Dentate gyrus basket cell GABAA receptors are blocked by Zn2+ via changes of their desensitization kinetics: an in situ patch-clamp and single-cell PCR study.
Although GABA type A receptors (GABAARs) in principal cells have been studied in detail, there is only limited information about GABAARs in interneurons. We have used the patch-clamp technique in acute rat hippocampal slices in combination with single-cell PCR to determine kinetic, pharmacological, and structural properties of dentate gyrus basket cell GABAARs. Application of 1 mM GABA (100 msec) to nucleated patches via a piezo-driven fast application device resulted in a current with a fast rise and a marked biexponential decay (time constants 2.4 and 61.8 msec). This decay could be attributed to strong receptor desensitization. Dose-response curves for the peak and the slow component yielded EC50 values of 139 and 24 microM, respectively. Zn2+ caused a marked blocking effect on both the peak and the slow component via a noncompetitive mechanism (IC50 values of 8 and 16 microM). This led to an acceleration of the slow component as well as a prolongation of recovery from desensitization. Zn2+ sensitivity was suggested to depend on the absence of gamma-subunits in GABAARs. To test this hypothesis we performed single-cell reverse transcription PCR that revealed primarily the presence of alpha2-, beta2-, beta3-, gamma1-, and gamma2-subunit mRNAs. In addition, flunitrazepam increased the receptor affinity for its agonist, indicating the presence of functional benzodiazepine binding sites, i.e., gamma-subunits. Thus, additional factors seem to co-determine the Zn2+ sensitivity of native GABAARs. The modulatory effects of Zn2+ on GABAAR desensitization suggest direct influences on synaptic integration via changes in inhibition and shunting at GABAergic synapses.
Berger
T
T
Institute of Physiology, University of Freiburg, D-79104 Freiburg, Germany.
Schwarz
C
C
Kraushaar
U
U
Monyer
H
H
D000818:Animals; D018891:Dentate Gyrus / Q000737:chemistry* / Q000166:cytology*; D004305:Dose-Response Relationship, Drug; D005445:Flunitrazepam / Q000494:pharmacology; D018682:GABA Agents / Q000494:pharmacology; D015870:Gene Expression; D015640:Ion Channel Gating / Q000187:drug effects*; D007700:Kinetics; D018408:Patch-Clamp Techniques; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D017208:Rats, Wistar; D011963:Receptors, GABA-A / Q000235:genetics* / Q000378:metabolism; D015032:Zinc / Q000494:pharmacology*; D005680:gamma-Aminobutyric Acid / Q000494:pharmacology
D018682:GABA Agents; D012333:RNA, Messenger; D011963:Receptors, GABA-A; D005680:gamma-Aminobutyric Acid; D005445:Flunitrazepam; D015032:Zinc