Biomedical subjects

S Mertens

Publications and source records attributed to S Mertens.

At least 19 recordsLinked to original sources

PubMed2001

Hemodynamic changes with right lateral decubitus body positioning in the tilted porcine heart.

BACKGROUND: In beating-heart coronary surgical procedures, exposure of posterior vessels through sternotomy causes cardiac function to deteriorate. We hypothesized that turning the subject to the right lateral decubitus position before cardiac retraction improves exposure of posterior vessels and preserves cardiac pump function on displacement. METHODS: Eight 80-kg open-chest pigs were instrumented with catheter-tip manometers. After a stepwise 60-degree turn to the right lateral decubitus position of the body, the heart was retracted anteriorly to 90 degrees with a suction stabilizer. RESULTS: Right lateral body positioning caused an approximately 45-degree right deviation of the apex, thereby exposing the left atrial groove. Stroke volume, mean arterial pressure, right atrial pressure, and right ventricular end-diastolic pressure increased to 106% +/- 5% (mean +/- standard error of the mean, p = 0.31), 106% +/- 3% (p = 0.01), 129% +/- 8% (p = 0.001), and 171% +/- 14% (p = 0.002), respectively, compared with control values. In contrast, left atrial pressure decreased to 73% +/- 6% (p = 0.007), whereas left ventricular preload remained unchanged (110% +/- 8%, p = 0.26). Additional anterior displacement to 90 degrees fully exposed the posterior vessels, and stroke volume decreased to 90% +/- 3% (p = 0.01) and mean arterial pressure to 93% +/- 5% (p = 0.07) at the expense of further increased right ventricular preload (256% +/- 28%, p < 0.001). CONCLUSIONS: By placing the subject in the right lateral decubitus position, exposure through sternotomy of posterior vessels in the beating porcine heart was facilitated while mean arterial pressure was maintained.

Animals↗

PubMed2000

Random costs in combinatorial optimization

The random cost problem is the problem of indentifying the minimum in a list of random numbers. By definition, this problem cannot be solved faster than by exhaustive search. It is shown that a classical NP-hard optimization problem, number partitioning, is essentially equivalent to the random cost problem. On the one hand this explains the bad performance of heuristic approaches to the number partitioning problem, but on the other hand it allows one to calculate the probability distributions of the optimum and suboptimum costs.

Journal Article↗

PubMed2000

Effect of perfluorodecalin on human retinal pigment epithelium and human corneal endothelium in vitro.

BACKGROUND: Perfluorocarbon liquids are useful intraoperative tools in complicated vitreoretinal surgery. They are usually removed at the end of the procedure, but small amounts may remain in the eye. Recently, contradictory results have been reported on the damage in association with residual perfluorocarbon liquids in the eye. This study examined the effects of perfluorodecalin on human retinal pigment epithelium and corneal endothelium in vitro. METHODS: Vitality and proliferative capacity of cell cultures were measured after incubation with perfluorodecalin. Vitality of cell cultures were measured using the Life-Dead assay. Cell proliferation was determined by measuring incorporation of 5-bromo-2'-deoxyuridine into cellular DNA. Furthermore, endothelium of organ-cultured human corneas was examined after incubation with perfluorodecalin by photodocumentation. RESULTS: Both cell types showed less extinctions in the Life-Dead assay after incubation with perfluorodecalin. After removing perfluorodecalin from the cultures, cells showed the same capacity of proliferation as the control cells. Compared to control corneas, perfluorodecalin induced a decrease in endothelial cell density. In four corneas, endothelial cell necrosis was observed. CONCLUSION: Decreasing extinctions in the Life-Dead assay after incubation with perfluorodecalin can be interpreted as showing a decreasing amount of vital cells. Because cell proliferation showed no significant changes the results suggest that perfluorodecalin may not be directly toxic to cells in vitro. It may exert an indirect or mechanical effect on cell function by impeding the normal metabolic exchange between endothelium and medium. Based on these results perfluorodecalin should be completely removed after operation.

Cell Count↗

PubMed1999

A conformational switch at the 3' end of a plant virus RNA regulates viral replication.

3' untranslated regions of alfamo- and ilar-virus RNAs fold into a series of stem-loop structures to which the coat protein binds with high affinity. This binding plays a role in initiation of infection ('genome activation') and has been thought to substitute for a tRNA-like structure that is found at the 3' termini of related plant viruses. We propose the existence of an alternative conformation of the 3' ends of alfamo- and ilar-virus RNAs, including a pseudoknot. Based on (i) phylogenetic comparisons, (ii) in vivo and in vitro functional analyses of mutants in which the pseudoknot has been disrupted or restored by compensatory mutations, (iii) competition experiments between coat protein and viral replicase, and (iv) investigation of the effect of magnesium, we demonstrate that this pseudoknot is required for replication of alfalfa mosaic virus. This conformation resembles the tRNA-like structure of the related bromo- and cucumo-viruses. A low but specific interaction with yeast CCA-adding enzyme was found. The existence of two mutually exclusive conformations for the 3' termini of alfamo- and ilar-virus RNAs could enable the virus to switch from translation to replication and vice versa. The role of coat protein in this modulation and in genome activation is discussed.

3' Untranslated Regions↗

PubMed1997

Wolfram (DIDMOAD) syndrome and Leber hereditary optic neuropathy (LHON) are associated with distinct mitochondrial DNA haplotypes.

Because Wolfram (or DIDMOAD) syndrome is supposed to be a mitochondrial (mt)-mediated disease, we investigated a group of eight DIDMOAD patients with respect to point mutations of the mtDNA thus far described as being associated with defined mitochondrial disorders such as MELAS, MERRF, and LHON. Furthermore, to screen DIDMOAD patients for other mtDNA defects we used Southern blot analysis to detect mtDNA length mutations and rearrangements as well as PCR-SSCP and direct sequencing to screen all ND genes (complex I of the respiratory chain), the 22 tRNAs, and a part of the cyt b gene for unknown mutations. As a disease control group, 17 LHON patients (harboring one of the primary LHON mutations) were included in this study because of the overlapping clinical symptoms (optic atrophy) in both syndromes. We compared mtDNA variants identified in DIDMOAD patients with those found in LHON patients as well as in a control group consisting of 67 healthy German blood donors. In total, the control group was characterized by 29 polymorphic sites in ND and tRNA genes that define certain major Caucasian haplotypes. We found that a cluster of nucleotide exchanges at nucleotide positions (nps) 4216 and 11,251 roughly discriminates controls (12/67 controls, 18%) from the disease groups (6/8 DIDMOAD patients, 75%; 10/17 LHON patients, 59%). All 4216-positive LHON patients (10 patients) were concentrated in a haplogroup defined by additional exchanges at nps 10,398, 12,612, and 13,708 (haplogroup A), while the bulk of 4216-positive DIDMOAD patients (5 patients) were found in a distinct haplogroup consisting of nucleotide exchanges at nps 4917, 10,463, 13,368, 14,233, and 15,928. The frequencies of both haplogroups were significantly lower in the control group versus the respective disease groups. A more detailed analysis was performed by sequencing the two hypervariable regions of the non-coding D-loop region from patients and controls and corroborated the ranging in the two major haplogroups. Thus, the different clinical features of the mitochondrial disease groups investigated here corresponded to different clusters of mtDNA variants, which might act as predisposing haplotypes, increasing the risk for disease.

Adult↗

PubMed1997

Population genetics and disease susceptibility: characterization of central European haplogroups by mtDNA gene mutations, correlation with D loop variants and association with disease.

Mitochondrial (mt)DNA haplogroups in a German control group (n = 67) were characterized by screening mitochondrial coding regions encompassing most of the ND, tRNA and cyt b genes. We used a PCR-SSCP screening approach followed by direct sequencing of polymorphic mtDNA fragments. Five major mtDNA lineages, diverging in at least nine different haplogroups, could be defined by characteristic polymorphic sites in mitochondrial genes. Additional sequencing of two hypervariable segments (HVS-I and II) of the non-coding displacement (D) loop in all control subjects revealed that certain D loop variants were strongly correlated with lineages and haplogroups, while others represented hotspots occurring frequently in different haplogroups. The existence of identified lineages and haplogroups received support from data in the literature, obtained by use of different approaches. Subsequently, we investigated four disease groups for association with these haplogroups: (i) LHON patients (n = 55) carrying at least one of the primary/intermediate LHON mutations at nt 3460, 11778, 14484 and/or 15257; (ii) patients suffering from Wolfram or DIDMOAD syndrome (n = 8); (iii) MELAS patients (n = 9); (iv) a group of children, who died from 'sudden infant death syndrome' (SIDS) (n = 9). The distribution patterns among the haplogroups of the disease groups (LHON, DIDMOAD and SIDS) differed considerably from the control population. LHON and DIDMOAD were significantly under-represented in the most frequent German haplogroup DC, but were concentrated in a mtDNA lineage defined by polymorphisms at nt 4216 + 11251 + 16126. As this lineage diverged into two precisely defined haplogroups, LHON and DIDMOAD could be assigned to the two haplogroups separately. Strikingly, SIDS was often found in association with two rare German haplogroups. MELAS patients were equally distributed among German haplogroups and, moreover, did not reveal any accumulation of specific D loop variants. We conclude that certain European mtDNA haplogroups define a genetic susceptibility basis for various disorders.

Child↗

PubMed1996

SAP kinase-3, a new member of the family of mammalian stress-activated protein kinases.

Stress-activated protein kinases are MAP kinase homologues that are activated by cellular stresses, bacterial endotoxin and inflammatory cytokines. They are activated by a dual threonine/tyrosine phosphorylation within a TPY sequence in the case of stress-activated protein kinase-1 and its isoforms (also called JNKs) or a TGY sequence in the case of stress-activated protein kinase-2 and its isoforms (also called p38, p40, RK, CSBPs, XMpk2 and Mxi2). Here we report the cloning and sequencing of a new protein kinase from rat with a TGY sequence in the activation domain. This stress-activated protein kinase-3 is 60% identical to mouse stress-activated protein kinase-2 and 45% identical to HOG1 from Saccharomyces cerevisiae. Transcripts encoding stress-activated protein kinase-3 are widely expressed, with high levels in skeletal muscle.

Amino Acid Sequence↗

PubMed1993

Molecular distinction of three N-methyl-D-aspartate-receptor subtypes in situ and developmental receptor maturation demonstrated with the photoaffinity ligand 125I-labeled CGP 55802A.

Activation of N-methyl-D-aspartate (NMDA) receptors is essential for synaptic plasticity in the central nervous system and contributes to neuronal death under various pathological conditions. Although several subunits have been cloned, the structure of NMDA receptors in situ is unresolved. By using a photoreactive antagonist with nanomolar affinity to the NMDA-binding site, three types of receptors were differentiated by their pattern of photoaffinity-labeled subunits. In adult brain, a protein of 175-kDa was photoreactive that displayed a profile of ligand binding and autoradiographical distribution corresponding to NMDA receptors. In contrast, in early postnatal brain, proteins of both 175 kDa and 115 kDa were photolabeled. This labeling pattern is switched to that of adult brain around postnatal day 10, pointing to a structural maturation of NMDA receptors. A third type of receptor could be identified in cerebellar granule cell cultures, where NMDA receptors mediate trophic effects and photolabeling was exclusively targeted to a 115-kDa protein. To identify the proteins labeled in situ, recombinant receptors were subjected to photolabeling. When the NR1 subunit was coexpressed with either the NR2A, NR2B, or NR2C subunit, only the combination of NR1/NR2A was photoreactive. Both the NR1 and NR2A subunits were photolabeled, corresponding in size to the proteins labeled in situ. However, the lack of subunit-selectivity in photolabeling the NR1/NR2A combination suggests the presence of additional receptor components in situ to explain the subunit-selective photoreactivity in adult brain (175 kDa) and in cerebellar granule cells (115 kDa). The subunit combination NR1/NR2A by itself appears insufficient to describe a major population of NMDA receptors, in particular, in adult brain.

Affinity Labels↗

PubMed1993

GABAA receptor populations with novel subunit combinations and drug binding profiles identified in brain by alpha 5- and delta-subunit-specific immunopurification.

The pharmacological significance and structural basis of the gamma-aminobutyrate (GABAA) receptor heterogeneity was investigated in situ by an immunobiochemical analysis of receptor populations characterized by the delta- and alpha 5-subunit. Using an antiserum specific for the delta-subunit, a population of GABAA receptors (21 +/- 2% of solubilized receptors) was immunoprecipitated from rat brain extracts which contained high affinity benzodiazepine binding sites. They were distinguished from those immunoprecipitated by the alpha 1- and alpha 3-subunit antisera by a 4-5-fold and 5-10-fold higher affinity for diazepam and beta CCM, respectively. Using the delta-antiserum in immunoaffinity chromatography the delta-subunit was found to be associated with the alpha 1-, alpha 3-, beta 2/3-, and gamma 2-subunits, suggesting that the latter conveys benzodiazepine receptor sensitivity also to GABAA receptors containing the delta-subunit. The receptor population immunoprecipitated by the alpha 5-subunit antiserum (10 +/- 2% of receptors solubilized from whole brain extracts) was characterized by affinities for zolpidem, beta CCM, and CL 218872 which distinguished it from all other known receptor populations. The alpha 5-subunit was associated with the alpha 1-, alpha 3-, beta 2/3- and gamma 2-subunits pointing to differential subunit combinations. Indeed, when receptor populations were immunoprecipitated by the alpha 5-subunit antiserum from different brain regions, zolpidem displayed striking differences in affinity pointing to the role of subunits other than alpha 5 in determining receptor affinity.

Animals↗

PubMed1993

Enoximone therapy as pharmacological bridging to cardiac transplantation.

Keeping pre-transplant patients alive while waiting for a suitable donor is still a major challenge. New pharmacological agents which can provide improved hemodynamics are urgently needed in patients with severe heart failure who are on the waiting list for cardiac transplantation. Intravenous enoximone therapy (an initial 0.5 mg/kg bolus, then 1.25-5.0 mcg/kg/min infusion) was administered to 35 transplant candidates with progressive heart failure despite optimal drug regimen including digoxin, diuretics, and ACE-inhibitors. In 18 out of 35 patients complete hemodynamic, echocardiographic, neurohumoral, and Holter-ECG studies were performed before and 24 hours after intravenous enoximone infusion. Patients were then continued on chronic oral therapy of 100 mg twice a day. Enoximone infusion increased the cardiac index (CI) (1.78 +/- 0.45 l/min/m2 vs 3.04 +/- 0.83 l/min/m2; p < 0.001) and stroke volume index (SVI)(22.33 +/- 9.45 ml/m2 vs 32.28 +/- 7.29 ml/m2; p < 0.05) and decreased wedge pressure (PCP)(24.1 +/- 11.98 mmHg vs 17.78 +/- 8.76 mmHg; p < 0.05) while mean arterial pressure (MAP) was unchanged. Left ventricular ejection time (LVET)(225.1 +/- 26.9 ms vs 242.2 +/- 25.8 ms; p < 0.05) was increased whereas other echocardiographic parameters were unchanged (Left ventricular end-diastolic dimension LVEDD, left ventricular end-systolic dimension LVESD, fractional shortening FS, early diastolic relaxation parameter Te). Plasma neurohumoral parameters did not change (Aldosterone, epinephrine, renin, atrial natriuretic factor) except for a significant drop in norepinephrine (936.7 +/- 443.2 pg/ml vs 522.4 +/- 287.6 pg/ml; p < 0.05). Holter-ECG parameters (ventricular premature beats VPB, couplets, ventricular tachycardia VT) were not influenced by enoximone infusion.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗

PubMed1992

Ischemia: reperfusion injury and restenosis after coronary angioplasty.

Percutaneous transluminal coronary angioplasty (PTCA) is a very effective technology that allows, without surgery, successful mechanical revascularization of acutely or chronically obstructed coronary arteries. The success of PTCA in patients with acute myocardial infarction or unstable angina is questioned by early coronary reocclusion and by so-called reperfusion injury. In a biochemical context, reperfusion injury occurs as a very complex interaction between the different tissues that build heart muscle. Free radicals play a pivotal role and initiate a deleterious cascade of events after reperfusion. Protective mechanisms such as superoxide dismutase, glutathione peroxidase, and catalase are normally present in the cell to prevent damage by free radicals. Endothelial cells have a greater number of specific physiologic and metabolic functions and influence the microcirculatory flow. In the presence of exogenous glucose, coronary endothelial cells show a pronounced lactate production under well-oxygenated conditions. Low energy demand and high glycolytic activity may be the cause of why the coronary endothelium is less severely injured than the cardiomyocytes in the ischemic and anoxic heart. The success of PTCA in patients with chronically obstructed coronary arteries (stable angina) is questioned by vessel occlusion and restenosis. Restenosis is a very complex process involving clinical, morphological, procedural, regional flow-dependent, and biological determinants. Early platelet deposition, formation of mural thrombus, coronary vasospasm, and elastic recoil forces of stretched vessel wall may contribute to early restenosis in the first days after PTCA, but the peak incidence of restenosis occurs between two and three months after PTCA. Intimal hyperplasia or proliferation of smooth muscle cells is believed to be the fundamental process of restenosis. To solve the problem of restenosis, much effort has been expended, which includes several technical and pharmacological approaches. Pharmacological strategies, systemically or locally administered, aim at increased vasomotor tone, platelet function, smooth muscle cell proliferation/migration, and fibrocollagenous healing. Up to now none of the proposed drugs has been able to reduce the restenosis rate. There is experimental evidence for a claim that the antioxidant functions of vitamins (E, C, and beta-carotene) may prevent restenosis post-PTCA. Until recently, in most post-PTCA restenosis trials the angiographic analyses were not performed using computerized measurement methods. In order to assess the efficacy of acute or long-term interventions on the natural course or acute complications of coronary artery disease, quantitative measures have been introduced and validated that make use of digital coronary angiography and computerized image processing techniques.(ABSTRACT TRUNCATED AT 400 WORDS)

Angioplasty, Balloon, Coronary↗

PubMed1992

Five subtypes of type A gamma-aminobutyric acid receptors identified in neurons by double and triple immunofluorescence staining with subunit-specific antibodies.

The extraordinary structural diversity of subunits forming type A gamma-aminobutyric acid (GABAA) receptors in the brain is expected to give rise to different modes of GABAergic synaptic inhibition and different profiles of modulatory drugs effective in anxiolytic, hypnotic, and antiepileptic therapy. To identify receptor subtypes in situ, the most prevalent subunits were visualized by double and triple immunofluorescence staining in rat brain, using polyclonal antibodies to the alpha 1, alpha 3, and gamma 2 subunits and a monoclonal antibody to locate both the beta 2 and the beta 3 subunit. At both cellular and subcellular levels five distinct patterns of subunit colocalization were identified: I, alpha 1 beta 2,3 gamma 2; II, alpha 3 beta 2,3 gamma 2; III, alpha 1 alpha 3 beta 2,3 gamma 2; IV, alpha 3 gamma 2; and V, alpha 1 alpha 3 gamma 2. As analyzed by confocal laser microscopy, different subunits displayed the same local variations of staining intensity ("hot spots") along the plasma membrane. The covisualized subunits appear therefore to be coassembled in receptor subtypes. Most neurons expressed only a single major receptor subtype with no apparent distinction between synaptic and extrasynaptic sites. However, in some neurons, most notably in Purkinje cells, the subunit composition varied between the soma and the dendrites, pointing to the existence of receptor heterogeneity within single neurons. Furthermore, different populations of neurons may be characterized by particular receptor subtypes. Cells displaying alpha 1-subunit immunoreactivity were mostly identified as GABAergic, whereas monoaminergic neurons displayed intense alpha 3-subunit immunoreactivity but virtually no alpha 1-subunit immunoreactivity. The allocation of defined GABAA receptor subtypes to identified neurons opens the way for a functional analysis of receptor heterogeneity.

Animals↗

PubMed1992

GABAA-receptor subtypes differing in alpha-subunit composition display unique pharmacological properties.

GABAA-receptor subtypes in rat brain were characterized using anti-peptide antisera specific for the alpha 1-, alpha 3- and alpha 5-subunits. While a high abundance of alpha 1-containing receptors was demonstrated by immunoprecipitation (80-90% of receptors), the receptors precipitated with the alpha 3- and the alpha 5-antiserum were less frequent (18-25% and 10-23%, respectively). The three receptor populations displayed unique pharmacological properties as shown by radioligand binding. Diazepam, flumazenil and flunitrazepam displayed similar displacing potencies in [3H]-flumazenil binding. However, the affinities of CL 218872, beta CCM and zolpidem were up to 10-fold lower in the alpha 3- than the alpha 1-receptor population while intermediate values were found for the alpha 5-receptor population. In many brain areas, a neuron-specific expression of receptors containing either alpha 1- or alpha 3-subunits could be visualized immunohistochemically. It will be of major interest to determine, whether ligands with differential affinities for receptor subtypes in situ will provide novel therapeutic profiles.

Animals↗

PubMed1991

Identification and immunohistochemical mapping of GABAA receptor subtypes containing the delta-subunit in rat brain.

Synaptic inhibition in brain is mainly mediated via GABAA receptors which display a striking structural heterogeneity. A novel type of GABAA receptor subunit, the delta-subunit, has recently been described based on molecular cloning of its cDNA. To identify the prevalence and distribution of GABAA receptors which contain the delta-subunit protein in situ, polyclonal site-directed antisera were developed against three synthetic peptides derived form the rat delta-subunit cDNA-sequence. All antisera specifically recognized a 54 kDa protein in GABAA receptor preparations. Nearly 30% of the GABAA receptors contained the delta-subunit immunoreactivity and displayed high affinity GABA and high affinity benzodiazepine binding sites as shown by immunoprecipitation. Receptors which contain the delta-subunit were immunohistochemically shown to be restricted to a few brain areas such as the cerebellum, thalamus and dentate gyrus of the hippocampal formation. Thus, those neurons which express GABAA receptors with a delta-subunit have now been visualized and made accessible for a functional analysis of this GABAA receptor subtype in situ.

Amino Acid Sequence↗

PubMed1991

GABAA receptors display association of gamma 2-subunit with alpha 1- and beta 2/3-subunits.

Recombinant GABAA (gamma-aminobutyrate-Type A) receptors that are sensitive to benzodiazepine receptor ligands can be generated by coexpression of alpha-, beta-, and gamma 2-subunit cDNAs (Pritchett, D. B., Sontheimer, H., Shivers, B. D., Ymer S., Kettenmann, H., Schofield, P. R., and Seeburg, P. H. (1989) Nature 338, 582-585; Pritchett, D. B., Lüddens, H., and Seeburg, P. H. (1989) Science 245, 1389-1392; Malherbe, P., Sigel, E., Baur, R., Perssohn, E., Richards, J. G., and Mohler, H. (1990) J. Neurosci. 10, 2330-2337). However, in brain tissue, only alpha- and beta-subunit proteins have so far been detected. To identify the size and distribution of the gamma 2-subunit protein in brain tissue, polyclonal antibodies were prepared against two synthetic peptides corresponding to amino acids 1-15 and 336-350 of the cDNA-derived rat gamma 2-subunit sequence. On Western blots, both anti-gamma 2-subunit antisera selectively labeled a 43-kDa protein. gamma 2-Subunit immunoreactivity was detected immunohistochemically in various brain regions, e.g. in the olfactory bulb, cerebral cortex, islands of Calleja, hippocampus, substantia nigra, and cerebellum. Immunoprecipitation with both antisera identified the gamma 2-subunit immunoreactivity in 40 and 50% of the native GABAA receptors purified from bovine and rat brains, respectively. Monoclonal antibody bd24 selectively recognizes the alpha 1-subunit, whereas bd17 recognizes both the beta 2- and beta 3-subunits (Ewert, M., Shivers, B. D., Lüddens, H., Mohler, H., and Seeburg, P. H. (1990) J. Cell Biol. 110, 2043-2048). Since either of these monoclonal antibodies (bd17 and bd24) precipitated approximately 90% of the GABAA receptors, the gamma 2-subunit is frequently associated with the alpha 1-subunit and the beta 2- and/or beta 3-subunit in vivo.

Amino Acid Sequence↗

PubMed1991

A computer controlled device to facilitate studies of the kinetics of ligand-binding: binding of diazepam to bovine brain membranes.

A device to facilitate kinetic receptor filtration assays is described. The receptor containing membranes and the labeled ligand are placed in two separate syringes and are rapidly mixed into a collecting syringe using a pneumatic ram. Shortly after the start of mixing, a pneumatically controlled valve switches the collecting syringe containing the receptor-ligand mixture to the filtration unit. Filtration is performed on glass/microfiber filters or equivalent by pushing the plunger of the collecting syringe by a stepper motor. A valve positioner controlling several valves allows the filtered membranes to be washed and dried by pressure in any user programmable sequence. Further filtration of the receptor-ligand mixture can be programmed at selected time points. The entire system is controlled by an IBM-PC. With this system, the association and dissociation of diazepam from crude bovine-brain membranes has been studied at 4 degrees C. The dissociation shows a biphasic pattern with half lifes of 1.3 and more than 23 minutes respectively. Association appears to be into a single compartment.

Animals↗

PubMed1991

Immunochemical identification of the alpha 1- and alpha 3-subunits of the GABAA-receptor in rat brain.

To identify subunit variants of the GABAA-receptor antisera were developed against specific cDNA-derived peptide sequences of the alpha 1- and alpha 3-subunits of rat brain. The alpha 1-subunit antiserum selectively recognized a protein of 50 +/- 1 kDa in rat and bovine GABAA-receptor preparations, while the alpha 3-subunit antiserum interacted with a protein doublet of 59 +/- 2 kDa and 61 +/- 3 kDa. The alpha 1-subunit immunoreactivity resides in a large population of GABAA-receptors as shown by immunoprecipitation of 63 +/- 6% of [3H]flumazenil binding sites with the alpha 1-subunit antiserum. In contrast, only 24 +/- 3% of receptor binding sites were precipitated with the alpha 3-subunit antiserum. Co-precipitation studies suggest that the alpha 1- and alpha 3-subunit immunoreactivities do not share the same receptor population while the gamma 2-subunit immunoreactivity is associated with the alpha 1-subunit immunoreactivity.

Amino Acid Sequence↗

PubMed1990

Identification of the gamma 2-subunit protein in native GABAA receptors in brain.

GABAA receptors with functional benzodiazepine receptors have been reconstituted by coexpression of alpha-, beta- and gamma-subunit cDNAs. In brain, proteins of the alpha- and beta-subunits, but not the gamma 2-subunit have been identified. Using an antipeptide antiserum, we now demonstrate that the gamma 2-subunit is a protein of 43 kDa. It is present in at least 50% of GABAA receptors, as shown by immunoprecipitation.

Amino Acid Sequence↗