Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:4.1.1.15 (glutamate decarboxylase)
 2,169 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The Belgian Diabetes Registry (BDR) has studied the epidemiology of Type 1 diabetes with clinical onset before age 40 years in the Antwerp district. Both in the age categories 0-14 years and 15-39 years, the incidence approximates 10 new cases per 100,000 inhabitants per years. Most patients are adults. Juvenile-onset diabetic patients who were so far more intensively studied must therefore not be considered as "prototypes" for the disease, but rather represent "atypical" cases with rapid evolution. Participation in international programs (EURODIAB ACE, DIAMOND) has characterized the incidence of Type 1 diabetes in Belgium as being intermediate between values in low-incidence regions (5 to 10 cases/100,000 inhabitants/yr such as in parts of Southern or Eastern Europe) and values in high-incidence regions 30 to 40 cases/100,000 inhabitants/yr such as in Finland and Sardinia). Type 1 diabetes is a heterogenous disease in terms of clinical presentation, etiological factors and biological markers. Clinical and fundamental research on Type 1 diabetes needs to study patient groups which faithfully reflect this heterogeneity. This is best achieved by recruiting patients through diabetes registries. In Belgium, the BDR presently registers about 40% of all new cases of Type 1 diabetes with onset before age 40 years. Until now, more than 1700 patients and about 1100 first degree relatives are registered. This group is representative of the Belgian population of Type 1 diabetic patients with onset before age 40 years. High quality-assays for immune and genetic markers of Type 1 diabetes were designed and validated through repeated participation in international quality control programs. By systematically performing these assays on the representative non-selected samples of BDR-patients, it became possible to further clarify the clinical biology of Type 1 diabetes and to demonstrate hitherto unrecognized associations among disease markers. Certain of these markers (insulin auto-antibodies, IAA; islet cell antibodies, ICA; HLA DQA1*0301-DQB1*0302 risk haplotype) are more frequent for clinical onset before age 10 years and appear associated in that age category. Other markers (glutamate decarboxylase antibodies; GAD-Ab) occur more frequently at onset between age 10 and 40 years, where they are preferentially associated with increased genetic risk. Yet another genetic marker (1/1 susceptibility genotype in the 5' polymorphic region of the insulin gene) occurs regardless of age and presence of auto-antibodies, preferentially in patients without the highest HLA-linked genetic risk. Age-dependent differences in marker frequency and -associations possibly reflect age-dependent differences in etiological factors, in order of appearance of biological markers, or in progression rate of the disease. Presently, more than 90% of the patients under age 40 years carry at least one diabetes-associated immune or genetic marker, which opens perspectives for a better classification of the disease, especially in adults. Preliminary follow-up studies in first-degree relatives of registered patients confirm the diabetes-predictive value of the markers studied. A group of subjects at high risk for the disease could thus be identified. These subjects qualify for participation in preventive intervention trials. In this respect BDR officially represents Belgium in several international programs: EURODI-AB ACE for marker-studies, ICARUS for diabetes prediction, ENDIT for diabetes-prevention and GETREM for optimal diabetes treatment. This collaboration will focus in the near future on neonatal screening and follow-up, objective classification criteria for diabetes in adults, refined diabetes-prediction and preventive intervention studies. BDR may also serve as a tool for systematic research on the complications, innovative treatments and socio-economical aspects of diabetes.
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PMID:The importance of diabetes registries and clinical biology for the study and treatment of type 1 (insulin-dependent) diabetes mellitus. 900 3

We have previously reported direct glutamate (Glu) synapses upon GnRH-containing neurons in the primate hypothalamus, and extensive interactions between Glu and aminobutyric acid (GABA) neurons in areas associated with reproductive function. Both Glu and GABA are known to affect peripubertal GnRH neurohormone release, but their relative roles remain unclear. In a developmental survey, estrogen receptors (ER) and progesterone receptors (PR) were virtually undetectable after immunostaining the hypothalamus of prepubertal monkeys, but were clearly evident in neurons of adults. We hypothesized, therefore, that Glu and GABA neurons which develop ER or PR expression during puberty may participate in reactivation of the hypothalamic-pituitary-gonadal axis. To identify those neurons in midpubertal female cynomolgus monkeys, we performed immunofluorescence staining for ER or for PR in separate sets of hypothalamic sections, and then immunostained for Glu or for glutamate decarboxylase (GAD, to identify GABA neurons) using a contrasting fluorophore. ER and PR were localized in the cytoplasm and nuclei of Glu and GAD neurons in nine hypothalamic and related brain regions. Quantitation revealed intranuclear ER in an average of 80% of the Glu neurons in all regions analyzed, and an average of 84% of the GAD neurons in all regions except the supraoptic nucleus (28%). Intranuclear PR expression was more variable, occurring in an average of 93% of the Glu neurons in seven regions, but in only 41% in the medial preoptic area, and 0% in the arcuate-periventicular zone. In addition, while intranuclear PR was seen in 96% of the GAD neurons in the septum, it appeared in 67% of the GAD neurons in the paraventricular nucleus, 47% in the medial preoptic area, 40% in the periventricular zone, and was absent from neurons in the supraoptic nucleus and mammillary bodies. In summary, certain subpopulations of Glu and GABA neurons in principal hypothalamic regions of the female monkey express ER and PR at midpuberty. Taken together with previous findings, these results suggest that Glu and GABA neurons which become sensitive to steroid hormones may help regulate GnRH neurohormone release and promote the onset of puberty. Since neuronal expression of ER or PR connotes sensitivity to gonadal feedback, and intranuclear translocation signals transcriptional activation, these results provide insights into the specific neuronal events involved in the peripubertal transition in primates.
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PMID:Expression of estrogen and progesterone receptors in glutamate and GABA neurons of the pubertal female monkey hypothalamus. 915 63

Autoantibodies to the islet-cell 65-kDa variant of glutamate decarboxylase (GAD65) are found in most insulin-dependent diabetes mellitus (IDDM) patients many years before the appearance of clinical symptoms of the disease. As IDDM-preventive therapies may be available in the future, an international effort is taking place to develop widely applicable anti-GAD immunochemical tests. These tests would help to detect individuals at risk before the full installation of the disease and to enroll them in prevention programs. Autoantibodies to GAD65 are mostly directed to conformational epitopes, and the enzyme is a complex molecule with a prosthetic group and 15 cysteine residues. Thus, the conformational integrity of GAD65 is essential for an appropriate anti-GAD assay. Isolation of large amounts of GAD65 from pancreas or other tissues is impractical, and no successful production of properly folded GAD65 has been reported in bacteria. Native recombinant GAD65 for immunochemical tests is usually obtained from eukaryotic expression systems. Since the large-scale production of a recombinant protein in an eukaryotic system is expensive and technically difficult, we investigated the expression of GAD65 in Escherichia coli as an alternative. A number of DNA constructs intended to export the enzyme to the periplasmic space or to improve its cytoplasmic solubility were designed and tested. Our results provide a solution to the two main problems associated with the expression of GAD65 in E. coli: misfolding, leading to the formation of inclusion bodies; and the presence of alternative initiation sites for translation that causes the preferential production of truncated variants of GAD65. We describe here the production of properly folded, fully active, and immunochemically competent GAD65 as an N-terminal fusion protein with thioredoxin. An account of the reactivity of the produced protein with sera of six IDDM patients is also presented.
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PMID:Expression of properly folded human glutamate decarboxylase 65 as a fusion protein in Escherichia coli. 920 24

The cellular distribution of the mRNAs encoding for the two isoforms of glutamate decarboxylase, GAD67 and GAD65, was analyzed by in situ hybridization histochemistry in the caudate nucleus and putamen of control and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated parkinsonian squirrel monkeys. On brain sections processed with a radioactive and a digoxigenin-labeled cRNA probe, the GAD67 and GAD65 mRNAs were colocalized in virtually all labeled neurons of the caudate nucleus and putamen, in both control and MPTP-treated monkeys. Furthermore, neurons labeled with the GAD cRNAs constituted at least 90% of all striatal neurons, as estimated on adjacent Nissl-stained sections. In the two groups of monkeys, double-labeling experiments using a combination of radioactive GAD67 or GAD65 and digoxigenin-labeled preproenkephalin (PPE) cRNA probes showed that roughly half of all neurons labeled with the GAD cRNAs were also labeled with the PPE cRNA probe. When compared to controls, GAD67 and GAD65 mRNA levels were higher in the putamen, and to a lesser extent in the caudate nucleus, of MPTP-treated monkeys. Further analysis of labeling at the cellular level in a dorsolateral sector of the putamen revealed that GAD67 and GAD65 mRNA levels in MPTP-treated monkeys were increased in PPE-labeled (presumed striato-pallidal) neurons but not in PPE-unlabeled (presumed striato-nigral) neurons. Our results demonstrate that most neurons in the caudate nucleus and putamen of squirrel monkeys contain the mRNAs encoding for the two GAD isoforms. In addition, the selective increase in GAD mRNA levels in PPE-labeled neurons provides further evidence that striato-pallidal GABAergic neurons are hyperactive in MPTP-treated parkinsonian monkeys.
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PMID:Glutamate decarboxylase (GAD67 and GAD65) gene expression is increased in a subpopulation of neurons in the putamen of Parkinsonian monkeys. 926 73

The bipolar cells in vertebrate retinas are considered to be excitatory in nature and use L-glutamate as their neurotransmitter. Our earlier studies have provided evidence demonstrating that a small but significant population of orthotopic bipolar cells in salamander retina may be gamma-aminobutyric acid (GABA)ergic. In this work, the stratification levels of axon terminals in the inner plexiform layer (IPL) of single L-glutamic acid decarboxylase-immunoreactive (GAD-IR) and GABA-immunoreactive (GABA-IR) bipolar cells in the salamander retinal slices were studied. GAD-IR and GABA-IR bipolar cells marked by a fluorescent probe, Texas Red, were injected with Lucifer Yellow (LY) through a patch pipette under visual control. A total number of 42 GAD-IR bipolar cells in 24 slices and 84 GABA-IR bipolar cells in 56 slices were injected. Among these, terminals of nine GAD-IR bipolar cells and 22 GABA-IR bipolar cells were sufficiently filled with LY for determination of the stratification levels in the IPL. The stratification patterns and levels of GAD-IR and GABA-IR bipolar cells were very similar. GAD-IR and GABA-IR orthotopic type I and type II bipolar cells (soma located in the most distal or middle of the inner nuclear layer [INL], respectively), had their axon terminals stratified in sublamina a and sublamina b of the IPL with comparable frequency. Axonal processes were restricted largely to either the distal or the proximal region within sublaminae a and b. In addition, three of the bipolar cells had their terminals located in the middle region of the IPL. The similarities of stratification patterns and levels between GAD-IR and GABA-IR type I and type II bipolar cells indicate that they represent the same population of presumed GABAergic bipolar cells. Based on comparative stratifications of GABA bipolar cells reported here and those derived from electrophysiological studies (Hensley et al. [1993] J. Neurophysiol. 69:2086-2098), it is suggested that putative GABAergic bipolar cells represent cone-dominated and rod-dominated ON- and OFF-bipolar cells and that they subserve a broad role in the ON- and OFF-visual pathways in the retina.
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PMID:L-glutamic acid decarboxylase- and gamma-aminobutyric acid-immunoreactive bipolar cells in tiger salamander retina are of ON- and OFF-response types as inferred from Lucifer Yellow injection. 930 11

Antibodies to the 40 kD antigen (identified as tyrosine phosphatase IA-2) and glutamate decarboxylase (GAD65) are strongly associated with insulin dependent diabetes mellitus (IDDM). However, antibodies to GAD (GADA) can appear in the absence of IDDM, particularly in stiff man syndrome (SMS) and in some individuals with autoimmune polyendocrine syndrome type II (APS II) and organ specific autoimmune diseases. The aim of this study was to compare the specificity of IA-2 antibodies (IA-2A) and GADA for IDDM by determining their frequency in different patient groups. IA-2A were present in 64/114 (56%) IDDM patients and 9/19 (47%) APS II patients with IDDM but in only 4/28 (14%) SMS patients. 1/24 (4%) APS II patients without IDDM and 1/113 (0.9%) patients with organ specific autoimmune disease had low level IA-2A. In contrast GADA were present in 77/114 (68%) IDDM patients and 17/19 (89%) APS II patients with IDDM, but also in 25/28 (89%) SMS patients, 5/24 (21%) APS II patients without IDDM and 22/113 (19%) patients with organ specific autoimmune diseases. Furthermore, within the group of new onset IDDM, IA-2A seemed to be associated with ICA and age: 63% of ICA positive IDDM patients had IA-2A (74% had GADA) increasing to 77% in the group below 20 years of age (69% for GADA). Our results demonstrate that IA-2A may be more specific for IDDM than GADA, as the latter are also present in patients with SMS, APS II without IDDM and organ specific autoimmune diseases. IA-2A were less frequent in older patients with IDDM than GADA or ICA. A combination of IA-2A and GADA detected 84% of total and 93% of ICA positive IDDM patients.
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PMID:Antibodies to the tyrosine phosphatase-like protein IA-2 are highly associated with IDDM, but not with autoimmune endocrine diseases or stiff man syndrome. 934 28

gamma-Aminobutyric acid (GABA), the major inhibitory neurotransmitter in the mammalian brain, is synthesized by two glutamate decarboxylase isoforms, GAD65 and GAD67. The separate role of the two isoforms is unknown, but differences in saturation with cofactor and subcellular localization suggest that GAD65 may provide reserve pools of GABA for regulation of inhibitory neurotransmission. We have disrupted the gene encoding GAD65 and backcrossed the mutation into the C57BL/6 strain of mice. In contrast to GAD67-/- animals, which are born with developmental abnormalities and die shortly after birth, GAD65-/- mice appear normal at birth. Basal GABA levels and holo-GAD activity are normal, but the pyridoxal 5' phosphate-inducible apo-enzyme reservoir is significantly decreased. GAD65-/- mice develop spontaneous seizures that result in increased mortality. Seizures can be precipitated by fear or mild stress. Seizure susceptibility is dramatically increased in GAD65-/- mice backcrossed into a second genetic background, the nonobese diabetic (NOD/LtJ) strain of mice enabling electroencephalogram analysis of the seizures. The generally higher basal brain GABA levels in this backcross are significantly decreased by the GAD65-/- mutation, suggesting that the relative contribution of GABA synthesized by GAD65 to total brain GABA levels is genetically determined. Seizure-associated c-fos-like immunoreactivity reveals the involvement of limbic regions of the brain. These data suggest that GABA synthesized by GAD65 is important in the dynamic regulation of neural network excitability, implicate at least one modifier locus in the NOD/LtJ strain, and present GAD65-/- animals as a model of epilepsy involving GABA-ergic pathways.
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PMID:Epilepsy in mice deficient in the 65-kDa isoform of glutamic acid decarboxylase. 939 Nov 52

The temporal and spatial distribution of glutamate decarboxylase 67 (GAD67) mRNA-containing neurons in the dentate gyrus was analyzed from embryonic day 20 (E20) to postnatal day 15 (PN15) with nonradioactive in situ hybridization methods. A major goal was to follow the development of an early-appearing population of gamma-aminobutyric acid (GABA) neurons within the developing molecular layer. At E20, GAD67 mRNA-containing neurons were highly concentrated slightly above the outer border of the developing granule cell layer. By PN3-PN5, many labeled cells were distributed within the developing granule cell layer; by PN15, labeled neurons occupied positions normally seen in the adult, such as along the inner border of the granule cell layer. This developmental pattern is unique and led to additional studies to determine the potential fate of the early-appearing GABA population. The possibility of apoptotic cell death was investigated with in situ end labeling techniques at developmental ages E21-PN15. Very few apoptotic cells were detected in the developing molecular layer at all ages examined. Birthdating studies of neurons labeled with bromodeoxyuridine revealed a changing pattern, similar to that of GAD67 mRNA, for cells with birthdates (E14) of many of the mature GAD-containing neurons in the dentate gyrus. The changes in the mRNA and birthdating patterns from E20-PN15 suggest that some of the early-appearing GABA neurons within the developing molecular layer of the dentate gyrus may alter their positions to eventually become the mature GABA population along the inner border of the granule cell layer.
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PMID:Developmental changes in GABA neurons of the rat dentate gyrus: an in situ hybridization and birthdating study. 941 3

The conformation, stability, cofactor interactions, and activation of a recombinant 65-kDa form of rat brain glutamate decarboxylase (GAD65) were investigated by using UV/visible spectrophotometry, fluorescence spectroscopy, circular dichroism, and differential scanning microcalorimetry. The enzyme was prepared from Sf9 insect cells infected with a recombinant baculovirus containing the entire GAD65 coding region. The UV/visible absorption spectrum of purified holoenzyme (holoGAD) exhibits two peaks in the range of 300-450 nm, which are due to the formation of a Schiff base when pyridoxal phosphate (pyridoxal-P) binds to GAD. Fluorescence emission intensity (excited at 295 or 280 nm) was substantially enhanced when pyridoxal-P was removed from holoGAD and quenched when pyridoxal-P was added to the apoenzyme (apoGAD). These observations implied that a significant enzyme conformational change occurs during the formation of holoGAD. Circular dichroism provided additional evidence for a conformational change, as the ellipticity of both negative (202-242 nm) and positive (188-202 nm) bands decreased when pyridoxal-P was removed from holoGAD. Secondary structure determination estimated that holoGAD contains a higher content of alpha-helix (34% versus 24%) and a lower content of beta-sheet (18% versus 30%) than apo-GAD. Differential scanning microcalorimetry indicated that holoGAD exhibits a much larger enthalpy and a 3 degrees C higher temperature of thermal unfolding than apoGAD, suggesting that holoGAD has a much tighter conformation and greater stability than apoGAD. A model describing the interaction of pyridoxal-P with GAD is presented, which proposes that an intermediate complex involving ionic interaction between the phosphate group of pyridoxal-P and the positive, charged residues in the active site of GAD maintains the pyridoxal-P molecule in an appropriate position in the active center. Simultaneously, this complex formation is accompanied by a moderate enzyme conformational change, providing a favorable configuration that enables the epsilon-amino of the active-site lysine to react with the aldehyde group of pyridoxal-P. The formation of active holoGAD involves a large enzyme conformational change, which leads to increased stability.
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PMID:Structural characteristics of brain glutamate decarboxylase in relation to its interaction and activation. 943 96

Increasing evidence indicates that the classical, fast-acting neurotransmitter gamma-amino butyric acid (GABA) may initially act as morphogen in cell proliferation and differentiation via specific receptors. In view of the potential roles for GABA in central nervous system development, we examined the expression of GABA, GABA(A) receptor beta1 and gamma1 subunits by immunocytochemistry and the expression of transcripts for two GABA-synthesizing enzymes, glutamate decarboxylase (GAD65, GAD67 mRNAs), and for alpha4, beta1, and gamma1 subunits of GABA(A) receptor by in situ hybridization in the developing neocortex. Tissue sections were taken from embryonic days (E) 17 and E20 embryos and newborn rats (P0). The embryos' mothers and newborn rats had been injected with 5-bromo-2'-deoxyuridine (BrdU) and had survived for 2 hours. At E17, BrdU-positive cells were largely restricted in the synthetic zone at the ventricular margin when cortical neurogenesis was still active. GAD mRNAs and GABA immunoreactivity were detected in the subventricular zone, while alpha4, beta1, and gamma1 subunits were abundant in the ventricular zone. At E20 and P0, when neurogenesis had largely ceased and gliogenesis had commenced, BrdU-positive cells were found throughout the ventricular zone with GABA, GAD mRNAs, and alpha4, beta1, and gamma1 subunits. GABA, GAD mRNAs and alpha4, beta1, and gamma1 subunit signals intensified in the ventricular zone from E17 to P0 as gliogenesis proceeded. Thus, specific components of a putative GABAergic circuit are expressed in cells of the ventricular zone during the late embryonic/early postnatal period coincident with gliogenesis, suggesting a role for GABA in glial cell proliferation.
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PMID:GABA, GAD, and GABA(A) receptor alpha4, beta1, and gamma1 subunits are expressed in the late embryonic and early postnatal neocortical germinal matrix and coincide with gliogenesis. 952 49


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