Biomedical subjects

J Vitorica

Publications and source records attributed to J Vitorica.

At least 19 recordsLinked to original sources

PubMed1997

GABAA receptor subunit expression changes in the rat cerebellum and cerebral cortex during aging.

Significant aging-related decreased expression of various GABAAR subunit mRNAs (alpha 1, gamma 2, beta 2, beta 3 and sigma) was found in both cerebellum and cerebral cortex using quantitative dot blot and in situ hybridization techniques. Contrary to the other subunits, the alpha 6 mRNA expression was significantly increased in the aged cerebellum. Parallel age-related changes in protein expression for gamma 2 and beta 2/3 (decrease) and alpha 6 (increase) were revealed in cerebellum by quantitative immunocytochemistry. However, no significant changes in alpha 1 protein expression nor in the number or affinity of [3H]zolpidem binding sites were detected in cerebellum even though alpha 1 mRNA expression was significantly decreased in the aged rat. Age-related increased expression of alpha 6 mRNA and protein in the cerebellum was accompanied by no significant changes in the number of diazepam-insensitive [3H]Ro15-4513 binding sites. In the cerebral cortex, no changes in the protein expression of the main GABAA receptor subunits (alpha 1, gamma 2 and beta 2/3) were observed which contrasted with the age-related decreased expression of the corresponding mRNAs. No significant changes in the number or affinity of [3H]zolpidem binding sites were observed in the cerebral cortex. Thus, age-related changes in the mRNA expression of a particular subunit does not necessarily lead to similar changes in protein or assembly into mature GABAA receptors. The results reveal the existence of complex regulatory mechanisms of GABAA receptor expression, at the transcriptional, translational and post-translational and/or assembly levels, which vary with the subunit and brain area.

Aging↗

PubMed1996

Molecular and pharmacological characterization of native cortical gamma-aminobutyric acidA receptors containing both alpha1 and alpha3 subunits.

We have investigated the existence, molecular composition, and benzodiazepine binding properties of native cortical alpha1-alpha3 gamma-aminobutyric acidA (GABAA) receptors using subunit-specific antibodies. The co-existence of alpha1 and alpha3 subunits in native GABAA receptors was demonstrated by immunoblot analysis of the anti-alpha1- or anti-alpha3-immunopurified receptors and by immunoprecipitation experiments of the [3H]zolpidem binding activity. Furthermore, immunodepletion experiments indicated that the alpha1-alpha3 GABAA receptors represented 54.7 +/- 5.0 and 23.6 +/- 3.3% of the alpha3 and alpha1 populations, respectively. Therefore, alpha1 and alpha3 subunits are associated in the same native GABAA receptor complex, but, on the other hand, these alpha1-alpha3 GABAA receptors from the cortex constitute a large proportion of the total alpha3 population and a relatively minor component of the alpha1 population. The pharmacological analysis of the alpha1- or alpha3-immunopurified receptors demonstrated the presence of two different benzodiazepine binding sites in each receptor population with high (type I binding sites) and low (type II binding sites) affinities for zolpidem and Cl 218,872. These results indicate the existence of native GABAA receptors possessing both alpha1 and alpha3 subunits, with alpha1 and alpha3 subunits expressing their characteristic benzodiazepine pharmacology. The molecular characterization of the anti-alpha1-anti-alpha3 double-immunopurified receptors demonstrated the presence of stoichiometric amounts of alpha1 and alpha3 subunits, associated with beta2/3, and gamma2 subunits. The pharmacological analysis of alpha1-alpha3 GABAA receptors demonstrated that, despite the fact that each alpha subunit retained its benzodiazepine binding properties, the relative proportion between type I and II binding sites or between 51- and 59-61-kDa [3H]Ro15-4513-photolabeled peptides was 70:30. Therefore, the alpha1 subunit is pharmacologically predominant over the alpha3 subunit. These results indicate the existence of active and nonactive alpha subunits in the native alpha1-alpha3 GABAA receptors from rat cortex.

Amino Acid Sequence↗

PubMed1996

Age-related modifications on the GABAA receptor binding properties from Wistar rat prefrontal cortex.

In the present communication we have investigated the pharmacological properties of the GABAA receptor from adult (3 months old) and aged (24 months old) Wistar rat prefrontal cortex. The prefrontal cortex is implicated in cognitive functions and stress and both processes seem to be altered during aging. These changes could be mediated by modifications in the GABAA receptor properties. Our results indicated the absence of generalized age-related modifications on the pharmacological properties of the GABAA receptor from prefrontal cortical membranes. Saturation experiments using the non-selective benzodiazepine [3H]flunitrazepam revealed that neither the Kd values or the Bmax were modified during aging. Moreover, Cl 218 872 displacement of [3H]flunitrazepam showed no age-related modifications on either the Kis or the relative proportion between the Type I and Type II benzodiazepine binding sites. Therefore, the benzodiazepine binding sites are well preserved in aged prefrontal cortex. On the other hand, saturation experiments using the GABA agonist [3H]muscimol demonstrated in the Bmax of the low affinity [3H]muscimol binding sites in aged rats (4.3 +/- 0.8 pmol/mg protein vs. 2.3 +/- 0.2 pmol/mg protein in adult and aged rats, respectively). However, no age-dependent modifications were observed in the allosteric interaction between GABA and benzodiazepine binding sites. These results demonstrate that the benzodiazepine binding sites and the GABA binding sites of the GABAA receptor complex from rat prefrontal cortical membranes are differentially affected by the aging process.

Aging↗

PubMed1996

Aging-related subunit expression changes of the GABAA receptor in the rat hippocampus.

Aging-related changes in the subunit expression of some hippocampal GABAA receptors have been found. Quantitative in situ hybridization has revealed that alpha 1, subunit messenger RNA expression was significantly increased in the hippocampus (34%) of old rats. The largest increases were observed in the dentate gyrus (76%) and in the CA1 field (30%). Quantitative immunocytochemistry also showed increased protein expression of the alpha 1 subunit in the dentate gyrus (19%) and CA1 (14%) of old rats. The increased alpha 1 messenger RNA and protein expression led to increased proportions of assembled GABAA receptors that contained alpha 1 subunits, as revealed by quantitative immunoprecipitation of (3H)flunitrazepam and (3H)muscimol binding. In contrast, there were no significant changes in the expression of beta 2, beta 3 and total gamma 2 (gamma 2S + gamma 2L) subunits, although a slightly increased expression of gamma 2L peptide was detected in the hippocampus proper (7%), but not in the dentate gyrus. The results are consistent with the notion that in the rat hippocampus there is an aging-related change in the subunit composition of some GABAA receptors.

Aging↗

PubMed1995

Implication of dopamine transporter system on 1-methyl-4-phenylpyridinium and rotenone effect in striatal synaptosomes.

The neurotoxic effect of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) seems to be produced by the inhibition of the respiratory chain by its metabolite 1-methyl-4-phenylpyridinium ion (MPP+). At the same time, its specific selectivity seems to be related especially to the dopamine uptake system. However, it is possible that other specific differences in dopaminergic neurons at the nigrostriatal system, such as constitutive metabolic deficiencies or other differences related to the energy capacity, could determine the greater vulnerability to MPP+. We have addressed this point by studying the effect of MPP+ and different inhibitors of the respiratory chain (rotenone, antimycin A and KCN) on the maximal respiratory rate from both synaptosomes and isolated synaptosomal mitochondria from different brain areas, i.e. cortex, hippocampus and striatum, and in isolated liver mitochondria. The results demonstrate the absence of differences in the effect of the inhibitors in isolated mitochondria. In contrast, a greater inhibition was found in striatal synaptosomes than in cortical or hippocampal synaptosomes when MPP+ and rotenone were used. Moreover, nomifensine or 1-[2-[bis(4-fluorophenyl)methoxy]ethyl]-4-(3-phenylpropyl) piperazine dihydrochloride (GBR-12909), inhibitor of the dopamine uptake system, has a protective effect in both cases. Our study indicates the great importance of the dopamine uptake system in the vulnerability of the dopamine striatum system. Moreover, our results show the low selectivity of this dopamine uptake system that is able to transport actively compounds with different chemical structures such as dopamine, MPP+ and rotenone.

1-Methyl-4-phenylpyridinium↗

PubMed1995

Aging-associated changes in the pharmacological properties of the benzodiazepine (omega) receptor isotypes in the rat hippocampus.

The aging-associated changes in hippocampal benzodiazepine (omega) receptor isotypes have been investigated in rats of the Wistar and Fischer 344 strains. Displacement experiments of [3H]flunitrazepam binding by zolpidem demonstrated that in hippocampal membranes from adult (3-month-old) Wistar strain rats, high (type I; omega 1)-, intermediate (type IIM; omega 2)-, and low (type IIL; omega 5)-affinity sites for this imidazopyridine account for 27.1 +/- 7.5, 44.2 +/- 7.5, and 28.8 +/- 5.1%, respectively. In hippocampal membranes from aged (24-month-old) rats of the same strain, the relative abundance of these sites was 42.8 +/- 9.3, 26.3 +/- 4, and 36.0 +/- 5.9%, respectively. In contrast, no significant difference was observed in the whole benzodiazepine (omega) binding site density between adult and aged rats. The increase in type I (omega 1) binding site density in the hippocampus of aged rats was also demonstrated in saturation experiments with [3H]zolpidem. This aging-induced increase in [3H]zolpidem binding was also observed in hippocampal membranes from Fischer 344 rats. Moreover, in both rat strains, GABA induced a greater enhancement of [3H]zolpidem (5 nM) binding to type I (omega 1) sites (GABA shift) in aged than in adult hippocampal membranes. Quantitative autoradiographic analysis of [3H]zolpidem binding to coronal brain sections from adult and aged Fischer 344 rats demonstrated that the aging-associated increases in the density of type I (omega 1) binding sites were restricted to the hippocampus. Moreover, increases in binding density were larger in the dentate gyrus and in the CA2 field than in the CA1 and CA3 fields.

Aging↗

PubMed1994

Molecular heterogeneity of the type I GABAA/benzodiazepine receptor complex.

We have investigated the presence of alpha 1, gamma 2 and beta 2-3 subunits as part of the type I (high affinity for [3H]zolpidem) GABAA/benzodiazepine receptor from rat cerebral cortex. Three subunit specific antibodies have been used (anti-alpha 1, to the C-terminal of rat alpha 1 subunit; anti-gamma 2, to the large intracellular loop of the gamma 2 subunit short form and the monoclonal 62-3G1, specific to beta 2-3 subunits) in immunoprecipitation experiments of the [3H]zolpidem binding activity or the 51,000 Da [3H]Ro 15-4513 photoaffinity labeled peptide (P51). The results indicated that alpha 1 subunit was present in the whole population (90%) of the GABAA/benzodiazepine receptors with high affinity for [3H]zolpidem. We cannot exclude the presence of other alpha subunits co-localized with alpha 1. On the one hand, 70-75% of type I GABAA/benzodiazepine receptor from rat cortex have co-existing alpha 1, beta 2-3 and gamma 2 subunits. We call this type Ia. On the other hand, 20-25% of the type I GABAA/benzodiazepine receptor complex should be constructed by the association of alpha 1 with subunits other than beta 2-3 and gamma 2. We call this type Ib. The identity of these subunits is currently unknown.

Animals↗

PubMed1994

Differential expression of the short and long forms of the gamma 2 subunit of the GABAA/benzodiazepine receptors.

The distribution of the mRNAs encoding the gamma 2S and gamma 2L subunits of the GABAA receptor in the rat brain has been revealed by in situ hybridization, northern blot and dot blot analysis using specific antisense oligonucleotides. In addition, the quantitative distribution of the gamma 2S and gamma 2L subunit peptides participating in the fully assembled GABAA receptors/benzodiazepine receptors has been mapped by immunoprecipitation with specific anti-gamma 2S and anti-gamma 2L antibodies. Several neuronal types and brain regions are enriched in gamma 2L such as neurons of the layer II of striate cortex and cerebellar Purkinje cells as well as the inferior colliculus, superior colliculus, deep cerebellar nuclei, medulla and pons. Other neuronal types and regions are enriched in gamma 2S such as the mitral cells of the olfactory bulb, pyramidal neurons of the pyriform cortex, layer VI of the neocortex, granule cells of the dentate gyrus and pyramidal cells of the hippocampus. Other cortical areas and cerebellar granule cells express both gamma 2S and gamma 2L in comparable amounts. There is a good correlation between the relative expression of gamma 2S and gamma 2L mRNAs and the relative presence of these protein subunits in fully assembled and mature receptors in the studied brain regions. The differential distribution of gamma 2S and gamma 2L might result in differential ethanol sensitivity of the neurons expressing these GABAA receptor subunits.

Animals↗

PubMed1994

Molecular characterization of type I GABAA receptor complex from rat cerebral cortex and hippocampus.

The molecular composition of the native gamma-aminobutyric acidA (GABAA) receptor complex is actually unknown. In the present communication we report a novel approach to characterize the minimal molecular conformation of the native GABAA receptor complex. This novel approach is based on the combination of subunit specific antibodies and specific 3H-labeled ligands in immunoprecipitation experiments. We have determined the presence of beta 2/3 and gamma 2 subunits in the Type I GABAA receptor complex, from rat cerebral cortex and hippocampus, by using two antibodies, the monoclonal 62-3G1 (specific for beta 2/3) and the polyclonal anti-gamma 2 (to the large intracellular loop of the gamma 2 short form) together with the Type I-specific ligand [3H]zolpidem. The association of gamma 2 and beta 2/3 subunits with the GABAA receptor complex was also tested using [3H]flumazenil. The results indicated that both gamma 2 and beta 2/3 were the most abundant subunits associated to either Type I or total benzodiazepine receptors from both cortex and hippocampus. Between 70-80% of Type I or total benzodiazepine binding activity was immunoprecipitated by either antibody. In addition, we have also investigated the coexistence of both subunits as part of the same population of Type I GABAA receptor complex by cross-immunoprecipitation experiments with 62-3G1 and anti-gamma 2. The results indicated that, in cerebral cortex, both gamma 2 and beta 2/3 subunits were part of the same population of Type I receptors. In hippocampus, an additional 20% of Type I receptors displayed either gamma 2 or beta 2/3 but not both subunits.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1993

Absence of modifications of the pharmacological properties of the GABAA receptor complex during aging, as assessed in 3- and 24-month-old rat cerebral cortex.

We have investigated the possible modifications of the pharmacological properties of Type I and Type II benzodiazepine binding sites of the gamma-aminobutyric acidA (GABAA) receptor complex in cortical membranes from 3- and 24-month-old Wistar or Fischer rats. No major changes were found in the binding parameters of [3H]zolpidem (a Type I-specific ligand) or [3H]flunitrazepam (a non-selective benzodiazepine). Neither the Kd values nor the Bmax for either ligand were modified during aging in cortical membranes from Wistar or Fischer rats. Consequently, the proportion of Type I binding sites was also unmodified in aged cortical membranes. The absence of modifications of Type I and Type II binding sites was also confirmed by Cl 218,872 displacement of [3H]flunitrazepam binding in aged cortical membranes from Wistar rats. Furthermore, the [3H]muscimol binding and the allosteric interactions between Type I or total benzodiazepine binding sites and GABA binding sites also remained unaltered with aging in cortical membranes from Wistar rats. Moreover, the pattern and proportion of the [3H]flunitrazepam photoaffinity labeled peptides were also unmodified by aging. These results demonstrate the absence of modifications in Type I or total benzodiazepine binding sites of the GABAA receptor complex from adult and aged cortical membranes in Fischer or Wistar rats.

Affinity Labels↗

PubMed1993

Comparative autoradiographic distribution of central omega (benzodiazepine) modulatory site subtypes with high, intermediate and low affinity for zolpidem and alpidem.

Pharmacological characterization of [3H]benzodiazepine binding to membrane preparations of adult rat hippocampus and neonatal rat brain have demonstrated, in addition to the omega 1 and omega 2 populations of central omega benzodiazepine binding sites associated with GABAA receptors, the existence of binding sites with microM affinity for the imidazopyridines zolpidem and alpidem. In the present study we have investigated their comparative autoradiographic distribution using [3H]flumazenil as a ligand. In the neonatal rat CNS, the imidazopyridine derivatives zolpidem and alpidem were found to discriminate two [3H]flumazenil binding site populations with an IC50 value ratio of more than 200-fold. In the different regions investigated (spinal cord, striatum, neocortex and inferior colliculus) the low affinity component had IC50 values of 20-40 microM (zolpidem) and 5-15 microM (alpidem) and accounted for ca. 50% of the total binding site population. In the adult rat, these imidazopyridine derivatives displayed a greater displacing potency in the cerebellum (IC50 = 6 and 36 nM, respectively) than in the hippocampus (IC50 = 37 and 403 nM, respectively). In the cerebellum, [3H]flumazenil binding was fully displaced by 1 microM of either compound and Hill coefficients of displacement curves were close to unity. In the hippocampus, 25% of [3H]flumazenil binding were resistant to 3 microM zolpidem or 1 microM alpidem, but were displaced by 100 microM of either compound. CL 218,872 also displayed a greater displacing potency in the cerebellum (IC50 = 83 nM) than in the hippocampus (IC50 = 711 nM) but [3H]flumazenil binding in the hippocampus was fully displaced by 10 microM of this compound. In adult rat hippocampus, zolpidem and alpidem were found to discriminate between three central omega site subtypes which display high (IC50 = 31 and 6.1 nM, for these imidazopyridine derivatives. In contrast, CL 218,872 discriminated between omega 1 and omega 2 sites but not between two omega 2 receptor subpopulations. omega 1 sites were mainly localized in layer IV of the sensorimotor cortex, cerebellum, substantia nigra, olfactory bulb and inferior colliculus. omega 2I sites were present in the cortical mantle (with higher levels in the cingulate and olfactory than in the sensorimotor cortex) and in subcortical (hippocampus, hypothalamus and nucleus accumbens) limbic structures. In the hippocampus, hypothalamus, spinal cord and nucleus accumbens, omega 2L sites accounted for more than 25% of the specific [3H]flumazenil binding; the density of these sites was minor in the cortex and in most pyramidal and extrapyramidal system structures.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

PubMed1993

Regional differences in the enhancement by GABA of [3H]zolpidem binding to omega 1 sites in rat brain membranes and sections.

The potential heterogeneity in the allosteric coupling between GABA and omega 1 binding sites within the native GABAA receptor complex has been evaluated in the rat by measuring the enhancement by GABA of [3H]zolpidem binding to omega 1 site in membranes from three rat brain structures (neocortex, cerebellum and hippocampus) and brain sections. The maximal stimulatory effect of GABA was significantly higher (265 +/- 47%) in cortical membranes than in cerebellar (165 +/- 48%) or in hippocampal (118 +/- 17%) membranes. These differences are not due either to the presence of different amounts of residual GABA in the membrane preparations or to the labeling, in presence of GABA, of binding sites other than omega 1 since: (1) the pharmacological properties of the [3H]zolpidem binding sites were similar in the three regions; (2) the degree of allosteric enhancement was unrelated to the relative proportion of omega 1 sites in each structure; and (3) GABA did not increase the Bmax for [3H]zolpidem. Regional differences in the effect of 100 microM GABA on [3H]zolpidem binding were also observed by quantitative autoradiography. Regions where the strongest (3-4-fold) effects of GABA in [3H]zolpidem binding were observed included the substantia nigra, lateral geniculate body, olfactory tubercule and red nucleus. A large increase in [3H]zolpidem binding was also demonstrated in the cingulate and frontoparietal cortices with higher effects in deep (4.2-fold) rather than in superficial layers (3.3-fold). Heterogeneous subregional increases in [3H]zolpidem binding in the presence of GABA were quantified within the cerebellum, hippocampus and superior colliculus.(ABSTRACT TRUNCATED AT 250 WORDS)

Analysis of Variance↗

PubMed1992

Heterogeneity in the allosteric interaction between the gamma-aminobutyric acid (GABA) binding site and three different benzodiazepine binding sites of the GABAA/benzodiazepine receptor complex in the rat nervous system.

In the present communication we have investigated the allosteric coupling between the gamma-aminobutyric acidA (GABAA) receptor and the pharmacologically different benzodiazepine (BZD) receptor subtypes in membranes from various rat nervous system regions. Two types of BZD receptors (type I and type II) have been classically defined using CL 218.872. However, using zolpidem, three different BZD receptors have been identified by binding displacement experiments in membranes. These BZD receptor subtypes displayed high, low, and very low affinity for zolpidem. The distribution of the high- and low-affinity binding sites for zolpidem was similar to that of type I and type II subtypes in cerebellum, prefrontal cortex, and adult cerebral cortex. On the other hand, the very-low-affinity binding site was localized in relative high proportion in spinal cord, hippocampus, and newborn cerebral cortex and, to a minor extent, in superior colliculus. The allosteric coupling between the GABAA receptor and the BZD receptor subtypes was different. The high- and low-affinity binding sites for zolpidem seemed to have a similar high degree of coupling, except in spinal cord. On the other hand, the very-low-affinity binding site for zolpidem displayed a low degree of coupling with the GABAA receptor. These results seem to indicate that the different efficacy of GABA in enhancing the [3H]flunitrazepam binding could be due to the different BZD receptor subtypes present in the GABAA/BZD receptor complex and, moreover, led us to speculate that the low GABA efficacy found in membranes from spinal cord, hippocampus, and newborn cerebral cortex might be due to the presence in relatively high proportion of the very-low-affinity binding site for zolpidem.

Allosteric Site↗

PubMed1991

Purification of the gamma-aminobutyric acidA/benzodiazepine receptor complex by immunoaffinity chromatography.

The bovine gamma-aminobutyric acidA/benzodiazepine receptor complex has been purified by a novel immunoaffinity chromatography method on immobilized monoclonal antibody 62-3G1. Immunopurification of the complex was achieved in a single step with an improved yield over affinity chromatography on the benzodiazepine Ro 7-1986/1. High-resolution sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) of the immunoaffinity-purified receptor revealed three major peptide bands of 51,000, 55,000, and 57,000 Mr which were also present in the Ro 7-1986/1 affinity-purified receptor. Peptide mapping, immunoblotting with subunit specific antibodies, and photoaffinity labeling with [3H]flunitrazepam and [3H]muscimol have been used for the identification of receptor subunits, including several which comigrated in a single band in SDS-PAGE.

Affinity Labels↗

PubMed1991

Pharmacologic characterization of GABAA/benzodiazepine receptor in rat hippocampus during aging.

We have characterized the pharmacologic properties of the gamma-aminobutyric acid (GABAA)/benzodiazepine receptor complex in hippocampal membranes from 3-month- and 24-month-old Wistar rats. No major changes were found in [3H]flunitrazepam or [3H]muscimol binding parameters. Neither the dissociation constant(s) nor the Bmax for either ligand seemed to be modified during aging in hippocampus. Furthermore, the allosteric interaction between the barbiturates binding site and the GABA binding site, determined by pentobarbital stimulation of [3H] muscimol binding, remained unaltered. However, there was a significant increase with aging in the efficacy of the GABA-enhancement of [3H]flunitrazepam binding. On the other hand, we have also detected a significant increase in the proportion of type I benzodiazepine receptor in 24-month-old hippocampal membranes. We propose that the age-related increase in the efficacy of GABA-enhancement of [3H]flunitrazepam binding could be correlated with the increase in the proportion of type I benzodiazepine receptor. Based on these results it is tempting to speculate that the age-dependent modifications on the GABAA/benzodiazepine receptor might reflect an age-dependent neuronal degeneration of the hippocampus or the hippocampal formation.

Aging↗

PubMed1990

The GABAA/benzodiazepine receptor complex in rat brain neuronal cultures. Characterization by immunoprecipitation.

The expression of the GABAA/benzodiazepine receptor (GABAR/BZDR) complex in primary neuronal cultures from rat brain embryos has been investigated. The GABAR/BZDR complex was photoaffinity labeled with [3H]flunitrazepam [3H]FNZ and immunoprecipitated with subunit specific antibodies. These were the mAb 62-3G1 which is specific for the 57-kDa GABA binding subunit, and the rabbit antiserum A which recognizes the 51-kDa [3H]FNZ binding subunit. The results indicate that the cultured neurons express 5 different peptides of 51, 53, 54, 57 and 59 kDa that can be photoaffinity labeled with [3H]FNZ and that all of them are physically coupled to the GABAA receptor. Most of the [3H]FNZ photolabeled peptides have similar mobilities to those found in the brain of the newborn rat. Nevertheless, some of the quantitative changes in the photolabeled peptides observed during the normal development of the rat brain were not observed or occurred at much slower pace in the cultured neurons.

Affinity Labels↗

PubMed1990

Characterization with antibodies of the gamma-aminobutyric acidA/benzodiazepine receptor complex during development of the rat brain.

The postnatal development of the gamma-aminobutyric acidA/benzodiazepine receptor (GABAR/BZDR) complex of the rat brain has been investigated using the monoclonal antibody 62-3G1 and the polyclonal rabbit antiserum A, specific for the 57,000 and 51,000 Mr receptor subunits, respectively. Both GABAR and BZDR binding activities co-precipitated during all postnatal ages. Adult rats showed a main 51,000 Mr[3H]flunitrazepam photoaffinity-labeled peptide, whereas newborn rats showed several photolabeled peptides of higher Mr. All the photolabeled peptides could be immunoprecipitated with each antibody regardless of the age of the rats. These results suggest that the physical coupling between the GABAR and the BZDR is already present in newborn animals and it is maintained afterwards during development. Glycosidase and peptidase treatments of the immunoprecipitated GABAR/BZDR complex indicated that all the [3H]flunitrazepam-photolabeled subunits are different peptides, although they seem to conserve a high degree of homology. In addition to the age-dependent heterogeneity, the results also suggest that for each age, there is heterogeneity in the subunit composition of the GABAR/BZDR complex.

Acetylglucosaminidase↗

PubMed1990

Rapid postnatal developmental changes in the passive proton permeability of the inner membrane in rat liver mitochondria.

Titration of mitochondrial respiration against the membrane potential with the inhibitor malonate has been carried out during the perinatal period in isolated rat liver mitochondria. Neonatal and adult mitochondria exhibited the characteristic "nonohmic" behavior for the proton conductance (CmH+). In contrast, fetal mitochondria exhibited an "anomalous" "ohmic" behavior for CmH+. The calculated passive proton permeability of the membrane undergoes a profound reduction during the first postnatal hour. The results reported demonstrate that the hypothesis [Pollak, J.K. & Sutton, R. (1980) Trends Biochem. Sci. 5, 23-27] of the existence of a "leaky" mitochondria in the fetal rat liver, and of its sudden neonatal change towards a state of higher energy conservation of the proton electrochemical gradient, is correct.

Animals↗