Robust photoregulation of GABA(A) receptors by allosteric modulation with a propofol analogue.
Photochemical switches represent a powerful method for improving pharmacological therapies and controlling cellular physiology. Here we report the photoregulation of GABA(A) receptors (GABA(A)Rs) by a derivative of propofol (2,6-diisopropylphenol), a GABA(A)R allosteric modulator, which we have modified to contain photoisomerizable azobenzene. Using α(1)β(2)γ(2) GABA(A)Rs expressed in Xenopus laevis oocytes and native GABA(A)Rs of isolated retinal ganglion cells, we show that the trans-azobenzene isomer of the new compound (trans-MPC088), generated by visible light (wavelengths ~440 nm), potentiates the γ-aminobutyric acid-elicited response and, at higher concentrations, directly activates the receptors. cis-MPC088, generated from trans-MPC088 by ultraviolet light (~365 nm), produces little, if any, receptor potentiation/activation. In cerebellar slices, MPC088 co-applied with γ-aminobutyric acid affords bidirectional photomodulation of Purkinje cell membrane current and spike-firing rate. The findings demonstrate photocontrol of GABA(A)Rs by an allosteric ligand, and open new avenues for fundamental and clinically oriented research on GABA(A)Rs, a major class of neurotransmitter receptors in the central nervous system.
PubMed ID: 23033071
PMC ID: PMC4023869
Article link: Nat Commun.
Grant support: AA01973 NIAAA NIH HHS , EY001792 NEI NIH HHS , EY016094 NEI NIH HHS , UL1RR029879 NCRR NIH HHS , UL1 RR029879 NCRR NIH HHS , P30 EY001792 NEI NIH HHS , R01 EY016094 NEI NIH HHS , R21 AA019737 NIAAA NIH HHS
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|Figure 3. Effect of MPC088 on α1β3γ2 and α4β3δ GABAAR-expressing oocytesData in each group obtained from 4 oocytes. a: Aggregate concentration-response data (mean ± SD) obtained from α1β3γ2 (black) and α4β3δ (red) receptors with co-applied GABA at similar ~EC8 dose (i.e., 3 μM for the former and 0.05 μM for the latter receptor type) and varying concentrations of trans-dominant MPC088. Response amplitudes obtained from each oocyte normalized to its saturating GABA response (i.e., responses obtained with 1,000 μM GABA for α1β3γ2 and 100 μM GABA for α4β3δ GABAARs). b: Aggregate concentration-response data (mean ± SD) obtained from α1β3γ2 (black) and α4β3δ (red) receptors with trans-dominant MPC088 alone. Data similarly normalized to the saturating GABA response obtained from each oocyte. Results for α1β2γ2 GABAARs in a and b (blue) reproduced from those of Figures 1c and 2b, respectively.|
|Figure 5. Action of MPC088 on retinal ganglion cellsa: Potentiation data obtained with trans- and cis-dominant MPC088; the cis-dominant preparation was obtained by pre-treating trans-dominant MPC088 with UV light for 5 min. Representative responses obtained from a single cell treated with 200 μM GABA (black), 2 μM GABA (red), 2 μM GABA co-applied with 10 μM of trans-dominant (blue) or cis-dominant (magenta) MPC088. Inset: Aggregate data (mean ± SD) for potentiation factor determined with trans- and cis-dominant MPC088 (left and right bars, respectively) (n=6, p = 0.003). b: Direct activation data obtained with trans- and cis-dominant MPC088. Representative responses obtained from a cell treated with 200 μM GABA (black), 60 μM of trans-dominant (blue) and 60 μM of cis-dominant (magenta) MPC088. Inset: Aggregate data (mean ± SD) for direct activation by MPC088. Left, middle and right bars show results obtained, respectively, with 30 μM trans-dominant (n=3), 60 μM trans-dominant (n=7), and 60 μM cis-dominant MPC088 (n=4).|
|Figure 8. Lack of agonist activity of MPC088 on PNsa: Averages of 10 recordings from a PN exposed to multiple Blue/UV light flashes, in the presence of cis-dominant MPC088 (30 μM) in the external solution, before (black) and after (red) the addition of gabazine (30 μM). There was no exogenous GABA present in the tissue. b: Summary of results from 8 experiments (8 PNs) including that described in a. The data indicate the magnitude of an outward current. The temporal sequence of illumination differed among the experiments (e.g., only the cell in a received multiple blue/UV exposures), but the data plotted are the amplitudes from the blue to UV transition (in the case of the cell in a, this is the blue/UV transition from the first of the three exposures). Yellow-filled data points indicate results from the experiment in a. Each data point is the average obtained from 4-10 consecutive recordings. The light-evoked membrane current was significantly reduced by gabazine (Wilcoxon Signed Rank Test, p=0.008). Error bars represent the SEM.|