Pharmacology of GABA(A) receptors of retinal dopaminergic neurons.
When the vertebrate retina is stimulated by light, a class of amacrine or interplexiform cells release dopamine, a modulator responsible for neural adaptation to light. In the intact retina, dopamine release can be pharmacologically manipulated with agonists and antagonists at GABA(A) receptors, and dopaminergic (DA) cells receive input from GABAergic amacrines. Because there are only 450 DA cells in each mouse retina and they cannot be distinguished in the living state from other cells on the basis of their morphology, we used transgenic technology to label DA cells with human placental alkaline phosphatase, an enzyme that resides on the outer surface of the cell membrane. We could therefore identify DA cells in vitro after dissociation of the retina and investigate their activity with whole cell voltage clamp. We describe here the pharmacological properties of the GABA(A) receptors of solitary DA cells. GABA application induces a large inward current carried by chloride ions. The receptors are of the GABA(A) type because the GABA-evoked current is blocked by bicuculline. Their affinity for GABA is very high with an EC(50) value of 7.4 microM. Co-application of benzodiazepine receptor ligands causes a strong increase in the peak current induced by GABA (maximal enhancement: CL-218872 220%; flunitrazepam 214%; zolpidem 348%) proving that DA cells express a type I benzodiazepine-receptor (BZ1). GABA-evoked currents are inhibited by Zn(2+) with an IC(50) of 58.9 +/- 8.9 microM. Furthermore, these receptors are strongly potentiated by the modulator alphaxalone with an EC(50) of 340 +/- 4 nM. The allosteric modulator loreclezole increases GABA receptor currents by 43% (1 microM) and by 107% (10 microM). Using outside-out patches, we measured in single-channel recordings a main conductance (29 pS) and two subconductance (20 and 9 pS) states. We have previously shown by single-cell RT-PCR and immunocytochemistry that DA cells express seven different GABA(A) receptor subunits (alpha1, alpha3, alpha4, beta1, beta3, gamma1, gamma2(S), and gamma2(L)) and by immunocytochemistry that all subunits are expressed in the intact retina. We show here that at least alpha1, beta3 and gamma2 subunits are assembled into functional receptors.
Feigenspan
A
A
Department of Neurobiology, Harvard Medical School, Boston, Massachusetts 02115, USA.
Gustincich
S
S
Raviola
E
E
D000469:Alkaline Phosphatase / Q000235:genetics; D000818:Animals; D001569:Benzodiazepines / Q000378:metabolism; D001640:Bicuculline / Q000494:pharmacology; D001665:Binding Sites; D004298:Dopamine / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D018756:GABA Antagonists / Q000494:pharmacology; D058787:GABA-A Receptor Antagonists; D006801:Humans; D051379:Mice; D008822:Mice, Transgenic / Q000235:genetics; D009474:Neurons / Q000378:metabolism*; D009994:Osmolar Concentration; D018408:Patch-Clamp Techniques; D010852:Picrotoxin / Q000031:analogs & derivatives* / Q000494:pharmacology; D011277:Pregnanediones / Q000494:pharmacology; D020033:Protein Isoforms / Q000378:metabolism; D011963:Receptors, GABA-A / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D012160:Retina / Q000166:cytology / Q000378:metabolism*; D054830:Sesterterpenes; D014230:Triazoles / Q000494:pharmacology; D015032:Zinc / Q000494:pharmacology; D005680:gamma-Aminobutyric Acid / Q000494:pharmacology
D000469:Alkaline Phosphatase; D001569:Benzodiazepines; D001640:Bicuculline; D004298:Dopamine; D018756:GABA Antagonists; D058787:GABA-A Receptor Antagonists; D010852:Picrotoxin; D011277:Pregnanediones; D020033:Protein Isoforms; D011963:Receptors, GABA-A; D054830:Sesterterpenes; D014230:Triazoles; D015032:Zinc; D005680:gamma-Aminobutyric Acid; C066440:loreclezole; C022961:picrotoxinin; C006477:alphaxalone