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D Bertrand

Publications and source records attributed to D Bertrand.

At least 145 records · Page 8Linked to original sources
PubMed1992

Unconventional pharmacology of a neuronal nicotinic receptor mutated in the channel domain.

The putative channel-forming MII domains of the nicotinic, gamma-aminobutyric acid type A, and glycine receptors contain a highly conserved leucine residue. Mutation of this hydrophobic amino acid in the neuronal nicotinic receptor alpha 7 (Leu-247), reconstituted in Xenopus oocytes, modifies the ionic response to acetylcholine and alters desensitization. Furthermore, the Leu----Thr (L247T) mutant has two conducting states (46 pS and 80 pS), in contrast with the wild-type (WT) receptor, which has only one (45 pS). We now show that this mutant possesses a rather paradoxical pharmacology: antagonists of the WT receptor such as dihydro-beta-erythroidin, hexamethonium, or (+)-tubocurarine elicit ionic currents when applied to the L247T alpha 7 mutant and these responses are blocked by alpha-bungarotoxin. Furthermore, prolonged application of acetylcholine causes desensitization in the WT but leads to a potentiation of the responses to acetylcholine or dihydro-beta-erythroidin in the mutant. These data are consistent with a scheme in which mutation of Leu-247 renders a desensitized state in the WT channel a conducting state. They also strengthen the proposal that, in the WT, some competitive antagonists may stabilize desensitized states. Finally, these observations may shed light on properties of other ion channels, in particular the glutamate receptors, which display multiple conductance levels associated with various pharmacological agents.

Acetylcholine↗
PubMed1992

Stratified organization of the nicotinic acetylcholine receptor channel.

Mutations of leucine 247 within the M2 channel domain of the alpha 7 neuronal nicotinic receptor, confer electrophysiological and pharmacological properties, which allow one of the desensitized states to become conductive. Here we show that, in Xenopus oocytes, the effects of the mutations were preserved when 1,2-bis(2-aminophenoxy)-ethane N,N,N',N'-tetraacetic acid (BAPTA) was injected in the cytoplasm to block Ca(2+)-dependent chloride currents, and that in agreement with the proposed interpretation, the ionic currents do not desensitize and rise slowly, in the time-scale of seconds, upon agonist application. Interestingly, similar effects were observed when the two rings (T244, V251) neighbouring L247 on the alpha-helix, but not the more distant ones (S240, L254/255), were mutated, thus supporting the proposal of a functional stratification of the channel domain.

Affinity Labels↗
PubMed1992

[Multifactorial urodynamic study of 115 patients with dysuria and benign prostatic hypertrophy].

Discriminant analyses based on preoperative urodynamic parameters have proved inaccurate in predicting functional results after prostatectomy. To investigate the cause of this failure, we studied a group of 115 patients consecutively referred for prostatism and selected for prostatic surgery only on clinical criteria and flow rate. A preoperative urodynamic work-up was performed, including cystometry and urethrometry. The patients were reexamined 2 months postoperatively and underwent control uroflowmetry. All preoperative urodynamic data were computed simultaneously using principal component analysis and canonical discriminant analysis. The significance of the diagrams obtained with these multifactorial analyses was more closely examined than in previous studies. The results of classification by the discriminant function demonstrating the best combination of preoperative urodynamic variables resulted in 44% of patients being correctly rated. Examination of the diagrams showed that the major cause of failure in previous studies was the great variety of urodynamic conditions in men with prostatism, which indicates a need for multifactorial interpretation of the results of urodynamic explorations.

Aged↗
PubMed1991

Functional significance of aromatic amino acids from three peptide loops of the alpha 7 neuronal nicotinic receptor site investigated by site-directed mutagenesis.

Three aromatic amino acids, Tyr92, Trp148 and Tyr187 belonging to three separate domains of the alpha 7-subunit of neuronal nicotinic acetylcholine receptor were mutated to phenylalanine, and the electrophysiological response of the resulting mutant receptors analyzed in the Xenopus oocyte expression system. All mutations significantly decreased the apparent affinities for acetylcholine and nicotine, and to a lesser extent, those for the competitive antagonists dihydro-beta-erythroidine and alpha-bungarotoxin. Other properties investigated, such as the voltage dependency of the ion response as well as its sensitivity to the open channel blocker QX222, were not significantly changed, indicating that the mutations affected selectively the recognition of cholinergic ligands by the receptor protein. The maximal rates for the rapid desensitization process were slightly modified, suggesting that the contribution of Tyr92, Trp148 and Tyr187 to the binding area might differ in the various conformations of the nicotinic receptor. Other mutations at nearby positions (S94N, W153F, G151D and G82E) did not affect the properties of the electrophysiological response. These data point to the functional significance of Tyr92, Trp148 and Tyr187 in the binding of cholinergic ligands and ion channel activation of the nicotinic receptor, thus supporting a multiple loop model [(1990) J. Biol. Chem. 265, 10430-10437] for the ligand binding area.

Acetylcholine↗
PubMed1991

Mutations in the channel domain alter desensitization of a neuronal nicotinic receptor.

A variety of ligand-gated ion channels undergo a fast activation process after the rapid application of agonist and also a slower transition towards desensitized or inactivated closed channel states when exposure to agonist is prolonged. Desensitization involves at least two distinct closed states in the acetylcholine receptor, each with an affinity for agonists higher than those of the resting or active conformations. Here we investigate how structural elements could be involved in the desensitization of the acetylcholine-gated ion channel from the chick brain alpha-bungarotoxin sensitive homo-oligomeric alpha 7 receptor, using site-directed mutagenesis and expression in Xenopus oocytes. Mutations of the highly conserved leucine 247 residue from the uncharged MII segment of alpha 7 suppress inhibition by the open-channel blocker QX-222, indicating that this residue, like others from MII, faces the lumen of the channel. But, unexpectedly, the same mutations decrease the rate of desensitization of the response, increase the apparent affinity for acetylcholine and abolish current rectification. Moreover, unlike wild-type alpha 7, which has channels with a single conductance level, the leucine-to-threonine mutant has an additional conducting state active at low acetylcholine concentrations. It is possible that mutation of Leu 247 renders conductive one of the high-affinity desensitized states of the receptor.

Acetylcholine↗
PubMed1991

Neuronal nicotinic acetylcholine receptors expressed in Xenopus oocytes: role of the alpha subunit in agonist sensitivity and desensitization.

Neuronal nicotinic acetylcholine receptors (nAChRs) were expressed in Xenopus laevis oocytes after nuclear injection of complementary deoxyribonucleic acid (cDNA) expression vectors. The two receptor subtypes alpha 4/n alpha 1 and alpha 3/n alpha 1 were readily distinguishable from one another by ACh sensitivity and desensitization. alpha 3/n alpha 1 receptors showed lower ACh sensitivity and stronger desensitization than alpha 4/n alpha 1 receptors. Furthermore, although the current/voltage relationship was very similar in both receptor subtypes, the voltage dependence of desensitization was found to be strikingly different. As the n alpha 1 subunit was unchanged, the alpha subunits must be responsible for these functional differences. Symmetric hybrid alpha cDNAs, alpha 4:alpha 3 and alpha 3:alpha 4, were constructed and functional receptors were obtained by co-injection with n alpha 1. These hybrid receptors displayed an ACh sensitivity that was mainly defined by the extracellular sequence of the alpha subunit. In contrast, no part of the alpha subunit was found fully to determine desensitization.

Acetylcholine↗
PubMed1991

Steroids inhibit nicotinic acetylcholine receptors.

Application of progesterone to Xenopus oocytes expressing a cloned neuronal nicotinic acetylcholine (nAChR) revealed two effects. The first effect was a fully reversible reduction of the current induced by acetylcholine (ACh), its onset being nearly instantaneous. The second effect, which developed in a few hours, was an irreversible suppression of ACh-evoked currents. The transient inhibition had an apparent Ki of 7 microM when tested with 50 nM ACh, but the percentage of inhibition was positively correlated to the ACh concentration. A reduction of ACh-induced currents which appeared immediately upon progesterone application was also observed with muscle nAChR expressed in oocytes and with nAChR on membrane patches isolated from ciliary ganglion neurons. Thus nAChRs are modulated by progesterone and steroids may play an important role in nicotinic cholinoception.

Animals↗
PubMed1990

Alpha 5, alpha 3, and non-alpha 3. Three clustered avian genes encoding neuronal nicotinic acetylcholine receptor-related subunits.

In vertebrates, neuronal nicotinic acetylcholine receptors (nAChRs) assemble in an unknown stoichiometry from two homologous subunits, an alpha and a non-alpha. How large is the repertoire of these subunits and how many subtypes of functionally different nAChRs can they constitute? We found in the avian genome a cluster of three closely linked genes spanning 28 kilobase pairs and encoding three proteins, n alpha 3, alpha 3, and alpha 5, that have the features expected of neuronal nAChR subunits. Gene n alpha 3 lies 5' of alpha 3 (whose role in cholinoception has already been established) and is transcribed from the same DNA strand, whereas alpha 5 lies 3' of alpha 3 and is transcribed from the opposite DNA strand. The structure of the n alpha 3 and alpha 5 genes consists of six exons with precisely conserved splice sites and is identical to the structure of the previously characterized avian neuronal receptor subunit genes alpha 2, alpha 3, alpha 4, and n alpha 1. alpha 3, n alpha 3, and alpha 5 transcripts are rare in the central nervous system, but alpha 3 and n alpha 3 are readily detectable in embryonic superior cervical and ciliary ganglia. In order to assay function, the gene encoding n alpha 3 and the cDNAs encoding alpha 3, alpha 4, alpha 5, and n alpha 1 were subcloned into an expression vector, and the constructs were injected into Xenopus oocyte nuclei, either singly or in pairwise combinations of one alpha and one non-alpha. One to five days later, ACh sensitivity of the injected oocytes was examined in voltage clamp. The n alpha 3 gene and n alpha 1 cDNA elicited assembly of nAChRs when coinjected with alpha 3 or alpha 4 cDNA and the electrophysiological properties of the four pairwise combinations were significantly different. alpha 5, however, did not direct the assembly of functional nAChRs when injected alone or in combination with n alpha 1 or n alpha 3.

Acetylcholine↗
PubMed1990

A neuronal nicotinic acetylcholine receptor subunit (alpha 7) is developmentally regulated and forms a homo-oligomeric channel blocked by alpha-BTX.

cDNA and genomic clones encoding alpha 7, a novel neuronal nicotinic acetylcholine receptor (nAChR) alpha subunit, were isolated and sequenced. The mature alpha 7 protein (479 residues) has moderate homology with all other alpha and non-alpha nAChR subunits and probably assumes the same transmembrane topology. alpha 7 transcripts transiently accumulate in the developing optic tectum between E5 and E16. They are present in both the deep and the superficial layers of E12 tectum. In Xenopus oocytes, the alpha 7 protein assembles into a homo-oligomeric channel responding to acetylcholine and nicotine. The alpha 7 channel desensitizes very rapidly, rectifies strongly above -20 mV, and is blocked by alpha-bungarotoxin. A bacterial fusion protein encompassing residues 124-239 of alpha 7 binds labeled alpha-bungarotoxin. We conclude that alpha-bungarotoxin binding proteins in the vertebrate nervous system can function as nAChRs.

Acetylcholine↗
PubMed1990

Activation and blocking of neuronal nicotinic acetylcholine receptor reconstituted in Xenopus oocytes.

Neuronal nicotinic acetylcholine receptor of the alpha 4/non-alpha (alpha 4/n alpha) type was reconstituted in Xenopus oocytes after nuclear injection of cDNA expression vectors. Functional neuronal receptor was only formed when the two subunits alpha 4 and n alpha were coinjected, neither alpha 4 nor n alpha alone being effective. Responses to bath application of acetylcholine (AcCho) have been measured in voltage clamp. AcCho doses as low as 10 nM induce currents of up to 50 nA. Dose-response studies indicate a Kd of about 0.77 x 10(-6) M and a Hill coefficient of 1.5, thus predicting more than one AcCho binding site per receptor molecule. The current-voltage relationship of AcCho-induced currents presents a strong inward rectification. Responses to AcCho were compared to those of three other agonists: L-nicotine, carbachol, and 1,1-dimethyl-4-phenylpiperazinium iodide (DMPP). Sensitivities to AcCho, nicotine, and DMPP are quite similar. Sensitivity to carbachol is much lower, but the currents are otherwise indistinguishable from those induced by AcCho. Five AcCho antagonists--neuronal bungarotoxin (kappa-bungarotoxin), tubocurarine (TC), hexamethonium bromide (Hex), decamethonium bromide (Dec), and mecamylamine (Mec)--have been tested. Neuronal bungarotoxin has no effect on the alpha 4/n alpha channel, whereas 2.5 microM TC reduces by half the current peak evoked by 1 microM AcCho. The block by TC is independent of membrane voltage. By contrast, the block of AcCho-induced currents by Hex or Dec is strongly voltage dependent, suggesting that these substances enter the channel. The block by Mec is detectable at concentrations as low as 100 nM when applied together with 1 microM AcCho and is voltage independent. Hex, Dec, and Mec are effective only when AcCho is present. While the effects of all other agents are fully reversible, the Mec block is persistent.

Acetylcholine↗
PubMed1990

Potassium current activated by intracellular sodium in quail trigeminal ganglion neurons.

Whole-cell voltage clamp and single-channel recordings were performed on cultured trigeminal ganglion neurons from quail embryos in order to study a sodium-activated potassium current (KNa). When KNa was activated by a step depolarization in voltage clamp, there was a proportionality between KNa and INa at all voltages between the threshold of INa and ENa. Single-channel recordings indicated that KNa could be activated already by 12 mM intracellular sodium and was almost fully activated at 50 mM sodium. 100 mM lithium, 100 mM choline, or 5 microM calcium did not activate KNa. The relationship between the probability for the channel to be open (Po) vs. the sodium concentration and the relationship of KNa open time-distributions vs. the sodium concentration suggest that two to three sodium ions bind cooperatively before KNa channels open. KNa channels were sensitive to depolarization; at 12 mM sodium, a 42-mV depolarization caused an e-fold increase in Po. Under physiological conditions, the conductance of the KNa channel was 50 pS. This conductance increased to 174 pS when the intra- and extracellular potassium concentrations were 75 and 150 mM, respectively.

Animals↗
PubMed1990

Child health care policy and delivery in France.

Medical and social protection of mothers, infants, and children began in France more than a century ago. A number of laws and regulations have improved the system, which is discussed in detail. The discussion includes an overview of health policy, service delivery, and the financing of care. Is the current French system of Maternal and Child Health responsible for the good health of today's children? This question is addressed through selected examples. Finally, failures and short-comings of the system are described, including the persistence of underserved groups, unequal access to care, and other problems. Solutions are feasible, and some are now being implemented.

Adolescent↗
PubMed1989

Transient expression of a Ca2+-activated Cl- current during development of quail sensory neurons.

The expression of a calcium-activated chloride current (ICl(Ca)) was studied during the development of the sensory neurons of quail trigeminal ganglia. This current is expressed in 20% of the neurons by the 5th day of embryonic development; it can be found in nearly all neurons by the 7th day and subsequently disappears in half of them. Similar results were obtained with dorsal root ganglion neurons. The disappearance of ICl(Ca) in part of the sensory neurons during development is not due to a selective death of the neurons possessing this current and our results suggest that it is mediated by an interaction of the sensory neurons with their target tissue.

Animals↗