Composition of the GABA(A) receptors of retinal dopaminergic neurons.
Transgenic technology, single-cell RT-PCR, and immunocytochemistry were combined to investigate the composition of the GABA(A) receptors of dopaminergic (interplexiform) amacrine (DA) cells. A mouse line was used in which these neurons were labeled with human placental alkaline phosphatase and could therefore be identified in vitro after dissociation of the retina. We performed single-cell RT-PCR on the isolated cells and showed that (1) DA cells contained the messages for alpha1, alpha3, alpha4, beta1, beta3, gamma1, gamma2(S), and gamma2(L) subunits; (2) this transcript repertory did not change on dissociation of the retina and throughout the time required for cell harvesting; and (3) all DA cells contained the entire transcript repertory. Immunocytochemistry with subunit-specific antibodies showed that all subunits were expressed and appeared homogeneously distributed throughout the cell membrane at a low concentration. In addition, with the exception of alpha4, the subunits formed clusters at the surface of the dendrites and on the inner pole of the cell body. Because of their size, shape, and topographic coincidence with GABAergic endings, the clusters were interpreted as postsynaptic active zones containing GABA(A) receptors. The composition of the synaptic receptors was not uniform: clusters distributed throughout the dendritic tree contained alpha3, beta3, and, less frequently, beta1 subunits, whereas clusters containing the alpha1 subunit were confined to large dendrites. Therefore, DA cells possess at least two types of GABA(A) receptors localized in different synapses. Furthermore, they exhibit multiple extrasynaptic GABA(A) receptors.
Gustincich
S
S
Department of Neurobiology, Harvard Medical School, Boston, Massachusetts 02115, USA.
Feigenspan
A
A
Sieghart
W
W
Raviola
E
E
D000469:Alkaline Phosphatase / Q000032:analysis / Q000235:genetics; D000818:Animals; D004298:Dopamine / Q000032:analysis; D006801:Humans; D007150:Immunohistochemistry; D046911:Macromolecular Substances; D051379:Mice; D008822:Mice, Transgenic; D009474:Neurons / Q000166:cytology / Q000502:physiology*; D011963:Receptors, GABA-A / Q000032:analysis* / Q000737:chemistry / Q000235:genetics*; D015183:Restriction Mapping; D012160:Retina / Q000166:cytology / Q000502:physiology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013569:Synapses / Q000502:physiology / Q000648:ultrastructure; D014158:Transcription, Genetic; D014446:Tyrosine 3-Monooxygenase / Q000032:analysis / Q000235:genetics
D046911:Macromolecular Substances; D011963:Receptors, GABA-A; D014446:Tyrosine 3-Monooxygenase; D000469:Alkaline Phosphatase; D004298:Dopamine