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:2.7.7.49 (reverse transcriptase)
31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The reduced progesterone metabolite tetrahydroprogesterone (3 alpha-hydroxy-5 alpha-pregnan-20-one; 3 alpha,5 alpha-THP) is a positive modulator of the gamma-aminobutyric acid type A (GABAA) receptor. Experiments performed in vitro with hypothalamic fragments have previously shown that GABA could modulate the release of gonadotropin-releasing hormone (GnRH). Using GT1-1 immortalized GnRH neurons, we investigated the role of GABAA receptor ligands, including 3 alpha,5 alpha-THP, on the release of GnRH. We first characterized the GABAA receptors expressed by these neurons. [3H]Muscimol, but not [3H]flunitrazepam, bound with high affinity to GT1-1 cell membranes (Kd = 10.9 +/- 0.3 nM; Bmax = 979 +/- 12 fmol/mg of protein), and [3H]muscimol binding was enhanced by 3 alpha,5 alpha-THP. mRNAs encoding the alpha 1 and beta 3 subunits of the GABAA receptor were detected by the reverse transcriptase polymerase chain reaction. In agreement with binding data, the benzodiazepine-binding gamma subunit mRNA was absent. GnRH release studies showed a dose-related stimulating action of muscimol. 3 alpha,5 alpha-THP not only modulated muscimol-induced secretion but also stimulated GnRH release when administered alone. Bicuculline and picrotoxin blocked the effects of 3 alpha,5 alpha-THP and muscimol. Finally, we observed that GT1-1 neurons convert progesterone to 3 alpha,5 alpha-THP. We propose that progesterone may increase the release of GnRH by a membrane mechanism, via its reduced metabolite 3 alpha,5 alpha-THP acting at the GABAA receptor.
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PMID:A progesterone metabolite stimulates the release of gonadotropin-releasing hormone from GT1-1 hypothalamic neurons via the gamma-aminobutyric acid type A receptor. 773 81

We have used quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) to analyze the expression of GABAA receptor subunit genes in cultured neurons from the chick embryo cerebral cortex. During maturation of the neurons between day 2 and day 8 in culture, levels of the alpha 1 subunit transcript (per ng total RNA) increased 3.8 +/- 0.3 fold, while those for the beta 2S and beta 4S subunits increased 2.4 +/- 0.4 and 1.8 +/- 0.2 fold, respectively. The accumulation of the beta 4 S subunit mRNA was more rapid than those encoding either the alpha 1 or beta 2S polypeptides. After 4 days in culture the beta 4S subunit transcript level reached 105 +/- 7.7% of that found after 8 days, while the corresponding amounts for the alpha 1 and beta 2S subunit mRNAs were 50 +/- 7.1% and 44 +/- 10.7%, respectively. On the other hand, no significant differences were observed in the level of either the gamma 1 or the gamma 2S subunit mRNA during development in vitro. Likewise, the ratios of the large/small splice variants (beta 2 = 0.16 +/- 0.02; beta 4 = 0.57 +/- 0.02; gamma 2 = 0.30 +/- 0.06) did not show detectable changes during this period. To study the down-regulation of the mRNAs, a single dose of 100 microM GABA was added to the culture medium. After 7 days of exposure to GABA, the levels of transcripts for the alpha 1, beta 2, beta 4, gamma 1, and gamma 2 subunits and their splice variants (where present) were all reduced by 47-65% compared to untreated controls.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Developmental up-regulation and agonist-dependent down-regulation of GABAA receptor subunit mRNAs in chick cortical neurons. 785 72

Levels of the GABA A receptor alpha1-, alpha6-, beta2-, beta3-, gamma2-, and delta-subunit mRNAs in cerebellar granule neurons rise concurrently during the second week of postnatal ontogeny. Previous studies in culture have suggested that extrinsic signals control these increases, but little is known about the nature of the regulatory cues. To determine when granule neurons become competent to express these six subunit mRNAs in mature patterns and to gain insight into their regulation, reverse transcriptase-PCR was used to examine transcript expression in cultured granule neurons prepared at 2-day intervals from postnatal days 2 through 10. Although only one pattern of expression was observed in vivo, three patterns were detected in culture. First, the levels of the alpha1- and alpha6-subunit mRNAs were constant in cultures prepared at all ages. Second, the levels of the beta2-, beta3-, and gamma2-subunit mRNAs were constant in cultures prepared at postnatal days 2-6 but increased in those prepared at days 8-10. Third, the delta-subunit mRNA level increased over time in culture regardless of cerebellar age at plating. Moreover, only delta-subunit transcript expression was modulated by cell density. These findings indicate that the subunit transcripts are differentially regulated by multiple environmental cues.
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PMID:Differential regulation of GABA A receptor subunit mRNAs in rat cerebellar granule neurons: importance of environmental cues. 862 85

Using patch-clamp recording in combination with reverse transcriptase-polymerase chain reaction (RT-PCR), we show in individual bipolar cells acutely dissociated from the adult rat retina a correlation between the expression of the GABAA receptor rho 1 subunit mRNA and a bicuculline-resistant, diazepam-insensitive component of the GABA-activated whole-cell current response. This "GABAC-like" response, contributing to approximately 42% of the GABA-activated whole-cell current and displaying variable sensitivity to picrotoxin, was found in bipolar cells but not in any of the ganglion cells examined. Expression profiling of GABAA receptor subunit mRNAs in individual electrophysiologically tested retinal neurons revealed that, while both bipolar cells and ganglion cells may express numerous GABAA receptor subunit isoforms, including that of rho 2, the expression of the rho 1 subunit was strictly limited to bipolar cells. We propose a possible link between the presence of a receptor with GABAC-like pharmacological profile and the expression of the retina-specific rho 1 subunit isoform. The results presented in this study constitute the first direct demonstration of such a correlation at the single-cell level.
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PMID:Correlation between a bicuculline-resistant response to GABA and GABAA receptor rho 1 subunit expression in single rat retinal bipolar cells. 873 79

The alpha6 subunit of the gamma-aminobutyric acid type A receptor (GABA(A)-R) has been implicated in mediating the intoxicating effects of ethanol and the motor ataxic effects of general anesthetics. To test this hypothesis, we used gene targeting in embryonic stem cells to create mice lacking a functional alpha6 gene. Homozygous mice are viable and fertile and have grossly normal cerebellar cytoarchitecture. Northern blot and reverse transcriptase-polymerase chain reaction analyses demonstrated that the targeting event disrupted production of functional alpha6 mRNA. Autoradiography of histological sections of adult brains demonstrated that diazepam-insensitive binding of [3H]Ro15-4513 to the cerebellar granule cell layer of wild-type mice was completely absent in homozygous mice. Cerebellar GABA(A)-R density was unchanged in the mutant mice; however, the apparent affinity for muscimol was markedly reduced. Sleep time response to injection of ethanol after pretreatment with vehicle or Ro15-4513 did not differ between genotypes. Sleep time response to injection of pentobarbital and loss of righting reflex and response to tail clamp stimulus in mice anesthetized with volatile anesthetics also did not differ between genotypes. Thus, the alpha6 subunit of the GABA(A)-R is not required for normal development, viability, and fertility and does not seem to be a critical or unique component of the neuronal pathway mediating the hypnotic effect of ethanol and its antagonism by Ro15-4513 in mice. Similarly, the alpha6 subunit does not seem to be involved in the behavioral responses to general anesthetics or pentobarbital.
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PMID:Gene knockout of the alpha6 subunit of the gamma-aminobutyric acid type A receptor: lack of effect on responses to ethanol, pentobarbital, and general anesthetics. 910 23

The single-locus mutant mouse tottering (tg) is an established model for absence seizures. We have previously reported an impairment in GABA-induced chloride uptake in tg brain [Tehrani and Barnes, Epilepsy Res. 1995;22:13-21]. In order to determine if this alteration in GABAA receptor function can be related to specific receptor isoforms, we examined the radioligand binding properties of GABAA receptors and the expression of GABAA receptor subunit mRNAs in the cerebral cortex. Saturation binding of [3H]flunitrazepam revealed a significantly lower Kd value in tg cortical tissues (1.77 +/- 0.05 nM) in comparison to that for the background C57BL/6J strain (3.23 +/- 0.23 nM), while the Bmax values were indistinguishable. Biphasic displacement of [3H]flunitrazepam binding by 2-oxoquazepam showed that low affinity binding sites account for 36 +/- 7.6 and 51 +/- 7.5% of the total in control and tg, respectively. The level of [35S]-t-butylbicyclophosphorothionate (TBPS) binding to tg cortical membranes was 73.6 +/- 5.8% of that in controls. Paired measurements by quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) revealed no significant differences in the levels of GABAA receptor alpha 1, alpha 3, alpha 5, beta 2, beta 3, gamma 2 or gamma 3 subunit mRNAs between tg and control cortex. However, tg tissues showed elevated levels of alpha 2- and beta 1-subunit mRNAs, representing 256 and 177%, respectively, those of controls. For the tg cortex, the enhanced expression of GABAA receptor alpha 2 and beta 1 subunits correlates with recombinant subtypes known to have low affinity for 2-oxoquazepam and impaired binding of TBPS. These aberrant properties of GABAA receptors could influence the development or propagation of phenotypic seizures in the tottering mouse.
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PMID:Aberrant expression of GABAA receptor subunits in the tottering mouse: an animal model for absence seizures. 933 86

Gamma-Aminobutyric acid-B (GABAB) receptor function and regulation in the nucleus of the solitary tract (NTS) was examined in Sprague-Dawley rats made chronically (4 to 5 weeks) hypertensive with the one-kidney, figure-8 renal wrap model of hypertension. NTS microinjection of the GABAB agonist baclofen produced a pressor response that was enhanced in hypertensive rats compared with the response observed in sham-operated normotensive rats (36+/-4 mm Hg increase in mean arterial pressure in 8 hypertensive rats compared with 21+/-2 mm Hg increase in 7 sham-operated normotensive rats, P=0. 03). Responses to microinjection of GABAB antagonists (CGP-55845A and SCH-90511), the GABAA agonist muscimol, the GABAA antagonist bicuculline, and the GABA reuptake inhibitor nipecotic acid were not different comparing normotensive sham-operated and hypertensive rats. Renal sympathetic nerve responses to NTS microinjection of these drugs were not different in hypertensive compared with normotensive rats. Micropunches of the NTS were homogenized and reverse transcriptase-polymerase chain reaction was performed to examine mRNA levels for the GABAB receptor. There was a 3-fold increase in GABAB receptor mRNA levels in the caudal NTS of 7 chronically hypertensive rats compared with levels measured in 8 sham-operated normotensive rats (P=0.01). In conclusion, chronic hypertension is associated with an upregulation of GABAB receptor function; however, the tonic activity of the system does not appear to be different between normotensive and hypertensive rats. The upregulation of GABAB receptor function might be due to an increased number of receptors, as suggested by the elevated levels of GABAB receptor mRNA measured in the NTS of hypertensive rats. All of these alterations suggest that hypertension is associated with dynamic changes in receptor-mediated mechanisms within the NTS, and these alterations could modify baroreflex regulation of cardiovascular function in hypertension.
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PMID:Enhanced gamma-aminobutyric acid-B receptor agonist responses and mRNA within the nucleus of the solitary tract in hypertension. 993 Nov 60

1. Molecular and biophysical properties of GABAA receptors of dopaminergic (DA) neurones of the pars compacta of the rat substantia nigra were studied in slices and after acute dissociation. 2. Single-cell reverse transcriptase-multiplex polymerase chain reaction confirmed that DA neurones contained mRNAs encoding for the alpha3 subunit of the GABAA receptor, but further showed the presence of alpha4 subunit mRNAs. alpha2, beta1 and gamma1 subunit mRNAs were never detected. Overall, DA neurones present a pattern of expression of GABAA receptor subunit mRNAs containing mainly alpha3/4beta2/3gamma3. 3. Outside-out patches were excised from DA neurones and GABAA single-channel patch-clamp currents were recorded under low doses (1-5 microM) of GABA or isoguvacine, a selective GABAA agonist. Recordings presented several conductance levels which appeared to be integer multiples of an elementary conductance of 4-5 pS. This property was shared by GABAA receptors of cerebellar Purkinje neurones recorded in slices (however, with an elementary conductance of 3 pS). Only the 5-6 lowest levels were analysed. 4. A progressive change in the distribution of occupancy of these levels was observed when increasing the isoguvacine concentration (up to 10 microM) as well as when adding zolpidem (20-200 nM), a drug acting at the benzodiazepine binding site: both treatments enlarged the occupancy of the highest conductance levels, while decreasing that of the smallest ones. Conversely, Zn2+ (10 microM), a negative allosteric modulator of GABAA receptor channels, decreased the occupancy of the highest levels in favour of the lowest ones. 5. These properties of alpha3/4beta2/3gamma3-containing GABAA receptors would support the hypothesis of either single GABAA receptor channels with multiple open states or that of a synchronous recruitment of GABAA receptor channels that could involve their clustering in the membranes of DA neurones.
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PMID:Incremental conductance levels of GABAA receptors in dopaminergic neurones of the rat substantia nigra pars compacta. 1020 Apr 21

GABA, the predominant inhibitory neurotransmitter present in the mammalian CNS, is also found in the periphery. GABA actions are mediated by the ionotropic GABA(A)/GABA(C) receptors, as well as the metabotropic GABA(B) receptor. The rat GABA(B) receptor has recently been cloned and two cDNA clones have been isolated encoding two isoforms of the receptor, GABA(B)R1a and R1b. Northern blot analysis revealed the presence of both transcripts in the rat brain using specific cDNA probes for GABA(B)R1a and R1b, respectively. However, Northern blot analysis, hybridized with a probe containing a sequence common to both isoforms, revealed specific RNAs in the rat brain and in testis, but not in other peripheral tissues. In the present study, by using the more sensitive reverse transcriptase-polymerase chain reaction with a specific set of primers for each isoform and Southern blot analysis, we found that both isoforms of the GABA(B) receptor are expressed not only throughout the brain but also in all peripheral organs examined, including heart, spleen, lung, liver, small intestine, large intestine, kidney, stomach, adrenal, testis, ovary and urinary bladder. The peripheral distribution of GABA(B)R1 mRNAs supports the notion of a physiological role for GABA in the control of a wide range of peripheral organs.
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PMID:Distribution of GABA(B) receptor mRNAs in the rat brain and peripheral organs. 1022 88

The patterns of expression of voltage gated potassium channel genes of the Shaker family have been mapped in identified neurons of the lobster (Homarus americanus) ventral nerve cord using a single cell reverse transcriptase polymerase chain reaction procedure. Using specific oligonucleotides derived from the sequences of the shaker, shab, and shaw genes of the spiny lobster, Panulirus interruptus, we detected the corresponding potassium channel DNA fragments from Homarus americanus. The Homarus DNA fragments are 87-98% identical at the nucleotide level to the Panulirus DNA fragments. We used the Panulirus primers to measure the complement of RNAs for shaker, shab, and shaw in single identified cells that use GABA, glutamate, octopamine or serotonin as chemical messengers. Shaker and shaw RNAs were found in all four identified neuron types but shab RNA was not detected in serotonin cells under the present experimental conditions. All cells expressed alpha-tubulin RNA, which serves as an internal control suggesting that cells are intact after dissection. In glial cells that surround the neuronal cell bodies, the potassium channel genes are expressed at low to non-detectable levels.
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PMID:Patterns of shaker family gene expression in single identified neurons of the American lobster, Homarus americanus. 1080 Jul 76


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