Differential acetylcholine and GABA release from cultured chick retina cells.
In the present work we investigated the mechanisms controlling the release of acetylcholine (ACh) and of gamma-aminobutyric acid (GABA) from cultures of amacrine-like neurons, containing a subpopulation of cells which are simultaneously GABAergic and cholinergic. We found that 81.2 +/- 2.8% of the cells present in the culture were stained immunocytochemically with an antibody against choline acetyltransferase, and 38.5 +/- 4.8% of the cells were stained with an antibody against GABA. Most of the cells containing GABA (87.0 +/- 2.9%) were cholinergic. The release of acetylcholine and GABA was mostly Ca2+-dependent, although a significant release of [3H]GABA occurred by reversal of its transporter. Potassium evoked the Ca2+-dependent release of [3H]GABA and [3H]acetylcholine, with EC50 of 31.0 +/- 1.0 mm and 21.6 +/- 1.1 mm, respectively. The Ca2+-dependent release of [3H]acetylcholine was significantly inhibited by 1 micrometer tetrodotoxin and by low (30 nm) omega-conotoxin GVIA (omega-CgTx GVIA) concentrations, or by high (300 nm) nitrendipine (Nit) concentrations. On the contrary, the release of [14C]GABA was reduced by 30 nm nitrendipine, or by 500 nm omega-CgTx GVIA, but not by this toxin at 30 nm. The release of either transmitters was unaffected by 200 nm omega-Agatoxin IVA (omega-Aga IVA), a toxin that blocks P/Q-type voltage-sensitive Ca2+ channels (VSCC). The results show that Ca2+-influx through omega-CgTx GVIA-sensitive N-type VSCC and through Nit-sensitive L-type VSCC induce the release of ACh and GABA. However, the significant differences observed regarding the Ca2+ channels involved in the release of each neurotransmitter suggest that in amacrine-like neurons containing simultaneously GABA and acetylcholine the two neurotransmitters may be released in distinct regions of the cells, endowed with different populations of VSCC.
Center for Neuroscience of Coimbra, Department of Zoology, University of Coimbra, 3000 Coimbra, Portugal.
P F
PF
Santos
A L
AL
Carvalho
A P
AP
Carvalho
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CB
Duarte
D000109:Acetylcholine / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000502:physiology; D002121:Calcium Channel Blockers / Q000494:pharmacology; D015220:Calcium Channels / Q000187:drug effects / Q000378:metabolism; D002478:Cells, Cultured; D002642:Chick Embryo; D002799:Cholinergic Fibers / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D008564:Membrane Potentials / Q000502:physiology; D011189:Potassium Chloride / Q000494:pharmacology; D012160:Retina / Q000378:metabolism*; D014316:Tritium; D005680:gamma-Aminobutyric Acid / Q000378:metabolism*
D002121:Calcium Channel Blockers; D015220:Calcium Channels; D014316:Tritium; D005680:gamma-Aminobutyric Acid; D011189:Potassium Chloride; D000109:Acetylcholine; D002118:Calcium