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R.F. Nelson, A.M. Bender, V.P. Connaughton; Transporter–Like GABA Excitation of Horizontal and Bipolar Cells in Zebrafish Distal Retina . Invest. Ophthalmol. Vis. Sci. 2006;47(13):390.
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© ARVO (1962-2015); The Authors (2016-present)
To identify GABA signaling pathways for horizontal and bipolar cells in the outer plexiform layer of the zebrafish retina.
Retinas removed from adult zebrafish eyes were papain–dissociated and plated on poly–D–lysine–coated plastic culture dishes. Dissociated horizontal cells (HCs) and bipolar cells (BCs) were identified morphologically (Connaughton & Dowling, 1998). Changes in membrane potential induced by GABA, selective GABAergic ligands, or treatments affecting GABA responses were obtained by photometry using the fluorescent voltage–sensitive dye oxonol (DiBaC4(5), 80 nM; Nelson et al, 2003). Only the dendro–somatic GABA responses of axotomized BCs were considered. HC recordings were mainly of the HB type (Connaughton et al, 2004).
Most BCs (194/289, 67%) hyperpolarized in response to GABA (1–20 µM, but HCs almost never did (2/73, 3%). A subpopulation of HCs (27/73, 37%) either depolarized and/or responded with after hyperpolarization (dep/AHP) in response to GABA, as did a minor group of BCs (13/289; 4%). In GABA depolarized BCs or HCs the mean fluorescence increase was .24 log units (N=27), about 30 mV (Nelson et al, 1999). Muscimol (10 µM), an iontropic GABA agonist, readily hyperpolarized BCs (56/91, 62%), but was ineffective at evoking dep/AHP in either HCs (3/44, 7%) or BCs (2/91, 2%). GABA–induced hyperpolarizations in BCs were blocked or partially blocked by picrotoxin (24/24, 100%); however, the dep/AHP GABA response, wherever seen in dissociated retinal neurons, was not (0/16, 0%). Dep/AHP responses required both extracellular Na+ and Cl–. They were reversibly blocked by Li+ substitution for Na+ (15/16; 94%). In some cases (8/16, 50%) a hyperpolarizing GABA response was unmasked by this substitution. Na isethionate substituted for NaCl resulted in irreversible loss of GABA dep/AHP (13/14, 93%). Among 14 dep/AHP responses studied with GABA concentration series, the median concentration for half saturation of response was 0.7 µM.
Ionotropic GABA receptors are expressed on most zebrafish BCs, but not HCs. Many HCs and rarely, BCs, express an Na+ –dependent, Cl– –dependent, picrotoxin–insensitive, and muscimol–resistant membrane transporter for GABA. A depolarized Na+ gradient drives GABA into these cells, depolarizing the plasma membrane and triggering Na+, K+–dependent ATPase generating AHP. In effect many HCs and a few BCs in zebrafish exhibit a GABA–selective, Na+–dependent excitation within the outer plexiform layer.
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