| 1. |
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| 2. |
- Dunham, I, et al.
(författare)
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The DNA sequence of human chromosome 22
- 1999
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Ingår i: Nature. - : Springer Science and Business Media LLC. - 0028-0836 .- 1476-4687. ; 402:6761, s. 489-495
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Tidskriftsartikel (refereegranskat)
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| 3. |
- Birnir, Bryndis, et al.
(författare)
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Rapid desensitization of alpha 1 beta 1 GABA A receptors expressed in Sf9 cells under optimized conditions.
- 1995
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Ingår i: Journal of Membrane Biology. - 0022-2631 .- 1432-1424. ; 148:2, s. 193-202
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Tidskriftsartikel (refereegranskat)abstract
- alpha 1 and beta 1 subunits of human GABA A receptors were expressed in Sf9 cells using the Sf9-baculovirus system. Better expression was obtained by manipulating the system. Cell growth phase at the time of infection determined the practical range of virus titre, the period postinfection during which cells were useful for signal detection and the maximal current obtained. Cells in the early exponential phase were relatively insensitive to multiplicity of infection (MOI) whereas cells in the mid- to late-exponential phase were highly dependent on MOI and they responded with the largest Cl- current generated by GABA. Channels activated by GABA were chloride-selective. Half the maximum peak whole-cell current was obtained with 11 microM GABA. The time course of Cl- currents activated by saturating GABA concentrations in cells infected with alpha 1 beta 1-recombinant viruses was examined employing a rapid perfusion system which allowed whole-cell solution exchange in less than 1 msec. The current decay could be fitted by 3 to 4 exponentials for the first 8 sec. The initial fast current decrease had a time constant of about 23 msec. No voltage dependence of time constants was detected but the whole-cell IV relation showed outward rectification. Currents were depressed by bicuculline, penicillin and picrotoxin and potentiated by pentobarbitone.
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| 4. |
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| 5. |
- Tierney, M L, et al.
(författare)
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Effects of mutating leucine to threonine in the M2 segment of alpha1 and beta1 subunits of GABAA alpha1beta1 receptors.
- 1996
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Ingår i: Journal of Membrane Biology. - 0022-2631 .- 1432-1424. ; 154:1, s. 11-21
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Tidskriftsartikel (refereegranskat)abstract
- The conserved leucine residues at the 9' positions in the M2 segments of alpha1 (L264) and beta1 (L259) subunits of the human GABAA receptor were replaced with threonine. Normal or mutant alpha1 subunits were co-expressed with normal or mutant beta1 subunits in Sf9 cells using the baculovirus/Sf9 expression system. Cells in which one or both subunits were mutated had a higher "resting" chloride conductance than cells expressing wild-type alpha1beta1 receptors. This chloride conductance was blocked by 10 mM penicillin, a recognized blocker of GABAA channels, but not by bicuculline (100 microm) or picrotoxin (100 microm) which normally inhibit the chloride current activated by GABA: nor was it potentiated by pentobarbitone (100 microM). In cells expressing wild-type beta1 with mutated alpha1 subunits, an additional chloride current could be elicited by GABA but the rise time and decay were slower than for wild-type alpha1beta1 receptors. In cells expressing mutated beta1 subunits with wild-type or mutated alpha1 subunits (alphabeta(L9'T) and alpha(L9'T)beta(L9'T)), no response to GABA could be elicited: this was not due to an absence of GABAA receptors in the plasmalemma because the cells bound [3H]-muscimol. It was concluded that in GABAA channels containing the L9'T mutation in the beta1 subunit, GABA-binding does not cause opening of channels, and that the L9'T mutation in either or both subunits gives an open-channel state of the GABAA receptor in the absence of ligand.
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| 6. |
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| 7. |
- Birnir, Bryndis, et al.
(författare)
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A structural determinant of desensitization and allosteric regulation by pentobarbitone of the GABAA receptor.
- 1997
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Ingår i: Journal of Membrane Biology. - 0022-2631 .- 1432-1424. ; 155:2, s. 157-66
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Tidskriftsartikel (refereegranskat)abstract
- Functional properties of the alpha1beta1 GABAA receptor changes in a subunit-specific manner when a threonine residue in the M2 region at the 12' position was mutated to glutamine. The rate and extent of desensitization increased in all mutants but the rate of activation was faster in the beta1 mutants. A negligible plateau current and abolition of potentiation by pentobarbitone of the GABA-activated current depended on the Thr 12' Gln mutation being present in the beta1 subunit. The Hill coefficient of the peak current response to GABA was reduced to less than one also in a beta1 subunit-specific manner. It was concluded that the beta1 subunit dominated conformational changes activated by GABA.
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| 8. |
- Birnir, Bryndis, et al.
(författare)
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Nature of the 5' residue in the M2 domain affects function of the human alpha 1 beta 1 GABAA receptor.
- 1997
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Ingår i: Synapse. - 0887-4476 .- 1098-2396. ; 26:3, s. 324-7
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Tidskriftsartikel (refereegranskat)abstract
- The effects on the functional properties of the alpha 1 beta 1 GABAA receptor when the 5' (alpha 1 Val260; beta 1 Ile255) hydrophobic amino acids in the second transmembrane (M2) region were changed to threonine were examined. In response to a saturating concentration of GABA, the current evoked in mutant receptors showed a decreased rate of desensitization and at equilibrium was a greater fraction of the peak current than in wild-type receptors. The half-saturation concentration of the peak current response to GABA in mutant receptors was comparable to that in wild-type receptors, but the Hill coefficient was reduced to less than one. It was concluded that the 5' amino acids in the M2 region have a role in the conformational changes that occur within the alpha 1 beta 1 GABAA receptor in response to GABA.
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| 9. |
- Dalziel, J E, et al.
(författare)
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A threonine residue in the M2 region of the beta1 subunit is needed for expression of functional alpha1beta1 GABA(A) receptors.
- 1999
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Ingår i: European Journal of Pharmacology. - 0014-2999 .- 1879-0712. ; 370:3, s. 345-8
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Tidskriftsartikel (refereegranskat)abstract
- Although there is a high degree of homology in the M2 transmembrane segments of alpha1 and beta1 subunits, subunit-specific effects were observed in alpha1beta1 GABA(A) receptors expressed in Spodoptera frugipedra (Sf9) cells when the conserved 13' threonine residue in the M2 transmembrane region was mutated to alanine. When threonine 263 (13') was mutated to alanine in the beta1 subunit, high-affinity muscimol binding and the response to GABA were abolished. This did not occur when the threonine 263 (13') was mutated to alanine in the alpha1 subunit, but the rate of desensitisation increased and the effect of bicuculline, a competitive inhibitor, was reduced. The results show differential effects of subunits on receptor function and support a role for M2 in desensitisation.
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| 10. |
- Dalziel, J E, et al.
(författare)
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Mutant human alpha(1)beta(1)(T262Q) GABA(A) receptors are directly activated but not modulated by pentobarbital.
- 1999
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Ingår i: European Journal of Pharmacology. - 0014-2999 .- 1879-0712. ; 385:2-3, s. 283-6
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Tidskriftsartikel (refereegranskat)abstract
- Pentobarbital activates GABA(A) receptors and enhances GABA-activated currents. A threonine residue (262) in the second membrane spanning region at the 12' position in the beta(1) subunit, alpha(1)beta(1)(T12'Q), is necessary for the potentiating action of pentobarbital. We examined whether T12'Q-mutated receptors expressed in Spodoptera frugipedra (Sf 9) cells responded to direct activation by pentobarbital. In both mutant and wild type receptors, pentobarbital (100 microM to 1 mM) evoked a current response. The pentobarbital EC(50) values were similar; 119 and 158 microM for alpha(1)beta(1) and alpha(1)beta(1)(T12'Q) receptors, respectively. The results show it is possible to discriminate between agonistic and potentiating effects of pentobarbital, suggesting these actions involve separate mechanisms.
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