the level of cell bodies and axons suggest functional interaction in between.

Animals↗

Effect of surgical isolation of the hypothalamus on its neurotransmitter content.

The concentrations of norepinephrine, dopamine-beta-hydroxylase, dopamine, tyrosine hydroxylase, phenylethanolamine-N-methyltransferase, serotonin, tryptophan hydroxylase, histamine, glutamic acid decarboxylase, and choline acetyltransferase were determined in selected hypothalamic nuclei and in the median eminence after deafferentation of the medial basal hypothalamus. Norepinephrine and dopamine-beta-hydroxylase fell markedly while dopamine and tyrosine hydroxylase did not. Serotonin also decreased in all regions studied; histamine decreased in none. Choline acetyltransferase, phenylethanolamine-N-methyltransferase, and glutamic acid decarboxylase declined in some areas, but not in others.

Animals↗

Phenotypic changes of AADC-only immunopositive premammillary neurons in the brain of laboratory shrew Suncus murinus by systemic administration of monoamine precursors.

After 5-hydroxy-L-tryptophan (5-HTP) and L-3,4-dihydroxyphenylalanine (L-DOPA) were injected i.p. in the laboratory shrew Suncus murinus, immunocytochemical and immunofluorescence studies were conducted on continuous or same sections of the brain, using specific anti-tyrosine hydroxylase (TH), anti-aromatic L-amino acid decarboxylase (AADC), anti-dopamine (DA) and anti-serotonin (5-HT) antisera which were produced in our laboratory. The results of double-staining by the immunofluorescence method as well as immunoelectron microscopy strongly indicate that the cells of the premammillary nucleus of the laboratory shrew brain (AADC-only-positive neurons) are capable of synthesizing DA and 5-HT simultaneously upon simultaneous administration of L-dopa and 5-HTP.

Animals↗

Transient appearance of GTP cyclohydrolase I--positive non-monoaminergic neurons in the ventral lateral geniculate nucleus of postnatal mice.

The transient appearance of GTP cyclohydrolase I (GCH)-immunoreactive (ir) cells in the ventral lateral geniculate nuclear region of mice was detected by use of an avidin-biotin peroxidase complex method with an antibody specific for an oligopeptide of rat GCH (residues from 12 to 23, GFPERELPRPGA). In this brain region, we found for the first time novel GCH-ir cells already at postnatal day 1 (P1). The numbers reached maximum at P14 and decreased until P29, and they had mostly disappeared by P56. These cells were tyrosine hydroxylase negative and aromatic L-amino acid decarboxylase negative, indicating a lack of dopamine or serotonin production, and thus do not belong to the monoaminergic neuron system.

Age Factors↗

Metabolism of 3-nitrotyrosine induces apoptotic death in dopaminergic cells.

Intrastriatal injection of 3-nitrotyrosine, which is a biomarker for nitrating oxidants, provokes dopaminergic neuronal death in rats by unknown mechanisms. Herein, we show that extracellular 3-nitrotyrosine is transported via the l-aromatic amino acid transporter in nondopaminergic NT2 cells, whereas in dopaminergic PC12 cells, it is transported by both the l-aromatic amino acid and the dopamine transporters. In both cell lines, 3-nitrotyrosine is a substrate for tyrosine tubulin ligase, resulting in its incorporation into the C terminus of alpha-tubulin. In NT2 cells, incorporation of 3-nitrotyrosine into alpha-tubulin induces a progressive, reversible reorganization of the microtubule architecture. In PC12 cells, 3-nitrotyrosine decreases intracellular dopamine levels and is metabolized by the concerted action of the aromatic amino acid decarboxylase and monoamine oxidase. Intracellular levels of 133 micromol of 3-nitrotyrosine per mole of tyrosine did not alter NT2 viability but induced PC12 apoptosis. The cell death was reversed by caspases and aromatic amino acid decarboxylase and monoamine oxidase inhibitors. 3-Nitrotyrosine induced loss of tyrosine hydroxylase-positive primary rat neurons, which was also prevented by an aromatic amino acid decarboxylase inhibitor. These findings provide a novel mechanism by which products generated by reactive nitrogen species induce dopaminergic neuron death and thus may contribute to the selective neurodegeneration in Parkinson's disease.

Amino Acid Transport Systems↗

Differential and tissue-specific expression of a gene family for tyrosine/dopa decarboxylase in opium poppy.

Two early and potential rate-limiting steps in the biosynthesis of isoquinoline alkaloids, such as morphine and codeine, in opium poppy (Papaver somniferum) involve decarboxylation of L-tyrosine and L-dihydroxyphenylalanine (L-dopa) to yield tyramine and dopamine, respectively. A DNA fragment was amplified by polymerase chain reaction (PCR) using degenerate primers designed to two highly conserved domains found in other aromatic amino acid decarboxylases. A poppy seedling cDNA library was screened with this PCR product and a cDNA (cTYDC1) for tyrosine/dopa decarboxylase (TYDC/DODC) was isolated. Two other independent cDNAs (cTYDC2 and cTYDC3) encoding TYDC/DODC were isolated by heterologous screening with a plant tryptophan decarboxylase (TDC) cDNA as probe. A poppy genomic library was screened with cTYDC1 and two intronless genomic clones (gTYDC1 and gTYDC4) were isolated. The deduced amino acid sequences of all poppy clones share extensive identity with other reported pyridoxal phosphate-dependent decarboxylases from both plants and animals. Based on sequence homology, members of the gene family were divided into two subsets (cTYDC1 and gTYDC4; cTYDC2 and cTYDC3) of proteins with predicted M(r) = 56,983 and 59,323, respectively. Within each subset the clones exhibit greater than 90% identity, whereas clones between subsets share less than 75% identity. Expression of gTYDC1 and cTYDC2 as beta-galactosidase fusion proteins in Escherichia coli resulted in catalytically active enzymes immunodetectable with TDC-specific polyclonal antibodies. Each enzyme showed marginally higher substrate specificity for L-dopa over L-tyrosine, but did not accept L-tryptophan and L-phenylalanine as substrates. Genomic DNA blot-hybridization analysis revealed 6 to 8 genes homologous to cTYDC1 and 4 to 6 genes homologous to cTYDC2 in the tetraploid poppy genome. A premature translation stop codon was found in the gTYDC4 clone suggesting that it may not encode a functional protein. RNA blot-hybridization with probes specific to the gTYDC1- or cTYDC2-like subsets showed that members of the TYDC gene family are differentially expressed in various plant tissues.

Amino Acid Sequence↗

Effect of 1-methyl-4-phenylpyridinium ion (MPP+) on catecholamine levels and activity of related enzymes in clonal rat pheochromocytoma PC12h cells.

1-Methyl-4-phenylpyridinium ion (MPP+), a metabolite of a neurotoxin, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine, was found to reduce dopamine (DA) level and the activity of enzymes related to its metabolism in clonal rat pheochromocytoma PC12h cells. After 6 days' culture in the presence of 1 mM and 100 microM MPP+, DA content in PC12h cells was reduced markedly, but with MPP+ at concentrations lower than 10 microM, DA levels in the cells did not change. The amounts of 3,4-dihydrophenylacetic acid (DOPAC), a metabolite of DA were reduced markedly in culture medium and in PC12h cells cultured with MPP+ at concentrations higher than 1 microM. MPP+ was found to reduce the enzyme activity of tyrosine hydroxylase (TH), monoamine oxidase (MAO) and aromatic L-aminoacid decarboxylase (AADC). In the presence of MPP+ at concentrations higher than 10 microM, reduction of TH activity in the cells was more pronounced than reduction of cell protein or of the activity of a non-specific enzyme, beta-galactosidase. With 1 mM and 100 microM MPP+, MAO activity was reduced to about 30% of that in control cells. Reduction was observed with MPP+ at concentrations higher than 1 microM. AADC was the most sensitive to MPP+ and its activity was reduced markedly in the cells cultured with 100 nM MPP+. These results indicate that MPP+ inhibits not only the biosynthesis of catecholamines, but also the enzyme participating in their catabolism in cells, and may thus perturb catecholamine levels in the brain.

1-Methyl-4-phenylpyridinium↗

Evidence concerning the anatomical location of the dopamine stimulated adenylate cyclase in the substantia nigra.

The dopamine (DA)-sensitive adenylate cyclase in the substantia nigra was assayed in rats which had been subjected to 3 different kinds of brain lesion: (1) unilateral 6-hydroxydopamine (6-OHDA) lesions of the medial forebrain bundle; (2) unilateral lesions of the descending strio-nigral and pallido-nigral projections; (3) total lesions of the serotoninergic raphe-nigral pathway. Lesions of the medial forebrain bundle causing 97% depletion of striatal DA, 72% depletion of nigral tyrosine hydroxylase, and no change in nigral glutamate decarboxylase (GAD), resulted in no change in basal or DA-stimulated cyclic AMP production ipsilateral to the injection. Lesions of the globus pallidus, causing 70% and 79% reductions in GAD and substance P respectively in the ipsilateral nigra, produced a reduction in basal cyclic AMP production and abolished the normal increase in cyclic AMP produced by DA on the side of the lesion. Lesions to the dorsal and median raphe nuclei did not affect the normal DA-sensitive adenylate cyclase response in the nigra. The results suggest that one of the neurotransmitter functions of DA in this brain region may be to modulate the release of psi-aminobutyric acid (GABA) or substance P from synaptic terminals afferent to the nigra.

Adenylyl Cyclases↗

Early events in the antiproliferative action of tumor necrosis factor are similar to the early events in epidermal growth factor growth stimulation.

The process of TNF-induced cytotoxicity is complex but appears to be mediated through a TNF-specific cell surface receptor. Recent evidence suggests that TNF action on tumor cells may be antagonized by epidermal growth factor (EGF) and other EGF-receptor modulatory peptides implicating a role for EGF-R in the process of TNF-induced cytotoxicity. In the present report, we investigated the biochemical actions of TNF on several biochemical events known to occur in the process of EGF signal transduction in intact cells. The actions of TNF were compared directly to those of EGF in both TNF-sensitive and -resistant tumor cell lines. In TNF-sensitive ME-180 cervical carcinoma cells, TNF (20 ng/ml) stimulated the tyrosine protein kinase activity of the EGF-receptor (EGF-R) fivefold when measured by receptor autophosphorylation in an immune complex kinase assay. TNF activation of EGF-R kinase activity in ME-180 was measurable 10 min after TNF incubation and enzymatic activity remained elevated 20 min after TNF addition. Activation of the receptor by TNF correlated with increased 32P incorporation into EGF-R protein when receptor was immunoprecipitated from 32P-equilibrated cells following a 20 min incubation with TNF. Acid hydrolysis of EGF-R protein isolated from TNF-treated ME-180 cells demonstrates an increase in the phosphotyrosine content of EGF-R when compared to receptor isolated from untreated cells. The results suggest that TNF increased EGF-R tyrosine protein kinase activity and the state of EGF-receptor tyrosine phosphorylation in a manner similar to that reported for EGF. However, TNF does not appear to be structurally related to EGF since TNF was unable to directly activate EGF-R when incubated with extensively washed immunoprecipitates of EGF-R. In TNF-resistant T24 bladder carcinoma cells, TNF failed to alter EGF-R tyrosine protein kinase activity although both EGF and phorbol ester were shown to modulate the enzymatic activity of the receptor in these cells. These results indicate that the ability of TNF to modulate EGF-R kinase in target cells may correlate with its cytotoxic actions on TNF-sensitive tumor cells. Other biochemical activities associated with the induction or regulation of cellular growth were examined in TNF- or EGF-treated tumor cells. EGF stimulated a rapid 8-16-fold increase in the expression of the proto-oncogene c-myc when analyzed by dot-blot analysis of total cellular RNA or Northern blot hybridization of polyadenylated RNA. TNF treatment failed to alter c-myc expression in ME-180 cells when analyzed by either technique.(ABSTRACT TRUNCATED AT 400 WORDS)

Cell Division↗

Blockade of alpha-and beta -adrenergic receptors can prevent stimulation of liver ornithine decarboxylase activity by glucocorticoid or laparatomy.

Rat liver ornithine decarboxylase induction by dexamethasone or laparatomy, which is dramatically impaired by catecholamine depletion, is not affected by alpha-and beta -adrenergic blockers administered simultaneously 1 h prior to steroid injection or operation. However, if blockade is maintained for 24 h, an effect comparable to that of catecholamine depletion is obtained. Reciprocally, the response of the decarboxylase to catecholamines is severely compromised in adrenalectomized rats. Under the same conditions, induction of tyrosine aminotransferase by dexamethasone is not significantly affected by catecholamine availability, which altogether demonstrates that rat liver ornithine decarboxylase activity is specifically governed by the interaction between glucocorticoids and catecholamines.

Animals↗

Isoflavone genistein inhibits the initiation and promotion of two-stage skin carcinogenesis in mice.

Isoflavone genistein is a specific inhibitor of protein tyrosine kinase (PTK) and has been shown to have a variety of anticancer activities in cultured cells and animal models. We report here that genistein significantly inhibits 7,12-dimethylbenz[a]anthracene (DMBA)-initiated and 12-O-tetradecanoyl phorbol-13-acetate (TPA)-promoted skin tumorigenesis in a two-stage carcinogenesis model. In an initiation study, 10 micromol genistein was applied daily to female SENCAR mouse skin for 1 week, followed by initiation with 10 nmol DMBA. Mice were then treated with twice weekly 4 microg TPA. Genistein was shown to reduce tumor incidence and multiplicity in DMBA-initiated skin tumors by approximately 20 (P < 0.05) and 50% (P < 0.01), respectively. Two promotion studies were conducted using CD-1 and SENCAR mice. In experiment 1, CD-1 mice were initiated with 100 nmol DMBA and followed by a twice weekly regimen of 1 and 5 micromol genistein/4 microg TPA. In experiment 2, SENCAR mice were initiated with 10 nmol DMBA and followed by a regimen of 5, 10 and 20 micromol genistein/2 microg TPA. Both studies consistently showed that genistein substantially inhibited TPA-promoted skin tumorigenesis by reducing the tumor multiplicity by approximately 60 and 75%, respectively (P < 0.01). However, the tumor incidence appeared to be less affected. Mechanistic studies showed that genistein inhibited DMBA-induced bulky DNA adduct formation and substantially suppressed TPA-stimulated H2O2 and inflammatory responses in mouse skin by >60% (P < 0.01). In contrast, genistein only exhibited a moderate inhibition of TPA-induced ornithine decarboxylase activity (P > 0.05). Our results suggest that genistein exerts its anti-initiational and anti-promotional effects on skin carcinogenesis probably through blockage of DNA adduct formation and inhibition of oxidative and inflammatory events in vivo.

9,10-Dimethyl-1,2-benzanthracene↗

Differential effects of prepubertal chronic oestrogen treatment on the synthesis of noradrenaline in uterine myometrial and perivascular sympathetic nerves.

Previous studies have shown that chronic administration of oestrogen to prepubertal rats reduces the total content of noradrenaline in the uterine horn, abolishes myometrial noradrenergic innervation and reduces noradrenaline-fluorescence intensity of intrauterine perivascular nerve fibres. The mechanisms underlying these changes are not known. In the present study we have analysed the effects of prepubertal chronic oestrogen treatment on the synthesis of noradrenaline in the rat uterine sympathetic nerves using biochemical and immunohistochemical approaches. Tyrosine hydroxylase activity was evaluated biochemically, by measuring the in vivo accumulation of dihydroxyphenylalanine (DOPA) in the presence of a DOPA-decarboxylase inhibitor. In addition, nerve fibres were visualised immunohistochemically using antibodies against tyrosine hydroxylase (TH), dopamine-beta-hydroxylase (DbetaH) and the general marker protein gene product 9.5 (PGP 9.5). After chronic oestrogen treatment, the total content of noradrenaline of the uterine horn was reduced, whereas the total content of DOPA was increased. In controls, TH-immunoreactive, DbetaH-immunoreactive and PGP 9.5-immunoreactive nerve fibres were distributed in both the circular and longitudinal myometrial layers and in the blood vessels of the intran-myometrial region. After chronic oestrogen treatment the only fibres recognised by the three antibodies were those associated with the blood vessels, but no myometrial-associated fibres could be recognised. These results suggest that noradrenaline synthesis is selectively reduced in myometrial-associated uterine sympathetic nerves, but is preserved in perivascular sympathetic nerves. The increased DOPA levels measured after chronic exposure to oestrogen was interpreted as the consequence of the substantial increase in size and number of blood vessels observed in the uterus of oestrogen-treated animals. A possible neurodegenerative effect of oestrogen on myometrial sympathetic fibres is discussed.

Animals↗

The effect of a single dose of reserpine administered prior to incubation on the development of tyrosine hydroxylase activity in chick sympathetic ganglia.

A single dose of reserpine administered into the yolk sac of chicken eggs prior to incubation produces two distinct periods of significant increase in tyrosine hydroxylase (TH) activity over controls. The first period is 21 days of incubation (55%) and the second is between day 14 and 30 after hatching (a.h.) (69%). Cholineacetyltransferase (ChAc) and dopadecarboxylase (DDC) are not modified in the two periods of increased TH activity. Reserpine had no effect on cholinergic parasympathetic synapses and neurons in the ciliary ganglion, as judged by ChAc activity. When reserpine was acutely administered in three different posthatching periods only the injection at the latest period (days 26 and 27) caused a significant (38%) increase in TH activity at day 30. Postsynaptic nicotinic receptors were blocked selectively by injecting chlorisondamine in the chick starting at hatching for one week. The administration of chlorisondamine almost completely abolished the reserpine induced increase of TH activity at day 15 a.h. The present results support the view that the development of enzyme activities specifically related to neurotransmitter biosynthesis in chick autonomic ganglia is regulated not only by transsynaptic influences but also by regulatory inputs originating in the periphery.

Animals↗

Autoantibodies predicting diabetes mellitus type I in celiac disease.

UNLABELLED: Celiac disease (CD) and diabetes mellitus type I (DM-I) are both autoimmune diseases. Abnormal first-phase insulin response (FPIR) is associated with the prediabetic phase. Glutamic acid decarboxylase (GAD) and islet cell antibodies (ICAs) - especially the tyrosine phosphatase-like protein IA-2 antibodies - are considered to be serological markers of DM-I future development. The aim of this study is to investigate the presence of autoantibodies (GAD, IA-2) in individuals with CD, on a gluten-free diet, who have normal intestinal morphology. Thirty patients with CD (4-22, mean 15 years), 30 newly diagnosed diabetic children (2.5-16, mean 10 years) and 30 healthy subjects (7-35, mean 18 years) were investigated. Serum GAD and IA-2 autoantibodies were assessed by a quantitative enzyme-linked immunosorbent assay (ELISA) method in all patients and controls. Seven CD patients (23%), 28 diabetic children (93%) and none in the control group had positive GAD and IA-2 antibodies. The FPIR was normal in CD patients (>/=46 mU/l). CONCLUSIONS: GAD and IA-2 antibodies are detected in 23% of patients with CD. These patients may be at risk to develop DM-I. Regular follow-up and determination of FPIR for the early diagnosis of the prediabetic phase in patients with CD having circulating autoantibodies is recommended.

Adolescent↗

Studies on tyrosine hydroxylase system in rat brain slices using high-performance liquid chromatography with electrochemical detection.

A new method for the measurement of tyrosine hydroxylase (TH; EC 1.14.16.2) activity in brain slices was developed by using high-performance liquid chromatography (HPLC) with electrochemical detection (ED). To estimate TH activity in brain slices containing all of the components of the enzyme system, tetrahydrobiopterin, dihydropteridine reductase, and TH itself, slices were incubated with NSD-1055, an inhibitor of aromatic L-amino acid decarboxylase, and 3,4-dihydroxyphenylalanine (DOPA) formed from endogenous tyrosine was measured using HPLC-ED. Hydroxylation of endogenous tyrosine to DOPA in striatal slices was linear up to 90 min at 37 degrees C, and increased by incubation with 20 mM K+ to depolarize the nerve cells. Furthermore, the formation of DOPA could be detected in all parts of brain regions examined, and the activity in this slice system was nearly parallel to the maximal velocity of the homogenate from the slices as enzyme in the presence of saturating concentrations of tyrosine and 6-methyltetrahydropterin as cofactor. This assay system should be useful to study the regulatory mechanisms of TH in relatively intact tissue preparations.

Animals↗

Localization of L-glutamate decarboxylase and GABA transaminase immunoreactivity in the sympathetic ganglia of the rat.

The location of L-glutamate decarboxylase and gamma-aminobutyrate (GABA)-transaminase immunoreactivity in the superior cervical ganglion and in the coeliac-superior mesenteric ganglion complex of the rat was studied by an indirect immunofluorescence method and by immunoelectron microscopy, with specific antisera raised in rabbits against the corresponding enzymes. In light microscopy, several glutamate decarboxylase- or GABA-transaminase-immunoreactive principal nerve cells were detected in the superior cervical ganglion and coeliac-superior mesenteric ganglion complex. In addition, numerous small cells in both the superior cervical ganglion and coeliac-superior mesenteric ganglion complex showed intense immunoreactivity to glutamate decarboxylase or GABA-transaminase. The small cells were 10-20 micron in diameter and resembled in size and morphology the small intensely fluorescent cells. In consecutive sections, the small glutamate decarboxylase-immunoreactive cell clusters also showed immunoreactivity to tyrosine hydroxylase, suggesting that these cells contain the enzymes for both GABA and catecholamine synthesis. In the superior cervical ganglion and in the coeliac-superior mesenteric ganglion complex, GABA-transaminase immunoreactivity was also localized in fibre-like processes around and between the principal nerve cells, in nerve trunks traversing the ganglia, and around or in close contact with ganglionic blood vessels. Furthermore, GABA-transaminase immunoreactivity was observed in fibre-like structures close to the capsule of the ganglia. Division of the preganglionic nerve trunk of the superior cervical ganglion caused no detectable change in GABA-transaminase immunoreactivity in the ganglion. In immunoelectron microscopy of the superior cervical ganglion, GABA-transaminase immunoreactivity was localized in nerve fibres in association with neurotubules. A large number of GABA-transaminase labelled principal nerve cells were detected, containing immunoreactivity evenly distributed in their cytoplasm. GABA-transaminase immunoreactivity was also observed in satellite cells and their processes in the superior cervical ganglion. The present immunocytochemical results provide evidence that the rat sympathetic ganglia contain an intrinsic neuronal system showing histochemical markers for GABA synthesis and inactivation, but its functional role in the modulation of ganglionic neurotransmission remains to be established.

4-Aminobutyrate Transaminase↗

Course of islet autoantibody titers during Ig-immunoadsorption in a patient with newly diagnosed type 1 diabetes.

The purpose of this study was to determine whether and to what extent diabetes-specific autoantibodies can be removed from the plasma by Ig-immunoadsorption therapy. We followed the course of islet cell antibodies (ICA), insulin antibodies (I[A]A), glutamic acid decarboxylase antibodies (GADA) and antibodies to the protein tyrosine phosphatase IA-2 (IA2A) in a patient with newly diagnosed insulin-dependent diabetes mellitus (IDDM) under multiple immunoadsorption treatments over 6 months. Autoantibodies were not removed from the plasma as efficiently as expected when compared to the removal of total immunoglobulin (IgG). Whereas IgG levels were lowered by 70-90% through each immunoadsorption treatment, antibodies to insulin were reduced by an average of 83%, IA2A by 36% and GADA by only 9% directly after treatment. ICA were > 320 JDF U at diabetes onset and remained above this level. During the 6 months of multiple immunoadsorption therapies, I[A]A levels showed a 24-fold increase due to stimulation of insulin antibody production by exogenous insulin substitution, IA2A levels remained unchanged (average 6% increase), and GADA levels were reduced by an average of 39% compared to antibody titers at onset. All four antibodies were highly positive in the eluate from the immunoadsorption columns. We showed that antibodies to pancreatic islet cells can be reduced by immunoadsorption, but as for plasmapheresis the effect is incomplete and transient for most of the antibodies. If there is clinical benefit through immunoadsorption therapy--as has been shown for newly diagnosed IDDM patients treated with plasmapheresis--our data suggest that this may be due to factors other than the sufficient removal of antibodies.

Adult↗

Augmentation of endogenous dopamine production increases lung liquid clearance.

We have previously reported that dopamine increased active Na+ transport in rat lungs by upregulating the alveolar epithelial Na,K-ATPase. Here we tested whether alveolar epithelial cells produce dopamine and whether increasing endogenous dopamine production by feeding rats a 4% tyrosine diet (TSD) would increase lung liquid clearance. Alveolar Type II cells express the enzyme aromatic-L-amino acid decarboxylase (AADC) and, when incubated with the dopamine precursor, 3-hydroxy-L-tyrosine (L-dopa), produce dopamine. Rats fed TSD, a precursor of L-dopa and dopamine, had increased urinary dopamine levels, which were inhibited by benserazide, an inhibitor of AADC. Rats fed TSD for 15, 24, and 48 hours had a 26, 46, and 45% increase in lung liquid clearance, respectively, as compared with controls. Also, dopaminergic D1 receptor antagonist--but not dopaminergic D2 receptor antagonist--inhibited the TSD-mediated increase in lung liquid clearance. Alveolar Type II cells isolated from the lungs of rats after they had been fed TSD for 24 hours demonstrated increased protein abundance of Na,K-ATPase alpha1 and beta1 subunits. Basolateral membranes isolated from peripheral lung tissue of tyrosine-fed rats had increased Na,K-ATPase activity and Na,K-ATPase alpha1 subunit. These data provide the first evidence that alveolar epithelial cells produce dopamine and that increasing endogenous dopamine increases lung liquid clearance.

Animals↗