Display Settings:

Items per page
We are sorry, but NCBI web applications do not support your browser and may not function properly. More information

Results: 6

1.
Figure 2

Figure 2. Alanine scanning mutagenesis reveals mutations in β-10 strand that reduce GABA sensitivity. From: The pre-M1 segment of the ?1 subunit is a transduction element in the activation of the GABAA receptor.

Representative recordings of GABA responses from HEK293 cells transfected with WT (A) and α1(V212A)β2γ2s (B) channels. The bars and numbers above the recordings indicate the duration and concentration (in μm) of applied GABA. C, GABA concentration–response curves for WT, and α1(K220A)β2γ2s and α1(V212A)β2γ2s mutant channels (rightward shifted). Values represent the pooled means of n = 8–25 cells.

Angelo Keramidas, et al. J Physiol. 2006 August 15;575(Pt 1):11-22.
2.
Figure 5

Figure 5. Desensitization and deactivation in α1(K220A)β2γ2s GABAARs. From: The pre-M1 segment of the ?1 subunit is a transduction element in the activation of the GABAA receptor.

A, normalized currents elicited by a 500-ms application of a saturating concentration of GABA (30 mm) for WT and α1(K220A)β2γ2s GABAARs. Note that there is no discernable difference in the rate of macroscopic desensitization. B, normalized currents elicited by a brief pulse (1–2 ms) of a saturating concentration of GABA (30 mm) for WT and α1(K220A)β2γ2s GABAARs. As indicated by the plot, the α1(K220A)β2γ2s GABAARs deactivate more rapidly than WT GABAARs. The currents normalized in A and B were originally of comparable magnitude, being 1 nA and 1.4 nA in A and 2 nA and 2.5 nA in B.

Angelo Keramidas, et al. J Physiol. 2006 August 15;575(Pt 1):11-22.
3.
Figure 6

Figure 6. Simulated currents in response to 1-ms pulse of GABA. From: The pre-M1 segment of the ?1 subunit is a transduction element in the activation of the GABAA receptor.

Normalized simulated (grey, continuous line) and real (black, dashed line) for WT (A) and α1(K220A)β2γ2s (B) GABAARs in response to a 1- to 2-ms application of 30 mm GABA. The rate constants required to best fit the individual currents shown were as follows. WT: β, 1200 s−1; α, 250 s−1; k−1, 200 s−1; k+1, 1.5 × 106m−1 s−1; α1(K220A)β2γ2s: β, 800 s−1; α, 330 s−1; k−1, 300 s−1; k+1, 0.9 × 106m−1 s−1. To the right of the current traces are the corresponding reaction schemes with the mean rate constants for the reaction obtained from the simulations.

Angelo Keramidas, et al. J Physiol. 2006 August 15;575(Pt 1):11-22.
4.
Figure 4

Figure 4. Concentration responses and current activation. From: The pre-M1 segment of the ?1 subunit is a transduction element in the activation of the GABAA receptor.

A, normalized currents showing the rising phase of the current evoked by 20 mm GABA for WT (kobs, 1488 s−1), α1(V212A)β2γ2s (kobs, 757 s−1), α1(K220A)β2γ2s (kobs, 717 s−1), and α1(K220D)β2γ2s (kobs, 377 s−1) GABAARs. Each current is an average of n = 6–10 consecutive recordings (sweeps) from the same cell. B, concentration–response relationship for WT and α1(K220A)β2γ2s GABAARs obtained using ultra-fast agonist application techniques to minimize the effects of current desensitization. The mean data points were fitted to the Hill equation (eqn (1)). C, plot of activation rate constant (kobs), obtained by fitting eqn (3) to the rising phase of the current as a function of agonist concentration.

Angelo Keramidas, et al. J Physiol. 2006 August 15;575(Pt 1):11-22.
5.
Figure 1

Figure 1. The β-10 strand. From: The pre-M1 segment of the ?1 subunit is a transduction element in the activation of the GABAA receptor.

A, the amino acid sequences of the β-10 strands of the GABAAR α1 and β2 subunits aligned with homologous sequences of the AChBP and the α1 subunits of the GlyR and Torpedo nAChR. All of the GABAAR α1 subunit amino acid residues shown were individually mutated to alanine in this study. The nomenclature for the loops was adopted from the paper by Brejc et al. (2001). The two residues, α1V212 and α1K220, that are implicated in channel activation are shown in bold type and boxed. B, schematic model of the GABAAR α1 subunit showing the β-10 strand. For reference, a critical residue of the GABA binding site (F64) and the M2 helix are also indicated. The intracellular segment linking M3 and M4 has been omitted from the schematic.

Angelo Keramidas, et al. J Physiol. 2006 August 15;575(Pt 1):11-22.
6.
Figure 3

Figure 3. Mutations to residues in the β-10 strand that reduce P4S relative efficacy. From: The pre-M1 segment of the ?1 subunit is a transduction element in the activation of the GABAA receptor.

A, bar graph showing the relative efficacy (ɛ = Imax(P4S)/Imax(GABA)) of the partial agonist, P4S, compared to GABA, for alanine substitutions of residues in the β strand. Values represent the pooled means of n = 6–25 cells. B, bar graph of ɛ for WT (n = 5), α1(V212A)β2γ2s (n = 7), α1(K220A)β2γ2s (n = 7), and α1(K220D)β2γ2s (n = 8) channels using the ultra-fast agonist application system. Saturating concentrations of GABA (20 mm) and P4S (20 mm) were used for these experiments. Note the order of magnitude lower values of ɛ compared to the graph in A. **Significantly different from WT (P < 0.01) calculated using a one-way ANOVA with a Dunnett's post hoc test.

Angelo Keramidas, et al. J Physiol. 2006 August 15;575(Pt 1):11-22.

Display Settings:

Items per page

Supplemental Content

Recent activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...
Write to the Help Desk